Long non-coding RNA (lncRNA) such as ANRIL and UFC1 have been verified as oncogenic genes in non-small cell lung cancer (NSCLC). It is well known that the tumor suppressor microRNA-34a (miR-34a) is downregulated in NSCLC. Furthermore, miR-34a induces senescence and apoptosis in breast, glioma, cervical cancer including NSCLC by targeting Myc. Recent evidence suggests that these two lncRNAs act as a miR-34a sponge in corresponding cancers. However, the biological functions between these two non-coding RNAs (ncRNAs) have not yet been studied in NSCLC. Therefore, we present a Boolean model to analyze the gene regulation between these two ncRNAs in NSCLC. We compared our model to several experimental studies involving gain- or loss-of-function genes in NSCLC cells and achieved an excellent agreement. Additionally, we predict three positive circuits involving miR-34a/E2F1/ANRIL, miR-34a/E2F1/UFC1, and miR-34a/Myc/ANRIL. Our circuit- perturbation analysis shows that these circuits are important for regulating cell-fate decisions such as senescence and apoptosis. Thus, our Boolean network permits an explicit cell-fate mechanism associated with NSCLC. Therefore, our results support that ANRIL and/or UFC1 is an attractive target for drug development in tumor growth and aggressive proliferation of NSCLC, and that a valuable outcome can be achieved through the miRNA-34a/Myc pathway.

The objective was to evaluate the efficacy of a single dose of exogenous galectin-1 in improving the pregnancy rate in inseminated cows, comparing the pregnancy rate of the two groups (treatment and control Groups) into 107 contemporary groups (YG) established. An ultrasound exam determined the pregnancy rate performed 25 to 35 days after the fixed-time artificial insemination (FTAI) of breeding beef cows (n = 3469). The pregnancy rate of cows that received a single dose of eGAL-1 (200 ± 10 µg), with an intrauterine administration (n = 1901), was compared with the pregnancy rate of cows inseminated using a conventional AI protocol (n = 1568), both comparing into the same YG. YGs were created considering the grouping of cows belonging to the same farm, with the same nutritional score and management, inseminated by the same inseminator and semen batch, and using the same estrus synchronization protocol). The statistical method used calculated the probability of obtaining pregnancy within each group. The administration of a single dose of eGAL-1 can increase the probability of obtaining pregnancy in beef cows by up to 8.68% (p < 0.0001), suggesting that a single dose of eGAL-1 during the FTAI procedure was reasonable in the beef cattle AI routine and can improve the pregnancy rate considerably.

In this study, the computational analysis of electronic circular dichroism (ECD) spectra and the employment of biphenyl chiroptical probes were compared in the absolute configuration assignment of (-)-1α,2α,3β-trihydroxy-p-menthane (1), taken as a representative example of a UV-transparent chiral natural product. The usefulness of chiroptical probes in the configurational assignments of natural products and their complementarity to the computational protocols is herein highlighted. The biphenyl probe approach proves to be straightforward, reliable, and suitable for conformationally mobile and ECD silent compounds, not treatable by computational analysis of chiroptical data.

Tackling neurodegeneration and neuroinflammation is particularly challenging due to the complexity of central nervous system (CNS) disorders, as well as the limited drug accessibility to the brain. The activation of tropomyosin-related kinase A (TRKA) receptor signaling by the nerve growth factor (NGF) or the neurosteroid dehydroepiandrosterone (DHEA) may combat neurodegeneration and regulate microglial function. In the present study, we synthesized a C-17-spiro-cyclopropyl DHEA derivative (ENT-A010), which was capable of activating TRKA. ENT-A010 protected PC12 cells against serum starvation-induced cell death, dorsal root ganglia (DRG) neurons against NGF deprivation-induced apoptosis and hippocampal neurons against Aβ-induced apoptosis. In addition, ENT-A010 pretreatment partially restored homeostatic features of microglia in the hippocampus of lipopolysaccharide (LPS)-treated mice, enhanced Aβ phagocytosis, and increased Ngf expression in microglia in vitro. In conclusion, the small molecule ENT-A010 elicited neuroprotective effects and modulated microglial function, thereby emerging as an interesting compound, which merits further study in the treatment of CNS disorders.

The negative impact of chronic kidney disease (CKD) on health status and quality of life in older patients has been well documented. However, data on frailty trajectories and long-term outcomes of older CKD patients undergoing structured Comprehensive Geriatric Assessment (CGA) with multidimensional frailty evaluation are sparse. Here, we analysed records from 375 CKD patients admitted to our university hospital (mean age 77.5 (SD 6.1) years, 36% female) who had undergone a CGA-based calculation of the frailty score with the multidimensional prognostic index (MPI) as well as follow-up evaluations at 3, 6 and 12 months after discharge. Based on the MPI score at admission, 21% of the patients were frail and 56% were prefrail. MPI values were significantly associated with KDIGO CKD stages (p = 0.003) and rehospitalisation after 6 months (p = 0.027) and mortality at 3, 6 and 12 months (p = 0.001), independent of chronological age. Kidney transplant recipients (KTR) showed a significantly lower frailty compared to patients with renal replacement therapy (RRT, p = 0.028). The association between frailty and mortality after 12 months appeared particularly strong for KTR (mean MPI 0.43 KTR vs. 0.52 RRT, p < 0.001) and for patients with hypoalbuminemia (p < 0.001). Interestingly, RRT was per se not significantly associated with mortality during follow up. However, compared to patients on RRT those with KTR had a significantly lower grade of care (p = 0.031) and lower rehospitalisation rates at 12 months (p = 0.010). The present analysis shows that the large majority of older CKD inpatients are prefrail or frail and that the risk for CKD-related adverse outcomes on the long term can be accurately stratified by CGA-based instruments. Further studies are needed to explore the prognostic and frailty-related signature of laboratory biomarkers in CKD.

Pre-eclampsia is a severe hypertensive disorder of pregnancy (HDP), mainly characterized by new-onset hypertension with proteinuria after 20-week gestation. Sirtuin1 (SIRT1), a class III histone deacetylase, is associated with the regulation of various pathophysiological processes, including inflammation, immune response, metabolism, and autophagy. However, the effect of SIRT1 in the pathogenesis of pre-eclampsia remains to be elucidated. In this study, we found that the expression of SIRT1 was relatively lower in the placentas and serum samples of pre-eclampsia patients. Typical pre-eclampsia-like symptoms, such as hypertension, proteinuria, fetal growth restriction, kidney injury, and a narrow placental labyrinth layer, were observed in SIRT1 knockdown (SIRT1+/-) mice. Of note, these performances could be improved after the intraperitoneal injection of SIRT1 agonist SRT2104. More importantly, we found that the efficacy of progesterone on attenuating symptoms of PE was profoundly better than that of metformin in SIRT1+/- mice. In addition, our results suggested that progesterone can promote the invasion and inhibit the apoptosis of trophoblasts. These data suggest that SIRT1 plays an important role in pre-eclampsia and that progesterone alleviates pre-eclampsia-like symptoms mediated by SIRT1 deficiency.

The immunosuppressive effect of adenosine in the microenvironment of a tumor is well established. Presently, researchers are developing approaches in immune therapy that target inhibition of adenosine or its signaling such as CD39 or CD73 inhibiting antibodies or adenosine A2A receptor antagonists. However, numerous enzymatic pathways that control ATP-adenosine balance, as well as understudied intracellular adenosine regulation, can prevent successful immunotherapy. This review contains the latest data on two adenosine-lowering enzymes: adenosine kinase (ADK) and adenosine deaminase (ADA). ADK deletes adenosine by its phosphorylation into 5'-adenosine monophosphate. Recent studies have revealed an association between a long nuclear ADK isoform and an increase in global DNA methylation, which explains epigenetic receptor-independent role of adenosine. ADA regulates the level of adenosine by converting it to inosine. The changes in the activity of ADA are detected in patients with various cancer types. The article focuses on the biological significance of these enzymes and their roles in the development of cancer. Perspectives of future studies on these enzymes in therapy for cancer are discussed.

Maintenance of protein homeostasis is crucial for virtually every aspect of eukaryotic biology. The ubiquitin-proteasome system (UPS) represents a highly regulated quality control machinery that protects cells from a variety of stress conditions as well as toxic proteins. A large body of evidence has shown that UPS dysfunction contributes to the pathogenesis of cardiovascular diseases. This review highlights the latest findings regarding the physiological and pathological roles of cullin-RING ubiquitin ligases (CRLs), an essential player in the UPS, in the cardiovascular system. To inspire potential therapeutic invention, factors regulating CRL activities are also discussed.

Mammalian genomic DNA methylation represents a key epigenetic modification and its dynamic regulation that fine-tunes the gene expression of multiple pathways during development. It maintains the gene expression of one generation of cells; particularly, the mitotic inheritance of gene-expression patterns makes it the key governing mechanism of epigenetic change to the next generation of cells. Convincing evidence from recent discoveries suggests that the dynamic regulation of DNA methylation is accomplished by the enzymatic action of TET dioxygenase, which oxidizes the methyl group of cytosine and activates transcription. As a result of aberrant DNA modifications, genes are improperly activated or inhibited in the inappropriate cellular context, contributing to a plethora of inheritable diseases, including cancer. We outline recent advancements in understanding how DNA modifications contribute to tumor suppressor gene silencing or oncogenic-gene stimulation, as well as dysregulation of DNA methylation in cancer progression. In addition, we emphasize the function of PARP1 enzymatic activity or inhibition in the maintenance of DNA methylation dysregulation. In the context of cancer remediation, the impact of DNA methylation and PARP1 pharmacological inhibitors, and their relevance as a combination therapy are highlighted.

As the third generation gene editing technology, Crispr/Cas9 has a wide range of applications. The success of Crispr depends on the editing of the target gene via a functional complex of sgRNA and Cas9 proteins. Therefore, highly specific and high on-target cleavage efficiency sgRNA can make this process more accurate and efficient. Although there are already many sophisticated machine learning or deep learning models to predict the on-target cleavage efficiency of sgRNA, prediction accuracy remains to be improved. XGBoost is good at classification as the ensemble model could overcome the deficiency of a single classifier to classify, and we would like to improve the prediction efficiency for sgRNA on-target activity by introducing XGBoost into the model. We present a novel machine learning framework which combines a convolutional neural network (CNN) and XGBoost to predict sgRNA on-target knockout efficacy. Our framework, called CNN-XG, is mainly composed of two parts: a feature extractor CNN is used to automatically extract features from sequences and predictor XGBoost is applied to predict features extracted after convolution. Experiments on commonly used datasets show that CNN-XG performed significantly better than other existing frameworks in the predicted classification mode.

Preeclampsia is a maternal hypertensive disease, complicating 2-8% of all pregnancies. It has been linked to a 2-7-fold increased risk for the development of cardiovascular disease, including heart failure, later in life. A total of 40% of formerly preeclamptic women develop preclinical heart failure, which may further deteriorate into clinical heart failure. Noninvasive cardiac imaging could assist in the early detection of myocardial abnormalities, especially in the preclinical stage, when these changes are likely to be reversible. Moreover, imaging studies can improve our insights into the relationship between preeclampsia and heart failure and can be used for monitoring. Cardiac ultrasound is used to assess quantitative changes, including the left ventricular cavity volume and wall thickness, myocardial mass, systolic and diastolic function, and strain. Cardiac magnetic resonance imaging may be of additional diagnostic value to assess diffuse and focal fibrosis and perfusion. After preeclampsia, sustained elevated myocardial mass along with reduced myocardial circumferential and longitudinal strain and decreased diastolic function is reported. These findings are consistent with the early phases of heart failure, referred to as preclinical (asymptomatic) or B-stage heart failure. In this review, we will provide an up-to-date overview of the potential of cardiac magnetic resonance imaging and echocardiography in identifying formerly preeclamptic women who are at high risk for developing heart failure. The potential contribution to early cardiac screening of women with a history of preeclampsia and the pros and cons of these imaging modalities are outlined. Finally, recommendations for future research are presented.

Cardiovascular (CV) and renal diseases are increasingly prevalent in the United States and globally. CV-related mortality is the leading cause of death in the United States, while renal-related mortality is the 8th. Despite advanced therapeutics, both diseases persist, warranting continued exploration of disease mechanisms to develop novel therapeutics and advance clinical outcomes for cardio-renal health. CV and renal diseases increase with age, and there are sex differences evident in both the prevalence and progression of CV and renal disease. These age and sex differences seen in cardio-renal health implicate sex hormones as potentially important regulators to be studied. One such regulator is G protein-coupled estrogen receptor 1 (GPER1). GPER1 has been implicated in estrogen signaling and is expressed in a variety of tissues including the heart, vasculature, and kidney. GPER1 has been shown to be protective against CV and renal diseases in different experimental animal models. GPER1 actions involve multiple signaling pathways: interaction with aldosterone and endothelin-1 signaling, stimulation of the release of nitric oxide, and reduction in oxidative stress, inflammation, and immune infiltration. This review will discuss the current literature regarding GPER1 and cardio-renal health, particularly in the context of aging. Improving our understanding of GPER1-evoked mechanisms may reveal novel therapeutics aimed at improving cardio-renal health and clinical outcomes in the elderly.

Covalent binding between nitric oxide (NO) and a protein's free thiol group (SH) is termed protein S-nitrosylation. Protein S-nitrosylation is involved in cellular regulation mechanisms that underlie a wide range of critical functions, such as apoptosis, alteration of enzyme activities, and transcription-factor stability. Impaired protein S-nitrosylation is associated with a growing list of pathophysiological conditions, such as cardiovascular disease, multiple sclerosis, pulmonary hypertension, and sickle cell disease. The enzyme paraoxonase 1 (PON1) binds to high-density lipoprotein to provide many of its antiatherogenic properties. The enzyme has a strong antioxidant capacity, which protects fats, lipids, and lipoproteins from oxidation, in addition to breaking down oxidized fats. We investigated the effect of S-S transnitrosylation on PON1 activities. Incubation of recombinant PON1 (rePON1) with nitrosylated human serum albumin (HSA-NO) resulted in S-nitrosylation of about 70% of the rePON1, as measured by Q-TOF LC/MS. S-nitrosylation significantly increased rePON1 hydrolytic activities. It also increased rePON1's ability to inhibit low-density lipoprotein oxidation induced by Cu2+. Finally, it increased the enzyme's penetration into macrophage cells by 31%. Our findings suggest that S-nitrosylation of rePON1 improves its biological functions which may positively affect atherosclerosis disease progression.

A prodigious increment of scientific evidence in both preclinical and clinical studies is narrowing a major gap in knowledge regarding sex-specific biological responses observed in numerous branches of clinical practices. Some paradigmatic examples include neurodegenerative and mental disorders, immune-related disorders such as pathogenic infections and autoimmune diseases, oncologic conditions, and cardiovascular morbidities. The male-to-female proportion in a population is expressed as sex ratio and varies eminently with respect to the pathophysiology, natural history, incidence, prevalence, and mortality rates. The factors that determine this scenario incorporate both sex-associated biological differences and gender-dependent sociocultural issues. A broad narrative review focused on the current knowledge about the role of hormone regulation in gender medicine and gender peculiarities across key clinical areas is provided. Sex differences in immune response, cardiovascular diseases, neurological disorders, cancer, and COVID-19 are some of the hints reported. Moreover, gender implications in occupational health and health policy are offered to support the need for more personalized clinical medicine and public health approaches to achieve an ameliorated quality of life of patients and better outcomes in population health.

One of the most desirable properties that biomaterials designed for tissue engineering or drug delivery applications should fulfill is biodegradation and resorption without toxicity. Therefore, there is an increasing interest in the development of biomaterials able to be enzymatically degraded once implanted at the injury site or once delivered to the target organ. In this paper, we demonstrate the protease sensitivity of self-assembling amphiphilic peptides, in particular, RAD16-I (AcN-RADARADARADARADA-CONH2), which contains four potential cleavage sites for trypsin. We detected that when subjected to thermal denaturation, the peptide secondary structure suffers a transition from β-sheet to random coil. We also used Matrix-Assisted Laser Desorption/Ionization-Time-Of-Flight (MALDI-TOF) to detect the proteolytic breakdown products of samples subjected to incubation with trypsin as well as atomic force microscopy (AFM) to visualize the effect of the degradation on the nanofiber scaffold. Interestingly, thermally treated samples had a higher extent of degradation than non-denatured samples, suggesting that the transition from β-sheet to random coil leaves the cleavage sites accessible and susceptible to protease degradation. These results indicate that the self-assembling peptide can be reduced to short peptide sequences and, subsequently, degraded to single amino acids, constituting a group of naturally biodegradable materials optimal for their application in tissue engineering and regenerative medicine.

Acute lung injury is the most common type of organ damage with high incidence and mortality in sepsis, which is a poorly understood syndrome of disordered inflammation. The aims of this study are to explore whether heat shock protein 70 (HSP70), as a molecular chaperone, attenuates the septic lung injury, and to understand the underlying mechanisms. In our study, treatment with HSP70 ameliorated the survival rate, dysfunction of lung, inflammation, and apoptosis in cecal ligation and puncture (CLP)-treated mice as well as in LPS-treated human alveolar epithelial cells. Furthermore, HSP70 interacted with KANK2, leading to reversed cell viability and reduced apoptosis-inducing factor (AIF) and apoptosis. Additionally, knockdown of KANK2 in epithelial cells and deletion of hsp70.1 gene in CLP mice aggravated apoptosis and tissue damage, suggesting that interaction of KANK2 and HSP70 is critical for protecting lung injury induced by sepsis. HSP70 plays an important role in protection of acute lung injury caused by sepsis through interaction with KANK2 to reduce AIF release and apoptotic cell. HSP70 is a novel potential therapeutic approach for attenuation of septic lung injury.

Metals and metal-based compounds have many implications in biological systems. They are involved in cellular functions, employed in the formation of metal-based drugs and present as pollutants in aqueous systems, with toxic effects for living organisms. Amphiphilic molecules also play important roles in the above bio-related fields as models of membranes, nanocarriers for drug delivery and bioremediating agents. Despite the interest in complex systems involving both metal species and surfactant aggregates, there is still insufficient knowledge regarding the quantitative aspects at the basis of their binding interactions, which are crucial for extensive comprehension of their behavior in solution. Only a few papers have reported quantitative analyses of the thermodynamic, kinetic, speciation and binding features of metal-based compounds and amphiphilic aggregates, and no literature review has yet addressed the quantitative study of these complexes. Here, we summarize and critically discuss the recent contributions to the quantitative investigation of the interactions of metal-based systems with assemblies made of amphiphilic molecules by calorimetric, spectrophotometric and computational techniques, emphasizing the unique picture and parameters that such an analytical approach may provide, to support a deep understanding and beneficial use of these systems for several applications.

Ghrelin receptor, a growth hormone secretagogue receptor (GHS-R), is expressed in the pancreas. Emerging evidence indicates that GHS-R is involved in the regulation of glucose-stimulated insulin secretion (GSIS), but the mechanism by which GHS-R regulates GSIS in the pancreas is unclear. In this study, we investigated the role of GHS-R on GSIS in detail using global Ghsr-/- mice (in vivo) and Ghsr-ablated pancreatic islets (ex vivo). GSIS was attenuated in both Ghsr-/- mice and Ghsr-ablated islets, while the islet morphology was similar between WT and Ghsr-/- mice. To elucidate the mechanism underpinning Ghsr-mediated GSIS, we investigated the key steps of the GSIS signaling cascade. The gene expression of glucose transporter 2 (Glut2) and the glucose-metabolic intermediate-glucose-6-phosphate (G6P) were reduced in Ghsr-ablated islets, supporting decreased glucose uptake. There was no difference in mitochondrial DNA content in the islets of WT and Ghsr-/- mice, but the ATP/ADP ratio in Ghsr-/- islets was significantly lower than that of WT islets. Moreover, the expression of pancreatic and duodenal homeobox 1 (Pdx1), as well as insulin signaling genes of insulin receptor (IR) and insulin receptor substrates 1 and 2 (IRS1/IRS2), was downregulated in Ghsr-/- islets. Akt is the key mediator of the insulin signaling cascade. Concurrently, Akt phosphorylation was reduced in the pancreas of Ghsr-/- mice under both insulin-stimulated and homeostatic conditions. These findings demonstrate that GHS-R ablation affects key components of the insulin signaling pathway in the pancreas, suggesting the existence of a cross-talk between GHS-R and the insulin signaling pathway in pancreatic islets, and GHS-R likely regulates GSIS via the Akt-Pdx1-GLUT2 pathway.

Sulforaphane and other natural isothiocyanates released from the respective plant glucosinolates by the plant enzyme myrosinase (β-thioglucoside glucohydrolase) show extensive anticancer and antimicrobial effects. In this study, myrosinase from garden cress (Lepidium sativum) seeds was purified to electrophoretic homogeneity by a fast and easy strategy consisting of fractionation by isoelectric precipitation with ammonium sulphate (AS) and affinity chromatography using sulforaphane (SFN) attached to cellulose resin. The overall purification of enzyme with respect to crude extract was 169-fold and recovery of 37%. Under non-reducing conditions, two protein bands exhibiting myrosinase activity with masses of about 114 and 122 kDa, respectively, and a 58 kDa protein band with no activity were detected by SDS-PAGE and zymography on polyacrylamide gel. MALDI-Tof/Tof of tryptic fragments obtained from the respective protein bands detected sequence motifs homologous to the regions responsible for glycoside-substrate binding and similarities to members of the enzyme subfamilies β-glucosidases and myrosinases GH. The enzyme hydrolyzed both the natural (sinigrin, sinalbin, glucoraphanin) and the synthetic (p-nitrophenol-β-D-glucopyranoside (pNPG)) substrates. The highest catalytic activity of purified enzyme was achieved against sinigrin. The KM and Vmax values of the enzyme for sinigrin were found to be 0.57 mM, and 1.3 mM/s, respectively. The enzyme was strongly activated by 30 μM ascorbic acid. The optimum temperature and pH for enzyme was 50 °C and pH 6.0, respectively. The purified enzyme could be stored at 4 °C and slightly acidic pH for at least 45 days without a significant decrease in specific activity.

We have synthesized a series of 10 new, PSMA-targeted, near-infrared imaging agents intended for use in vivo for fluorescence-guided surgery (FGS). Compounds were synthesized from the commercially available amine-reactive active NHS ester of DyLight800. We altered the linker between the PSMA-targeting urea moiety and the fluorophore with a view to improve the pharmacokinetics. Chemical yields for the conjugates ranged from 51% to 86%. The Ki values ranged from 0.10 to 2.19 nM. Inclusion of an N-bromobenzyl substituent at the ε-amino group of lysine enhanced PSMA+ PIP tumor uptake, as did hydrophilic substituents within the linker. The presence of a polyethylene glycol chain within the linker markedly decreased renal uptake. In particular, DyLight800-10 demonstrated high specific uptake relative to background signal within kidney, confirmed by immunohistochemistry. These compounds may be useful for FGS in prostate, renal or other PSMA-expressing cancers.

Primary tumours, particularly from major solid organs, are able to disseminate into the blood and lymphatic system and spread to distant sites. These secondary metastases to other major organs are the most lethal aspect of cancer, accounting for the majority of cancer deaths. The brain is a frequent site of metastasis, and brain metastases are often fatal due to the critical role of the nervous system and the limited options for treatment, including surgery. This creates a need to further understand the complex cell and molecular biology associated with the establishment of brain metastasis, including the changes to the environment of the brain to enable the arrival and growth of tumour cells. Local changes in the vascular network, immune system and stromal components all have the potential to recruit and foster metastatic tumour cells. This review summarises our current understanding of brain vascular microenvironments, fluid circulation and drainage in the context of brain metastases, as well as commenting on current cutting-edge experimental approaches used to investigate changes in vascular environments and alterations in specialised subsets of blood and lymphatic vessel cells during cancer spread to the brain.

Congenital Disorders of Glycosylation (CDG) are multisystemic metabolic disorders showing highly heterogeneous clinical presentation, molecular etiology, and laboratory results. Here, we present different transferrin isoform patterns (obtained by isoelectric focusing) from three female patients harboring the ALG13 c.320A>G mutation. Contrary to other known variants of type I CDGs, where transferrin isoelectric focusing revealed notably increased asialo- and disialotransferrin fractions, a normal glycosylation pattern was observed in the probands. To verify this data and give novel insight into this variant, we modeled the human Alg13 protein and analyzed the dynamics of the apo structure and the complex with the UDP-GlcNAc substrate. We also modeled the Alg13-Alg14 heterodimer and ran multiple simulations of the complex in the presence of the substrate. Finally, we proposed a plausible complex formation mechanism.

The Epstein-Barr Virus (EBV) is a gammaherpesvirus involved in the etiopathogenesis of a variety of human cancers, mostly of lymphoid and epithelial origin. The EBV infection participates in both cell transformation and tumor progression, also playing an important role in subverting immune responses against cancers. The homeostasis of the immune system is tightly regulated by inhibitory mechanisms affecting key immune effectors, such as T lymphocytes and NK cells. Collectively known as immune checkpoints, these mechanisms rely on a set of cellular receptors and ligands. These molecules may be candidate targets for immune checkpoints blockade-an emergent and promising modality of immunotherapy already proven to be valuable for a variety of human cancers. The EBV was lately suspected to interfere with the expression of immune checkpoint molecules, notably PD-1 and its ligands, found to be overexpressed in cases of Hodgkin lymphoma, nasopharyngeal, and gastric adenocarcinomas associated with the viral infection. Even though there is compelling evidence showing that the EBV interferes with other immune checkpoint regulators (e.g., CTLA-4, LAG-3, TIM-3, and VISTA), the published data are still scarce. Herein, we discuss the current state of the knowledge on how the EBV interferes with the activity of immune checkpoints regulators, as well as its implications considering the immune checkpoints blockade for clinical management of the EBV-associated malignancies, notably lymphomas.

Cells have adapted to mechanical forces early in evolution and have developed multiple mechanisms ensuring sensing of, and adaptation to, the diversity of forces operating outside and within organisms. The nucleus must necessarily adapt to all types of mechanical signals, as its functions are essential for virtually all cell processes, many of which are tuned by mechanical cues. To sense forces, the nucleus is physically connected with the cytoskeleton, which senses and transmits forces generated outside and inside the cell. The nuclear LINC complex bridges the cytoskeleton and the nuclear lamina to transmit mechanical information up to the chromatin. This system creates a force-sensing macromolecular complex that, however, is not sufficient to regulate all nuclear mechanoadaptation processes. Within the nucleus, additional mechanosensitive structures, including the nuclear envelope and the nuclear pore complex, function to regulate nuclear mechanoadaptation. Similarly, extra nuclear mechanosensitive systems based on plasma membrane dynamics, mechanotransduce information to the nucleus. Thus, the nucleus has the intrinsic structural components needed to receive and interpret mechanical inputs, but also rely on extra nuclear mechano-sensors that activate nuclear regulators in response to force. Thus, a network of mechanosensitive cell structures ensures that the nucleus has a tunable response to mechanical cues.

Podocytopathies are kidney diseases that are driven by podocyte injury with proteinuria and proteinuria-related symptoms as the main clinical presentations. Albeit podocytopathies are the major contributors to end-stage kidney disease, the underlying molecular mechanisms of podocyte injury remain to be elucidated. Mitochondrial oxidative stress is associated with kidney diseases, and increasing evidence suggests that oxidative stress plays a vital role in the pathogenesis of podocytopathies. Accumulating evidence has placed mitochondrial oxidative stress in the focus of cell death research. Excessive generated reactive oxygen species over antioxidant defense under pathological conditions lead to oxidative damage to cellular components and regulate cell death in the podocyte. Conversely, exogenous antioxidants can protect podocyte from cell death. This review provides an overview of the role of mitochondrial oxidative stress in podocytopathies and discusses its role in the cell death of the podocyte, aiming to identify the novel targets to improve the treatment of patients with podocytopathies.

Rapeseed (Brassica napus L.) is mainly used for oil production and industrial purposes. A high photosynthetic efficiency is the premise of a high yield capable of meeting people's various demands. Chlorophyll-deficient mutants are ideal materials for studying chlorophyll biosynthesis and photosynthesis. In a previous study, we obtained the mutant yl1 for leaf yellowing throughout the growth period by ethyl methanesulfonate mutagenesis of B. napus. A genetic analysis showed that the yl1 chlorophyll-deficient phenotype was controlled by one incompletely dominant gene, which was mapped on chromosome A03 by a quantitative trait loci sequencing analysis and designated as BnA03.Chd in this study. We constructed an F2 population containing 5256 individuals to clone BnA03.Chd. Finally, BnA03.Chd was fine-mapped to a 304.7 kb interval of the B. napus 'ZS11' genome containing 58 annotated genes. Functional annotation, transcriptome, and sequence variation analyses confirmed that BnaA03g0054400ZS, a homolog of AT5G13630, was the most likely candidate gene. BnaA03g0054400ZS encodes the H subunit of Mg-chelatase. A sequence analysis revealed a single-nucleotide polymorphism (SNP), causing an amino-acid substitution from glutamic acid to lysine (Glu1349Lys). In addition, the molecular marker BnaYL1 was developed based on the SNP of BnA03.Chd, which perfectly cosegregated with the chlorophyll-deficient phenotype in two different F2 populations. Our results provide insight into the molecular mechanism underlying chlorophyll synthesis in B. napus.

Passive immunotherapy is a very promising approach for the treatment of Alzheimer's disease (AD). Among the different antibodies under development, those targeting post-translationally modified Aβ peptides might combine efficient reduction in beta-amyloid accompanied by lower sequestration in peripheral compartments and thus anticipated and reduced treatment-related side effects. In that regard, we recently demonstrated that the antibody-mediated targeting of isoD7-modified Aβ peptides leads to the attenuation of AD-like amyloid pathology in 5xFAD mice. In order to assess novel strategies to enhance the efficacy of passive vaccination approaches, we investigated the role of CD33 for Aβ phagocytosis in transgenic mice treated with an isoD7-Aβ antibody. We crossbred 5xFAD transgenic mice with CD33 knock out (CD33KO) mice and compared the amyloid pathology in the different genotypes of the crossbreds. The knockout of CD33 in 5xFAD mice leads to a significant reduction in Aβ plaques and concomitant rescue of behavioral deficits. Passive immunotherapy of 5xFAD/CD33KO showed a significant increase in plaque-surrounding microglia compared to 5xFAD treated with the antibody. Additionally, we observed a stronger lowering of Aβ plaque load after passive immunotherapy in 5xFAD/CD33KO mice. The data suggest an additive effect of passive immunotherapy and CD33KO in terms of lowering Aβ pathology. Hence, a combination of CD33 antagonists and monoclonal antibodies might represent a strategy to enhance efficacy of passive immunotherapy in AD.

The 5-year survival rate for oral squamous cell carcinoma (OSCC), one of the most common head and neck cancers, has not improved in the last 20 years. Poor prognosis of OSCC is the result of failure in early and precise diagnosis. Metabolic reprogramming, including the alteration of the uptake and utilisation of glucose, amino acids and lipids, is an important feature of OSCC and can be used to identify its biomarkers for early and precise diagnosis. In this review, we summarise how recent findings of rewired metabolic networks in OSCC have facilitated early and precise diagnosis of OSCC.

β-cells are insulin-producing cells in the pancreas that maintain euglycemic conditions. Pancreatic β-cell maturity and function are regulated by a variety of transcription factors that enable the adequate expression of the cellular machinery involved in nutrient sensing and commensurate insulin secretion. One of the key factors in this regulation is MAF bZIP transcription factor A (MafA). MafA expression is decreased in type 2 diabetes, contributing to β-cell dysfunction and disease progression. The molecular biology underlying MafA is complex, with numerous transcriptional and post-translational regulatory nodes. Understanding these complexities may uncover potential therapeutic targets to ameliorate β-cell dysfunction. This article will summarize the role of MafA in normal β-cell function and disease, with a special focus on known transcriptional and post-translational regulators of MafA expression.

Intercellular communication between monocytes/macrophages and cells involved in tissue regeneration, such as mesenchymal stromal cells (MSCs) and primary tissue cells, is essential for tissue regeneration and recovery of homeostasis. Typically, in the final phase of the inflammation-resolving process, this intercellular communication drives an anti-inflammatory immunomodulatory response. To obtain a safe and effective treatment to counteract the cytokine storm associated with a disproportionate immune response to severe infections, including that associated with COVID-19, by means of naturally balanced immunomodulation, our group has standardized the production under GMP-like conditions of a secretome by coculture of macrophages and MSCs. To characterize this proteome, we determined the expression of molecules related to cellular immune response and tissue regeneration, as well as its possible toxicity and anti-inflammatory potency. The results show a specific molecular pattern of interaction between the two cell types studied, with an anti-inflammatory and regenerative profile. In addition, the secretome is not toxic by itself on human PBMC or on THP-1 monocytes and prevents lipopolysaccharide (LPS)-induced growth effects on those cell types. Finally, PRS CK STORM prevents LPS-induced TNF-A and IL-1Β secretion from PBMC and from THP-1 cells at the same level as hydrocortisone, demonstrating its anti-inflammatory potency.

We hypothesized that the vitamin A (VA) status regulates type 2 diabetes (T2D) development in Zucker diabetic fatty (ZDF) rats. Zucker Lean and ZDF rats at weaning were fed a VA deficient with basal fat (VAD-BF, no VA and 22.1% fat energy), VA marginal with BF (VAM-BF, 0.35 mg retinyl palmitate (RP)/kg), VA sufficient with BF (VAS-BF, 4.0 mg RP/kg), VAD with high fat (VAD-HF, 60% fat energy), VAM-HF or VAS-HF diet for 8 weeks, including an oral glucose tolerance test (OGTT) at week 7.5. The hepatic mRNA and proteins levels were determined using real-time PCR and Western blot, respectively. The VAD-BF/HF and VAM-BF/HF diets prevented peripheral hyperglycemia and attenuated obesity in ZDF rats, which occurred in the presence of the VAS-BF/HF diets. This lowered VA status reduced venous blood hyperglycemia, hyperinsulinemia and hyperlipidemia, and improved OGTT and homeostasis model assessment for insulin resistance results in ZDF rats. The expression levels of key hepatic genes for glucose and fat metabolism were regulated by VA status and dietary fat contents. An interaction between VA and HF condition was also observed. We conclude that the reduction in the dietary VA status in both BF and HF conditions prevents T2D and obesity in ZDF rats.

Epidemiological evidence indicates that stress and aversive psychological conditions can affect cancer progression, while well-being protects against it. Although a large set of studies have addressed the impact of stress on cancer, not much is known about the mechanisms that protect from cancer in healthy psychological conditions. C57BL/6J mouse pups were exposed to an environmental enrichment condition consisting of being raised until weaning by the biological lactating mother plus a non-lactating virgin female (LnL = Lactating and non-Lactating mothers). The Control group consisted of mice raised by a single lactating mother (L = Lactating). Four months after weaning, mice from LnL and L conditions were exposed to intramuscular injection of 3-methylcolantrene (3MCA), a potent tumorigenic drug, and onset and progression of 3MCA-induced fibrosarcomas were monitored over time. Pups from the LnL compared to the L group received more parental care and were more resilient to stressful events during the first week of life. In association, the onset of tumors in LnL adults was significantly delayed. At the molecular level, we observed increased levels of wild-type p53 protein in tumor samples of LnL compared to L adults and higher levels of its target p21 in healthy muscles of LnL mice compared to the L group, supporting the hypothesis of potential involvement of p53 in tumor development. Our study sustains the model that early life care protects against tumor susceptibility.

While Polo-like kinase 1 (PLK1) inhibitors have shown promise in clinical settings for treating triple-negative breast cancer tumors and other solid tumors, they are limited by their ability to bind non-selectively to the ATP kinase domain. Therefore, we sought to develop a PLK1 allosteric inhibitor targeting the PLK1 T-loop (a switch responsible for activation) and evaluate its effects in triple-negative breast cancer cells. A novel compound, RK-10, was developed based on an in silico model, and its effects on specificity, viability, migration, and cell cycle regulation in MCF-10A and MDA-MB 231 cells were evaluated. When MDA-MB 231 cells were treated with 0-50 µg/mL RK-10, phospho-PLK1 (Thr-210) was decreased in cells cultured adherently and cells cultured as mammospheres. RK-10 significantly inhibited viability after 24 h; however, by 48 h, 25-50 µM RK-10 caused >50% reduction. RK-10 attenuated wound healing by up to 99.7% and caused S and G2/M cell cycle arrest, which was associated with increased p21 expression. We developed a novel allosteric inhibitor which mediates anti-proliferative and anti-migratory properties through targeting phospho-PLK1 (Thr-210) in mammospheres and causing S phase and G2/M cell cycle arrest. Further development of PLK1 allosteric inhibitors may be a promising approach for TNBC treatment.

Cochlear implantation initiates an inflammatory cascade in which both acute insertion trauma and chronic foreign body reaction lead to intracochlear fibrosis and loss of residual hearing. Several strategies have been proposed to attenuate the local reactive process after implantation, including intracochlear drug delivery. The present study gives an overview of what is being investigated in the field of inner ear therapeutics and cochlear implant surgery. The aim is to evaluate its potential benefit in clinical practice. A systematic search was conducted in PubMed, Embase, and Cochrane Library databases identifying comparative prospective studies examining the effect of direct inner ear drug application on mechanical cochlear trauma. Both animal and human studies were considered and all studies were assessed for quality according to the validated risk of bias tools. Intracochlear administration of drugs is a feasible method to reduce the local inflammatory reaction following cochlear implantation. In animal studies, corticosteroid use had a significant effect on outcome measures including auditory brainstem response, impedance, and histological changes. This effect was, however, only durable with prolonged drug delivery. Significant differences in outcome were predominantly seen in studies where the cochlear damage was extensive. Six additional reports assessing non-steroidal agents were found. Overall, evidence of anti-inflammatory effects in humans is still scarce.

Nutrient supply dictates cell signaling changes, which in turn regulate membrane protein trafficking. To better exploit nutrients, cells relocalize membrane transporters via selective protein trafficking. Key in this reshuffling are the α-arrestins, selective protein trafficking adaptors conserved from yeast to man. α-Arrestins bind membrane proteins, controlling the ubiquitination and endocytosis of many transporters. To prevent the spurious removal of membrane proteins, α-arrestin-mediated endocytosis is kept in check through phospho-inhibition. This phospho-regulation is complex, with up to 87 phospho-sites on a single α-arrestin and many kinases/phosphatases targeting α-arrestins. To better define the signaling pathways controlling paralogous α-arrestins, Aly1 and Aly2, we screened the kinase and phosphatase deletion (KinDel) library, which is an array of all non-essential kinase and phosphatase yeast deletion strains, for modifiers of Aly-mediated phenotypes. We identified many Aly regulators, but focused our studies on the TORC1 kinase, a master regulator of nutrient signaling across eukaryotes. We found that TORC1 and its signaling effectors, the Sit4 protein phosphatase and Npr1 kinase, regulate the phosphorylation and stability of Alys. When Sit4 is lost, Alys are hyperphosphorylated and destabilized in an Npr1-dependent manner. These findings add new dimensions to our understanding of TORC1 regulation of α-arrestins and have important ramifications for cellular metabolism.

Brain aquaporins facilitate the movement of water between the four water compartments: blood, cerebrospinal fluid, interstitial fluid, and intracellular fluid. This work analyzes the expression of the four most abundant aquaporins (AQPs) (AQP1, AQP4, AQP9, and AQP11) in the brains of mice and discuss their contribution to hydrocephalus. We analyzed available data from single-cell RNA sequencing of the central nervous system of mice to describe the expression of aquaporins and compare their distribution with that based on qPCR, western blot, and immunohistochemistry assays. Expression of AQP1 in the apical cell membrane of choroid plexus epithelial cells and of AQP4 in ependymal cells, glia limitans, and astrocyte processes in the pericapillary end foot is consistent with the involvement of both proteins in cerebrospinal fluid homeostasis. The expression of both aquaporins compensates for experimentally induced hydrocephalus in the animals. Recent data demonstrate that hypoxia in aged animals alters AQP4 expression in the choroidal plexus and cortex, increasing the ventricle size and intraventricular pressure. Cerebral distensibility is reduced in parallel with a reduction in cerebrospinal fluid drainage and cognitive deterioration. We propose that aged mice chronically exposed to hypoxia represent an excellent experimental model for studying the pathophysiological characteristics of idiopathic normal pressure hydrocephalus and roles for AQPs in such disease.

The development of antimicrobial agents against multidrug-resistant bacteria is an important medical challenge. Antimicrobial peptides (AMPs), human cathelicidin LL-37 and its derivative OP-145, possess a potent antimicrobial activity and were under consideration for clinical trials. In order to overcome some of the challenges to their therapeutic potential, a very promising AMP, SAAP-148 was designed. Here, we studied the mode of action of highly cationic SAAP-148 in comparison with OP-145 on membranes of Enterococcus hirae at both cellular and molecular levels using model membranes composed of major constituents of enterococcal membranes, that is, anionic phosphatidylglycerol (PG) and cardiolipin (CL). In all assays used, SAAP-148 was consistently more efficient than OP-145, but both peptides displayed pronounced time and concentration dependences in killing bacteria and performing at the membrane. At cellular level, Nile Red-staining of enterococcal membranes showed abnormalities and cell shrinkage, which is also reflected in depolarization and permeabilization of E. hirae membranes. At the molecular level, both peptides abolished the thermotropic phase transition and induced disruption of PG/CL. Interestingly, the membrane was disrupted before the peptides neutralized the negative surface charge of PG/CL. Our results demonstrate that SAAP-148, which kills bacteria at a significantly lower concentration than OP-145, shows stronger effects on membranes at the cellular and molecular levels.

Calprotectin, a heterodimer of S100A8 and S100A9 EF-hand calcium-binding proteins, is an integral part of the innate immune response. Calprotectin (CP) serves as a ligand for several pattern recognition cell surface receptors including the receptor for advanced glycation end products (RAGE), toll-like receptor 4 (TLR4), and cluster of differentiation 33 (CD33). The receptors initiate kinase signaling cascades that activate inflammation through the NF-kB pathway. Receptor activation by CP leads to upregulation of both receptor and ligand, a positive feedback loop associated with specific chronic inflammatory syndromes. Hence, CP and its two constituent homodimers have been viewed as potential targets to suppress certain chronic inflammation pathologies. A variety of inhibitors of CP and other S100 proteins have been investigated for more than 30 years, but no candidates have advanced significantly into clinical trials. Here, current knowledge of the interactions of CP with its receptors is reviewed along with recent progress towards the development of CP-directed chemotherapeutics.

Wasps, members of the order Hymenoptera, use their venom for predation and defense. Accordingly, their venoms contain various constituents acting on the circulatory, immune and nervous systems. Wasp venom possesses many allergens, enzymes, bioactive peptides, amino acids, biogenic amines, and volatile matters. In particular, some peptides show potent antimicrobial, anti-inflammatory, antitumor, and anticoagulant activity. Additionally, proteinous components from wasp venoms can cause tissue damage or allergic reactions in organisms. These bioactive peptides and proteins involved in wasp predation and defense may be potential sources of lead pharmaceutically active molecules. In this review, we focus on the advances in bioactive peptides and protein from the venom of wasps and their biological effects, as well as the allergic reactions and immunotherapy induced by the wasp venom.

Signs of impaired thiamine (vitamin B1) status in feeding-migrating Atlantic salmon (Salmo salar) were studied in three Baltic Sea areas, which differ in the proportion and nutritional composition of prey fish sprat (Sprattus sprattus) and herring (Clupea harengus). The concentration of n-3 polyunsaturated fatty acids (n-3 PUFAs) increased in salmon with dietary lipids and n-3 PUFAs, and the hepatic peroxidation product malondialdehyde (MDA) concentration increased exponentially with increasing n-3 PUFA and docosahexaenoic acid (DHA, 22:6n-3) concentration, whereas hepatic total thiamine concentration, a sensitive indicator of thiamine status, decreased with the increase in both body lipid and n-3 PUFA or DHA concentration. The hepatic glucose 6-phosphate dehydrogenase activity was suppressed by high dietary lipids. In salmon muscle and in prey fish, the proportion of thiamine pyrophosphate increased, and that of free thiamine decreased, with increasing body lipid content or PUFAs, or merely DHA. The thiamine status of salmon was impaired mainly due to the peroxidation of n-3 PUFAs, whereas lipids as a source of metabolic energy had less effect. Organochlorines or general oxidative stress did not affect the thiamine status. The amount of lipids, and, specifically, their long-chain n-3 PUFAs, are thus responsible for generating thiamine deficiency, and not a prey fish species per se.

Cinnamomum cassia is a natural product found in plants that has been used as a folk remedy for inflammation. In this study, we investigated the mechanism underlying the anti-inflammatory and antioxidant properties of C. cassia extract (ECC) in lipopolysaccharide (LPS)-induced murine RAW 264.7 cells, in comparison with 4-hydroxycinnamaldehyde, a C. cassia extract component. ECC and 4-hydroxycinnamaldehyde inhibited the production of nitrite oxide in a dose-dependent manner and did not show any change in cellular toxicity when treated with the same dose as that used in the nitrite assay. Moreover, they attenuated ROS accumulation after lipopolysaccharide (LPS) stimulation. ECC and 4-hydroxycinnamaldehyde decreased the mRNA and protein expression levels of inflammatory mediators (iNOS and COX-2) and cytokines such as TNF and IL-6. We also found that ECC and 4-hydroxycinnamaldehyde mitigated the phosphorylation of ERK, JNK, and transcription factors, such as NF-κB and STAT3, suppressing NF-κB nuclear translocation in LPS-activated macrophages. In addition, administration of ECC in a Sprague Dawley rat model of acute gastric injury caused by indomethacin significantly increased the gastric mucus volume. Analysis of serum and tissue levels of inflammatory mediators revealed a significant decrease in serum PGE2 and myeloperoxidase levels and a reduction in gastric iNOS, COX-2, and p65 protein levels. Collectively, these results suggest that ECC has antioxidant and anti-inflammatory effects and is a potential candidate for curing gastritis.

Finding, exploring and filtering frequent sentence-based associations between a disease and a biomedical entity, co-mentioned in disease-related PubMed literature, is a challenge, as the volume of publications increases. Darling is a web application, which utilizes Name Entity Recognition to identify human-related biomedical terms in PubMed articles, mentioned in OMIM, DisGeNET and Human Phenotype Ontology (HPO) disease records, and generates an interactive biomedical entity association network. Nodes in this network represent genes, proteins, chemicals, functions, tissues, diseases, environments and phenotypes. Users can search by identifiers, terms/entities or free text and explore the relevant abstracts in an annotated format.

In archaeal microorganisms, the compaction and organization of the chromosome into a dynamic but condensed structure is mediated by diverse chromatin-organizing proteins in a lineage-specific manner. While many archaea employ eukaryotic-type histones for nucleoid organization, this is not the case for the crenarchaeal model species Sulfolobus acidocaldarius and related species in Sulfolobales, in which the organization appears to be mostly reliant on the action of small basic DNA-binding proteins. There is still a lack of a full understanding of the involved proteins and their functioning. Here, a combination of in vitro and in vivo methodologies is used to study the DNA-binding properties of Sul12a, an uncharacterized small basic protein conserved in several Sulfolobales species displaying a winged helix-turn-helix structural motif and annotated as a transcription factor. Genome-wide chromatin immunoprecipitation and target-specific electrophoretic mobility shift assays demonstrate that Sul12a of S. acidocaldarius interacts with DNA in a non-sequence specific manner, while atomic force microscopy imaging of Sul12a-DNA complexes indicate that the protein induces structural effects on the DNA template. Based on these results, and a contrario to its initial annotation, it can be concluded that Sul12a is a novel chromatin-organizing protein.

Type 2 (T2) inflammation plays an important role in the pathogenesis of allergic diseases such as asthma, eosinophilic chronic rhinosinusitis (ECRS), or eosinophilic otitis media (EOM). Currently, in severe asthma with the T2 phenotype, biologics targeting mediators of T2 inflammation dramatically improve the management of severe asthma. While treatment with a single biologic is common, little is known about cases of the sequential use of two biologics. Here, we report a case of severe asthma with refractory ECRS and EOM in which total control of these allergic diseases could not be achieved with a single biologic but could be achieved via the sequential use of the anti-IL-5 receptor antibody and human anti-IL-4/13 receptor monoclonal antibody. It is suggested that it is necessary to control multiple T2 inflammatory pathways to achieve total control of severe allergic diseases. Sequential biotherapy may help solve the clinical challenges associated with single-agent molecular-targeted therapies.

Campylobacter jejuni represents an eminent zoonotic germ responsible for foodborne infections causing campylobacteriosis. In addition, infections with C. jejuni constitute a risk factor for the occurrence of inflammatory bowel disease (IBD). In the latter case, patients show inflammatory reactions not only against C. jejuni, but also against the non-infectious microbiota. However, the involved mechanisms and molecular basis are still largely unclear. We recently reported that C. jejuni breaches the intestinal epithelial barrier by secretion of serine protease HtrA (high temperature requirement A), which cleaves several major tight and adherens junction proteins. In the present study, we aimed to study if HtrA-expressing C. jejuni may also trigger the transepithelial migration of non-pathogenic gastrointestinal microbiota. Using confocal immunofluorescence and scanning electron microscopy, we demonstrate that C. jejuni wild-type (wt) as well as the isogenic ∆htrA mutant bind to the surface of polarized intestinal Caco-2 epithelial cells, but do not invade them at the apical side. Instead, C. jejuni wt, but not ∆htrA mutant, disrupt the cellular junctions and transmigrate using the paracellular route between neighboring cells. Using transwell filter systems, we then co-incubated the cells with C. jejuni and non-invasive microbiota strains, either Escherichia coli or Lactococcus lactis. Interestingly, C. jejuni wt, but not ∆htrA mutant, induced the efficient transmigration of these microbiota bacteria into the basal compartment. Thus, infection of the intestinal epithelium with C. jejuni causes local opening of cellular junctions and paracellular translocation in an HtrA-dependent manner, which paves the way for transmigration of microbiota that is otherwise non-invasive. Taken together, these findings may have impacts on various Campylobacter-associated diseases such as IBD, which are discussed here.

Several methods to isolate monocytes from whole blood have been previously published, with different advantages and disadvantages. For the purpose of cytokine release assessment upon external stimulation, the use of monocyte preparations consisting of non-activated cells is prerequisite. Affinity-isolated monocyte preparations from peripheral blood mononuclear cells (PBMCs), obtained via positive or negative selection using magnetic beads, released pro-inflammatory cytokines such as TNF-α and IL-6 even without adding external stimuli, hindering any assessment of an effect of bacterial lipoproteins on cell stimulation. Hence, the cell preparation protocol was modified by adding a quiescence step on repellent surface culture plates, dampening any monocyte pre-activation. This protocol now provides a robust method to prepare silent yet fully activatable, pure monocyte populations for further use in stimulus-elicited activation experiments.

Despite substance use disorders (SUD) being one of the leading causes of disability and mortality globally, available therapeutic approaches remain ineffective. The difficulty in accurately characterizing the neurobiological mechanisms involved with a purely qualitative diagnosis is an obstacle to improving the classification and treatment of SUD. In this regard, identifying central and peripheral biomarkers is essential to diagnosing the severity of drug dependence, monitoring therapeutic efficacy, predicting treatment response, and enhancing the development of safer and more effective pharmacological tools. In recent years, the crucial role that the endocannabinoid system (ECS) plays in regulating the reinforcing and motivational properties of drugs of abuse has been described. This has led to studies characterizing ECS alterations after exposure to various substances to identify biomarkers with potential diagnostic, prognostic, or therapeutic utility. This review aims to compile the primary evidence available from rodent and clinical studies on how the ECS components are modified in the context of different substance-related disorders, gathering data from genetic, molecular, functional, and neuroimaging experimental approaches. Finally, this report concludes that additional translational research is needed to further characterize the modifications of the ECS in the context of SUD, and their potential usefulness in the necessary search for biomarkers.

Leptin is a pleiotropic hormone known for regulating appetite and metabolism. To characterize the role of leptin signaling in rainbow trout, we used CRISPR/Cas9 genome editing to disrupt the leptin receptor (LepR) genes, lepra1 and lepra2. We compared wildtype (WT) and mutant fish that were either fed to satiation or feed deprived for six weeks. The LepR mutants exhibited a hyperphagic phenotype, which led to heavier body weight, faster specific growth rate, increased viscero- and hepatosomatic indices, and greater condition factor. Muscle glycogen, plasma leptin, and leptin transcripts (lepa1) were also elevated in fed LepR mutant fish. Expression levels of several hypothalamic genes involved in feed regulation were analyzed (agrp, npy, orexin, cart-1, cart-2, pomc-a1, pomc-b). No differences were detected between fed WT and mutants except for pomc-b (proopiomelanocortin-b), where levels were 7.5-fold higher in LepR fed mutants, suggesting that pomc-b expression is regulated by leptin signaling. Fatty acid (FA) content did not statistically differ in muscle of fed mutant fish compared to WT. However, fasted mutants exhibited significantly lower muscle FA concentrations, suggesting that LepR mutants exhibit increased FA mobilization during fasting. These data demonstrate a key role for leptin signaling in lipid and energy mobilization in a teleost fish.

Ghrelin and obestatin are two "sibling proteins" encoded by the same preproghrelin gene but possess an array of diverse and complex functions. While there are ample literature documenting ghrelin's functions, the roles of obestatin are less clear and controversial. Ghrelin and obestatin have been perceived to be antagonistic initially; however, recent studies challenge this dogma. While they have opposing effects in some systems, they function synergistically in other systems, with many functions remaining debatable. In this review, we discuss their functional relationship under three "C" categories, namely complex, complementary, and contradictory. Their functions in food intake, weight regulation, hydration, gastrointestinal motility, inflammation, and insulin secretion are complex. Their functions in pancreatic beta cells, cardiovascular, muscle, neuroprotection, cancer, and digestive system are complementary. Their functions in white adipose tissue, thermogenesis, and sleep regulation are contradictory. Overall, this review accumulates the multifaceted functions of ghrelin and obestatin under both physiological and pathological conditions, with the intent of contributing to a better understanding of these two important gut hormones.

Allergen immunotherapy is a promising treatment for allergic diseases that induce immune tolerance through the administration of specific allergens. In this study, we investigate the efficacy of sublingual immunotherapy (SLIT) in asthmatic patients with SAR-JCP and the dynamics of the parameters before and after treatment in a real-world setting. This was a prospective single-center observational study. Patients with asthma and SAR-JCP (n = 24) were recruited for this study and assessed using symptom questionnaires before SLIT and a year after the SLIT. In addition, a respiratory function test, forced oscillation technique, and blood sampling test were performed during the off-season before and after SLIT. The one-year SLIT for asthma patients with SAR-JCP significantly improved not only allergic rhinitis symptoms, but also asthma symptoms during the JCP dispersal season, and significantly improved airway resistance during the off-season. The change in the asthma control test and the visual analog scale score during the season before and after SLIT was negatively and positively correlated with the change in peripheral blood γδ T cells off-season before and after SLIT, respectively. It was suggested that improvement in asthma symptoms during the JCP dispersal season after SLIT was associated with reduced peripheral blood γδ T cells.

Prostate cancer is the most frequent malignant tumour among males (19%), often clinically silent and of difficult prognosis. Although several studies have highlighted the diagnostic and prognostic role of circulating biomarkers, such as PSA, their measurement does not necessarily allow the detection of the disease. Within this context, many authors suggest that the evaluation of circulating polyamines could represent a valuable tool, although several analytical problems still counteract their clinical practice. In particular, agmatine seems particularly intriguing, being a potential inhibitor of polyamines commonly derived from arginine. The aim of the present work was to evaluate the potential role of agmatine as a suitable biomarker for the identification of different classes of patients with prostate cancer (PC). For this reason, three groups of human patients-benign prostatic hyperplasia (BPH), precancerous lesion (PL), and prostate cancer (PC)-were recruited from a cohort of patients with suspected prostate cancer (n = 170), and obtained plasma was tested using the LC-HRMS method. Statistics on the receiver operating characteristics curve (ROC), and multivariate analysis were used to examine the predictive value of markers for discrimination among the three patient groups. Statistical analysis models revealed good discrimination using polyamine levels to distinguish the three classes of patients. AUC above 0.8, sensitivity ranging from 67% to 89%, specificity ranging from 74% to 89% and accuracy from 73% to 86%, considering the validation set, were achieved. Agmatine plasma levels were measured in PC (39.9 ± 12.06 ng/mL), BPH (77.62 ± 15.05 ng/mL), and PL (53.31 ± 15.27 ng/mL) patients. ROC analysis of the agmatine panel showed an AUC of 0.959 and p ≤ 0.001. These results could represent a future tool able to discriminate patients belonging to the three different clinical groups.

Charcot-Marie-Tooth (CMT) syndrome is the most common progressive human motor and sensory peripheral neuropathy. CMT type 1E is a demyelinating neuropathy affecting Schwann cells due to peripheral-myelin-protein-22 (PMP22) mutations, modelized by Trembler-J mice. Curcumin, a natural polyphenol compound obtained from turmeric (Curcuma longa), exhibits dose- and time-varying antitumor, antioxidant and neuroprotective properties, however, the neurotherapeutic actions of curcumin remain elusive. Here, we propose curcumin as a possible natural treatment capable of enhancing cellular detoxification mechanisms, resulting in an improvement of the neurodegenerative Trembler-J phenotype. Using a refined method for obtaining enriched Schwann cell cultures, we evaluated the neurotherapeutic action of low dose curcumin treatment on the PMP22 expression, and on the chaperones and autophagy/mammalian target of rapamycin (mTOR) pathways in Trembler-J and wild-type genotypes. In wild-type Schwann cells, the action of curcumin resulted in strong stimulation of the chaperone and macroautophagy pathway, whereas the modulation of ribophagy showed a mild effect. However, despite the promising neuroprotective effects for the treatment of neurological diseases, we demonstrate that the action of curcumin in Trembler-J Schwann cells could be impaired due to the irreversible impact of ethanol used as a common curcumin vehicle necessary for administration. These results contribute to expanding our still limited understanding of PMP22 biology in neurobiology and expose the intrinsic lability of the neurodegenerative Trembler-J genotype. Furthermore, they unravel interesting physiological mechanisms of cellular resilience relevant to the pharmacological treatment of the neurodegenerative Tremble J phenotype with curcumin and ethanol. We conclude that the analysis of the effects of the vehicle itself is an essential and inescapable step to comprehensibly assess the effects and full potential of curcumin treatment for therapeutic purposes.

It is well known that ginsenosides-major bioactive constituents of Panax ginseng-are attracting more attention due to their beneficial pharmacological activities. Ginsenoside Rd, belonging to protopanaxadiol (PPD)-type ginsenosides, exhibits diverse and powerful pharmacological activities. In recent decades, nearly 300 studies on the pharmacological activities of Rd-as a potential treatment for a variety of diseases-have been published. However, no specific, comprehensive reviews have been documented to date. The present review not only summarizes the in vitro and in vivo studies on the health benefits of Rd, including anti-cancer, anti-diabetic, anti-inflammatory, neuroprotective, cardioprotective, ischemic stroke, immunoregulation, and other pharmacological effects, it also delves into the inclusion of potential molecular mechanisms, providing an overview of future prospects for the use of Rd in the treatment of chronic metabolic diseases and neurodegenerative disorders. Although biotransformation, pharmacokinetics, and clinical studies of Rd have also been reviewed, clinical trial data of Rd are limited; the only data available are for its treatment of acute ischemic stroke. Therefore, clinical evidence of Rd should be considered in future studies.

Ménière's disease is a chronic illness characterized by intermittent episodes of vertigo associated with fluctuating sensorineural hearing loss, tinnitus and aural pressure. This pathology strongly correlates with a dilatation of the fluid compartment of the endolymph, so-called hydrops. Dexamethasone is one of the therapeutic approaches recommended when conventional antivertigo treatments have failed. Several mechanisms of actions have been hypothesized for the mode of action of dexamethasone, such as the anti-inflammatory effect or as a regulator of inner ear water homeostasis. However, none of them have been experimentally confirmed so far. Aquaporins (AQPs) are transmembrane water channels and are hence central in the regulation of transcellular water fluxes. In the present study, we investigated the hypothesis that dexamethasone could impact water fluxes in the inner ear by targeting AQP2. We addressed this question through molecular dynamics simulations approaches and managed to demonstrate a direct interaction between AQP2 and dexamethasone and its significant impact on the channel water permeability. Through compartmentalization of sodium and potassium ions, a significant effect of Na+ upon AQP2 water permeability was highlighted as well. The molecular mechanisms involved in dexamethasone binding and in its regulatory action upon AQP2 function are described.

Abdominal aortic aneurysm (AAA), defined as a focal dilation of the abdominal aorta beyond 50% of its normal diameter, is a common and potentially life-threatening vascular disease. The molecular and cellular mechanisms underlying AAA pathogenesis remain unclear. Healthy endothelial cells (ECs) play a critical role in maintaining vascular homeostasis by regulating vascular tone and maintaining an anti-inflammatory, anti-thrombotic local environment. Increasing evidence indicates that endothelial dysfunction is an early pathologic event in AAA formation, contributing to both oxidative stress and inflammation in the degenerating arterial wall. Recent studies utilizing single-cell RNA sequencing revealed heterogeneous EC sub-populations, as determined by their transcriptional profiles, in aortic aneurysm tissue. This review summarizes recent findings, including clinical evidence of endothelial dysfunction in AAA, the impact of biomechanical stress on EC in AAA, the role of endothelial nitric oxide synthase (eNOS) uncoupling in AAA, and EC heterogeneity in AAA. These studies help to improve our understanding of AAA pathogenesis and ultimately may lead to the generation of EC-targeted therapeutics to treat or prevent this deadly disease.

Environmental factors, including pollutants and lifestyle, constitute a significant role in severe, chronic pathologies with an essential societal, economic burden. The measurement of all environmental exposures and assessing their correlation with effects on individual health is defined as the exposome, which interacts with our unique characteristics such as genetics, physiology, and epigenetics. Epigenetics investigates modifications in the expression of genes that do not depend on the underlying DNA sequence. Some studies have confirmed that environmental factors may promote disease in individuals or subsequent progeny through epigenetic alterations. Variations in the epigenetic machinery cause a spectrum of different disorders since these mechanisms are more sensitive to the environment than the genome, due to the inherent reversible nature of the epigenetic landscape. Several epigenetic mechanisms, including modifications in DNA (e.g., methylation), histones, and noncoding RNAs can change genome expression under the exogenous influence. Notably, the role of long noncoding RNAs in epigenetic processes has not been well explored in the context of exposome-induced tumorigenesis. In the present review, our scope is to provide relevant evidence indicating that epigenetic alterations mediate those detrimental effects caused by exposure to environmental toxicants, focusing mainly on a multi-step regulation by diverse noncoding RNAs subtypes.

Cocaine use disorder has been reported to cause transgenerational effects. However, due to the lack of standardized biomarkers, the effects of cocaine use during pregnancy on postnatal development and long-term neurobiological and behavioral outcomes have not been investigated thoroughly. Therefore, in this study, we examined extracellular vesicles (EVs) in adult (~12 years old) female and male rhesus monkeys prenatally exposed to cocaine (n = 11) and controls (n = 9). EVs were isolated from the cerebrospinal fluid (CSF) and characterized for the surface expression of specific tetraspanins, concentration (particles/mL), size distribution, and cargo proteins by mass spectrometry (MS). Transmission electron microscopy following immunogold labeling for tetraspanins (CD63, CD9, and CD81) confirmed the successful isolation of EVs. Nanoparticle tracking analyses showed that the majority of the particles were <200 nm in size, suggesting an enrichment for small EVs (sEV). Interestingly, the prenatally cocaine-exposed group showed ~54% less EV concentration in CSF compared to the control group. For each group, MS analyses identified a number of proteins loaded in CSF-EVs, many of which are commonly listed in the ExoCarta database. Ingenuity pathway analysis (IPA) demonstrated the association of cargo EV proteins with canonical pathways, diseases and disorders, upstream regulators, and top enriched network. Lastly, significantly altered proteins between groups were similarly characterized by IPA, suggesting that prenatal cocaine exposure could be potentially associated with long-term neuroinflammation and risk for neurodegenerative diseases. Overall, these results indicate that CSF-EVs could potentially serve as biomarkers to assess the transgenerational adverse effects due to prenatal cocaine exposure.

The concept of molecular similarity has been commonly used in rational drug design, where structurally similar molecules are examined in molecular databases to retrieve functionally similar molecules. The most used conventional similarity methods used two-dimensional (2D) fingerprints to evaluate the similarity of molecules towards a target query. However, these descriptors include redundant and irrelevant features that might impact the performance of similarity searching methods. Thus, this study proposed a new approach for identifying the important features of molecules in chemical datasets based on the representation of the molecular features using Autoencoder (AE), with the aim of removing irrelevant and redundant features. The proposed approach experimented using the MDL Data Drug Report standard dataset (MDDR). Based on experimental findings, the proposed approach performed better than several existing benchmark similarity methods such as Tanimoto Similarity Method (TAN), Adapted Similarity Measure of Text Processing (ASMTP), and Quantum-Based Similarity Method (SQB). The results demonstrated that the performance achieved by the proposed approach has proven to be superior, particularly with the use of structurally heterogeneous datasets, where it yielded improved results compared to other previously used methods with the similar goal of improving molecular similarity searching.

α-synuclein (α-syn) is a presynaptic, lipid-binding protein strongly associated with the neuropathology observed in Parkinson's disease (PD), dementia with Lewy bodies (DLB), and Alzheimer's Disease (AD). In normal physiology, α-syn plays a pivotal role in facilitating endocytosis and exocytosis. Interestingly, mutations and modifications of precise α-syn domains interfere with α-syn oligomerization and nucleation that negatively affect presynaptic vesicular dynamics, protein expressions, and mitochondrial profiles. Furthermore, the integration of the α-syn oligomers into the presynaptic membrane results in pore formations, ion influx, and excitotoxicity. Targeted therapies against specific domains of α-syn, including the use of small organic molecules, monoclonal antibodies, and synthetic peptides, are being screened and developed. However, the prospect of an effective α-syn targeted therapy is still plagued by low permeability across the blood-brain barrier (BBB), and poor entry into the presynaptic axon terminals. The present review proposes a modification of current strategies, which includes the use of novel encapsulation technology, such as lipid nanoparticles, to bypass the BBB and deliver such agents into the brain.

Previous studies have shown that atypical dopamine-transporter-inhibitors such as modafinil and its analogues modify behavioral and cognitive functions in rodents. Here, we tested potential promnestic effects of the novel, more dopamine-transporter selective modafinil analogue CE-158 in the social discrimination memory task in male mice. Systemic administration of CE-158 1 h before the social learning event prevented the impairment of social-recognition memory following retroactive interference 3 h after the learning session of a juvenile conspecific. This effect was dose-dependent, as mice treated with 10 mg/kg, but not with 1 mg/kg CE-158, were able to discriminate between the novel and familiar conspecific despite the presentation of an interference stimulus, both 3 h and 6 h post learning. However, when 10 mg/kg of the drug was administered after learning, CE-158 failed to prevent social memory from interference. Paralleling these behavioral effects, the systemic administration of 10 mg/kg CE-158 caused a rapid and sustained elevation of extracellular dopamine in the nucleus accumbens, a brain area where dopaminergic signaling plays a key role in learning and memory function, of freely moving mice, while 1 mg/kg was not sufficient for altering dopamine levels. Taken together, our findings suggest promnestic effects of the novel dopamine-transporter-inhibitor CE-158 in a social recognition memory test that may be in part mediated via increased dopamine-neurotransmission in the nucleus accumbens. Thus, selective-dopamine-transporter-inhibitors such as CE-158 may represent interesting drug candidates for the treatment of memory complaints observed in humans with cognitive impairments and dementia.

Tau mutations promote the formation of tau oligomers and filaments, which are neuropathological signs of several tau-associated dementias. Types of neurons in the CNS are spared of tau pathology and are surrounded by a specialized form of extracellular matrix; called perineuronal nets (PNs). Aggrecan, the major PN proteoglycans, is suggested to mediate PNs neuroprotective function by forming an external shield preventing the internalization of misfolded tau. We recently demonstrated a correlation between aggrecan amount and the expression and phosphorylation of tau in a TauP310L-acan mouse model, generated by crossbreeding heterozygous aggrecan mice with a significant reduction of aggrecan and homozygous TauP301L mice. Neurodegenerative processes have been associated with changes of PN structure and protein signature. In this study, we hypothesized that the structure and protein expression of PNs in this TauP310L-acan mouse is regulated by tau. Immunohistochemical and biochemical analyses demonstrate that protein levels of PN components differ between TauP301LHET-acanWT and TauP301LHET-acanHET mice, accompanied by changes in the expression of protein phosphatase 2 A. In addition, tau can modulate PN components such as brevican. Co-immunoprecipitation experiments revealed a physical connection between PN components and tau. These data demonstrate a complex, mutual interrelation of tau and the proteoglycans of the PN.

Cyanobacteria are a promising source for the sustainable production of biodegradable bioplastics such as poly(3-hydroxybutyrate) (PHB). The auto-phototrophic biomass formation is based on light and CO2, which is an advantage compared to heterotrophic PHB-producing systems. So far, only a handful of cyanobacterial species suitable for the high-yield synthesis of PHB have been reported. In the present study, the PHB formation, biomass, and elemental composition of Leptolyngbya sp. NIVA-CYA 255 were investigated. Therefore, a three-stage cultivation process was applied, consisting of a growth stage; an N-, P-, and NP-depleted phototrophic stage; and a subsequent mixotrophic deficiency stage, initiated by sodium acetate supplementation. The extracted cyanobacterial PHB was confirmed by FTIR- and GC-MS analyses. Furthermore, the fluorescent dyes LipidGreen2 and Nile red were used for fluorescence-based monitoring and the visualization of PHB. LipidGreen2 was well suited for PHB quantification, while the application of Nile red was limited by fluorescence emission crosstalk with phycocyanin. The highest PHB yields were detected in NP- (325 mg g-1) and N-deficiency (213 mg g-1). The glycogen pool was reduced in all cultures during mixotrophy, while lipid composition was not affected. The highest glycogen yield was formed under N-deficiency (217 mg g-1). Due to the high carbon storage capacity and PHB formation, Leptolyngbya sp. NIVA-CYA 255 is a promising candidate for PHB production. Further work will focus on upscaling to a technical scale and monitoring the formation by LipidGreen2-based fluorometry.

The plasminogen activation system regulates the activity of the serine protease, plasmin. The role of plasminogen receptors in cancer progression is being increasingly appreciated as key players in modulation of the tumor microenvironment. The interaction of plasminogen with cells to promote plasminogen activation requires the presence of proteins exposing C-terminal lysines on the cell surface. Plg-RKT is a structurally unique plasminogen receptor because it is an integral membrane protein that is synthesized with and binds plasminogen via a C-terminal lysine exposed on the cell surface. Here, we have investigated the expression of Plg-RKT in human breast tumors and human breast cancer cell lines. Breast cancer progression tissue microarrays were probed with anti-Plg-RKT mAB and we found that Plg-RKT is widely expressed in human breast tumors, that its expression is increased in tumors that have spread to draining lymph nodes and distant organs, and that Plg-RKT expression is most pronounced in hormone receptor (HR)-positive tumors. Plg-RKT was detected by Western blotting in human breast cancer cell lines. By flow cytometry, Plg-RKT cell surface expression was highest on the most aggressive tumor cell line. Future studies are warranted to address the functions of Plg-RKT in breast cancer.

Cells' ability to track chemical gradients is integral to many biological phenomena, including fertilization, development, accessing nutrients, and combating infection. Mating of the yeast Saccharomyces cerevisiae provides a tractable model to understand how cells interpret the spatial information in chemical gradients. Mating yeast of the two different mating types secrete distinct peptide pheromones, called a-factor and α-factor, to communicate with potential partners. Spatial gradients of pheromones are decoded to guide mobile polarity sites so that polarity sites in mating partners align towards each other, as a prerequisite for cell-cell fusion and zygote formation. In ascomycetes including S. cerevisiae, one pheromone is prenylated (a-factor) while the other is not (α-factor). The difference in physical properties between the pheromones, combined with associated differences in mechanisms of secretion and extracellular pheromone metabolism, suggested that the pheromones might differ in the spatial information that they convey to potential mating partners. However, as mating appears to be isogamous in this species, it is not clear why any such signaling difference would be advantageous. Here we report assays that directly track movement of the polarity site in each partner as a way to understand the spatial information conveyed by each pheromone. Our findings suggest that both pheromones convey very similar information. We speculate that the different pheromones were advantageous in ancestral species with asymmetric mating systems and may represent an evolutionary vestige in yeasts that mate isogamously.

Leaf spot disease caused by Lasiodiplodia theobromae and Alternaria tenuissima is a seriously fungal disease in kiwifruit production. In this study, the co-application of tetramycin and chitosan against leaf spot disease in kiwifruit and its effects on the disease resistance, photosynthesis, quality and amino acids of kiwifruit were investigated. The results show that tetramycin exhibited an excellent antifungal activity against L. theobromae and A. tenuissima with EC50 values of 2.37 and 0.16 mg kg-1. In the field, the foliar co-application of tetramycin and chitosan could effectively control leaf spot disease with control efficacy of 89.44% by spraying 0.3% tetramycin aqueous solutions (AS) 5000 time liquid + chitosan 100 time liquid, which was significantly (ANOVA, p < 0.01) higher than 79.80% of 0.3% tetramycin AS 5000 time liquid and 56.61% of chitosan 100 time liquid. Simultaneously, the co-application of tetramycin and chitosan was more effective than tetramycin or chitosan alone in enhancing the disease resistance and photosynthesis of kiwifruit leaves, as well as improving the quality and amino acids of kiwifruit fruits. This work highlights that chitosan is a practicable, cost-effective and eco-friendly adjuvant of tetramycin for controlling leaf spot disease of kiwifruit, enhancing resistance and photosynthesis of leaves and improving quality and amino acids of fruits.

Polyamines (PAs) are small, versatile molecules with two or more nitrogen-containing positively charged groups and provide widespread biological functions. Most of these aspects are well known and covered by quite a number of excellent surveys. Here, the present review includes novel aspects and questions: (1) It summarizes the role of most natural and some important synthetic PAs. (2) It depicts PA uptake from nutrition and bacterial production in the intestinal system following loss of PAs via defecation. (3) It highlights the discrepancy between the high concentrations of PAs in the gut lumen and their low concentration in the blood plasma and cerebrospinal fluid, while concentrations in cellular cytoplasm are much higher. (4) The present review provides a novel and complete scheme for the biosynthesis of Pas, including glycine, glutamate, proline and others as PA precursors, and provides a hypothesis that the agmatine pathway may rescue putrescine production when ODC knockout seems to be lethal (solving the apparent contradiction in the literature). (5) It summarizes novel data on PA transport in brain glial cells explaining why these cells but not neurons preferentially accumulate PAs. (6) Finally, it provides a novel and complete scheme for PA interconversion, including hypusine, putreanine, and GABA (unique gliotransmitter) as end-products. Altogether, this review can serve as an updated contribution to understanding the PA mystery.

Apolipoprotein E4 (ApoE4) is thought to increase the risk of developing Alzheimer's disease. Several studies have shown that ApoE4-Amyloid β (Aβ) interactions can increment amyloid depositions in the brain and that this can be augmented at low pH values. On the other hand, experimental studies in transgenic mouse models have shown that treatment with enoxaparin significantly reduces cortical Aβ levels, as well as decreases the number of activated astrocytes around Aβ plaques. However, the interactions between enoxaparin and the ApoE4-Aβ proteins have been poorly explored. In this work, we combine molecular dynamics simulations, molecular docking, and binding free energy calculations to elucidate the molecular properties of the ApoE4-Aβ interactions and the competitive binding affinity of the enoxaparin on the ApoE4 binding sites. In addition, we investigated the effect of the environmental pH levels on those interactions. Our results showed that under different pH conditions, the closed form of the ApoE4 protein, in which the C-terminal domain folds into the protein, remains stabilized by a network of hydrogen bonds. This closed conformation allowed the generation of six different ApoE4-Aβ interaction sites, which were energetically favorable. Systems at pH5 and 6 showed the highest energetic affinity. The enoxaparin molecule was found to have a strong energetic affinity for ApoE4-interacting sites and thus can neutralize or disrupt ApoE4-Aβ complex formation.

Age-related hearing loss (ARHL) or presbycusis is a prevalent condition associated with social isolation, cognitive impairment, and dementia. Age-related changes in the cochlea, the auditory portion of the inner ear, are the primary cause of ARHL. Unfortunately, there are currently no pharmaceutical approaches to treat ARHL. To examine the biological processes underlying age-related changes in the cochlea and identify candidate drugs for rapid repurposing to treat ARHL, we utilized bulk RNA sequencing to obtain transcriptomes from the functional substructures of the cochlea-the sensorineural structures, including the organ of Corti and spiral ganglion neurons (OC/SGN) and the stria vascularis and spiral ligament (SV/SL)-in young (6-week-old) and old (2-year-old) C57BL/6 mice. Transcriptomic analyses revealed both overlapping and unique patterns of gene expression and gene enrichment between substructures and with ageing. Based on these age-related transcriptional changes, we queried the protein products of genes differentially expressed with ageing in DrugBank and identified 27 FDA/EMA-approved drugs that are suitable to be repurposed to treat ARHL. These drugs target the protein products of genes that are differentially expressed with ageing uniquely in either the OC/SGN or SV/SL and that interrelate diverse biological processes. Further transcriptomic analyses revealed that most genes differentially expressed with ageing in both substructures encode protein products that are promising drug target candidates but are, nevertheless, not yet linked to approved drugs. Thus, with this study, we apply a novel approach to characterize the druggable genetic landscape for ARHL and propose a list of drugs to test in pre-clinical studies as potential treatment options for ARHL.

Endocan is a soluble dermatan sulfate proteoglycan expressed by endothelial cells and detected in serum/plasma. Its expression is increased in tumors/tumor vessels in several human malignancies, and high expression (high serum/plasma levels or tumor levels) has an adverse prognostic impact in several malignancies. The p14 endocan degradation product can also be detected in serum/plasma, but previous clinical studies as well as previously unpublished results presented in this review suggest that endocan and p14 endocan fragment levels reflect different biological characteristics, and the endocan levels seem to reflect the disease heterogeneity in acute leukemia better than the p14 fragment levels. Furthermore, decreased systemic endocan levels in previously immunocompetent sepsis patients are associated with later severe respiratory complications, but it is not known whether this is true also for immunocompromised acute leukemia patients. Finally, endocan is associated with increased early nonrelapse mortality in (acute leukemia) patients receiving allogeneic stem cell transplantation, and this adverse prognostic impact seems to be independent of the adverse impact of excessive fluid overload. Systemic endocan levels may also become important to predict cytokine release syndrome after immunotherapy/haploidentical transplantation, and in the long-term follow-up of acute leukemia survivors with regard to cardiovascular risk. Therapeutic targeting of endocan is now possible, and the possible role of endocan in acute leukemia should be further investigated to clarify whether the therapeutic strategy should also be considered.

Cancer is a complex family of diseases affecting millions of people worldwide. Gliomas are primary brain tumors that account for ~80% of all malignant brain tumors. Glioblastoma multiforme (GBM) is the most common, invasive, and lethal subtype of glioma. Therapy resistance and intra-GBM tumoral heterogeneity are promoted by subpopulations of glioma stem cells (GSCs). Cannabis sativa produces hundreds of secondary metabolites, such as flavonoids, terpenes, and phytocannabinoids. Around 160 phytocannabinoids have been identified in C. sativa. Cannabis is commonly used to treat various medical conditions, and it is used in the palliative care of cancer patients. The anti-cancer properties of cannabis compounds include cytotoxic, anti-proliferative, and anti-migratory activities on cancer cells and cancer stem cells. The endocannabinoids system is widely distributed in the body, and its dysregulation is associated with different diseases, including various types of cancer. Anti-cancer activities of phytocannabinoids are mediated in glioma cells, at least partially, by the endocannabinoid receptors, triggering various cellular signaling pathways, including the endoplasmic reticulum (ER) stress pathway. Specific combinations of multiple phytocannabinoids act synergistically against cancer cells and may trigger different anti-cancer signaling pathways. Yet, due to scarcity of clinical trials, there remains no solid basis for the anti-cancer therapeutic potential of cannabis compounds.

There is a sex bias in tuberculosis's severity, prevalence, and pathogenesis, and the rates are higher in men. Immunological and physiological factors are fundamental contributors to the development of the disease, and sex-related factors could play an essential role in making women more resistant to severe forms of the disease. In this study, we evaluated sex-dependent differences in inflammatory markers. Serum samples were collected from 34 patients diagnosed with pulmonary TB (19 male and 15 female) and 27 healthy controls (18 male and 9 female). Cytokines IL2, IL4, IL6, IL8, IL10, IFNγ, TNFα, and GM-CSF, and eicosanoids PGE2, LTB4, RvD1, and Mar1 were measured using commercially available immunoassays. The MDA, a product of lipidic peroxidation, was measured by detecting thiobarbituric-acid-reactive substances (TBARS). Differential inflammation patterns between men and women were observed. Men had higher levels of IL6, IL8, and TNFα than women. PGE2 and LTB4 levels were higher in patients than healthy controls, but there were no differences for RvD1 and Mar1. Women had higher RvD1/PGE2 and RvD1/LTB4 ratios among patients. RvD1 plays a vital role in resolving the inflammatory process of TB in women. Men are the major contributors to the typical pro-inflammatory profile observed in the serum of tuberculosis patients.

CHMP2B is a protein that coordinates membrane scission events as a core component of the ESCRT machinery. Mutations in CHMP2B are an uncommon cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), two neurodegenerative diseases with clinical, genetic, and pathological overlap. Different mutations have now been identified across the ALS-FTD spectrum. Disruption of the neuromuscular junction is an early pathogenic event in ALS. Currently, the links between neuromuscular junction functionality and ALS-associated genes, such as CHMP2B, remain poorly understood. We have previously shown that CHMP2B transgenic mice expressing the CHMP2Bintron5 mutant specifically in neurons develop a progressive motor phenotype reminiscent of ALS. In this study, we used complementary approaches (behavior, histology, electroneuromyography, and biochemistry) to determine the extent to which neuron-specific expression of CHMP2Bintron5 could impact the skeletal muscle characteristics. We show that neuronal expression of the CHMP2Bintron5 mutant is sufficient to trigger progressive gait impairment associated with structural and functional changes in the neuromuscular junction. Indeed, CHMP2Bintron5 alters the pre-synaptic terminal organization and the synaptic transmission that ultimately lead to a switch of fast-twitch glycolytic muscle fibers to more oxidative slow-twitch muscle fibers. Taken together these data indicate that neuronal expression of CHMP2Bintron5 is sufficient to induce a synaptopathy with molecular and functional changes in the motor unit reminiscent of those found in ALS patients.

Pulmonary hypertension (PH) is a fatal and untreatable disease, ultimately leading to right heart failure and eventually death. microRNAs are small, non-coding endogenous RNA molecules that can regulate gene expression and influence various biological processes. Changes in microRNA expression levels contribute to various cardiovascular disorders, and microRNAs have been shown to play a critical role in PH pathogenesis. In recent years, numerous studies have explored the role of microRNAs in PH, focusing on the expression profiles of microRNAs and their signaling pathways in pulmonary artery smooth muscle cells (PASMCs) or pulmonary artery endothelial cells (PAECs), PH models, and PH patients. Moreover, certain microRNAs, such as miR-150 and miR-26a, have been identified as good candidates of diagnosis biomarkers for PH. However, there are still several challenges for microRNAs as biomarkers, including difficulty in normalization, specificity in PH, and a lack of longitudinal and big sample-sized studies. Furthermore, microRNA target drugs are potential therapeutic agents for PH treatment, which have been demonstrated in PH models and in humans. Nonetheless, synthetic microRNA mimics or antagonists are susceptible to several common defects, such as low drug efficacy, inefficient drug delivery, potential toxicity and especially, off-target effects. Therefore, finding clinically safe and effective microRNA drugs remains a great challenge, and further breakthrough is urgently needed.

In DNA computing, the implementation of complex and stable logic operations in a universal system is a critical challenge. It is necessary to develop a system with complex logic functions based on a simple mechanism. Here, the strategy to control the secondary structure of assembled DNAzymes' conserved domain is adopted to regulate the activity of DNAzymes and avoid the generation of four-way junctions, and makes it possible to implement basic logic gates and their cascade circuits in the same system. In addition, the purpose of threshold control achieved by the allosteric secondary structure implements a three-input DNA voter with one-vote veto function. The scalability of the system can be remarkably improved by adjusting the threshold to implement a DNA voter with 2n + 1 inputs. The proposed strategy provides a feasible idea for constructing more complex DNA circuits and a highly integrated computing system.

Parkinson disease (PD) is attributed to a proteostasis disorder mediated by α-synuclein accumulating in a specific brain region. PD manifestation is often related to extraneuronal alterations, some of which could be used as diagnostic or prognostic PD biomarkers. In this work, we studied the shifts in the expression of proteostasis-associated chaperones of the HSP70 family and autophagy-dependent p62 protein values in the peripheral blood mononuclear cells (PBMC) of mild to moderate PD patients. Although we did not detect any changes in the intracellular HSP70 protein pool in PD patients compared to non-PD controls, an increase in the transcriptional activity of the stress-associated HSPA1A/B and HSPA6 genes was observed in these cells. Basal p62 content was found to be increased in PD patients' PBMC, similarly to the p62 level in substantia nigra neural cells in PD. Moreover, the spontaneous apoptosis level was increased among PBMC and positively correlated with the p62 intracellular level in the PD group. A combined HSPA6- and p62-based analysis among 26 PD patients and 36 age-matched non-PD controls pointed out the diagnostic significance of these markers, with intermediate sensitivity and high specificity of this combination when observing patients diagnosed with PD.

In 1957, Lionel Penrose built the first man-made self-replicating mechanical device and illustrated its function in a series of machine prototypes, prefiguring our current view of the genesis and the proliferation of amyloid fibrils. He invented and demonstrated, with the help of his son Roger, the concepts that decades later, would become the fundamentals of prion and prion-like neurobiology: nucleation, seeding and conformational templating of monomers, linear polymer elongation, fragmentation, and spread. He published his premonitory discovery in a movie he publicly presented at only two conferences in 1958, a movie we thus reproduce here. By making a 30-year-jump in the early 90's, we evoke the studies performed by Peter Lansbury and his group in which α-Synuclein (α-Syn) was for the first time (i) compared to a prion; (ii) shown to contain a fibrillization-prone domain capable of seeding its own assembly into fibrils; (iii) identified as an intrinsically disordered protein (IDP), and which, in the early 2000s, (iv) was described by one of us as a protein chameleon. We use these temporally distant breakthroughs to propose that the combination of the chameleon nature of α-Syn with the rigid gear of the Penrose machine is sufficient to account for a phenomenon that is of current interest: the emergence and the spread of a variety of α-Syn fibril strains in α-Synucleinopathies.

We conducted a case-control study in order to evaluate whether ABO gene polymorphisms were associated with a high risk of developing COVID-19 in a cohort of patients. Six ABO gene polymorphisms (rs651007 T/C, rs579459 T/C, rs495828 T/G, rs8176746 A/C, rs8176740 T/A, and rs512770 T/C) were determined using TaqMan genotyping assays in a group of 415 COVID-19 patients and 288 healthy controls. The distribution of rs651007 T/C, rs579459 T/C, rs495828 T/G, and rs8176746 A/C polymorphisms was similar in patients and healthy controls. Nonetheless, under co-dominant (OR = 1.89, pCCo-dominant = 6 × 10-6), recessive (OR = 1.98, pCRecessive = 1 × 10-4), and additive (OR = 1.36, pCAdditive = 3 × 10-3) models, the TT genotype of the rs8176740 T/A polymorphism increased the risk of developing COVID-19. In the same way, under co-dominant, recessive, and additive models, the TT genotype of the rs512770 T/C polymorphism was associated with a high risk of developing COVID-19 (OR = 1.87, pCCo-dominant = 2 × 10-3; OR = 1.87, pCRecessive = 5 × 10-4; and OR = 1.35, pCAdditive = 4 × 10-3, respectively). On the other hand, the GTC and GAT haplotypes were associated with a high risk of COVID-19 (OR = 5.45, pC = 1 × 10-6 and OR = 6.33, pC = 1 × 10-6, respectively). In addition, the rs8176740 TT genotype was associated with high-platelet plasma concentrations in patients with COVID-19. Our data suggested that the ABO rs512770 T/C and rs8176740 T/A polymorphisms increased the risk of developing COVID-19 and the plasma concentration of platelets.

Most of the research on bioremediation and estimation of microbial diversity in waste contaminated sites is focused on the domain Bacteria, whereas details on the relevance of Archaea are still lacking. The present study examined the archaeal diversity and predicted metabolic pathways in two discrete sites (SITE1 and SITE2) contaminated with explosives (RDX and HMX) by amplicon-targeted sequencing of 16S rRNA genes. In total, 14 soil samples were processed, and 35,758 OTUs were observed, among which 981 OTUs were classified as Archaea, representing ~2.7% of the total microbial diversity in our samples. The majority of OTUs belonged to phyla Euryarchaeota (~49%), Crenarchaeota (~24%), and Thaumarchaeota (~23%), while the remaining (~4%) OTUs were affiliated to Candidatus Parvarchaeota, Candidatus Aenigmarchaeota, and Candidatus Diapherotrites. The comparative studies between explosives contaminated and agricultural soil samples (with no history of explosives contamination) displayed significant differences between the compositions of the archaeal communities. Further, the metabolic pathways pertaining to xenobiotic degradation were presumably more abundant in the contaminated sites. Our data provide a first comprehensive report of archaeal communities in explosives contaminated sites and their putative degradation role in such ecosystems which have been as yet unexplored.

The sarcoplasmic reticulum of skeletal muscle cells is a highly ordered structure consisting of an intricate network of tubules and cisternae specialized for regulating Ca2+ homeostasis in the context of muscle contraction. The sarcoplasmic reticulum contains several proteins, some of which support Ca2+ storage and release, while others regulate the formation and maintenance of this highly convoluted organelle and mediate the interaction with other components of the muscle fiber. In this review, some of the main issues concerning the biology of the sarcoplasmic reticulum will be described and discussed; particular attention will be addressed to the structure and function of the two domains of the sarcoplasmic reticulum supporting the excitation-contraction coupling and Ca2+-uptake mechanisms.

Breast cancer is an ancient disease recognized first by the Egyptians as early as 1600 BC. The first cancer-causing gene in a chicken tumor virus was found in 1970. The United States signed the National Cancer Act in 1971, authorizing federal funding for cancer research. Irrespective of multi-disciplinary approaches, diverting a great deal of public and private resources, breast cancer remains at the forefront of human diseases, affecting as many as one in eight women during their lifetime. Because of overarching challenges and changes in the breast cancer landscape, five-year disease-free survival is no longer considered adequate. The absence of a cure, and the presence of drug resistance, severe side effects, and destruction of the patient's quality of life, as well as the fact that therapy is often expensive, making it unaffordable to many, have created anxiety among patients, families, and friends. One of the reasons for the failure of cancer therapeutics is that the approaches do not consider cancer holistically. Characteristically, all breast cancer cells and their microenvironmental capillary endothelial cells express asparagine-linked (N-linked) glycoproteins with diverse structures. We tested a small biological molecule, Tunicamycin, that blocks a specific step of the protein N-glycosylation pathway in the endoplasmic reticulum (ER), i.e., the catalytic activity of N-acetylglusosaminyl 1-phosphate transferase (GPT). The outcome was overwhelmingly exciting. Tunicamycin quantitatively inhibits angiogenesis in vitro and in vivo, and inhibits the breast tumor progression of multiple subtypes in pre-clinical mouse models with "zero" toxicity. Mechanistic details support ER stress-induced unfolded protein response (upr) signaling as the cause for the apoptotic death of both cancer and the microvascular endothelial cells. Additionally, it interferes with Wnt signaling. We therefore conclude that Tunicamycin can be expected to supersede the current therapeutics to become a glycotherapy for treating breast cancer of all subtypes.

Ghrelin, a hormone produced and secreted from the stomach, is prim arily known as an appetite stimulant. Recently, it has emerged as a potential regulator/biomarker of cancer progression. Inconsistent results on this subject make this body of literature difficult to interpret. Here, we attempt to identify commonalities in the relationships between ghrelin and various cancers, and summarize important considerations for future research. The main players in the ghrelin family axis are unacylated ghrelin (UAG), acylated ghrelin (AG), the enzyme ghrelin O-acyltransferase (GOAT), and the growth hormone secretagogue receptor (GHSR). GOAT is responsible for the acylation of ghrelin, after which ghrelin can bind to the functional ghrelin receptor GHSR-1a to initiate the activation cascade. Splice variants of ghrelin also exist, with the most prominent being In1-ghrelin. In this review, we focus primarily on the potential of In1-ghrelin as a biomarker for cancer progression, the unique characteristics of UAG and AG, the importance of the two known receptor variants GHSR-1a and 1b, as well as the possible mechanisms through which the ghrelin axis acts. Further understanding of the role of the ghrelin axis in tumor cell proliferation could lead to the development of novel therapeutic approaches for various cancers.

The merging of distinct computational approaches has become a powerful strategy for discovering new biologically active compounds. By using molecular modeling, significant efforts have recently resulted in the development of new molecules, demonstrating high efficiency in reducing the replication of severe acute respiratory coronavirus 2 (SARS-CoV-2), the agent responsible for the COVID-19 pandemic. We have focused our interest on non-structural protein Nsp13 (NTPase/helicase), as a crucial protein, embedded in the replication-transcription complex (RTC), that controls the virus life cycle. To assist in the identification of the most druggable surfaces of Nsps13, we applied a combination of four computational tools: FTMap, SiteMap, Fpocket and LigandScout. These software packages explored the binding sites for different three-dimensional structures of RTC complexes (PDB codes: 6XEZ, 7CXM, 7CXN), thus, detecting several hot spots, that were clustered to obtain ensemble consensus sites, through a combination of four different approaches. The comparison of data provided new insights about putative druggable sites that might be employed for further docking simulations on druggable surfaces of Nsps13, in a scenario of repurposing drugs.

Endothelial dysfunction is a key player in advancing vascular pathology in pulmonary arterial hypertension (PAH), a disease essentially characterized by intense remodeling of the pulmonary vasculature, vasoconstriction, endothelial dysfunction, inflammation, oxidative stress, and thrombosis in situ. These vascular features culminate in an increase in pulmonary vascular resistance, subsequent right heart failure, and premature death. Over the past years, there has been a great development in our understanding of pulmonary endothelial biology related to the genetic and molecular mechanisms that modulate the endothelial response to direct or indirect injury and how their dysregulation can promote PAH pathogenesis. Ion channels are key regulators of vasoconstriction and proliferative/apoptotic phenotypes; however, they are poorly studied at the endothelial level. The current review will describe and categorize different expression, functions, regulation, and remodeling of endothelial ion channels (K+, Ca2+, Na+, and Cl- channels) in PAH. We will focus on the potential pathogenic role of ion channel deregulation in the onset and progression of endothelial dysfunction during the development of PAH and its potential therapeutic role.

Immunotherapy is one of the most promising forms of cancer treatment. In particular, immune checkpoint blockers (ICBs) represent some of the leading candidates which many drug developers have heavily invested in. During pre-clinical development and prior to human clinical trials, animal tests are a critical component for determining the safety and efficacy of newly developed ICBs for cancer treatment. In this study, we strive to demonstrate the feasibility of using hollow fiber assay microtube array membrane (MTAM-HFA) in the screening of anti-cancer ICBs. The MTAM-HFA process was carried out by encapsulating peripheral blood mononuclear cells (PBMCs) and the target cancer cells (cell lines or primary cells) and subcutaneously implanting them into Balb/C mice. At predetermined time points combination regimens of PD-1/PD-L1+ were administered accordingly and at a predetermined time point, the MTAMs were retrieved, and cell viability assays were carried out. The outcomes of the MTAM-HFA were compared against the clinical outcome of patients. Clinical comparison demonstrated excellent correlation between the screening outcome of MTAM-HFA of PD-1/PD-L1+ combination therapy and the clinical outcome of the lung cancer patients. Basic cell studies revealed that the utilization of MTAM-HFA in PD-1/PD-L1+ combination therapy revealed enhanced T-cell activity upon the administration of the PD-1/PD-L1 drug; thereby resulting in the reduction of tumor cell viability by up to 70%, and the cytotoxic effects by 82%. The outcome was echoed in the in vivo cell studies. This suggested that the MTAM-HFA system is suitable for use in PD-1/PD-L1+ screening and the accuracy, rapidity and cost effectiveness made it extremely suitable for application as a companion diagnostic system in both personalized medicine for cancer treatment and could potentially be applied to screen for candidate compounds in the development of next generation PD-1/PD-L1+ combination therapies.

Chromatin compaction and regulation are essential processes for the normal function of all organisms, yet knowledge on how archaeal chromosomes are packed into higher-order structures inside the cell remains elusive. In this study, we investigated the role of archaeal architectural proteins Alba and Cren7 in chromatin folding and dynamics. Atomic force microscopy revealed that Sulfolobus solfataricus chromatin is composed of 28 nm fibers and 60 nm globular structures. In vitro reconstitution showed that Alba can mediate the formation of folded DNA structures in a concentration-dependent manner. Notably, it was demonstrated that Alba on its own can form higher-order structures with DNA. Meanwhile, Cren7 was observed to affect the formation of Alba-mediated higher-order chromatin structures. Overall, the results suggest an interplay between Alba and Cren7 in regulating chromatin compaction in archaea.

Aldose reductase, classified within the aldo-keto reductase family as AKR1B1, is an NADPH dependent enzyme that catalyzes the reduction of hydrophilic as well as hydrophobic aldehydes. AKR1B1 is the first enzyme of the so-called polyol pathway that allows the conversion of glucose into sorbitol, which in turn is oxidized to fructose by sorbitol dehydrogenase. The activation of the polyol pathway in hyperglycemic conditions is generally accepted as the event that is responsible for a series of long-term complications of diabetes such as retinopathy, cataract, nephropathy and neuropathy. The role of AKR1B1 in the onset of diabetic complications has made this enzyme the target for the development of molecules capable of inhibiting its activity. Virtually all synthesized compounds have so far failed as drugs for the treatment of diabetic complications. This failure may be partly due to the ability of AKR1B1 to reduce alkenals and alkanals, produced in oxidative stress conditions, thus acting as a detoxifying agent. In recent years we have proposed an alternative approach to the inhibition of AKR1B1, suggesting the possibility of a differential inhibition of the enzyme through molecules able to preferentially inhibit the reduction of either hydrophilic or hydrophobic substrates. The rationale and examples of this new generation of aldose reductase differential inhibitors (ARDIs) are presented.

In recent years, researchers have leveraged the ubiquitin-proteasome system (UPS) to induce selective degradation of proteins by E3 ubiquitin ligases, which has great potential as novel therapeutics for human diseases, including cancer and neurodegenerative disorders. However, despite extensive efforts, only a handful of ~600 human E3 ligases were utilized, and numerous protein-protein interaction surfaces on E3 ligases were not explored. To tackle these problems, we leveraged a structure-based protein engineering technology to develop a multi-domain fusion protein bringing functional E3 ligases to the proximity of a target protein to trigger its proteasomal degradation, which we termed Ubiquitin Variant Induced Proximity (UbVIP). We first generated non-inhibitory synthetic UbV binders for a selected group of human E3 ligases. With these UbVs employed as E3 ligase engagers, we designed a library of UbVIPs targeting a DNA damage response protein 53BP1. We observed that two UbVIPs recruiting RFWD3 and NEDD4L could effectively induce proteasome degradation of 53BP1 in human cell lines. This provides a proof-of-principle that UbVs can act as a means of targeted degradation for nucleus-localized proteins. Our work demonstrated that UbV technology is suitable to develop protein-based molecules for targeted degradation and can help identify novel E3 ligases for future therapeutic development.

Comment on Biomolecules. 2021 Aug 03;11(8): Biomolecules. 2021 Oct 14;11(10): Biomolecules. 2022 Jan 16;12(1): Biomolecules. 2022 Feb 23;12(3):

Epidemiological data suggest that various noncommunicable diseases develop as a result of altered maternal metabolic and physiological status due to exposure to several adverse factors during pregnancy. However, evidence for intrauterine exposure factors and mechanisms underlying the origin of early cartilage disease in chronic osteoarthritic disease is still lacking. In this study, we found that persistent overnutrition during pregnancy in obese mothers led to cartilage damage in neonatal male mice. This was mainly characterized by increased apoptosis with decreased expression of chondrocyte collagen II and low expression of Runx family transcription factor 2 (RUNX2) and SRY-box transcription factor 9 (SOX9). This reduction was also found to be associated with high leptin expression in newborn male mice of obese maternal offspring. Furthermore, the administration of leptin and mitogen-activated protein kinase (MAPK)-extracellular signal-regulated kinase (ERK) inhibitors in primary chondrocytes showed that leptin mediated MAPK/ERK signaling activation and thus affected the key regulators of cartilage matrix metallopeptidase 1 (MMP1) and tissue inhibitor of metalloproteinase 1 (TIMP1), thereby altering the expression of collagen II in mouse cartilage. Altogether, this study provided insights into the molecular mechanisms of cartilage-related disease development and also new clues and evidence for the fetogenetic origin of cartilage diseases.

In this study, circular RNAs (circRNAs) from Holstein cow mammary tissues were identified and compared between early lactation and non-lactation. After analysis, 10,684 circRNAs were identified, ranging from 48 to 99,406 bp, and the average size was 882 bp. The circRNAs were mainly distributed on chromosomes 1 to 11, and 89.89% of the circRNAs belonged to sense-overlapping circRNA. The exons contained with circRNAs ranged from 1 to 47 and were concentrated from 1 to 5. Compared with the non-lactating cows, 87 circRNAs were significantly differentially expressed in the peak lactation cows. There were 68 upregulated circRNAs and 19 downregulated circRNAs. Enrichment analysis of circRNAs showed that GO analysis mainly focused on immune response, triglyceride transport, T cell receptor signaling pathway, etc. Pathway analysis mainly focused on cytokine-cytokine receptor interaction, T helper 17 cell differentiation, fatty acid biosynthesis, the JAK-STAT signaling pathway, etc. Specific primers were designed for two proximal ends of the circRNA junction sites to allow for PCR validation of four randomly selected circRNAs and carry out circRNA-miRNA interaction research. This study revealed the expression profile and characteristics of circRNAs in mammary tissue from Holstein cows at early lactation and non-lactation, thus providing rich information for the study of circRNA functions and mechanisms, as well as potential candidate miRNA genes for studying lactation in Holstein cows.

Proteases and protease inhibitors (P/PIs) are involved in many biological processes in human skin, yet often only specific families or related groups of P/PIs are investigated. Proteomics approaches, such as mass spectrometry, can define proteome signatures (including P/PIs) in tissues; however, they struggle to detect low-abundance proteins. To overcome these issues, we aimed to produce a comprehensive proteome of all P/PIs present in normal and diseased human skin, in vivo, by carrying out a modified systematic review using a list of P/PIs from MEROPS and combining this with key search terms in Web of Science. Resulting articles were manually reviewed against inclusion/exclusion criteria and a dataset constructed. This study identified 111 proteases and 77 protease inhibitors in human skin, comprising the serine, metallo-, cysteine and aspartic acid catalytic families of proteases. P/PIs showing no evidence of catalytic activity or protease inhibition, were designated non-peptidase homologs (NPH), and no reported protease inhibitory activity (NRPIA), respectively. MMP9 and TIMP1 were the most frequently published P/PIs and were reported in normal skin and most skin disease groups. Normal skin and diseased skin showed significant overlap with respect to P/PI profile; however, MMP23 was identified in several skin disease groups, but was absent in normal skin. The catalytic profile of P/PIs in wounds, scars and solar elastosis was distinct from normal skin, suggesting that a different group of P/PIs is responsible for disease progression. In conclusion, this study uses a novel approach to provide a comprehensive inventory of P/PIs in normal and diseased human skin reported in our database. The database may be used to determine either which P/PIs are present in specific diseases or which diseases individual P/PIs may influence.

Melanocortin Receptor Accessory Protein 2 (MRAP2) modulates the trafficking and signal transduction of several G-protein-coupled receptors (GPCRs) involved in the control of energy homeostasis, such as Prokineticin receptors (PKRs). They bind the endogenous ligand prokineticin 2 (PK2), a novel adipokine that has an anorexic effect and modulates thermoregulation and energy homeostasis. In the present work, we used biochemical techniques to analyze the mechanism of interaction of MRAP2 with PKR2 and we identified the specific amino acid regions involved in the complex formation. Our results indicate that MRAP2 likely binds to the N-terminal region of PKR2, preventing glycosylation and consequently the correct receptor localization. We also identified a C-terminal region of MRAP2 that is critical for the interaction with PKR2. Consequently, we analyzed the role of the prokineticin transduction system in the regulation of MRAP2 expression in tissues involved in the control of food intake: at the central level, in hypothalamic explants, and at the peripheral level, in adipocytes. We demonstrated the modulation of MRAP2 expression by the prokineticin transduction system.

Adipose tissue and its regenerative products which are isolated with enzymatic or mechanical processing of the harvested fat have been studied in a wide range of degenerative diseases, including osteoarthritis of the knee and hip. Intra-articular injection of these products can provide symptomatic relief of pain and postpone surgery. However, their use in the treatment of thumb carpometacarpal joint (CMCJ) osteoarthritis is limited and just a few studies have been published on that topic. For this reason, a review of the literature was performed by a thorough search of eight terms using the Pubmed database. In total, seven human studies met the selection criteria, including case-control studies, case-series and one case report. In all studies, intra-articular injection of autologous fat in osteoarthritic thumb CMCJ provided reduction in pain and improvement in hand function. Grip and pinch strength showed variable results, from no change to significant improvement. Fat-processing techniques were based on centrifugation and mechanical homogenization but biological characterization of the injected cells was not performed in any study. Although the results are encouraging, a uniformly standardized method of fat processing and the conduction of randomized controlled trials in the future could better evaluate the effectiveness of this procedure for thumb CMCJ osteoarthritis.

Cardiovascular diseases (CVDs) are still a major cause of global mortality and disability, seriously affecting people's lives. Due to the severity and complexity of these diseases, it is important to find new regulatory mechanisms to treat CVDs. Ferroptosis is a new kind of regulatory cell death currently being investigated. Increasing evidence showed that ferroptosis plays an important role in CVDs, such as in ischemia/reperfusion injury, heart failure, cardiomyopathy, and atherosclerosis. Protecting against CVDs by targeting ferroptosis is a promising approach; therefore, in this review, we summarized the latest regulatory mechanism of ferroptosis and the current studies related to each CVD, followed by critical perspectives on the ferroptotic treatment of CVDs and the future direction of this intriguing biology.

Desmopressin (dDAVP) is the best characterized analogue of vasopressin, the endocrine regulator of water balance endowed with potent vasoconstrictive effects. Despite the use of dDAVP in clinical practice, ranging from the treatment of nephrogenic diabetes insipidus to bleeding disorders, much remains to be understood about the impact of the drug on endothelial phenotype. The aim of this study was, thus, to evaluate the effects of desmopressin on the viability and function of human pulmonary microvascular endothelial cells (HLMVECs). The results obtained demonstrate that the vasopressor had no cytotoxic effect on the endothelium; similarly, no sign of endothelial activation was induced by dDAVP, indicated by the lack of effect on the expression of inflammatory cytokines and adhesion molecules. Conversely, the drug significantly stimulated the production of nitric oxide (NO) and the expression of the inducible isoform of nitric oxide synthase, NOS2/iNOS. Since the intracellular level of cAMP also increased, we can hypothesize that NO release is consequent to the activation of the vasopressin receptor 2 (V2R)/guanylate cyclase (Gs)/cAMP axis. Given the multifaceted role of NOS2-deriving NO for many physio-pathological conditions, the meanings of these findings in HLMVECs appears intriguing and deserves to be further addressed.

Heat stress (HS) poses a significant threat to production and survival in the global swine industry. However, the molecular regulatory effects of heat stress on maternal endometrial cells are poorly understood in pigs during early embryo implantation. In this study, we systematically examined morphological changes in the endometrium and the corresponding regulation mechanism in response to HS by combining scanning electron microscopy (SEM), hematoxylin/eosin (H&E) staining, western blot, and RNA-seq analyses. Our results showed that HS led to porcine endometrium damage and endometrial thinness during embryo implantation. The expression levels of cell adhesion-related proteins, including N-cadherin and E-cadherin, in the uterus were significantly lower in the heat stress group (39 ± 1 °C, n = 3) than in the control group (28 ± 1 °C, n = 3). A total of 338 up-regulated genes and 378 down-regulated genes were identified in porcine endometrium under HS. The down-regulated genes were found to be mainly enriched in the pathways related to the microtubule complex, immune system process, and metalloendopeptidase activity, whereas the up-regulated genes were mainly involved in calcium ion binding, the extracellular region, and molecular function regulation. S100A9 was found to be one of the most significant differentially expressed genes (DEGs) in the endometrium under HS, and this gene could promote proliferation of endometrial cells and inhibit their apoptosis. Meanwhile, HS caused endometrial epithelial cell (EEC) damage and inhibited its proliferation. Overall, our results demonstrated that HS induced uterine morphological change and tissue damage by regulating the expression of genes associated with calcium ions and amino acid transport. These findings may provide novel molecular insights into endometrial damage under HS during embryo implantation.

Extracellular matrix (ECM)-based bioinks have been steadily gaining interest in the field of bioprinting to develop biologically relevant and functional tissue constructs. Herein, we propose the use of supercritical carbon dioxide (scCO2) technology to extract the ECM components of cell-sheets that have shown promising results in creating accurate 3D microenvironments replicating the cell's own ECM, to be used in the preparation of bioinks. The ECM extraction protocol best fitted for cell sheets was defined by considering efficient DNA removal with a minor effect on the ECM. Cell sheets of human dermal fibroblasts (hDFbs) and adipose stem cells (hASCs) were processed using a customised supercritical system by varying the pressure of the reactor, presence, exposure time, and type of co-solvent. A quantification of the amount of DNA, protein, and sulfated glycosaminoglycans (sGAGs) was carried out to determine the efficiency of the extraction in relation to standard decellularization methodologies. The bioinks containing the extracted ECM were fabricated by combining them with alginate as a support polymer. The influence of the alginate (1%, 2% w/vol) and ECM (0.5% and 1.5% w/vol) amounts on the printability of the blends was addressed by analysing the rheological behaviour of the suspensions. Finally, 3D printed constructs were fabricated using an in-house built extrusion-based bioprinter, and the impact of the extrusion process on cell viability was assessed. The optimised scCO2 protocol allowed efficient removal of DNA while preserving a higher number of proteins and sGAGs than the standard methodologies. The characterization of extract's composition also revealed that the ECM produced by hDFbs (fECM) and hASCs (aECM) is distinctively affected by the extraction protocols. Furthermore, rheological analysis indicated an increase in viscosity with increasing ECM composition, an effect even more prominent in samples containing aECM. 3D printing of alginate/ECM constructs demonstrated that cell viability was only marginally affected by the extrusion process, and this effect was also dependent on the ECM source. Overall, this work highlights the benefits of supercritical fluid-based methods for ECM extraction and strengthens the relevance of ECM-derived bioinks in the development of printed tissue-like constructs.

Most individuals diagnosed with alcohol use disorders smoke cigarettes. Large concentrations of malondialdehyde and acetaldehyde are found in lungs co-exposed to cigarette smoke and alcohol. Aldehydes directly injure lungs and form aldehyde protein adducts, impacting epithelial functions. Recently, 2-(3-Amino-6-chloroquinolin-2-yl)propan-2-ol (ADX-102) was developed as an aldehyde-trapping drug. We hypothesized that aldehyde-trapping compounds are protective against lung injury derived from cigarette smoke and alcohol co-exposure. To test this hypothesis, we pretreated mouse ciliated tracheal epithelial cells with 0-100 µM of ADX-102 followed by co-exposure to 5% cigarette smoke extract and 50 mM of ethanol. Pretreatment with ADX-102 dose-dependently protected against smoke and alcohol induced cilia-slowing, decreases in bronchial epithelial cell wound repair, decreases in epithelial monolayer resistance, and the formation of MAA adducts. ADX-102 concentrations up to 100 µM showed no cellular toxicity. As protein kinase C (PKC) activation is a known mechanism for slowing cilia and wound repair, we examined the effects of ADX-102 on smoke and alcohol induced PKC epsilon activity. ADX-102 prevented early (3 h) activation and late (24 h) autodownregulation of PKC epsilon in response to smoke and alcohol. These data suggest that reactive aldehydes generated from cigarette smoke and alcohol metabolism may be potential targets for therapeutic intervention to reduce lung injury.

Tumor-associated macrophages (TAMs) promote tumor proliferation, invasion, angiogenesis, stemness, therapeutic resistance, and immune tolerance in a protein-dependent manner. Therefore, the traditional target paradigms are often insufficient to exterminate tumor cells. These pro-tumoral functions are mediated by the subsets of macrophages that exhibit canonical protein markers, while simultaneously having unique transcriptional features, which makes the proteins expressed on TAMs promising targets during anti-tumor therapy. Herein, TAM-associated protein-dependent target strategies were developed with the aim of either reducing the numbers of TAMs or inhibiting the pro-tumoral functions of TAMs. Furthermore, the recent advances in TAMs associated with tumor metabolism and immunity were extensively exploited to repolarize these TAMs to become anti-tumor elements and reverse the immunosuppressive tumor microenvironment. In this review, we systematically summarize these current studies to fully illustrate the TAM-associated protein targets and their inhibitors, and we highlight the potential clinical applications of targeting the crosstalk among TAMs, tumor cells, and immune cells in anti-tumor therapy.

BACKGROUND: Collagen type XI α1 (COL11A1) is associated with tumorigenesis and development in many human malignancies. Previous reports indicate that COL11A1 may be a significant diagnostic marker for pancreatic ductal adenocarcinoma (PDAC); however, its biological role in PDAC progression remains unclear. In this study, we investigated the influence of COL11A1 on the invasion and migration abilities of pancreatic cancer cells and explored its potential molecular mechanisms. METHODS: Cell migration and invasion were assessed using Transwell assays in pancreatic cancer cells transfected with siCOL11A1 and pCNV3-COL11A1 plasmids. The protein and mRNA expression levels of N-cadherin, E-cadherin, Vimentin, cluster of differentiation (CD)-24, CD44, serine-threonine kinase (AKT), glycogen synthase kinase (GSK)-3β, phospho (p)-AKTSer473, p-GSK-3βSer9, and Snail were analyzed using Western blotting and real-time polymerase chain reaction (PCR). The effect of COL11A1 on cell stemness was tested using flow cytometry and clone formation assays. RESULTS: These results demonstrated that COL11A1 significantly promoted the invasion and migration abilities of PDAC cells. Furthermore, COL11A1 facilitated the occurrence of epithelial-mesenchymal transition (EMT) and cell stemness by upregulating the expression levels of p-AKTSer473, p-GSK-3βSer9, and Snail. CONCLUSIONS: This study suggests that the activation of the AKT/GSK-3β/Snail signaling pathway induced by COL11A1 plays a major role in the progression of PDAC. Therefore, COL11A1 could serve as a potential target for PDAC treatment.

Eukaryotic cells monitor and regulate metabolism through the atypical protein kinase target of rapamycin (TOR) regulatory hub. TOR is activated by amino acids in animals and fungi through molecular signaling pathways that have been extensively defined in the past ten years. Very recently, several studies revealed that TOR is also acutely responsive to amino acid metabolism in plants, but the mechanisms of amino acid sensing are not yet established. In this review, we summarize these discoveries, emphasizing the diversity of amino acid sensors in human cells and highlighting pathways that are indirectly sensitive to amino acids, i.e., how TOR monitors changes in amino acid availability without a bona fide amino acid sensor. We then discuss the relevance of these model discoveries to plant biology. As plants can synthesize all proteinogenic amino acids from inorganic precursors, we focus on the possibility that TOR senses both organic metabolites and inorganic nutrients. We conclude that an evolutionary perspective on nutrient sensing by TOR benefits both agricultural and biomedical science, contributing to ongoing efforts to generate crops for a sustainable agricultural future.

Glucose oxidase (GOx) is an important oxidoreductase enzyme with many important roles in biological processes. It is considered an "ideal enzyme" and is often called an oxidase "Ferrari" because of its fast mechanism of action, high stability and specificity. Glucose oxidase catalyzes the oxidation of β-d-glucose to d-glucono-δ-lactone and hydrogen peroxide in the presence of molecular oxygen. d-glucono-δ-lactone is sequentially hydrolyzed by lactonase to d-gluconic acid, and the resulting hydrogen peroxide is hydrolyzed by catalase to oxygen and water. GOx is presently known to be produced only by fungi and insects. The current main industrial producers of glucose oxidase are Aspergillus and Penicillium. An important property of GOx is its antimicrobial effect against various pathogens and its use in many industrial and medical areas. The aim of this review is to summarize the structure, function, production strains and biophysical and biochemical properties of GOx in light of its various industrial, biotechnological and medical applications.

Cystic fibrosis (CF) is a rare genetic disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR), an epithelial anion channel expressed in several vital organs. Absence of functional CFTR results in imbalanced osmotic equilibrium and subsequent mucus build up in the lungs-which increases the risk of infection and eventually causes death. CFTR is an ATP-binding cassette (ABC) transporter family protein composed of two transmembrane domains (TMDs), two nucleotide binding domains (NBDs), and an unstructured regulatory domain. The most prevalent patient mutation is the deletion of F508 (F508del), making F508del CFTR the primary target for current FDA approved CF therapies. However, no experimental multi-domain F508del CFTR structure has been determined and few studies have modeled F508del using multi-domain WT CFTR structures. Here, we used cryo-EM density data and Rosetta comparative modeling (RosettaCM) to compare a F508del model with published experimental data on CFTR NBD1 thermodynamics. We then apply this modeling method to generate multi-domain WT and F508del CFTR structural models. These models demonstrate the destabilizing effects of F508del on NBD1 and the NBD1/TMD interface in both the inactive and active conformation of CFTR. Furthermore, we modeled F508del/R1070W and F508del bound to the CFTR corrector VX-809. Our models reveal the stabilizing effects of VX-809 on multi-domain models of F508del CFTR and pave the way for rational design of additional drugs that target F508del CFTR for treatment of CF.

Ubiquitination is controlled by a series of E1, E2, and E3 enzymes that can ligate ubiquitin to cellular proteins and dictate the turnover of a substrate and the outcome of signalling events such as DNA damage repair and cell cycle. This process is complex due to the combinatorial power of ~35 E2 and ~1000 E3 enzymes involved and the multiple lysine residues on ubiquitin that can be used to assemble polyubiquitin chains. Recently, mass spectrometric methods have identified that most enzymes in the ubiquitination cascade can be further modified through acetylation or phosphorylation under particular cellular conditions and altered modifications have been noted in different cancers and neurodegenerative diseases. This review provides a cohesive summary of ubiquitination, acetylation, and phosphorylation sites in ubiquitin, the human E1 enzyme UBA1, all E2 enzymes, and some representative E3 enzymes. The potential impacts these post-translational modifications might have on each protein function are highlighted, as well as the observations from human disease.

Methods for glucagon analysis suffered in the past from lack of specificity and a narrow sensitivity range, which has led to inaccurate results and to the suggestion that type 1 diabetes (T1D) and type 2 diabetes (T2D) patients have elevated fasting glucagon levels. However, the availability of more specific and more sensitive methods to detect intact glucagon has shown that actual glucagon levels are lower than previously assumed. This study aimed to characterize fasting plasma glucagon levels in healthy individuals and T1D and T2D patients with two different glucagon assays. The study included 20 healthy individuals, 20 T1D and 20 T2D patients. Blood was collected under fasting conditions. A double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) and a conventional radioimmunoassay (RIA) were used. A significant difference in fasting glucagon levels between healthy individuals and T2D was observed by ELISA, but not by RIA. ELISA also yielded lower glucagon levels in healthy individuals than in T1D and T2D patients which RIA did not. RIA produced significantly (p = 0.0001) higher overall median glucagon values than ELISA in a pooled analysis. These results underline the notion that the choice of selective laboratory methods is highly relevant for mechanistic endocrine research.

In chronic hepatitis B virus (HBV) infection, virus-specific T cells are scarce and partially dysfunctional. Therapeutic vaccination is a promising strategy to induce and activate new virus-specific T cells. In long-term or high-level HBV carriers, however, therapeutic vaccination by itself may not suffice to cure HBV. One reason is the impairment of antiviral T cells by immune checkpoints. In this study, we used small-interfering RNA (siRNA) in combination with a heterologous prime-boost therapeutic vaccination scheme (TherVacB) to interfere with a major immune checkpoint, the interaction of programmed death protein-1 (PD-1) and its ligand (PDL-1). In mice persistently replicating HBV after infection with an adeno-associated virus harboring the HBV genome, siRNA targeting PD-L1 resulted in a higher functionality of HBV-specific CD8+ T cells after therapeutic vaccination, and allowed for a more sustained antiviral effect and control of HBV in peripheral blood and in the liver. The antiviral effect was more pronounced if PD-L1 was down-regulated during prime than during boost vaccination. Thus, targeting PD-L1 using siRNA is a promising approach to enhance the efficacy of therapeutic vaccination and finally cure HBV.

Parkinson's disease (PD) is associated with motor and non-motor symptoms and characterized by aggregates of alpha-synuclein (αSyn). Naturally occurring antibodies (nAbs) are part of the innate immune system, produced without prior contact to their specific antigen, and polyreactive. The abundance of nAbs against αSyn is altered in patients with PD. In this work, we biophysically characterized nAbs against αSyn (nAbs-αSyn) and determined their biological effects. nAbs-αSyn were isolated from commercial intravenous immunoglobulins using column affinity purification. Biophysical properties were characterized using a battery of established in vitro assays. Biological effects were characterized in HEK293T cells transiently transfected with fluorescently tagged αSyn. Specific binding of nAbs-αSyn to monomeric αSyn was demonstrated by Dot blot, ELISA, and Surface Plasmon Resonance. nAbs-αSyn did not affect viability of HEK293T cells as reported by Cell Titer Blue and LDH Assays. nAbs-αSyn inhibited fibrillation of αSyn reported by the Thioflavin T aggregation assay. Altered fibril formation was confirmed with atomic force microscopy. In cells transfected with EGFP-tagged αSyn we observed reduced formation of aggresomes, perinuclear accumulations of αSyn aggregates. The results demonstrate that serum of healthy individuals contains nAbs that specifically bind αSyn and inhibit aggregation of αSyn in vitro. The addition of nAbs-αSyn to cultured cells affects intracellular αSyn aggregates. These findings help understanding the role of the innate immune systems for the pathogenesis of PD and suggest that systemic αSyn binding agents could potentially affect neuronal αSyn pathology.

For single-molecule studies in solution, very small concentrations of dye-labelled molecules are employed in order to achieve single-molecule sensitivity. In typical studies with confocal microscopes, often concentrations in the pico-molar regime are required. For various applications that make use of single-molecule Förster resonance energy transfer (smFRET) or two-color coincidence detection (TCCD), the molecule concentration must be set explicitly to targeted values and furthermore needs to be stable over a period of several hours. As a consequence, specific demands must be imposed on the surface passivation of the cover slides during the measurements. The aim of having only one molecule in the detection volume at the time is not only affected by the absolute molecule concentration, but also by the rate of diffusion. Therefore, we discuss approaches to control and to measure absolute molecule concentrations. Furthermore, we introduce an approach to calculate the probability of chance coincidence events and demonstrate that measurements with challenging smFRET samples require a strict limit of maximal sample concentrations in order to produce meaningful results.

Relapse after surgery for oral squamous cell carcinoma (OSCC) contributes significantly to morbidity, mortality and poor outcomes. The current histopathological diagnostic techniques are insufficiently sensitive for the detection of oral cancer and minimal residual disease in surgical margins. We used whole-transcriptome gene expression and small noncoding RNA profiles from tumour, close margin and distant margin biopsies from 18 patients undergoing surgical resection for OSCC. By applying multivariate regression algorithms (sPLS-DA) suitable for higher dimension data, we objectively identified biomarker signatures for tumour and marginal tissue zones. We were able to define molecular signatures that discriminated tumours from the marginal zones and between the close and distant margins. These signatures included genes not previously associated with OSCC, such as MAMDC2, SYNPO2 and ARMH4. For discrimination of the normal and tumour sampling zones, we were able to derive an effective gene-based classifying model for molecular abnormality based on a panel of eight genes (MMP1, MMP12, MYO1B, TNFRSF12A, WDR66, LAMC2, SLC16A1 and PLAU). We demonstrated the classification performance of these gene signatures in an independent validation dataset of OSCC tumour and marginal gene expression profiles. These biomarker signatures may contribute to the earlier detection of tumour cells and complement existing surgical and histopathological techniques used to determine clear surgical margins.

Epigenetic mechanisms are known to play a key role in cancer progression. Specifically, histone methylation involves reversible post-translational modification of histones that govern chromatin structure remodelling, genomic imprinting, gene expression, DNA damage repair, and meiotic crossover recombination, among other chromatin-based activities. Demethylases are enzymes that catalyse the demethylation of their substrate using a flavin adenine dinucleotide-dependent amine oxidation process. Lysine-specific demethylase 1 (LSD1) and its homolog, lysine-specific demethylase 2 (LSD2), are overexpressed in a variety of human cancer types and, thus, regulate tumour progression. In this review, we focus on the literature from the last 5 years concerning the role of LSD1 and LSD2 in the main gastrointestinal cancers (i.e., gastric cancer, liver cancer, pancreatic cancer, and colorectal cancer).

RNA editing contributes to transcriptome diversification through RNA modifications in relation to genome-encoded information (RNA-DNA differences, RDDs). The deamination of Adenosine (A) to Inosine (I) or Cytidine (C) to Uridine (U) is the most common type of mammalian RNA editing. It occurs as a nuclear co- and/or post-transcriptional event catalyzed by ADARs (Adenosine deaminases acting on RNA) and APOBECs (apolipoprotein B mRNA editing enzyme catalytic polypeptide-like genes). RNA editing may modify the structure, stability, and processing of a transcript. This review focuses on RNA editing in psychiatric, neurological, neurodegenerative (NDs), and autoimmune brain disorders in humans and rodent models. We discuss targeted studies that focus on RNA editing in specific neuron-enriched transcripts with well-established functions in neuronal activity, and transcriptome-wide studies, enabled by recent technological advances. We provide comparative editome analyses between human disease and corresponding animal models. Data suggest RNA editing to be an emerging mechanism in disease development, displaying common and disease-specific patterns. Commonly edited RNAs represent potential disease-associated targets for therapeutic and diagnostic values. Currently available data are primarily descriptive, calling for additional research to expand global editing profiles and to provide disease mechanistic insights. The potential use of RNA editing events as disease biomarkers and available tools for RNA editing identification, classification, ranking, and functional characterization that are being developed will enable comprehensive analyses for a better understanding of disease(s) pathogenesis and potential cures.

Cisplatin has long been a first-line chemotherapeutic agent in the treatment of cancer, largely for solid tumors. During the course of the past two decades, autophagy has been identified in response to cancer treatments and almost uniformly detected in studies involving cisplatin. There has been increasing recognition of autophagy as a critical factor affecting tumor cell death and tumor chemoresistance. In this review and commentary, we introduce four mechanisms of resistance to cisplatin followed by a discussion of the factors that affect the role of autophagy in cisplatin-sensitive and resistant cells and explore the two-sided outcomes that occur when autophagy inhibitors are combined with cisplatin. Our goal is to analyze the potential for the combinatorial use of cisplatin and autophagy inhibitors in the clinic.

Nuclear factor erythroid 2 (NF-E2) p45-related factor 2 (NRF2) protein is the master regulator of oxidative stress, which is at the basis of various chronic diseases including cancer. Hyperactivation of NRF2 in already established cancers can promote cell proliferation and resistance to therapies, such as in colorectal cancer (CRC), one of the most lethal and prevalent malignancies in industrialized countries with limited patient overall survival due to its escape mechanisms in both chemo- and targeted therapies. In this study, we generated stable NRF2 knockout colon cancer cells (NRF2-Cas9) to investigate the cell response to chemotherapeutic drugs with regard to p53 oncosuppressor, whose inhibition we previously showed to correlate with NRF2 pathway activation. Here, we found that NRF2 activation by sulforaphane (SFN) reduced cisplatin (CDDP)-induced cell death only in NRF2-proficient cells (NRF2-ctr) compared to NRF2-Cas9 cells. Mechanistically, we found that NRF2 activation protected NRF2-ctr cells from the drug-induced DNA damage and the apoptotic function of the unfolded protein response (UPR), in correlation with reduction of p53 activity, effects that were not observed in NRF2-Cas9 cells. Finally, we found that ZnCl2 supplementation rescued the cisplatin cytotoxic effects, as it impaired NRF2 activation, restoring p53 activity. These findings highlight NRF2's key role in neutralizing the cytotoxic effects of chemotherapeutic drugs in correlation with reduced DNA damage and p53 activity. They also suggest that NRF2 inhibition could be a useful strategy for efficient anticancer chemotherapy and support the use of ZnCl2 to inhibit NRF2 pathway in combination therapies.

The heart is a hierarchical dynamic system consisting of molecules, cells, and tissues, and acts as a pump for blood circulation. The pumping function depends critically on the preceding electrical activity, and disturbances in the pattern of excitation propagation lead to cardiac arrhythmia and pump failure. Excitation phenomena in cardiomyocytes have been modeled as a nonlinear dynamical system. Because of the nonlinearity of excitation phenomena, the system dynamics could be complex, and various analyses have been performed to understand the complex dynamics. Understanding the mechanisms underlying proarrhythmic responses in the heart is crucial for developing new ways to prevent and control cardiac arrhythmias and resulting contractile dysfunction. When the heart changes to a pathological state over time, the action potential (AP) in cardiomyocytes may also change to a different state in shape and duration, often undergoing a qualitative change in behavior. Such a dynamic change is called bifurcation. In this review, we first summarize the contribution of ion channels and transporters to AP formation and our knowledge of ion-transport molecules, then briefly describe bifurcation theory for nonlinear dynamical systems, and finally detail its recent progress, focusing on the research that attempts to understand the developing mechanisms of abnormal excitations in cardiomyocytes from the perspective of bifurcation phenomena.

Tau aggregates accumulate in the Alzheimer's disease (AD) brain according to the established Braak staging scheme and spread from transentorhinal over limbic regions to the neocortex. To impact the management of AD patients, an in vivo tool for tau Braak staging is needed. First-generation tau tracers have limited performance in detecting early stages of tau. Therefore, we tested the corresponding capability of the next-generation tau tracer, 18F-PI-2620. We analyzed 18F-PI-2620 multicenter PET data from 37 beta-amyloid-positive AD dementia patients and those from 26 healthy controls. We applied kinetic modeling of the 0-60 min p.i. PET data using MRTM2 with the lower cerebellum as the reference region to extract Braak stage-dependent distribution volume ratios, whereas controls were used to define Braak stage PET positivity thresholds. Stage-dependent PET positivity widely followed the Braak scheme (except Braak stage III) presenting descending frequency of PET positivity from Braak I (43%), II (38%), III (49%), IV (35%), V (30%) to VI (14%). A strictly hierarchical model was met by 64% of AD dementia cases. Nineteen percent showed a hippocampal sparing tauopathy pattern. Thus, we could assign 87% to the six-stage hierarchical Braak model including tauopathy variants. 18F-PI-2620 PET appears to be able to perform Braak tau staging of AD in vivo.

Myelination of the peripheral nervous system requires Schwann cells (SC) differentiation into the myelinating phenotype. The peripheral myelin protein-22 (PMP22) is an integral membrane glycoprotein, expressed in SC. It was initially described as a growth arrest-specific (gas3) gene product, up-regulated by serum starvation. PMP22 mutations were pathognomonic for human hereditary peripheral neuropathies, including the Charcot-Marie-Tooth disease (CMT). Trembler-J (TrJ) is a heterozygous mouse model carrying the same pmp22 point mutation as a CMT1E variant. Mutations in lamina genes have been related to a type of peripheral (CMT2B1) or central (autosomal dominant leukodystrophy) neuropathy. We explore the presence of PMP22 and Lamin B1 in Wt and TrJ SC nuclei of sciatic nerves and the colocalization of PMP22 concerning the silent heterochromatin (HC: DAPI-dark counterstaining), the transcriptionally active euchromatin (EC), and the nuclear lamina (H3K4m3 and Lamin B1 immunostaining, respectively). The results revealed that the number of TrJ SC nuclei in sciatic nerves was greater, and the SC volumes were smaller than those of Wt. The myelin protein PMP22 and Lamin B1 were detected in Wt and TrJ SC nuclei and predominantly in peripheral nuclear regions. The level of PMP22 was higher, and those of Lamin B1 lower in TrJ than in Wt mice. The level of PMP22 was higher, and those of Lamin B1 lower in TrJ than in Wt mice. PMP22 colocalized more with Lamin B1 and with the transcriptionally competent EC, than the silent HC with differences between Wt and TrJ genotypes. The results are discussed regarding the probable nuclear role of PMP22 and the relationship with TrJ neuropathy.

The objective of this study was to investigate the vasoregulatory role of perivascular adipose tissue (PVAT) and its mutual interaction with endogenous and exogenous H2S in the thoracic aorta (TA) of adult normotensive Wistar rats and spontaneously hypertensive rats (SHRs). In SHRs, hypertension was associated with cardiac hypertrophy and increased contractility. Regardless of the strain, PVAT revealed an anticontractile effect; however, PVAT worsened endothelial-dependent vasorelaxation. Since H2S produced by both the vascular wall and PVAT had a pro-contractile effect in SHRs, H2S decreased the sensitivity of adrenergic receptors to noradrenaline in Wistar rats. While H2S had no contribution to endothelium-dependent relaxation in Wistar rats, in SHRs, H2S produced by the vascular wall had a pro-relaxant effect. We observed a larger vasorelaxation induced by exogenous H2S donor in SHRs than in Wistar rats. Additionally, in the presence of PVAT, this effect was potentiated. We demonstrated that PVAT of the TA aggravated endothelial function in SHRs. However, H2S produced by the TA vascular wall had a pro-relaxation effect, and PVAT revealed anti-contractile activity mediated by the release of an unknown factor and potentiated the vasorelaxation induced by exogenous H2S. All these actions could represent a form of compensatory mechanism to balance impaired vascular tone regulation.

Hyperglycaemia and its resulting glucotoxicity are among the most prominent hallmarks of diabetes mellitus (DM) development. Persistent hyperglycaemia further leads to oxidative stress via mitochondrial dysfunction and subsequent ER stress onset, while associated hyperlipidaemia triggers the adipose tissue to secrete pro-inflammatory cytokines. In this study, the effect of calystegines has been investigated in an experimental model of hyperglycaemia induced on human ASCs cells. Different cellular pathways including apoptosis, oxidative and ER stress, inflammation as well as Pi3K/AKT/mTOR metabolic-associated axis have been evaluated by means on RT-qPCR, western blot, and flow cytometry techniques. Treatment of HuASCs cells with calystegines strongly promoted the hyperglycaemic cells survival and significantly diminished oxidative stress, mitochondrial dynamics failure and ER stress, while improving the endogenous cellular antioxidant defenses. Interestingly, nortropane alkaloids efficiently prevented the hyperglycaemia-mediated inflammatory response, as evidenced by the regulation of the pro- and anti-inflammatory response in HuASCs cells. Finally, we evidenced that calystegines may exert their protective effect on HuASCs cells metabolic functions through the restoration of the defective PI3K/AKT/mTOR pathway. Overall, the present investigation demonstrated that calystegines possess important abilities to protect HuASCs against hyperglycaemia-induced cellular dysfunction, and it evidenced that the observed effects are associated to the promotion of PI3K/AKT/mTOR pathway.

Targeting cancer hallmarks is a cardinal strategy to improve antineoplastic treatment. However, cross-talk between signaling pathways and key oncogenic processes frequently convey resistance to targeted therapies. The p53 and Wnt pathway play vital roles for the biology of many tumors, as they are critically involved in cancer onset and progression. Over recent decades, a high level of interaction between the two pathways has been revealed. Here, we provide a comprehensive overview of molecular interactions between the p53 and Wnt pathway discovered in cancer, including complex feedback loops and reciprocal transactivation. The mutational landscape of genes associated with p53 and Wnt signaling is described, including mutual exclusive and co-occurring genetic alterations. Finally, we summarize the functional consequences of this cross-talk for cancer phenotypes, such as invasiveness, metastasis or drug resistance, and discuss potential strategies to pharmacologically target the p53-Wnt interaction.

With recent technical advances and diminishing sequencing costs, single-cell sequencing modalities have become commonplace. These tools permit analysis of RNA expression, DNA sequence, chromatin structure, and cell surface antigens at single-cell resolution. Simultaneous measurement of numerous parameters can resolve populations including rare cells, thus revealing cellular diversity within organs and permitting lineage reconstruction in developing tissues. Application of these methods to the enteric nervous system has yielded a wealth of data and biological insights. We review recent papers applying single-cell sequencing tools to the nascent neural crest and to the developing and mature enteric nervous system. These studies have shown significant diversity of enteric neurons and glia, suggested paradigms for neuronal specification, and revealed signaling pathways active during development. As technology evolves and multiome techniques combining two or more of transcriptomic, genomic, epigenetic, and proteomic data become prominent, we anticipate these modalities will become commonplace in ENS research and may find a role in diagnostic testing and personalized therapeutics.

Inherited retinal degenerations (IRDs) are a group of blinding diseases, typically involving a progressive loss of photoreceptors. The IRD pathology is often based on an accumulation of cGMP in photoreceptors and associated with the excessive activation of calpain and poly (ADP-ribose) polymerase (PARP). Inhibitors of calpain or PARP have shown promise in preventing photoreceptor cell death, yet the relationship between these enzymes remains unclear. To explore this further, organotypic retinal explant cultures derived from wild-type and IRD-mutant mice were treated with inhibitors specific for calpain, PARP, and voltage-gated Ca2+ channels (VGCCs). The outcomes were assessed using in situ activity assays for calpain and PARP and immunostaining for activated calpain-2, poly (ADP-ribose), and cGMP, as well as the TUNEL assay for cell death detection. The IRD models included the Pde6b-mutant rd1 mouse and rd1*Cngb1-/- double-mutant mice, which lack the beta subunit of the rod cyclic nucleotide-gated (CNG) channel and are partially protected from rd1 degeneration. We confirmed that an inhibition of either calpain or PARP reduces photoreceptor cell death in rd1 retina. However, while the activity of calpain was decreased by the inhibition of PARP, calpain inhibition did not alter the PARP activity. A combination treatment with calpain and PARP inhibitors did not synergistically reduce cell death. In the slow degeneration of rd1*Cngb1-/- double mutant, VGCC inhibition delayed photoreceptor cell death, while PARP inhibition did not. Our results indicate that PARP acts upstream of calpain and that both are part of the same degenerative pathway in Pde6b-dependent photoreceptor degeneration. While PARP activation may be associated with CNG channel activity, calpain activation is linked to VGCC opening. Overall, our data highlights PARP as a target for therapeutic interventions in IRD-type diseases.

Bacterial extracellular proteins participate in the host cell communication by virtue of the modulation of pathogenicity, commensalism and mutualism. Studies on the microbiome of cervical mucus of the water buffalo (Bubalus bubalis) have shown the occurrence of Staphylococcus pasteuri and that the presence of this bacterium is indicative of various physiological and reproductive states in the host. Recently, S. pasteuri has been isolated from the cervical mucus of the buffalo during the different phases of estrous cycle, and has proved to be much more pronounced during the estrus phase. The basis underlying the availability of a significantly increased S. pasteuri population, specifically during the estrus phase, is not known. Consequently, it is important to determine the significance of the specific abundance of S. pasteuri during the estrus phase of the buffalo host, particularly from the perspective of whether this bacterial species is capable of contributing to sexual communication via its extracellular proteins and volatiles. Therefore, the relevance of S. pasteuri exoproteome in the buffalo cervical mucus during the estrus phase was analyzed using LC-MS/MS. As many as 219 proteins were identified, among which elongation factor Tu (EF-Tu), 60-kDa chaperonin (Cpn60), enolase, fructose-bisphosphate aldolase class 1 (FBP aldolase), enoyl-[acyl-carrier-protein] reductase [NADPH] (ENR) and lipoprotein (Lpp) were the functionally important candidates. Most of the proteins present in the exoproteome of S. pasteuri were those involved in cellular-metabolic functions, as well as catalytic- and binding activities. Moreover, computational studies of Lpp have shown enhanced interaction with volatiles such as acetic-, butanoic-, isovaleric- and valeric acids, which were identified in the cervical mucus S. pasteuri culture supernatant. The present findings suggest that S. pasteuri extracellular proteins may play an important role in buffalo sexual communication during the estrus phase.

Histamine intolerance (HIT) is a common disorder associated with impaired histamine metabolism. Notwithstanding, it is often misdiagnosed as other diseases because of its lack of specific clinical manifestations. HIT did not gain traction until the early 21st century. In this review, we will focus on the latest research and elaborate on the clinical manifestations of HIT, including its manifestations in special populations such as atopic dermatitis (AD) and chronic urticaria (CU), as well as the latest understanding of its etiology and pathogenesis. In addition, we will explore the latest treatment strategies for HIT and the treatment of specific cases.

Dementia-a syndrome affecting human cognition-is a major public health concern given to its rising prevalence worldwide. Though multiple research studies have analyzed disorders such as Alzheimer's disease and Frontotemporal dementia using a systems biology approach, a similar approach to dementia syndrome as a whole is required. In this study, we try to find the high-impact core regulating processes and factors involved in dementia's protein-protein interaction network. We also explore various aspects related to its stability and signal propagation. Using gene interaction databases such as STRING and GeneMANIA, a principal dementia network (PDN) consisting of 881 genes and 59,085 interactions was achieved. It was assortative in nature with hierarchical, scale-free topology enriched in various gene ontology (GO) categories and KEGG pathways, such as negative and positive regulation of apoptotic processes, macroautophagy, aging, response to drug, protein binding, etc. Using a clustering algorithm (Louvain method of modularity maximization) iteratively, we found a number of communities at different levels of hierarchy in PDN consisting of 95 "motif-localized hubs", out of which, 7 were present at deepest level and hence were key regulators (KRs) of PDN (HSP90AA1, HSP90AB1, EGFR, FYN, JUN, CELF2 and CTNNA3). In order to explore aspects of network's resilience, a knockout (of motif-localized hubs) experiment was carried out. It changed the network's topology from a hierarchal scale-free topology to scale-free, where independent clusters exhibited greater control. Additionally, network experiments on interaction of druggable genome and motif-localized hubs were carried out where UBC, EGFR, APP, CTNNB1, NTRK1, FN1, HSP90AA1, MDM2, VCP, CTNNA1 and GRB2 were identified as hubs in the resultant network (RN). We finally concluded that stability and resilience of PDN highly relies on motif-localized hubs (especially those present at deeper levels), making them important therapeutic intervention candidates. HSP90AA1, involved in heat shock response (and its master regulator, i.e., HSF1), and EGFR are most important genes in pathology of dementia apart from KRs, given their presence as KRs as well as hubs in RN.

Signal transducer and activator of transcription 6 (STAT6) promotes an anti-inflammatory process by inducing the development of M2 macrophages. We investigated whether modulating STAT6 activity in macrophages using AS1517499, the specific STAT6 inhibitor, affects the restoration of homeostasis after an inflammatory insult by regulating PPARγ expression and activity. Administration of AS1517499 suppressed the enhanced STAT6 phosphorylation and nuclear translocation observed in peritoneal macrophages after zymosan injection. In addition, AS1517499 delayed resolution of acute inflammation as evidenced by enhanced secretion of pro-inflammatory cytokines, reduced secretion of anti-inflammatory cytokines in PLF and supernatants from peritoneal macrophages, and exaggerated neutrophil numbers and total protein levels in PLF. We demonstrate temporal increases in annexin A1 (AnxA1) protein and mRNA levels in peritoneal lavage fluid (PLF), peritoneal macrophages, and spleen in a murine model of zymosan-induced acute peritonitis. In vitro priming of mouse bone marrow-derived macrophages (BMDM) and peritoneal macrophages with AnxA1 induced STAT6 activation with enhanced PPARγ expression and activity. Using AS1517499, we demonstrate that inhibition of STAT6 activation delayed recovery of PPARγ expression and activity, as well as impaired efferocytosis. Taken together, these results suggest that activation of the STAT6 signaling pathway mediates PPARγ expression and activation in macrophages to resolve acute inflammation.

BACKGROUND: COVID-19 is an infectious disease caused by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Many COVID-19 patients require invasive mechanical ventilation (IMV) while others, even with acute respiratory failure, do not (NIMV). Therefore, we aimed to evaluate serum levels of MMP-7 and molecules related to exhausted T-cells as potential biomarkers to differentiate between IMV and NIMV patients. METHODS: 105 patients diagnosed with COVID-19 and confirmed by RT-PCR for SARS-CoV-2 were divided into two groups according to the requirement for IMV. Serum levels of sPD-L1, sPD-L2, sTIM-3, sGal-9 and sMMP-7 were quantified by ELISA and correlated with clinical data. Twelve patients were followed up after eight months to compare the levels of the biomarkers between acute disease and post-COVID-19. RESULTS: IMV patients experienced a lower PaO2/FiO2 (p < 0.0001) and a longer hospital stay (p < 0.0001), and exhibited higher levels of sPD-L1 (p < 0.05), sTIM-3 (p < 0.01) and sMMP-7 (p < 0.0001) when compared with NIMV patients. According to a ROC analysis, sMMP-7 had the highest sensitivity (78%) and specificity (76%) with a cut point of 4.5 ng/mL, followed by sTIM-3 and sPD-L1. Eight months post-COVID-19, IMV patients displayed a significant decrease in the initially high levels of sPD-L1, sTIM-3 and sGal-9, while sPD-L2 was increased, and sMMP-7 was unchanged. CONCLUSION: Circulating levels of sPD-L1, sTIM-3 and sMMP-7 are potential biomarkers of disease severity to distinguish patients requiring IMV. MMP-7 could also be a marker for the persistence of lung lesions post-COVID-19.

Aldose reductase (ALR2) is the enzyme in charge of developing cellular toxicity caused by diabetic hyperglycemia, which in turn leads to the generation of reactive oxygen species triggering oxidative stress. Therefore, inhibiting ALR2 while pursuing a concomitant anti-oxidant activity through dual-acting agents is now recognized as the gold standard treatment for preventing or at least delaying the progression of diabetic complications. Herein we describe a novel series of (E)-benzaldehyde O-benzyl oximes 6a-e, 7a-e, 8a-e, and 9-11 as ALR2 inhibitors endowed with anti-oxidant properties. Inspired by the natural products, the synthesized derivatives are characterized by a different polyhydroxy substitution pattern on their benzaldehyde fragment, which proved crucial for both the enzyme inhibitory activity and the anti-oxidant capacity. Derivatives (E)-2,3,4-trihydroxybenzaldehyde O-(3-methoxybenzyl) oxime (7b) and (E)-2,3,4-trihydroxybenzaldehyde O-(4-methoxybenzyl) oxime (8b) turned out to be the most effective dual-acting products, proving to combine the best ALR2 inhibitory properties with significant anti-oxidant efficacy.

Amyloid formation is a pathological process associated with a wide range of degenerative disorders, including Alzheimer's disease, Parkinson's disease, and diabetes mellitus type 2. During disease progression, abnormal accumulation and deposition of proteinaceous material are accompanied by tissue degradation, inflammation, and dysfunction. Agents that can interfere with the process of amyloid formation or target already formed amyloid assemblies are consequently of therapeutic interest. In this context, a few endogenous proteins have been associated with an anti-amyloidogenic activity. Here, we review the properties of transthyretin, apolipoprotein E, clusterin, and BRICHOS protein domain which all effectively interfere with amyloid in vitro, as well as displaying a clinical impact in humans or animal models. Their involvement in the amyloid formation process is discussed, which may aid and inspire new strategies for therapeutic interventions.

This study aimed to determine the effect of age on CVLM C1 neuron glucoregulatory proteins in the feeding pathway. Male Sprague Dawley rats aged 3 months and 24 months old were divided into two subgroups: the treatment group with 2-deoxy-d-glucose (2DG) and the control group. Rat brains were dissected to obtain the CVLM region of the brainstem. Western blot was used to determine protein expression of tyrosine hydroxylase (TH), phosphorylated TH at Serine40 (pSer40TH), AMP-activated protein kinase (AMPK), phosphorylated AMPK (phospho AMPK), and neuropeptide Y Y5 receptors (NPY5R) in CVLM samples. Immunofluorescence was used to determine TH-, AMPK-, and NPY5R-like immunoreactivities among other brain coronal sections. Results obtained denote a decrease in basal TH phosphorylation levels and AMPK proteins and an increase in TH proteins among aged CVLM neurons. Increases in the basal immunoreactivity of TH+, AMPK+, NPY5R+, TH+/AMPK+, and TH+/NPY5R+ were also observed among old rats. Young treatment-group rats saw a decrease in TH phosphorylation and AMPK proteins following 2DG administration, while an increase in AMPK phosphorylation and a decrease in TH proteins were found among the old-treatment-group rats. These findings suggest the participation of CVLM C1 neurons in counter-regulatory responses among young and old rats. Altering protein changes in aged CVLM C1 neurons may attenuate responses to glucoprivation, thus explaining the decline in food intake among the elderly.

Cellular functions are regulated through the gene expression program by the transcription of new messenger RNAs (mRNAs), alternative RNA splicing, and protein synthesis. To this end, the post-translational modifications (PTMs) of proteins add another layer of complexity, creating a continuously fine-tuned regulatory network. ADP-ribosylation (ADPr) is an ancient reversible modification of cellular macromolecules, regulating a multitude of key functional processes as diverse as DNA damage repair (DDR), transcriptional regulation, intracellular transport, immune and stress responses, and cell survival. Additionally, due to the emerging role of ADP-ribosylation in pathological processes, ADP-ribosyltransferases (ARTs), the enzymes involved in ADPr, are attracting growing interest as new drug targets. In this review, an overview of human ARTs and their related biological functions is provided, mainly focusing on the regulation of ADP-ribosyltransferase Diphtheria toxin-like enzymes (ARTD)-dependent RNA functions. Finally, in order to unravel novel gene functional relationships, we propose the analysis of an inventory of human gene clusters, including ARTDs, which share conserved sequences at 3' untranslated regions (UTRs).

Severe COVID-19 disease leads to hypoxemia, inflammation and lymphopenia. Viral infection induces cellular stress and causes the activation of the innate immune response. The ubiquitin-proteasome system (UPS) is highly implicated in viral immune response regulation. The main function of the proteasome is protein degradation in its active form, which recognises and binds to ubiquitylated proteins. Some proteasome subunits have been reported to be upregulated under hypoxic and hyperinflammatory conditions. Here, we conducted a prospective cohort study of COVID-19 patients (n = 44) and age-and sex-matched controls (n = 20). In this study, we suggested that hypoxia could induce the overexpression of certain genes encoding for subunits from the α and β core of the 20S proteasome and from regulatory particles (19S and 11S) in COVID-19 patients. Furthermore, the gene expression of proteasome subunits was associated with lymphocyte count reduction and positively correlated with inflammatory molecular and clinical markers. Given the importance of the proteasome in maintaining cellular homeostasis, including the regulation of the apoptotic and pyroptotic pathways, these results provide a potential link between COVID-19 complications and proteasome gene expression.

In the sericulture and silk production industry, sericin is discharged in the degumming wastewater, resulting in a large amount of wasted natural protein and environmental pollution. This study investigated the effect of degraded sericin recovered by the Ca(OH)2-ultrasound degumming method (a green process) on liver injury in T2D rats. After 4 weeks of dietary sericin supplementation, the liver masses and organ coefficients of the T2D rats improved compared with those of the model rats that were not fed sericin. Oral sericin activated the damaged PI3K/AKT/AMPK pathway to enhance glycogen synthesis, accelerate glycolysis, and inhibit gluconeogenesis. The protein expression levels of the inflammatory factors NF-κB, IL-6, and TNF-α in the T2D model group were up to two times higher than in the normal group. However, all three T2D groups that received oral sericin showed significant decreases in these factors to the level found in the normal group, indicating that inflammation in the body was significantly reduced. These results show that the sericin protein might improve glycogen synthesis, accelerate glycolysis, and inhibit gluconeogenesis by enhancing the anti-oxidation capability and reducing inflammatory reactions. Therefore, sericin recovered by Ca(OH)2 degradation has potential use in the development of functional health foods that can lower blood sugar.

The p53 family has the following three members: p53, p63 and p73. p53 is a tumor suppressor gene that frequently exhibits mutation in head and neck cancer. Most p53 mutants are loss-of-function (LoF) mutants, but some acquire some oncogenic function, such as gain of function (GoF). It is known that the aggregation of mutant p53 can induce p53 GoF. The p73 activators RETRA and NSC59984 have an anti-cancer effect in p53 mutation cells, but we found that p73 activators were not effective in all head and neck squamous cell carcinoma (HNSCC) cell lines, with different p53 mutants. A comparison of the gene expression profiles of several regulator(s) in mutant HNSCC cells with or without aggregation of p53 revealed that nicotinamide phosphoribosyltransferase (NAMPT) is a key regulator of mutant p53 aggregation. An NAMPT inhibitor, to reduce abnormal aggregation of mutant p53, used in combination with a p73 activator, was able to effectively repress growth in HNSCC cells with p53 GoF mutants. This study, therefore, suggests a potential combination therapy approach for HNSCC with a p53 GoF mutation.

Recently, disease-associated variants of the TUBA4A gene were identified in patients with amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Here, we present the neuropathological report of a patient with the semantic variant of primary progressive aphasia with a family history of Parkinsonism, harboring a novel frameshift mutation c.187del (p.Arg64Glyfs*90) in TUBA4A. Immunohistochemistry showed abundant TAR DNA-binding protein 43 kDa (TDP-43) dystrophic neurite pathology in the frontal and temporal cortex and the dentate gyrus of the hippocampus, consistent with frontotemporal lobar degeneration (FTLD). The observed pathology pattern fitted best with that of FTLD-TDP Type C. qPCR showed the presence of mutant TUBA4A mRNA. However, no truncated TUBA4A was detected at the protein level. A decrease in total TUBA4A mRNA and protein levels suggests loss-of-function as a potential pathogenic mechanism. This report strengthens the idea that N-terminal TUBA4A mutations are associated with FTLD-TDP. These N-terminal mutations possibly exert their pathogenic effects through haploinsufficiency, contrary to C-terminal TUBA4A mutations which are thought to disturb the microtubule network via a dominant-negative mechanism.

Cell adhesion molecule L1 is a cell surface glycoprotein that promotes neuronal cell migration, fosters regeneration after spinal cord injury and ameliorates the consequences of neuronal degeneration in mouse and zebrafish models. Counter-indicative features of L1 were found in tumor progression: the more L1 is expressed, the more tumor cells migrate and increase their metastatic potential. L1's metastatic potential is further evidenced by its promotion of epithelial-mesenchymal transition, endothelial cell transcytosis and resistance to chemo- and radiotherapy. These unfortunate features are indicated by observations that cells that normally do not express L1 are induced to express it when becoming malignant. With the aim to ameliorate the devastating functions of L1 in tumors, we designed an alternative approach to counteract tumor cell migration. Libraries of small organic compounds were screened using the ELISA competition approach similar to the one that we used for identifying L1 agonistic mimetics. Whereas in the former approach, a function-triggering monoclonal antibody was used for screening libraries, we here used the function-inhibiting monoclonal antibody 324 that reduces the migration of neurons. We now show that the L1 antagonistic mimetics anagrelide, 2-hydroxy-5-fluoropyrimidine and mestranol inhibit the migration of cultured tumor cells in an L1-dependent manner, raising hopes for therapy.

Metalloproteins are involved in key cell processes such as photosynthesis, respiration, and oxygen transport. However, the presence of transition metals (notably iron as a component of [Fe-S] clusters) often makes these proteins sensitive to oxygen-induced degradation. Consequently, their study usually requires strict anaerobic conditions. Although X-ray crystallography has been the method of choice for solving macromolecular structures for many years, recently electron microscopy has also become an increasingly powerful structure-solving technique. We have used our previous experience with cryo-crystallography to develop a method to prepare cryo-EM grids in an anaerobic chamber and have applied it to solve the structures of apoferritin and the 3 [Fe4S4]-containing pyruvate ferredoxin oxidoreductase (PFOR) at 2.40 Å and 2.90 Å resolution, respectively. The maps are of similar quality to the ones obtained under air, thereby validating our method as an improvement in the structural investigation of oxygen-sensitive metalloproteins by cryo-EM.

Within this review, sex-specific differences in alveolar epithelial functions are discussed with special focus on preterm infants and the respiratory disorders associated with premature birth. First, a short overview about fetal lung development, the challenges the lung faces during perinatal lung transition to air breathing and respiratory distress in preterm infants is given. Next, clinical observations concerning sex-specific differences in pulmonary morbidity of human preterm infants are noted. The second part discusses potential sex-specific causes of pulmonary complications, including pulmonary steroid receptors and local lung steroid metabolism. With regard to pulmonary steroid metabolism, it is important to highlight which steroidogenic enzymes are expressed at which stage during fetal lung development. Thereafter, we review the knowledge concerning sex-specific aspects of lung growth and maturation. Special focus is given to alveolar epithelial Na+ transport as a driver of perinatal lung transition and the sex differences that were noted in this process.

The distinct neuropathological features of the different α-Synucleinopathies, as well as the diversity of the α-Synuclein (α-Syn) intracellular inclusion bodies observed in post mortem brain sections, are thought to reflect the strain diversity characterizing invasive α-Syn amyloids. However, this "one strain, one disease" view is still hypothetical, and to date, a possible disease-specific contribution of non-amyloid factors has not been ruled out. In Multiple System Atrophy (MSA), the buildup of α-Syn inclusions in oligodendrocytes seems to result from the terminal storage of α-Syn amyloid aggregates first pre-assembled in neurons. This assembly occurs at the level of neuronal cytoplasmic inclusions, and even earlier, within neuronal intranuclear inclusions (NIIs). Intriguingly, α-Syn NIIs are never observed in α-Synucleinopathies other than MSA, suggesting that these inclusions originate (i) from the unique molecular properties of the α-Syn fibril strains encountered in this disease, or alternatively, (ii) from other factors specifically dysregulated in MSA and driving the intranuclear fibrillization of α-Syn. We report the isolation and structural characterization of a synthetic human α-Syn fibril strain uniquely capable of seeding α-Syn fibrillization inside the nuclear compartment. In primary mouse cortical neurons, this strain provokes the buildup of NIIs with a remarkable morphology reminiscent of cat's eye marbles (see video abstract). These α-Syn inclusions form giant patterns made of one, two, or three lentiform beams that span the whole intranuclear volume, pushing apart the chromatin. The input fibrils are no longer detectable inside the NIIs, where they become dominated by the aggregation of endogenous α-Syn. In addition to its phosphorylation at S129, α-Syn forming the NIIs acquires an epitope antibody reactivity profile that indicates its organization into fibrils, and is associated with the classical markers of α-Syn pathology p62 and ubiquitin. NIIs are also observed in vivo after intracerebral injection of the fibril strain in mice. Our data thus show that the ability to seed NIIs is a strain property that is integrally encoded in the fibril supramolecular architecture. Upstream alterations of cellular mechanisms are not required. In contrast to the lentiform TDP-43 NIIs, which are observed in certain frontotemporal dementias and which are conditional upon GRN or VCP mutations, our data support the hypothesis that the presence of α-Syn NIIs in MSA is instead purely amyloid-strain-dependent.

Different approaches to develop engineered scaffolds for periodontal tissues regeneration have been proposed. In this review, innovations in stem cell technology and scaffolds engineering focused primarily on Periodontal Ligament (PDL) regeneration are discussed and analyzed based on results from pre-clinical in vivo studies and clinical trials. Most of those developments include the use of polymeric materials with different patterning and surface nanotopography and printing of complex and sophisticated multiphasic composite scaffolds with different compartments to accomodate for the different periodontal tissues' architecture. Despite the increased effort in producing these scaffolds and their undoubtable efficiency to guide and support tissue regeneration, appropriate source of cells is also needed to provide new tissue formation and various biological and mechanochemical cues from the Extraccellular Matrix (ECM) to provide biophysical stimuli for cell growth and differentiation. Cell sheet engineering is a novel promising technique that allows obtaining cells in a sheet format while preserving ECM components. The right combination of those factors has not been discovered yet and efforts are still needed to ameliorate regenerative outcomes towards the functional organisation of the developed tissues.

Uncontrolled complement activation contributes to multiple immune pathologies. Although synthetic compstatin derivatives targeting C3 and C3b are robust inhibitors of complement activation, their physicochemical and molecular properties may limit access to specific organs, development of bifunctional moieties, and therapeutic applications requiring transgenic expression. Complement-targeting therapeutics containing only natural amino acids could enable multifunctional pharmacology, gene therapies, and targeted delivery for underserved diseases. A Nanofitin library of hyperthermophilic protein scaffolds was screened using ribosome display for C3/C3b-targeting clones mimicking compstatin pharmacology. APL-1030, a recombinant 64-residue Nanofitin, emerged as the lead candidate. APL-1030 is thermostable, binds C3 (KD, 1.59 nM) and C3b (KD, 1.11 nM), and inhibits complement activation via classical (IC50 = 110.8 nM) and alternative (IC50 = 291.3 nM) pathways in Wieslab assays. Pharmacologic activity (determined by alternative pathway inhibition) was limited to primate species of tested sera. C3b-binding sites of APL-1030 and compstatin were shown to overlap by X-ray crystallography of C3b-bound APL-1030. APL-1030 is a novel, high-affinity inhibitor of primate C3-mediated complement activation developed from natural amino acids on the hyperthermophilic Nanofitin platform. Its properties may support novel drug candidates, enabling bifunctional moieties, gene therapy, and tissue-targeted C3 pharmacologics for diseases with high unmet need.

Proinflammatory joint environment, coupled with impeded chondrogenic differentiation of mesenchymal stromal cells (MSCs), led to inferior cartilage repair outcomes. Nuclear translocation of phosphorylated-NFκB downregulates SOX9 and hinders the chondrogenesis of MSCs. Strategies that minimize the deleterious effects of NFκB, while promoting MSC chondrogenesis, are of interest. This study establishes the ability of continuous low-intensity ultrasound (cLIUS) to preserve MSC chondrogenesis in a proinflammatory environment. MSCs were seeded in alginate:collagen hydrogels and cultured for 21 days in an ultrasound-assisted bioreactor (5.0 MHz, 2.5 Vpp; 4 applications/day) in the presence of IL1β and evaluated by qRT-PCR and immunofluorescence. The differential expression of markers associated with the NFκB pathway was assessed upon a single exposure of cLIUS and assayed by Western blotting, qRT-PCR, and immunofluorescence. Mitochondrial potential was evaluated by tetramethylrhodamine methyl ester (TMRM) assay. The chondroinductive potential of cLIUS was noted by the increased expression of SOX9 and COLII. cLIUS extended its chondroprotective effects by stabilizing the NFκB complex in the cytoplasm via engaging the IκBα feedback mechanism, thus preventing its nuclear translocation. cLIUS acted as a mitochondrial protective agent by restoring the mitochondrial potential and the mitochondrial mRNA expression in a proinflammatory environment. Altogether, our results demonstrated the potential of cLIUS for cartilage repair and regeneration under proinflammatory conditions.

The composition of microbiota and the gut-brain axis is increasingly considered a factor in the development of various pathological conditions. The etiology of multiple sclerosis (MS), a chronic autoimmune disease affecting the CNS, is complex and interactions within the gut-brain axis may be relevant in the development and the course of MS. In this article, we focus on the relationship between gut microbiota and the pathophysiology of MS. We review the contribution of germ-free mouse studies to our understanding of MS pathology and its implications for treatment strategies to modulate the microbiome in MS. This summary highlights the need for a better understanding of the role of the microbiota in patients' responses to disease-modifying drugs in MS and disease activity overall.

Steady state peripheral blood (SSPB) contains hematopoietic stem and progenitor cells (HSPCs) presenting characteristics of real hematopoietic stem cells, and thus represents an interesting alternative cell supply for hematopoietic cell transplantation. Development of ex vivo expansion strategies could overcome the low HSPC numbers usually rescued from SSPB. We investigated the effect of alpha lipoic acid (ALA) on ex vivo culture of SSPB CD34 positive (CD34pos) cells on primitive cell expansion, cell cycle, and oxidative metabolism as estimated by determining the ROS and GSH content. ALA increased the ex vivo expansion of total CD34pos cells and of phenotypically defined CD34pos HSPCs subpopulations that retained in vivo repopulating capacity, concomitantly to a decreased expansion of differentiating cells. ALA did not modify cell cycle progression nor the proliferation of ex vivo expanded CD34pos cells, and coherently did not affect the ROS level. On the contrary, ALA decreased the proliferation and disturbed cell cycle progression of cells reaching a differentiated status, a phenomenon that seems to be associated with a drop in ROS level. Nonetheless, ALA affected the redox status of hematopoietic primitive cells, as it reproducibly increased GSH content. In conclusion, ALA represents an interesting molecule for the improvement of ex vivo expansion strategies and further clinical application in hematopoietic cell transplantation (HCT).

The issue of tolerance to continuous or repeated administration of opioids should be addressed. The ability of ketamine to improve opioid tolerance has been reported in clinical studies, and its mechanism of tolerance may involve improved desensitization of μ-opioid receptors (MORs). We measured changes in MOR activity and intracellular signaling induced by repeated fentanyl and morphine administration and investigated the effects of ketamine on these changes with human embryonic kidney 293 cells expressing MOR using the CellKey™, cADDis cyclic adenosine monophosphate, and PathHunter® β-arrestin recruitment assays. Repeated administration of fentanyl or morphine suppressed the second MOR responses. Administration of ketamine before a second application of opioids within clinical concentrations improved acute desensitization and enhanced β-arrestin recruitment elicited by fentanyl but not by morphine. The effects of ketamine on fentanyl were suppressed by co-treatment with an inhibitor of G-protein-coupled receptor kinase (GRK). Ketamine may potentially reduce fentanyl tolerance but not that of morphine through modulation of GRK-mediated pathways, possibly changing the conformational changes of β-arrestin to MOR.

Pulmonary hypertension is treated with drugs that stimulate cGMP or cAMP signalling. Both nucleotides can activate Kv7 channels, leading to smooth muscle hyperpolarisation, reduced Ca2+ influx and relaxation. Kv7 activation by cGMP contributes to the pulmonary vasodilator action of nitric oxide, but its contribution when dilation is evoked by the atrial natriuretic peptide (ANP) sensitive guanylate cyclase, or cAMP, is unknown. Small vessel myography was used to investigate the ability of Kv7 channel blockers to interfere with pulmonary artery relaxation when cyclic nucleotide pathways were stimulated in different ways. The pan-Kv7 blockers, linopirdine and XE991, caused substantial inhibition of relaxation evoked by NO donors and ANP, as well as endothelium-dependent dilators, the guanylate cyclase stimulator, riociguat, and the phosphodiesterase-5 inhibitor, sildenafil. Maximum relaxation was reduced without a change in sensitivity. The blockers had relatively little effect on cAMP-mediated relaxation evoked by forskolin, isoprenaline or treprostinil. The Kv7.1-selective blocker, HMR1556, had no effect on cGMP or cAMP-dependent relaxation. Western blot analysis demonstrated the presence of Kv7.1 and Kv7.4 proteins, while selective activators of Kv7.1 and Kv7.4 homomeric channels, but not Kv7.5, caused pulmonary artery relaxation. It is concluded that Kv7.4 channels contribute to endothelium-dependent dilation and the effects of drugs that act by stimulating cGMP, but not cAMP, signalling.

Ischemic stroke is a common cerebrovascular disease and recovering blood flow as early as possible is essential to reduce ischemic damage and maintain neuronal viability, but the reperfusion process usually causes additional damage to the brain tissue in the ischemic area, namely ischemia reperfusion injury. The accumulated studies have revealed that transplantation of exogenous neural stem cells (NSCs) is an ideal choice for the treatment of ischemia reperfusion injury. At present, the source and efficacy of exogenous NSCs after transplantation is still one of the key issues that need to be resolved. In this study, human umbilical cord mesenchymal stem cells (hUC-MSCs) were obtained and induced into NSCs byadding growth factor and neuregulin1β (NRG1β) was introduced during the differentiation process of NSCs. Then, the rat middle cerebral artery occlusion/reperfusion (MCAO/R) models were established, and the therapeutic effects were evaluated among groups treated by NRG1β, NSCs and NSCs pretreated with 10 nM NRG1β (NSCs-10 nM NRG1β) achieved through intra-arterial injection. Our data show that the NSCs-10 nM NRG1β group significantly improves neurobehavioral function and infarct volume after MCAO/R, as well as cerebral cortical neuron injury, ferroptosis-related indexes and mitochondrial injury. Additionally, NSCs-10 nM NRG1β intervention may function through regulating the p53/GPX4/SLC7A11 pathway, and reducing the level of ferroptosis in cells, further enhance the neuroprotective effect on injured cells.

Stents are lifesaving mechanical devices that re-establish essential blood flow to the coronary circulation after significant vessel occlusion due to coronary vessel disease or thrombolytic blockade. Improvements in stent surface engineering over the last 20 years have seen significant reductions in complications arising due to restenosis and thrombosis. However, under certain conditions such as diabetes mellitus (DM), the incidence of stent-mediated complications remains 2-4-fold higher than seen in non-diabetic patients. The stents with the largest market share are designed to target the mechanisms behind neointimal hyperplasia (NIH) through anti-proliferative drugs that prevent the formation of a neointima by halting the cell cycle of vascular smooth muscle cells (VSMCs). Thrombosis is treated through dual anti-platelet therapy (DAPT), which is the continual use of aspirin and a P2Y12 inhibitor for 6-12 months. While the most common stents currently in use are reasonably effective at treating these complications, there is still significant room for improvement. Recently, inflammation and redox stress have been identified as major contributing factors that increase the risk of stent-related complications following percutaneous coronary intervention (PCI). The aim of this review is to examine the mechanisms behind inflammation and redox stress through the lens of PCI and its complications and to establish whether tailored targeting of these key mechanistic pathways offers improved outcomes for patients, particularly those where stent placement remains vulnerable to complications. In summary, our review highlights the most recent and promising research being undertaken in understanding the mechanisms of redox biology and inflammation in the context of stent design. We emphasize the benefits of a targeted mechanistic approach to decrease all-cause mortality, even in patients with diabetes.

Mitochondria, the cell's major energy producers, also act as signaling hubs, interacting with other organelles both directly and indirectly. Despite having its own circular genome, the majority of mitochondrial proteins are encoded by nuclear DNA. To respond to changes in cell physiology, the mitochondria must send signals to the nucleus, which can, in turn, upregulate gene expression to alter metabolism or initiate a stress response. This is known as retrograde signaling. A variety of stimuli and pathways fall under the retrograde signaling umbrella. Mitochondrial dysfunction has already been shown to have severe implications for human health. Disruption of retrograde signaling, whether directly associated with mitochondrial dysfunction or cellular environmental changes, may also contribute to pathological deficits. In this review, we discuss known signaling pathways between the mitochondria and the nucleus, examine the possibility of direct contacts, and identify pathological consequences of an altered relationship.

Group 1 (Dur-19, PF00477, LEA_5) Late Embryogenesis Abundant (LEA) proteins are present in organisms from all three domains of life, Archaea, Bacteria, and Eukarya. Surprisingly, Artemia is the only genus known to include animals that express group 1 LEA proteins in their desiccation-tolerant life-history stages. Bioinformatics analysis of circular dichroism data indicates that the group 1 LEA protein AfLEA1 is surprisingly ordered in the hydrated state and undergoes during desiccation one of the most pronounced disorder-to-order transitions described for LEA proteins from A. franciscana. The secondary structure in the hydrated state is dominated by random coils (42%) and β-sheets (35%) but converts to predominately α-helices (85%) when desiccated. Interestingly, AfLEA1 interacts with other proteins and nucleic acids, and RNA promotes liquid-liquid phase separation (LLPS) of the protein from the solvent during dehydration in vitro. Furthermore, AfLEA1 protects the enzyme lactate dehydrogenase (LDH) during desiccation but does not aid in restoring LDH activity after desiccation-induced inactivation. Ectopically expressed in D. melanogaster Kc167 cells, AfLEA1 localizes predominantly to the cytosol and increases the cytosolic viscosity during desiccation compared to untransfected control cells. Furthermore, the protein formed small biomolecular condensates in the cytoplasm of about 38% of Kc167 cells. These findings provide additional evidence for the hypothesis that the formation of biomolecular condensates to promote water stress tolerance during anhydrobiosis may be a shared feature across several groups of LEA proteins that display LLPS behaviors.

Infrapatellar fat pad-derived mesenchymal stem cells (IPFP-MSCs) are a type of adipose-derived stem cell (ADSC). They potentially contribute to cartilage regeneration and modulation of the immune microenvironment in patients with osteoarthritis (OA). The ability of IPFP-MSCs to increase chondrogenic capacity has been reported to be greater, less age dependent, and less affected by inflammatory changes than that of other MSCs. Transcription-regulatory factors strictly regulate the cartilage differentiation of MSCs. However, few studies have explored the effect of transcriptional factors on IPFP-MSC-based neocartilage formation, cartilage engineering, and tissue functionality during and after chondrogenesis. Instead of intact MSCs, MSC-derived extracellular vesicles could be used for the treatment of OA. Furthermore, exosomes are increasingly being considered the principal therapeutic agent in MSC secretions that is responsible for the regenerative and immunomodulatory functions of MSCs in cartilage repair. The present study provides an overview of advancements in enhancement strategies for IPFP-MSC chondrogenic differentiation, including the effects of transcriptional factors, the modulation of released exosomes, delivery mechanisms for MSCs, and ethical and regulatory points concerning the development of MSC products. This review will contribute to the understanding of the IPFP-MSC chondrogenic differentiation process and enable the improvement of IPFP-MSC-based cartilage tissue engineering.