The function of retinoic acid (RA) during limb development is still debated, as loss and gain of function studies led to opposite conclusions. With regard to limb initiation, genetic studies demonstrated that activation of FGF10 signaling is required for the emergence of limb buds from the trunk, with Tbx5 and RA signaling acting upstream in the forelimb field, whereas Tbx4 and Pitx1 act upstream in the hindlimb field. Early studies in chick embryos suggested that RA as well as Meis1 and Meis2 (Meis1/2) are required for subsequent proximodistal patterning of both forelimbs and hindlimbs, with RA diffusing from the trunk, functioning to activate Meis1/2 specifically in the proximal limb bud mesoderm. However, genetic loss of RA signaling does not result in loss of limb Meis1/2 expression and limb patterning is normal, although Meis1/2 expression is reduced in trunk somitic mesoderm. More recent studies demonstrated that global genetic loss of Meis1/2 results in a somite defect and failure of limb bud initiation. Other new studies reported that conditional genetic loss of Meis1/2 in the limb results in proximodistal patterning defects, and distal FGF8 signaling represses Meis1/2 to constrain its expression to the proximal limb. In this review, we hypothesize that RA and Meis1/2 both function in the trunk to initiate forelimb bud initiation, but that limb Meis1/2 expression is activated proximally by a factor other than RA and repressed distally by FGF8 to generate proximodistal patterning.

For a long time, the pharmaceutical industry focused on natural biologically active molecules due to their unique properties, availability and significantly less side-effects. Mangiferin is a naturally occurring C-glucosylxantone that has substantial potential for the treatment of various diseases thanks to its numerous biological activities. Many research studies have proven that mangiferin possesses antioxidant, anti-infection, anti-cancer, anti-diabetic, cardiovascular, neuroprotective properties and it also increases immunity. It is especially important that it has no toxicity. However, mangiferin is not being currently applied to clinical use because its oral bioavailability as well as its absorption in the body are too low. To improve the solubility, enhance the biological action and bioavailability, mangiferin integrated polymer systems have been developed. In this paper, we review molecular mechanisms of anti-cancer action as well as a number of designed polymer-mangiferin systems. Taking together, mangiferin is a very promising anti-cancer molecule with excellent properties and the absence of toxicity.

Zebrafish are an instrumental system for the generation of photoreceptor degeneration models, which can be utilized to determine underlying causes of photoreceptor dysfunction and death, and for the analysis of potential therapeutic compounds, as well as the characterization of regenerative responses. We review the wealth of information from existing zebrafish models of photoreceptor disease, specifically as they relate to currently accepted taxonomic classes of human rod and cone disease. We also highlight that rich, detailed information can be derived from studying photoreceptor development, structure, and function, including behavioural assessments and in vivo imaging of zebrafish. Zebrafish models are available for a diversity of photoreceptor diseases, including cone dystrophies, which are challenging to recapitulate in nocturnal mammalian systems. Newly discovered models of photoreceptor disease and drusenoid deposit formation may not only provide important insights into pathogenesis of disease, but also potential therapeutic approaches. Zebrafish have already shown their use in providing pre-clinical data prior to testing genetic therapies in clinical trials, such as antisense oligonucleotide therapy for Usher syndrome.

RNA modifications are essential for proper RNA processing, quality control, and maturation steps. In the last decade, some eukaryotic DNA repair enzymes have been shown to have an ability to recognize and process modified RNA substrates and thereby contribute to RNA surveillance. Single-strand-selective monofunctional uracil-DNA glycosylase 1 (SMUG1) is a base excision repair enzyme that not only recognizes and removes uracil and oxidized pyrimidines from DNA but is also able to process modified RNA substrates. SMUG1 interacts with the pseudouridine synthase dyskerin (DKC1), an enzyme essential for the correct assembly of small nucleolar ribonucleoproteins (snRNPs) and ribosomal RNA (rRNA) processing. Here, we review rRNA modifications and RNA quality control mechanisms in general and discuss the specific function of SMUG1 in rRNA metabolism. Cells lacking SMUG1 have elevated levels of immature rRNA molecules and accumulation of 5-hydroxymethyluridine (5hmU) in mature rRNA. SMUG1 may be required for post-transcriptional regulation and quality control of rRNAs, partly by regulating rRNA and stability.

Rett Syndrome (RTT) is a severe, rare, and progressive developmental disorder with patients displaying neurological regression and autism spectrum features. The affected individuals are primarily young females, and more than 95% of patients carry de novo mutation(s) in the Methyl-CpG-Binding Protein 2 (MECP2) gene. While the majority of RTT patients have MECP2 mutations (classical RTT), a small fraction of the patients (atypical RTT) may carry genetic mutations in other genes such as the cyclin-dependent kinase-like 5 (CDKL5) and FOXG1. Due to the neurological basis of RTT symptoms, MeCP2 function was originally studied in nerve cells (neurons). However, later research highlighted its importance in other cell types of the brain including glia. In this regard, scientists benefitted from modeling the disease using many different cellular systems and transgenic mice with loss- or gain-of-function mutations. Additionally, limited research in human postmortem brain tissues provided invaluable findings in RTT pathobiology and disease mechanism. MeCP2 expression in the brain is tightly regulated, and its altered expression leads to abnormal brain function, implicating MeCP2 in some cases of autism spectrum disorders. In certain disease conditions, MeCP2 homeostasis control is impaired, the regulation of which in rodents involves a regulatory microRNA (miR132) and brain-derived neurotrophic factor (BDNF). Here, we will provide an overview of recent advances in understanding the underlying mechanism of disease in RTT and the associated genetic mutations in the MECP2 gene along with the pathobiology of the disease, the role of the two most studied protein variants (MeCP2E1 and MeCP2E2 isoforms), and the regulatory mechanisms that control MeCP2 homeostasis network in the brain, including BDNF and miR132.

Developmental and environmental signaling networks often converge during plant growth in response to changing conditions. Stress-induced hormones, such as jasmonates (JAs), can influence growth by crosstalk with other signals like brassinosteroids (BRs) and ethylene (ET). Nevertheless, it is unclear how avoidance of an abiotic stress triggers local changes in development as a response. It is known that stress hormones like JAs/ET and BRs can regulate the division rate of cells from the first asymmetric cell divisions (ACDs) in meristems, suggesting that stem cell activation may take part in developmental changes as a stress-avoidance-induced response. The root system is a prime responder to stress conditions in soil. Together with the primary root and lateral roots (LRs), adventitious roots (ARs) are necessary for survival in numerous plant species. AR and LR formation is affected by soil pollution, causing substantial root architecture changes by either depressing or enhancing rooting as a stress avoidance/survival response. Here, a detailed overview of the crosstalk between JAs, ET, BRs, and the stress mediator nitric oxide (NO) in auxin-induced AR and LR formation, with/without cadmium and arsenic, is presented. Interactions essential in achieving a balance between growth and adaptation to Cd and As soil pollution to ensure survival are reviewed here in the model species Arabidopsis and rice.

Renal injury observed in several pathologies has been associated with lipid accumulation in the kidney. While it has been suggested that the accumulation of renal lipids depends on free fatty acids released from adipose tissue, it is not known whether in situ renal lipogenesis due to endoplasmic reticulum (ER) stress contributes to kidney injury. The aim of the present study was to elucidate the role of pharmacological ER stress in renal structure and function and its effect on renal lipid metabolism of C57BL/6 mice. ER stress increased serum creatinine and induced kidney structural abnormalities. Tunicamycin-administered mice developed hyperinsulinemia, augmented lipolysis and increased circulating leptin and adiponectin. Renal unfolded protein response (UPR) gene expression markers, the lipogenic transcription factor SREBP1 and the phosphorylation of eIF2α increased 8 h after tunicamycin administration. At 24 h, an increase in BiP protein content was accompanied by a reduction in p-eIF2α and increased SREBP-1 and FASn protein content, in addition to a significant increase in triglyceride content and a reduction in AMPK. Thus, ER stress induces in situ lipid synthesis, leading to renal lipid accumulation and functional alterations. Future pharmacological and/or dietary strategies must target renal ER stress to prevent kidney damage and the progression of metabolic diseases.

The coronavirus disease 2019 (COVID-19) pandemic is caused by a novel coronavirus; the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2). Millions of cases and deaths to date have resulted in a global challenge for healthcare systems. COVID-19 has a high mortality rate, especially in elderly individuals with pre-existing chronic comorbidities. There are currently no effective therapeutic approaches for the prevention and treatment of COVID-19. Therefore, the identification of effective therapeutics is a necessity. Terpenes are the largest class of natural products that could serve as a source of new drugs or as prototypes for the development of effective pharmacotherapeutic agents. In the present study, we discuss the antiviral activity of these natural products and we perform simulations against the Mpro and PLpro enzymes of SARS-CoV-2. Our results strongly suggest the potential of these compounds against human coronaviruses, including SARS-CoV-2.

Staphylococcus aureus is a Gram-positive bacterium responsible for a variety of mild to life-threatening infections including bone infections such as osteomyelitis. This bacterium is able to invade and persist within non-professional phagocytic cells such as osteoblasts. In the present study, four different S. aureus strains, namely, 2SA-ST239-III (ST239), 5SA-ST5-II (ST5), 10SA-ST228-I (ST228), and 14SA-ST22-IVh (ST22), were tested for their ability to modulate cell viability in MG-63 osteoblast-like cells following successful invasion and persistence. Methicillin-sensitive S. aureus (MSSA) ATCC-12598-ST30 (ST30) was used as control strain. Despite being proven that ST30, ST239, and ST22 have a similar ability to internalize and persist in MG-63 osteoblast-like cells under our experimental conditions, we demonstrated that the observed decrease in cell viability was due to the different behavior of the considered strains, rather than the number of intracellular bacteria. We focused our attention on different biochemical cell functions related to inflammation, cell metabolism, and oxidative stress during osteoblast infections. We were able to show the following: (1) ST30 and ST239 were the only two clones able to persist and maintain their number in the hostile environment of the cell during the entire period of infection; (2) ST239 was the only clone able to significantly increase gene expression (3 and 24 h post-infection (p.i.)) and protein secretion (24 h p.i.) of both interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α) in MG-63 osteoblast-like cells; (3) the same clone determined a significant up-regulation of the transforming growth factorbeta 1 (TGF-β1) and of the metabolic marker glyceraldehyde 3-phosphate dehydrogenase (GAPDH) mRNAs at 24 h p.i.; and (4) neither the MSSA nor the four methicillin-resistant S. aureus (MRSA) strains induced oxidative stress phenomena in MG-63 cells, although a high degree of variability was observed for the different clones with regard to the expression pattern of nuclear factor E2-related factor 2 (Nrf2) and its downstream gene heme oxygenase 1 (HO-1) activation. Our results may pave the way for an approach to S. aureus-induced damage, moving towards individualized therapeutic strategies that take into account the differences between MSSA and MRSA as well as the distinctive features of the different clones. This approach is based on a change of paradigm in antibiotic therapy involving a case-based use of molecules able to counteract pro-inflammatory cytokines activity such as selective cytokine signaling inhibitors (IL-6, TNF-α).

Endothelial dysfunction precedes atherosclerosis and is an independent predictor of cardiovascular events. Cholesterol levels and oxidative stress are key contributors to endothelial damage, whereas high levels of plasma high-density lipoproteins (HDL) could prevent it. Cholesteryl ester transfer protein (CETP) is one of the most potent endogenous negative regulators of HDL-cholesterol. However, whether and to what degree CETP expression impacts endothelial function, and the molecular mechanisms underlying the vascular effects of CETP on endothelial cells, have not been addressed. Acetylcholine-induced endothelium-dependent relaxation of aortic rings was impaired in human CETP-expressing transgenic mice, compared to their non-transgenic littermates. However, endothelial nitric oxide synthase (eNOS) activation was enhanced. The generation of superoxide and hydrogen peroxide was increased in aortas from CETP transgenic mice, while silencing CETP in cultured human aortic endothelial cells effectively decreased oxidative stress promoted by all major sources of ROS: mitochondria and NOX2. The endoplasmic reticulum stress markers, known as GADD153, PERK, and ARF6, and unfolded protein response effectors, were also diminished. Silencing CETP reduced endothelial tumor necrosis factor (TNF) α levels, intercellular cell adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) expression, diminishing monocyte adhesion. These results support the notion that CETP expression negatively impacts endothelial cell function, revealing a new mechanism that might contribute to atherosclerosis.

Apolipoprotein A-I (ApoA-I) concentrations are decreased during inflammation, which may reduce high-density lipoprotein (HDL) functionality. Thus, rescuing ApoA-I concentrations during inflammation might help to prevent atherosclerosis. Recent studies have shown that butyric acid (C4) has anti-inflammatory effects and rescues ApoA-I production. However, whether intestinal short chain fatty acids (SCFAs) are able to influence hepatic processes is unknown. Therefore, we investigated C4 anti-inflammatory effects on ApoA-I transcription in the intestine-liver co-culture model. C4 dose-response experiments in the presence or absence of cytokines were performed in a co-culture system including Caco-2 cells, HepG2 cells, or both. Changes in ApoA-I transcription in Caco-2 cells and HepG2 cells were analyzed using qPCR. C4 increased ApoA-I expression in HepG2 cells that cultured alone. When both cells were cultured together, C4 decreased ApoA-I expression in Caco-2 cells and increased ApoA-I expression in HepG2 cells. However, adding C4 to apical Caco-2 cells resulted in a smaller effect in HepG2 cells compared with adding C4 directly to the hepatocytes. Moreover, C4 rescued ApoA-I expression in inflamed HepG2 cells. These findings suggests that intestinal SCFAs can affect hepatic processes. However, the smaller effect in the co-culture experiment indicates cross-talk between intestine and liver.

Silk fibroin (SF)-based materials are exposed to both natural and artificial ultraviolet (UV) light during preparation or administration. However, the effects of UV irradiation on SF films prepared under different conditions have not yet been described in detail. In this study, four SF films with different molecular weight (MW) distribution were fabricated using SF solutions, which were prepared by dissolving degummed SF for 0.5-24 h. We observed UV (365 nm) irradiation on SF films induced the increase of yellowness and absorbance at 310 nm of SF films, indicating the formation of new photo-products and di-tyrosine bonds by photo-oxidation. Due to di-tyrosine cross-links between SF chains, UV-irradiated SF films were not fully dissociated in urea solution. In addition to formation of new products, UV reduced the crystallinity of SF films by breaking hydrogen bonds of β-sheet conformation. Unlike the UV-induced decomposition of physical interactions, UV did not affect the covalent bonds (i.e., peptide bonds). Through these experiments, we could expect that SF with higher MW was more susceptible and SF with lower MW was more resistant to UV-induced photo-oxidation and photo-degradation. These results provide useful information about UV-induced aging of SF-based materials under natural sunlight and UV irradiating conditions.

Diesel exhaust particulates (DEP) adversely affect the respiratory system and exacerbate lung diseases, resulting in high mortality rates. However, its pathogenesis is complicated, and the mechanisms involved are incompletely understood. We investigated the effects of DEP pre-exposure on lipopolysaccharide (LPS)-induced acute lung injury (ALI) and identified the roles of interleukin (IL)-17 in mice. Mice were divided into vehicle control, DEP, LPS, and DEP pre-exposed and LPS-instilled groups. Pre-exposure to DEP enhanced the number of total cells, neutrophils, and lymphocytes in the BAL fluid of LPS-instilled mice. Pre-exposure to DEP synergistically exacerbated pulmonary acute lung inflammation and granulomatous inflammation/pulmonary fibrosis, concomitant with the enhanced expression of inflammatory cytokines in the BAL fluid and of collagen I and TGF-β1 in the lungs of LPS-instilled mice. The number of TGF-β1-positive cells in the DEP pre-exposed and LPS-instilled group was higher than that in the LPS group. The expression of NLR family pyrin domain containing 3 (NLRP3) inflammasome components was markedly increased in the DEP pre-exposed and LPS-instilled group. IL-17 levels in the BAL fluid and IL-17-positive cells in the lungs were significantly increased by pre-exposure to DEP in the LPS-induced group compared to that in the DEP or LPS group. These results suggest that DEP predominantly contributes to fibrotic lung disease in LPS-related acute lung injury by upregulating IL-17 cytokine-mediated collagen I and TGF-β1 and, at least in part, by activating LPS-induced NLRP3 inflammasome signaling. The study should be useful in devising better strategies for prevention and management of ALI.

Oncomodulin (Ocm), or parvalbumin β, is an 11-12 kDa Ca2+-binding protein found inside and outside of vertebrate cells, which regulates numerous processes via poorly understood mechanisms. Ocm consists of two active Ca2+-specific domains of the EF-hand type ("helix-loop-helix" motif), covered by an EF-hand domain with inactive EF-hand loop, which contains a highly conservative cysteine with unknown function. In this study, we have explored peculiarities of the microenvironment of the conservative Cys18 of recombinant rat Ocm (rWT Ocm), redox properties of this residue, and structural/functional sensitivity of rWT Ocm to the homologous C18S substitution. We have found that pKa of the Cys18 thiol lays beyond the physiological pH range. The measurement of redox dependence of rWT Ocm thiol-disulfide equilibrium (glutathione redox pair) showed that redox potential of Cys18 for the metal-free and Ca2+-loaded protein is of -168 mV and -176 mV, respectively. Therefore, the conservative thiol of rWT Ocm is prone to disulfide dimerization under physiological redox conditions. The C18S substitution drastically reduces α-helices content of the metal-free and Mg2+-bound Ocm, increases solvent accessibility of its hydrophobic residues, eliminates the cooperative thermal transition in the apo-protein, suppresses Ca2+/Mg2+ affinity of the EF site, and accelerates Ca2+ dissociation from Ocm. The distinct structural and functional consequences of the minor structural modification of Cys18 indicate its possible redox sensory function. Since some other EF-hand proteins also contain a conservative redox-sensitive cysteine located in an inactive EF-hand loop, it is reasonable to suggest that in the course of evolution, some of the EF-hands attained redox sensitivity at the expense of the loss of their Ca2+ affinity.

Every living cell is covered with a dense and complex layer of glycans on the cell surface, which have important functions in the interaction between cells and their environment. Glycosphingolipids (GSLs) are glycans linked to lipid molecules that together with sphingolipids, sterols, and proteins form plasma membrane lipid rafts that contribute to membrane integrity and provide specific recognition sites. GSLs are subdivided into three major series (globo-, ganglio-, and neolacto-series) and are synthesized in a non-template driven process by enzymes localized in the ER and Golgi apparatus. Altered glycosylation of lipids are known to be involved in tumor development and metastasis. Metastasis is frequently linked with reversible epithelial-to-mesenchymal transition (EMT), a process involved in tumor progression, and the formation of new distant metastatic sites (mesenchymal-to-epithelial transition or MET). On a single cell basis, cancer cells lose their epithelial features to gain mesenchymal characteristics via mechanisms influenced by the composition of the GSLs on the cell surface. Here, we summarize the literature on GSLs in the context of reversible and cancer-associated EMT and discuss how the modification of GSLs at the cell surface may promote this process.

Lysosomes, the degradative endpoints and sophisticated cellular signaling hubs, are emerging as intracellular Ca2+ stores that govern multiple cellular processes. Dys-homeostasis of lysosomal Ca2+ is intimately associated with a variety of human diseases including cancer. Recent studies have suggested that the Ca2+-permeable channels Transient Receptor Potential (TRP) Mucolipins (TRPMLs, TRPML1-3) integrate multiple processes of cell growth, division and metabolism. Dysregulation of TRPMLs activity has been implicated in cancer development. In this review, we provide a summary of the latest development of TRPMLs in cancer. The expression of TRPMLs in cancer, TRPMLs in cancer cell nutrient sensing, TRPMLs-mediated lysosomal exocytosis in cancer development, TRPMLs in TFEB-mediated gene transcription of cancer cells, TRPMLs in bacteria-related cancer development and TRPMLs-regulated antitumor immunity are discussed. We hope to guide readers toward a more in-depth discussion of the importance of lysosomal TRPMLs in cancer progression and other human diseases.

Propolis is a bee product with known medical properties, including antioxidant activity. The scope of the study is profiling 19 different Eurasian propolis samples (mostly from Russia and Kazakhstan, Kyrgyzstan, Poland, Ukraine, and Slovakia). Profiles of propolises were investigated by ultra-high-performance liquid chromatography-diode array detector-mass spectrometry (UPLC-DAD-MS). Classical antioxidant properties, which are based on electron donation mechanism, were assessed by DPPH, ferric reducing antioxidant power (FRAP), and oxygen radical absorbance capacity (ORAC) assays. Total phenolic and flavonoid contents were also evaluated by colorimetric tests. Most of the samples exhibited significant content of polyphenols (from 30.28 to 145.24 mg GAE/g of propolis) and flavonoids (from 10.45 to 82.71 mg GAE/g of propolis). Most of the propolis samples exhibited potent antiradical (DPPH test-from 8.83 to 64.47 mg GAE/g of propolis) and reducing activity (FRAP test-from 0.08 to 1.17 mmol Fe2+/g of propolis). Based on the occurrence of marker compounds, propolis samples were classified as poplar, aspen-birch, aspen-poplar, and aspen-birch-poplar type. Main markers present in propolis of poplar (e.g., chrysin, pinocembrin, galangin, and 3-O-acetyl-pinobanksin), birch (ermanin and acacetin) and aspen (2-acetyl-1,3-di-p-coumaroylglycerol) origin were used. DPPH, FRAP, and ORAC tests results were correlated with flavonoids, total polyphenols, or the polyphenols other than flavonoids content. In term of activity, poplar propolis type was variable, while aspen-birch-poplar type usually exhibited high DPPH and FRAP activity.

Royal jelly (RJ), a natural honeybee product, has a wide range of antibacterial activities. N-glycosylated major royal jelly protein 2 (N-MRJP2), purified from RJ, can inhibit the growth of Paenibacillus larvae (P. larvae, Gram-positive), a contagious etiological agent of the American foulbrood disease of honeybees. However, the inhibitory mechanism is largely unknown. Antibacterial assay and membrane proteome were conducted to investigate the inhibition capacity of RJ from different instar larvae and P. larvae treated by N-MRJP2, respectively. The similar antibacterial efficiency of RJ from different larval instar indicates that RJ is vital for the adaptive immune defense of small larvae. The killing of P. larvae by N-MRJP2 is achieved by disturbing the cell wall biosynthesis, increasing the permeability of cell membrane, hindering aerobic respiration, restraining cell division and inducing cell death. This demonstrates that RJ is critical for the passive immunity of immature larvae and N-MRJP2 can be used as natural antibiotic substance to resist P. larvae, even for other gram-positive bacteria. This constitutes solid evidence that RJ and N-MRJP2 have potentials as novel antibacterial agents.

Cell plasma membrane proteins are considered as gatekeepers of the cell and play a major role in regulating various processes. Transport proteins constitute a subclass of cell plasma membrane proteins enabling the exchange of molecules and ions between the extracellular environment and the cytosol. A plethora of human pathologies are associated with the altered expression or dysfunction of cell plasma membrane transport proteins, making them interesting therapeutic drug targets. However, the search for therapeutics is challenging, since many drug candidates targeting cell plasma membrane proteins fail in (pre)clinical testing due to inadequate selectivity, specificity, potency or stability. These latter characteristics are met by nanobodies, which potentially renders them eligible therapeutics targeting cell plasma membrane proteins. Therefore, a therapeutic nanobody-based strategy seems a valid approach to target and modulate the activity of cell plasma membrane transport proteins. This review paper focuses on methodologies to generate cell plasma membrane transport protein-targeting nanobodies, and the advantages and pitfalls while generating these small antibody-derivatives, and discusses several therapeutic nanobodies directed towards transmembrane proteins, including channels and pores, adenosine triphosphate-powered pumps and porters.

Mutations in the GJB2 gene encoding transmembrane protein connexin 26 (Cx26) are the most common cause for hearing loss worldwide. Cx26 plays a crucial role in the ionic and metabolic homeostasis in the inner ear, indispensable for normal hearing process. Different pathogenic mutations in the GJB2 gene can affect all stages of the Cx26 life cycle and result in nonsyndromic autosomal recessive (DFNB1) or dominant (DFNA3) deafness and syndromes associating hearing loss with skin disorders. This study aims to elucidate the functional consequences of a rare GJB2 variant c.516G>C (p.Trp172Cys) found with high frequency in deaf patients from indigenous populations of Southern Siberia (Russia). The substitution c.516G>C leads to the replacement of tryptophan at a conserved amino acid position 172 with cysteine (p.Trp172Cys) in the second extracellular loop of Cx26 protein. We analyzed the subcellular localization of mutant Cx26-p.Trp172Cys protein by immunocytochemistry and the hemichannels permeability by dye loading assay. The GJB2 knockout HeLa cell line has been generated using CRISPR/Cas9 genome editing tool. Subsequently, the HeLa transgenic cell lines stably expressing different GJB2 variants (wild type and mutations associated with hearing loss) were established based on knockout cells and used for comparative functional analysis. The impaired trafficking of mutant Cx26-p.Trp172Cys protein to the plasma membrane and reduced hemichannels permeability support the pathogenic effect of the c.516G>C (p.Trp172Cys) variant and its association with nonsyndromic hearing loss. Our data contribute to a better understanding of the role of mutations in the second extracellular loop of Cx26 protein in pathogenesis of deafness.

Clays attributed to have medicinal properties have been used since prehistoric times and are still used today as complementary medicines, which has given rise to unregulated "bioceutical" clays to treat skin conditions. Recently, clays with antibacterial characteristics have been proposed as alternatives to antibiotics, potentially overcoming modern day antibiotic resistance. Clays with suggested antibacterial properties were examined to establish their effects on common wound-infecting bacteria. Geochemical, microscopical, and toxicological characterization of clay particulates, their suspensions and filtered leachates was performed on THP-1 and HaCaT cell lines. Cytoskeletal toxicity, cell proliferation/viability (MTT assays), and migration (scratch wounds) were further evaluated. Clays were assayed for antibacterial efficacy using minimum inhibitory concentration assays. All clays possessed a mineral content with antibacterial potential; however, clay leachates contained insufficient ions to have any antibacterial effects. All clay leachates displayed toxicity towards THP-1 monocytes, while clay suspensions showed less toxicity, suggesting immunogenicity. Reduced clay cytotoxicity on HaCaTs was shown, as many leachates stimulated wound-healing responses. The "Green" clay exhibited antibacterial effects and only in suspension, which was lost upon neutralization. pH and its interaction with clay particle surface charge is more significant than previously understood to emphasize dangers of unregulated marketing and unsubstantiated bioceutical claims.

The ubiquitin-proteasome system (UPS) is the major intracellular and non-lysosomal protein degradation system. Thanks to its unique capacity of eliminating old, damaged, misfolded, and/or regulatory proteins in a highly specific manner, the UPS is virtually involved in almost all aspects of eukaryotic life. The critical importance of the UPS is particularly visible in immune cells which undergo a rapid and profound functional remodelling upon pathogen recognition. Innate and/or adaptive immune activation is indeed characterized by a number of substantial changes impacting various cellular processes including protein homeostasis, signal transduction, cell proliferation, and antigen processing which are all tightly regulated by the UPS. In this review, we summarize and discuss recent progress in our understanding of the molecular mechanisms by which the UPS contributes to the generation of an adequate immune response. In this regard, we also discuss the consequences of UPS dysfunction and its role in the pathogenesis of recently described immune disorders including cancer and auto-inflammatory diseases.

Today, the production of wine and beer is a worldwide industry worth millions of euros annually, with breweries and wineries throughout the globe [...].

The gene coding for a novel cold-active esterase PMGL3 was previously obtained from a Siberian permafrost metagenomic DNA library and expressed in Escherichia coli. We elucidated the 3D structure of the enzyme which belongs to the hormone-sensitive lipase (HSL) family. Similar to other bacterial HSLs, PMGL3 shares a canonical α/β hydrolase fold and is presumably a dimer in solution but, in addition to the dimer, it forms a tetrameric structure in a crystal and upon prolonged incubation at 4 °C. Detailed analysis demonstrated that the crystal tetramer of PMGL3 has a unique architecture compared to other known tetramers of the bacterial HSLs. To study the role of the specific residues comprising the tetramerization interface of PMGL3, several mutant variants were constructed. Size exclusion chromatography (SEC) analysis of D7N, E47Q, and K67A mutants demonstrated that they still contained a portion of tetrameric form after heat treatment, although its amount was significantly lower in D7N and K67A compared to the wild type. Moreover, the D7N and K67A mutants demonstrated a 40 and 60% increase in the half-life at 40 °C in comparison with the wild type protein. Km values of these mutants were similar to that of the wt PMGL3. However, the catalytic constants of the E47Q and K67A mutants were reduced by ~40%.

The natural triterpene celastrol (CE) is here used as lead compound for the design and synthesis of a panel of eleven CE carboxamides that were tested in vitro for their growth inhibitory activity against Leishmania infantum and L.tropica parasites. Among them, in vitro screening identified four basic CE carboxamides endowed with nanomolar leishmanicidal activity, against both the promastigotes and the intramacrophage Leishmania amastigotes forms. These compounds also showed low toxicity toward two human (HMEC-1 and THP-1) and one murine (BMDM) cell lines. Interestingly, the most selective CE analogue (compound 3) was also endowed with the ability to inhibit the ATPase activity of the Leishmania protein chaperone Hsp90 as demonstrated by the in vitro assay conducted on a purified, full-length recombinant protein. Preliminary investigations by comparing it with the naturally occurring Hsp90 active site inhibitor Geldanamycin (GA) in two different in vitro experiments were performed. These promising results set the basis for a future biochemical investigation of the mode of interaction of celastrol and CE-inspired compounds with Leishmania Hsp90.

Pseudomonas aeruginosa is an opportunistic bacterial pathogen that employs its type III secretion system (T3SS) during the acute phase of infection to translocate cytotoxins into the host cell cytoplasm to evade the immune system. The PcrV protein is located at the tip of the T3SS, facilitates the integration of pore-forming proteins into the eukaryotic cell membrane, and is required for translocation of cytotoxins into the host cell. In this study, we used surface plasmon resonance screening to identify small molecule binders of PcrV. A follow-up structure-activity relationship analysis resulted in PcrV binders that protect macrophages in a P. aeruginosa cell-based infection assay. Treatment of P. aeruginosa infections is challenging due to acquired, intrinsic, and adaptive resistance in addition to a broad arsenal of virulence systems such as the T3SS. Virulence blocking molecules targeting PcrV constitute valuable starting points for development of next generation antibacterials to treat infections caused by P. aeruginosa.

Protein homeostasis, or proteostasis, is crucial for the functioning of a cell, as proteins that are mislocalized, present in excessive amounts, or aberrant due to misfolding or other type of damage can be harmful. Proteostasis includes attaining the correct protein structure, localization, and the formation of higher order complexes, and well as the appropriate protein concentrations. Consequences of proteostasis imbalance are evident in a range of neurodegenerative diseases characterized by protein misfolding and aggregation, such as Alzheimer's, Parkinson's, and amyotrophic lateral sclerosis. To protect the cell from the accumulation of aberrant proteins, a network of protein quality control (PQC) pathways identifies the substrates and direct them towards refolding or elimination via regulated protein degradation. The main pathway for degradation of misfolded proteins is the ubiquitin-proteasome system. PQC pathways have been first described in the cytoplasm and the endoplasmic reticulum, however, accumulating evidence indicates that the nucleus is an important PQC compartment for ubiquitination and proteasomal degradation of not only nuclear, but also cytoplasmic proteins. In this review, we summarize the nuclear ubiquitin-proteasome pathways involved in proteostasis maintenance in yeast, focusing on inner nuclear membrane-associated degradation (INMAD) and San1-mediated protein quality control.

Heliotropium is one of the most important plant genera to have conventional folklore importance, and hence is a potential source of bioactive compounds. Thus, the present study was designed to explore the therapeutic potential of Heliotropium crispum Desf., a relatively under-explored medicinal plant species. Methanolic extracts prepared from a whole plant of H. crispum were studied for phytochemical composition and possible in vitro and in silico biological properties. Antioxidant potential was assessed via six different assays, and enzyme inhibition potential against key clinical enzymes involved in neurodegenerative diseases (acetylcholinesterase (AChE) and butyrylcholinesterase (BChE)), diabetes (α-amylase and α-glucosidase), and skin problems (tyrosinase) was assayed. Phytochemical composition was established via determination of the total bioactive contents and reverse phase ultra-high performance liquid chromatography mass spectrometry (RP-UHPLC-MS) analysis. Chemical profiling revealed the tentative presence of 50 secondary metabolites. The plant extract exhibited significant inhibition against AChE and BChE enzymes, with values of 3.80 and 3.44 mg GALAE/g extract, respectively. Further, the extract displayed considerable free radical scavenging activity against DPPH and ABTS radicals, with potential values of 43.19 and 41.80 mg TE/g extract, respectively. In addition, the selected compounds were then docked against the tested enzymes, which have shown high inhibition affinity. To conclude, H. crispum was found to harbor bioactive compounds and showed potent biological activities which could be further explored for potential uses in nutraceutical and pharmaceutical industries, particularly as a neuroprotective agent.

Green synthesis of metal nanoparticles using plant extracts as capping and reducing agents for the biomedical applications has received considerable attention. Moreover, emergence and spread of multidrug resistance among bacterial pathogens has become a major health concern and lookout for novel alternative effective drugs has gained momentum. In current study, we synthesized gold nanoparticles using the seed extract of Trachyspermum ammi (TA-AuNPs), assessed its efficacy against drug resistant biofilms of Listeria monocytogenes and Serratia marcescens, and evaluated its anticancer potential against HepG2 cancer cell lines. Microwave-assisted green synthesis of gold nanoparticles was carried out and characterization was done using UV-vis spectroscopy, X-ray diffraction (XRD), transmission electron microscopy (TEM), and dynamic light scattering (DLS). Most nanoparticles were observed as spherical and spheroidal with few anisotropies with an average crystalline size of 16.63 nm. Synthesized TA-AuNPs demonstrated significant biofilm inhibitory activity against L. monocytogenes (73%) as well as S. marcescens (81%). Exopolysaccharide (EPS), motility, and CSH, key elements that facilitate the formation and maintenance of biofilm were also inhibited significantly at the tested sub-minimum inhibitory concentrations (sub-MICs). Further, TA-AuNPs effectively obliterated preformed mature biofilms of S. marcescens and L. monocytogenes by 64% and 58%, respectively. Induction of intracellular ROS production in TA-AuNPs treated bacterial cells could be the plausible mechanism for the reduced biofilm formation in test pathogens. Administration of TA-AuNPs resulted in the arrest of cellular proliferation in a concentration-dependent manner. TA-AuNPs decrease the intracellular GSH in HepG2 cancer cell lines, cells become more prone to ROS generation, hence induce apoptosis. Thus, this work proposes a new eco-friendly and rapid approach for fabricating NPs which can be exploited for multifarious biomedical applications.

There are currently no validated biomarkers which can be used to accurately diagnose Alzheimer's disease (AD) or to distinguish it from other dementia-causing neuropathologies. Moreover, to date, only symptomatic treatments exist for this progressive neurodegenerative disorder. In the search for new, more reliable biomarkers and potential therapeutic options, epigenetic modifications have emerged as important players in the pathogenesis of AD. The aim of the article was to provide a brief overview of the current knowledge regarding the role of epigenetics (including mitoepigenetics) in AD, and the possibility of applying these advances for future AD therapy. Extensive research has suggested an important role of DNA methylation and hydroxymethylation, histone posttranslational modifications, and non-coding RNA regulation (with the emphasis on microRNAs) in the course and development of AD. Recent studies also indicated mitochondrial DNA (mtDNA) as an interesting biomarker of AD, since dysfunctions in the mitochondria and lower mtDNA copy number have been associated with AD pathophysiology. The current evidence suggests that epigenetic changes can be successfully detected, not only in the central nervous system, but also in the cerebrospinal fluid and on the periphery, contributing further to their potential as both biomarkers and therapeutic targets in AD.

The unfolded protein response (UPR) is an evolutionarily conserved adaptive signaling pathway triggered by a stress of the endoplasmic reticulum (ER) lumen compartment, which is initiated by the accumulation of unfolded proteins. This response, mediated by three sensors-Inositol Requiring Enzyme 1 (IRE1), Activating Transcription Factor 6 (ATF6), and Protein Kinase RNA-Like Endoplasmic Reticulum Kinase (PERK)-allows restoring protein homeostasis and maintaining cell survival. UPR represents a major cytoprotective signaling network for cancer cells, which frequently experience disturbed proteostasis owing to their rapid proliferation in an usually unfavorable microenvironment. Increased basal UPR also participates in the resistance of tumor cells against chemotherapy. UPR activation also occurs during hematopoiesis, and growing evidence supports the critical cytoprotective role played by ER stress in the emergence and proliferation of leukemic cells. In case of severe or prolonged stress, pro-survival UPR may however evolve into a cell death program called terminal UPR. Interestingly, a large number of studies have revealed that the induction of proapoptotic UPR can also strongly contribute to the sensitization of leukemic cells to chemotherapy. Here, we review the current knowledge on the consequences of the deregulation of UPR signaling in leukemias and their implications for the treatment of these diseases.

Self-aggregation of amyloid-β (Aβ) peptides has been known to play a vital role in the onset stage of neurodegenerative diseases, indicating the necessity of understanding the aggregation process of Aβ peptides. Despite previous studies on the aggregation process of Aβ peptides, the aggregation pathways of Aβ isoforms (i.e., Aβ40 and Aβ42) and their related structures have not been fully understood yet. Here, we study the aggregation pathways of Aβ40 and Aβ42, and the structures of Aβ40 and Aβ42 aggregates during the process, based on fluorescence and atomic force microscopy (AFM) experiments. It is shown that in the beginning of aggregation process for both Aβ40 and Aβ42, a number of particles (i.e., spherical oligomers) are formed. These particles are subsequently self-assembled together, resulting in the formation of different shapes of amyloid fibrils. Our finding suggests that the different aggregation pathways of Aβ isoforms lead to the amyloid fibrils with contrasting structure.

Over the past decade, the one strain many compounds (OSMAC) approach has been established for the activation of biosynthetic gene clusters (BGCs), which mainly encode the enzymes of secondary metabolite (SM) biosynthesis pathways. These BGCs were successfully activated by altering various culture conditions, such as aeration rate, temperature, and nutrient composition. Here, we determined the biosynthetic potential of 43 bacteria using the genome mining tool antiSMASH. Based on the number of BGCs, biological safety, availability of deposited cultures, and literature coverage, we selected five promising candidates: Bacillus amyloliquefaciens DSM7, Corallococcus coralloides DSM2259, Pyxidicoccus fallax HKI727, Rhodococcus jostii DSM44719, and Streptomyces griseochromogenes DSM40499. The bacteria were cultivated under a broad range of OSMAC conditions (nutrient-rich media, minimal media, nutrient-limited media, addition of organic solvents, addition of biotic additives, and type of culture vessel) to fully assess the biosynthetic potential. In particular, we investigated so far scarcely applied OSMAC conditions to enhance the diversity of SMs. We detected the four predicted compounds bacillibactin, desferrioxamine B, myxochelin A, and surfactin. In total, 590 novel mass features were detected in a broad range of investigated OSMAC conditions, which outnumber the predicted gene clusters for all investigated bacteria by far. Interestingly, we detected mass features of the bioactive compounds cyclo-(Tyr-Pro) and nocardamin in extracts of DSM7 and DSM2259. Both compounds were so far not reported for these strains, indicating that our broad OSMAC screening approach was successful. Remarkably, the infrequently applied OSMAC conditions in defined medium with and without nutrient limitation were demonstrated to be very effective for BGC activation and for SM discovery.

The epidemic due to the novel coronavirus (SARS-CoV-2) is now a global concern, posing a severe threat to the health of populations. At present, all countries in the world are stepping up the development of vaccines and antiviral agents to prevent the infection and further transmission of SARS-CoV-2. An in-depth investigation of the target organs and pathogenesis regarding SARS-CoV-2 infection will be beneficial for virus therapy. Besides pulmonary injury, SARS-CoV-2 also causes cardiac injury, but the exact mechanisms are unclear. This review summarizes the essential structural characteristics of SARS-CoV-2 and angiotensin-converting enzyme 2 (ACE2), describes the cardiac manifestations following SARS-CoV-2 infection, and explores the mechanisms of cardiac injury targeting ACE2 after the viral invasion. We aim to help the timely detection of related symptoms and implementation of therapeutic measures by clinicians for SARS-CoV-2 infection.

Metalloproteinases (MPs) are proteolytic enzymes involved in extracellular matrix deposition, regulation of cellular signals of inflammation, proliferation, and apoptosis. Metalloproteinases are classified into three families: Matrix-MPs (MMPs), A-Disintegrin-and-Metalloprotease (ADAMs), and the A-Disintegrin-and-Metalloproteinase-with-Thrombospondin-1-like-Domains (ADAMTS). Previous studies showed that MPs are involved in the development of aortic aneurysms (AA) and, concomitantly, in the onset of chronic kidney disease (CKD). CKD has been, per se, associated with an increased risk for AA. The aim of this review is to examine the pathways that may associate MPs with CKD and AA. Several MMPs, such as MMP-2, -8, -9, and TIMP-1 have been shown to damage the AA wall and to have a toxic effect on renal tubular cells, leading to fibrosis. Similarly, ADAM10 and 17 have been shown to degrade collagen in the AA wall and to worsen kidney function via pro-inflammatory stimuli, the impairment of the Renin-Angiotensin-Aldosterone System, and the degradation of structural proteins. Moreover, MMP-2 and -9 inhibitors reduced aneurysm growth and albuminuria in experimental and human studies. It would be important, in the future, to expand research on MPs from both a prognostic, namely, to refine risk stratification in CKD patients, and a predictive perspective, likely to improve prognosis in response to targeted treatments.

The expression of glypicans in different hair follicle (HF) compartments and their potential roles during hair shaft growth are still poorly understood. Heparan sulfate proteoglycan (HSPG) distribution in HFs is classically investigated by conventional histology, biochemical analysis, and immunohistochemistry. In this report, a novel approach is proposed to assess hair histology and HSPG distribution changes in HFs at different phases of the hair growth cycle using infrared spectral imaging (IRSI). The distribution of HSPGs in HFs was probed by IRSI using the absorption region relevant to sulfation as a spectral marker. The findings were supported by Western immunoblotting and immunohistochemistry assays focusing on the glypican-1 expression and distribution in HFs. This study demonstrates the capacity of IRSI to identify the different HF tissue structures and to highlight protein, proteoglycan (PG), glycosaminoglycan (GAG), and sulfated GAG distribution in these structures. The comparison between anagen, catagen, and telogen phases shows the qualitative and/or quantitative evolution of GAGs as supported by Western immunoblotting. Thus, IRSI can simultaneously reveal the location of proteins, PGs, GAGs, and sulfated GAGs in HFs in a reagent- and label-free manner. From a dermatological point of view, IRSI shows its potential as a promising technique to study alopecia.

Continuously increasing energy demand and growing concern about energy resources has attracted much research in the field of clean and sustainable energy sources. In this context, zero-emission fuels are required for energy production to reduce the usage of fossil fuel resources. Here, we present the synthesis of Pd-Ag-decorated reduced graphene oxide (rGO) nanostructures using a green chemical approach with stevia extract for hydrogen production and antibacterial studies under light irradiation. Moreover, bimetallic nanostructures are potentially lime lighted due to their synergetic effect in both scientific and technical aspects. Structural characteristics such as crystal structure and morphological features of the synthesized nanostructures were analyzed using X-ray diffraction and transmission electron microscopy. Analysis of elemental composition and oxidation states was carried out by X-ray photoelectron spectroscopy. Optical characteristics of the biosynthesized nanostructures were obtained by UV-Vis absorption spectroscopy, and Fourier transform infrared spectroscopy was used to investigate possible functional groups that act as reducing and capping agents. The antimicrobial activity of the biosynthesized Pd-Ag-decorated rGO nanostructures was excellent, inactivating 96% of Escherichia coli cells during experiments over 150 min under visible light irradiation. Hence, these biosynthesized Pd-Ag-decorated rGO nanostructures can be utilized for alternative nanomaterial-based drug development in the future.

Kaempferitrin is extracted in significantly high quantities from the leaves of Cinnamomum osmophloeum, which belongs to a group of plant species that comes under the genus Cinnamomum, well-known for its established anti-diabetic property in Chinese medicine. Oral administration of kaempferitrin and Cinnamomum osmophloeum extract reduced blood sugar in alloxan-induced diabetic rats and improved the lipid profile in hamsters respectively. In this paper we studied the differential protein expression profile using mass spectrometry approach in the kaempferitrin-treated conditioned medium of liver cancer cell line HepG2. We discovered that 33 genes were up/down-regulated consistently between two biological samples. A slightly different version of the analysis software selected 28 genes, and the final 18 genes that appeared in both lists were selected. Interestingly, 5 proteins out of 18 were either exosomal markers or reported in high frequency of occurrence in exosome/secreted vesicles. We also examined the extracellular particles with atomic force microscopy (AFM), which showed that the conditioned medium of kaempferitrin treated had larger vesicles and fewer small vesicles. Expression of some lipid-regulating genes were also altered. Our data suggested that extracellular vesicle secretions may be regulated by kaempferitrin, and regulation of lipid profile by kampeferitrin involves multiple mechanisms.

Lectins are a class of proteins responsible for several biological roles such as cell-cell interactions, signaling pathways, and several innate immune responses against pathogens. Since lectins are able to bind to carbohydrates, they can be a viable target for targeted drug delivery systems. In fact, several lectins were approved by Food and Drug Administration for that purpose. Information about specific carbohydrate recognition by lectin receptors was gathered herein, plus the specific organs where those lectins can be found within the human body.

Plants, due to their sessile nature, have an innate immune system that helps them to defend against different pathogen infections. The defense response of plants is composed of a highly regulated and complex molecular network, involving the extensive reprogramming of gene expression during the presence of pathogenic molecular signatures. Plants attain proper defense against pathogens through the transcriptional regulation of genes encoding defense regulatory proteins and hormone signaling pathways. Small RNAs are emerging as versatile regulators of plant development and act in different tiers of plant immunity, including pathogen-triggered immunity (PTI) and effector-triggered immunity (ETI). The versatile regulatory functions of small RNAs in plant growth and development and response to biotic and abiotic stresses have been widely studied in recent years. However, available information regarding the contribution of small RNAs in plant immunity against pathogens is more limited. This review article will focus on the role of small RNAs in innate immunity in plants.

Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) or Systemic Exertion Intolerance Disease (SEID) is a chronic multisystem illness of unconfirmed etiology. There are currently no biomarkers and/or signatures available to assist in the diagnosis of the syndrome and while numerous mechanisms have been hypothesized to explain the pathology of ME/CFS, the triggers and/or drivers remain unknown. Initial studies suggested a potential role of the human herpesviruses especially Epstein-Barr virus (EBV) in the disease process but inconsistent and conflicting data led to the erroneous suggestion that these viruses had no role in the syndrome. New studies using more advanced approaches have now demonstrated that specific proteins encoded by EBV could contribute to the immune and neurological abnormalities exhibited by a subgroup of patients with ME/CFS. Elucidating the role of these herpesvirus proteins in ME/CFS may lead to the identification of specific biomarkers and the development of novel therapeutics.

Mucopolysaccharidosis type I (MPS I) is a lysosomal disease, caused by a deficiency of the enzyme alpha-L-iduronidase (IDUA). IDUA catalyzes the degradation of the glycosaminoglycans dermatan and heparan sulfate (DS and HS, respectively). Lack of the enzyme leads to pathologic accumulation of undegraded HS and DS with subsequent disease manifestations in multiple organs. The disease can be divided into severe (Hurler syndrome) and attenuated (Hurler-Scheie, Scheie) forms. Currently approved treatments consist of enzyme replacement therapy (ERT) and/or hematopoietic stem cell transplantation (HSCT). Patients with attenuated disease are often treated with ERT alone, while the recommended therapy for patients with Hurler syndrome consists of HSCT. While these treatments significantly improve disease manifestations and prolong life, a considerable burden of disease remains. Notably, treatment can partially prevent, but not significantly improve, clinical manifestations, necessitating early diagnosis of disease and commencement of treatment. This review discusses these standard therapies and their impact on common disease manifestations in patients with MPS I. Where relevant, results of animal models of MPS I will be included. Finally, we highlight alternative and emerging treatments for the most common disease manifestations.

The bacterial flagellum and the related virulence-associated injectisome system of pathogenic bacteria utilize a type III secretion system (T3SS) to export substrate proteins across the inner membrane in a proton motive force-dependent manner. The T3SS is composed of an export gate (FliPQR/FlhA/FlhB) located in the flagellar basal body and an associated soluble ATPase complex in the cytoplasm (FliHIJ). Here, we summarise recent insights into the structure, assembly and protein secretion mechanisms of the T3SS with a focus on energy transduction and protein transport across the cytoplasmic membrane.

Axonal trajectories and neural circuit activities strongly rely on a complex system of molecular cues that finely orchestrate the patterning of neural commissures. Several of these axon guidance molecules undergo continuous recycling during brain development, according to incompletely understood intracellular mechanisms, that in part rely on endocytic and autophagic cascades. Based on their pivotal role in both pathways, lysosomes are emerging as a key hub in the sophisticated regulation of axonal guidance cue delivery, localization, and function. In this review, we will attempt to collect some of the most relevant research on the tight connection between lysosomal function and axon guidance regulation, providing some proof of concepts that may be helpful to understanding the relation between lysosomal storage disorders and neurodegenerative diseases.

Extracellular signal-regulated kinase (ERK5) is an essential regulator of cancer progression, tumor relapse, and poor patient survival. Epithelial to mesenchymal transition (EMT) is a complex oncogenic process, which drives cell invasion, stemness, and metastases. Activators of ERK5, including mitogen-activated protein kinase 5 (MEK5), tumor necrosis factor α (TNF-α), and transforming growth factor-β (TGF-β), are known to induce EMT and metastases in breast, lung, colorectal, and other cancers. Several downstream targets of the ERK5 pathway, such as myocyte-specific enhancer factor 2c (MEF2C), activator protein-1 (AP-1), focal adhesion kinase (FAK), and c-Myc, play a critical role in the regulation of EMT transcription factors SNAIL, SLUG, and β-catenin. Moreover, ERK5 activation increases the release of extracellular matrix metalloproteinases (MMPs), facilitating breakdown of the extracellular matrix (ECM) and local tumor invasion. Targeting the ERK5 signaling pathway using small molecule inhibitors, microRNAs, and knockdown approaches decreases EMT, cell invasion, and metastases via several mechanisms. The focus of the current review is to highlight the mechanisms which are known to mediate cancer EMT via ERK5 signaling. Several therapeutic approaches that can be undertaken to target the ERK5 pathway and inhibit or reverse EMT and metastases are discussed.

A protein isolate (85.5%) was obtained from the Vicia faba L. seeds. The main protein fraction, typical for the seeds of this plant, was found to be most numerous: Legumin (35 kDa) and Vicilin (45 kDa). The hydrodynamic and surface properties of isolate aqueous solutions were studied with the help of dynamic light scattering, ζ-potential, and tensometry in a wide range of concentrations and pH conditions. Selected functional properties, like foaming and emulsifying abilities, were studied. An increase of water solubility was shown with a raising pH, as well as a water holding capacity (WHC). The protein isolate showed a tendency to decrease the surface tension of water solutions, with high hydrophobicity and a negative charge of the isolate enhancing the foaming and emulsifying properties. The analysis of the concentration and the pH influence on selected functional properties indicated alkaline conditions as favorable for good foaming and emulsifying properties of the isolate and affected on their rheological properties.

Propolis, a product of the honey bee, has been used in traditional medicine for many years. A hydrophobic bioactive polyphenolic ester, caffeic acid phenethyl ester (CAPE), is one of the most extensively investigated active components of propolis. Several studies have indicated that CAPE has a broad spectrum of pharmacological activities as anti-oxidant, anti-inflammatory, anti-viral, anti-fungal, anti-proliferative, and anti-neoplastic properties. This review largely describes CAPE neuroprotective effects in many different conditions and summarizes its molecular mechanisms of action. CAPE was found to have a neuroprotective effect on different neurodegenerative disorders. At the basis of these effects, CAPE has the ability to protect neurons from several underlying causes of various human neurologic diseases, such as oxidative stress, apoptosis dysregulation, and brain inflammation. CAPE can also protect the nervous system from some diseases which negatively affect it, such as diabetes, septic shock, and hepatic encephalopathy, while numerous studies have demonstrated the neuroprotective effects of CAPE against adverse reactions induced by different neurotoxic substances. The potential role of CAPE in protecting the central nervous system (CNS) from secondary injury following various CNS ischemic conditions and CAPE anti-cancer activity in CNS is also reviewed. The structure-activity relationship of CAPE synthetic derivatives is discussed as well.

Microalgae have been attracting attention as feedstock for biorefinery because they have various advantages, such as carbon fixation, high growth rate and high energy yield. The bioactive compounds and lutein contained in microalgae are known to be beneficial for human health, especially eye and brain health. In this study, in order to improve the recovery of bioactive extracts including lutein from Tetraselmis suecica with higher efficiency, an effective solvent was selected, and the extraction parameters such as temperature, time and solid loading were optimized by response surface methodology. The most effective solvent for lutein recovery was identified as 100% methanol, and the optimum condition was determined (42.4 °C, 4.0 h and 125 g/L biomass loading) by calculation of the multiple regression model. The maximum content of recovered lutein was found to be 2.79 mg/mL, and the ABTS radical scavenging activity (IC50) and ferric reducing antioxidant power (FRAP) value were about 3.36 mg/mL and 561.9 μmol/L, respectively. Finally, the maximum lutein recovery from T. suecica through statistical optimization was estimated to be 22.3 mg/g biomass, which was 3.1-fold improved compared to the control group.

Fluorescently labeled lectins are useful tools for in vivo and in vitro studies of the structure and function of tissues and various pathogens such as viruses, bacteria, and fungi. For the evaluation of high-mannose glycans present on various glycoproteins, a three-dimensional (3D) model of the chimera was designed from the crystal structures of recombinant banana lectin (BanLec, Protein Data Bank entry (PDB): 5EXG) and an enhanced green fluorescent protein (eGFP, PDB 4EUL) by applying molecular modeling and molecular mechanics and expressed in Escherichia coli. BanLec-eGFP, produced as a soluble cytosolic protein of about 42 kDa, revealed β-sheets (41%) as the predominant secondary structures, with the emission peak maximum detected at 509 nm (excitation wavelength 488 nm). More than 65% of the primary structure was confirmed by mass spectrometry. Competitive BanLec-eGFP binding to high mannose glycans of the influenza vaccine (Vaxigrip®) was shown in a fluorescence-linked lectin sorbent assay (FLLSA) with monosaccharides (mannose and glucose) and wild type BanLec and H84T BanLec mutant. BanLec-eGFP exhibited binding to mannose residues on different strains of Salmonella in flow cytometry, with especially pronounced binding to a Salmonella Typhi clinical isolate. BanLec-eGFP can be a useful tool for screening high-mannose glycosylation sites on different microorganisms.

Tourette syndrome (TS) is a disabling neurodevelopmental disorder characterized by multiple, recurrent tics. The pharmacological treatment of TS is currently based on dopaminergic antagonists; however, these drugs are associated with extrapyramidal symptoms and other serious adverse events. Recent evidence suggests that positive allosteric modulators (PAMs) of GABAA receptors containing α6 subunits (α6 GABAARs) oppose the behavioral effects of dopamine. Building on this evidence, in the present study, we tested the efficacy of DK-I-56-1, a highly selective PAM for α6 GABAARs, in mouse models of TS exhibiting tic-related responses. DK-I-56-1 significantly reduced tic-like jerks and prepulse inhibition (PPI) deficits in D1CT-7 transgenic mice, a well-documented mouse model of TS. DK-I-56-1 also prevented the exacerbation of spontaneous eyeblink reflex induced by the potent dopamine D1 receptor agonist SKF 82958, a proxy for tic-like responses. We also showed that both systemic and prefrontal cortical administration of DK-I-56-1 countered the PPI disruption caused by SKF 82958. Although the effects of DK-I-56-1 were akin to those elicited by dopaminergic antagonists, this drug did not elicit extrapyramidal effects, as measured by catalepsy. These results point to α6 GABAAR PAMs as promising TS therapies with a better safety profile than dopaminergic antagonists.

Several pathogens continuously threaten viticulture worldwide. Until now, the investigation on resistance loci has been the main trend to understand the interaction between grapevine and the mildew causal agents. Dominantly inherited gene-based resistance has shown to be race-specific in some cases, to confer partial immunity, and to be potentially overcome within a few years since its introgression. Recently, on the footprint of research conducted in Arabidopsis, putative genes associated with downy mildew susceptibility have been discovered also in the grapevine genome. In this work, we deep-sequenced four putative susceptibility genes-namely VvDMR6.1, VvDMR6.2, VvDLO1, VvDLO2-in 190 genetically diverse grapevine genotypes to discover new sources of broad-spectrum and recessively inherited resistance. Identified Single Nucleotide Polymorphisms were screened in a bottleneck analysis from the genetic sequence to their impact on protein structure. Fifty-five genotypes showed at least one impacting mutation in one or more of the scouted genes. Haplotypes were inferred for each gene and two of them at the VvDMR6.2 gene were found significantly more represented in downy mildew resistant genotypes. The current results provide a resource for grapevine and plant genetics and could corroborate genomic-assisted breeding programs as well as tailored gene editing approaches for resistance to biotic stresses.

Besides the fetal period, the suckling period is a critical time window in determining long-term metabolic health. We undertook the present study to elucidate the impact of a diabetic suckling environment alone or associated with an in utero diabetic environment on beta cell mass development and the risk of diabetes in the offspring in the long term. To that end, we have compared two experimental settings. In setting 1, we used Wistar (W) rat newborns resulting from W ovocytes (oW) transferred into diabetic GK rat mothers (pGK). These oW/pGK neonates were then suckled by diabetic GK foster mothers (oW/pGK/sGK model) and compared to oW/pW neonates suckled by normal W foster mothers (oW/pW/sW model). In setting 2, normal W rat newborns were suckled by diabetic GK rat foster mothers (nW/sGK model) or normal W foster mothers (nW/sW model). Our data revealed that the extent of metabolic disorders in term of glucose intolerance and beta cell mass are similar between rats which have been exposed to maternal diabetes both pre- and postnatally (oW/pGK/sGK model) and those which have been exposed only during postnatal life (nW/sW model). In other words, being nurtured by diabetic GK mothers from birth to weaning was sufficient to significantly alter the beta cell mass, glucose-induced insulin secretion and glucose homeostasis of offspring. No synergistic deleterious effects of pre-and postnatal exposure was observed in our setting.

Neuroblastoma (NBL) is a pediatric cancer responsible for more than 15% of cancer deaths in children, with 800 new cases each year in the United States alone. Genomic amplification of the MYC oncogene family member MYCN characterizes a subset of high-risk pediatric neuroblastomas. Several cellular models have been implemented to study this disease over the years. Two of these, SK-N-BE-2-C (BE2C) and Kelly, are amongst the most used worldwide as models of MYCN-Amplified human NBL. Here, we provide a transcriptome-wide quantitative measurement of gene expression and transcriptional network activity in BE2C and Kelly cell lines at an unprecedented single-cell resolution. We obtained 1105 Kelly and 962 BE2C unsynchronized cells, with an average number of mapped reads/cell of roughly 38,000. The single-cell data recapitulate gene expression signatures previously generated from bulk RNA-Seq. We highlight low variance for commonly used housekeeping genes between different cells (ACTB, B2M and GAPDH), while showing higher than expected variance for metallothionein transcripts in Kelly cells. The high number of samples, despite the relatively low read coverage of single cells, allowed for robust pathway enrichment analysis and master regulator analysis (MRA), both of which highlight the more mesenchymal nature of BE2C cells as compared to Kelly cells, and the upregulation of TWIST1 and DNAJC1 transcriptional networks. We further defined master regulators at the single cell level and showed that MYCN is not constantly active or expressed within Kelly and BE2C cells, independently of cell cycle phase. The dataset, alongside a detailed and commented programming protocol to analyze it, is fully shared and reusable.

Ethylene, salicylic acid (SA), and jasmonic acid are the key phytohormones involved in plant immunity, and other plant hormones have been demonstrated to interact with them. The classic phytohormone cytokinins are important participants of plant defense signaling. Crosstalk between ethylene and cytokinins has not been sufficiently studied as an aspect of plant immunity and is addressed in the present research. We compared expression of the genes responsible for hormonal metabolism and signaling in wheat cultivars differing in resistance to Stagonospora nodorum in response to their infection with fungal isolates, whose virulence depends on the presence of the necrotrophic effector SnTox3. Furthermore, we studied the action of the exogenous cytokinins, ethephon (2-chloroethylphosphonic acid, ethylene-releasing agent) and 1-methylcyclopropene (1-MCP, inhibitor of ethylene action) on infected plants. Wheat susceptibility was shown to develop due to suppression of reactive oxygen species production and decreased content of active cytokinins brought about by SnTox3-mediated activation of the ethylene signaling pathway. SnTox3 decreased cytokinin content most quickly by its activated glucosylation in an ethylene-dependent manner and, furthermore, by oxidative degradation and inhibition of biosynthesis in ethylene-dependent and ethylene-independent manners. Exogenous zeatin application enhanced wheat resistance against S. nodorum through inhibition of the ethylene signaling pathway and upregulation of SA-dependent genes. Thus, ethylene inhibited triggering of SA-dependent resistance mechanism, at least in part, by suppression of the cytokinin signaling pathway.

Recent studies undoubtedly show the importance of inter organellar connections to maintain cellular homeostasis. In normal physiological conditions or in the presence of cellular and environmental stress, each organelle responds alone or in coordination to maintain cellular function. The Endoplasmic reticulum (ER) and mitochondria are two important organelles with very specialized structural and functional properties. These two organelles are physically connected through very specialized proteins in the region called the mitochondria-associated ER membrane (MAM). The molecular foundation of this relationship is complex and involves not only ion homeostasis through the shuttling of calcium but also many structural and apoptotic proteins. IRE1alpha and PERK are known for their canonical function as an ER stress sensor controlling unfolded protein response during ER stress. The presence of these transmembrane proteins at the MAM indicates its potential involvement in other biological functions beyond ER stress signaling. Many recent studies have now focused on the non-canonical function of these sensors. In this review, we will focus on ER mitochondrial interdependence with special emphasis on the non-canonical role of ER stress sensors beyond ER stress.

Eugenyl-β-D-glucopyranoside, also referred to as Citrusin C, is a natural glucoside found among others in cloves, basil and cinnamon plants. Eugenol in a form of free aglycone is used in perfumeries, flavourings, essential oils and in medicinal products. Synthetic Citrusin C was incubated with human saliva in several in vitro models together with substrate-specific enzyme and antibiotics (clindamycin, ciprofloxacin, amoxicillin trihydrate and potassium clavulanate). Citrusin C was detected using liquid chromatography with tandem mass spectrometry (LC-MS/MS). Citrusin C was completely degraded only when incubated with substrate-specific A. niger glucosidase E.C 3.2.1.21 (control sample) and when incubated with human saliva (tested sample). The addition of antibiotics to the above-described experimental setting, stopped Citrusin C degradation, indicating microbiologic origin of hydrolysis observed. Our results demonstrate that Citrusin C is subjected to complete degradation by salivary/oral cavity microorganisms. Extrapolation of our results allows to state that in the human oral cavity, virtually all β-D-glucosides would follow this type of hydrolysis. Additionally, a new method was developed for an in vivo rapid test of glucosidase activity in the human mouth on the tongue using fluorescein-di-β-D-glucoside as substrate. The results presented in this study serve as a proof of concept for the hypothesis that microbial hydrolysis path of β-D-glucosides begins immediately in the human mouth and releases the aglycone directly into the gastrointestinal tract.

Mechanistic Target of Rapamycin (mTOR) signaling is a key regulator of cellular metabolism, integrating nutrient sensing with cell growth. Over the past two decades, studies on the mTOR pathway have revealed that mTOR complex 1 controls life span, health span, and aging by modulating key cellular processes such as protein synthesis, autophagy, and mitochondrial function, mainly through its downstream substrates. Thus, the mTOR pathway regulates both physiological and pathological processes in the heart from embryonic cardiovascular development to maintenance of cardiac homeostasis in postnatal life. In this regard, the dysregulation of mTOR signaling has been linked to many age-related pathologies, including heart failure and age-related cardiac dysfunction. In this review, we highlight recent advances of the impact of mTOR complex 1 pathway and its regulators on aging and, more specifically, cardiac aging and heart failure.

Poisoning with organophosphorus compounds used as pesticides or misused as chemical weapons remains a serious threat to human health and life. Their toxic effects result from irreversible blockade of the enzymes acetylcholinesterase and butyrylcholinesterase, which causes overstimulation of the cholinergic system and often leads to serious injury or death. Treatment of organophosphorus poisoning involves, among other strategies, the administration of oxime compounds. Oximes reactivate cholinesterases by breaking the covalent bond between the serine residue from the enzyme active site and the phosphorus atom of the organophosphorus compound. Although the general mechanism of reactivation has been known for years, the exact molecular aspects determining the efficiency and selectivity of individual oximes are still not clear. This hinders the development of new active compounds. In our research, using relatively simple and widely available molecular docking methods, we investigated the reactivation of acetyl- and butyrylcholinesterase blocked by sarin and tabun. For the selected oximes, their binding modes at each step of the reactivation process were identified. Amino acids essential for effective reactivation and those responsible for the selectivity of individual oximes against inhibited acetyl- and butyrylcholinesterase were identified. This research broadens the knowledge about cholinesterase reactivation and demonstrates the usefulness of molecular docking in the study of this process. The presented observations and methods can be used in the future to support the search for new effective reactivators.

Nanobody (Nb), a new type of biorecognition element generally from Camelidae, has the characteristics of small molecular weight, high stability, great solubility and high expression level in E. coli. In this study, with 19-nortestosterone (19-NT), an anabolic androgenic steroid as target drug, three specific Nbs against 19-NT were selected from camel immune library by phage display technology. The obtained Nbs showed excellent thermostability and organic solvent tolerance. The nanobody Nb2F7 with the best performance was used to develop a sensitive indirect competitive enzyme-linked immunosorbent assay (ic-ELISA) for 19-NT detection. Under optimized conditions, the standard curve of ic-ELISA was fitted with a half-maximal inhibitory concentration (IC50) of 1.03 ng/mL and a detection limit (LOD) of 0.10 ng/mL for 19-NT. Meanwhile, the developed assay had low cross- reactivity with analogs and the recoveries of 19-NT ranged from 82.61% to 99.24% in spiked samples. The correlation coefficient between ic-ELISA and the ultra-performance liquid chromatography/mass spectrometry (UPLC-MS/MS) method was 0.9975, which indicated that the nanobody-based ic-ELISA could be a useful tool for a rapid analysis of 19-NT in animal urine samples.

High-fat diet (HFD) is a major problem causing neuronal damage. Thymoquinone (TQ) could regulate oxidative stress and the inflammatory process. Hence, the present study elucidated the significant role of TQ on oxidative stress, inflammation, as well as morphological changes in the cerebellum of rats with HFD. Rats were divided into three groups as (1) control, (2) saturated HFD for eight weeks and (3) HFD supplementation (four weeks) followed by TQ 300 mg/kg/day treated (four weeks). After treatment, blood samples were collected to measure oxidative stress markers glutathione (GSH), malondialdehyde (MDA), superoxide dismutase (SOD), and inflammatory cytokines. Furthermore, neuronal morphological changes were also observed in the cerebellum of the rats. HFD rats show higher body weight (286.5 ± 7.4 g) as compared with the control group (224.67 ± 1.78 g). TQ treatment significantly (p < 0.05) lowered the body weight (225.83 ± 13.15 g). TQ produced a significant (p < 0.05) reduction in cholesterol, triglycerides, high-density lipoprotein (HDL), and low-density lipoprotein (LDL). The antioxidative enzymes significantly reduced in HFD rats (GSH, 1.46 ± 0.36 mol/L and SOD, 99.13 ± 5.41 µmol/mL) as compared with the control group (GSH, 6.25 ± 0.36 mol/L and SOD, 159.67 ± 10.67 µmol/mL). MDA was increased significantly in HFD rats (2.05 ± 0.25 nmol/L) compared to the control group (0.695 ± 0.11 nmol/L). Surprisingly, treatment with TQ could improve the level of GSH, MDA, and SOD. TQ treatment significantly (p < 0.05) reduced the inflammatory markers as compared with HFD alone. TQ treatment minimizes neuronal damage as well as reduces inflammation and improves antioxidant enzymes. TQ can be considered as a promising agent in preventing the neuronal morphological changes in the cerebellum of obese populations.

Inflammatory response plays an essential role in the resolution of infections. However, inflammation can be detrimental to an organism and cause irreparable damage. For example, during sepsis, a cytokine storm can lead to multiple organ failures and often results in death. One of the strongest triggers of the inflammatory response is bacterial lipopolysaccharides (LPS), acting mostly through Toll-like receptor 4 (TLR4). Paradoxically, while exposure to LPS triggers a robust inflammatory response, repeated or prolonged exposure to LPS can induce a state of endotoxin tolerance, a phenomenon where macrophages and monocytes do not respond to new endotoxin challenges, and it is often associated with secondary infections and negative outcomes. The cellular mechanisms regulating this phenomenon remain elusive. We used metabolic measurements to confirm differences in the cellular metabolism of naïve macrophages and that of macrophages responding to LPS stimulation or those in the LPS-tolerant state. In parallel, we performed an unbiased secretome survey using quantitative mass spectrometry during the induction of LPS tolerance, creating the first comprehensive secretome profile of endotoxin-tolerant cells. The secretome changes confirmed that LPS-tolerant macrophages have significantly decreased cellular metabolism and that the proteins secreted by LPS-tolerant macrophages have a strong association with cell survival, protein metabolism, and the metabolism of reactive oxygen species.

The aim of the present study was to examine the protective effect of γ-mangostin, a component of the mangosteen shell, against oxidative damage to nerve cells induced by excessive glutamate, a known excitatory neurotransmitter. To investigate the effect of γ-mangostin on apoptosis, 5 mM of glutamate was used to induce apoptotic cell death in mouse hippocampal HT22 cells. In this study, γ-mangostin was found to exert a stronger protection than N-acetyl cysteine against glutamate-induced cell damage. γ-Mangostin showed prevented glutamate-induced apoptosis in HT22 cells by reducing the production of reactive oxygen species and stimulating the expression of heme oxygenase-1 protein. In addition, glutamate significantly induced the accumulation of intracellular calcium ions, whereas treatment with γ-mangostin markedly reduced it. Hoechst 33342 staining showed an improvement in glutamate-induced nuclear condensation following γ-mangostin treatment. Furthermore, the number of annexin V-positive cells was significantly reduced following treatment with γ-mangostin. Western blot analysis showed the inhibition of glutamate-induced mitogen-activated protein kinase phosphorylation by γ-mangostin. γ-mangostin also inhibited the regulation of the intrinsic mitochondrial apoptotic pathway. Thus, the results of this study suggest that γ-mangostin is an active ingredient of mangosteen and exerts neuroprotective activities in HT22 cells.

Acute intense exercise causes significant oxidative stress and consequently an increase in total antioxidant capacity; however, the mechanisms and combined effects of intense exercise and smoking on oxidative stress among active and non-active smokers are not clear. The aim of this study was to investigate the effect of acute intense exercise on antioxidant enzyme activity responses in active and non-active individuals exposed to cigarette smoke. The study included 40 subjects who were equally classified as: smokers that did exercise (SE), smokers that did not do exercise (SnE), non-smokers that did exercise (NSE), and non-smokers that did not do exercise (NSnE). The adjusted Astrand test was used to exhaust the subjects. Salivary enzymes of peroxidase (POX), catalase (CAT), and superoxide dismutase (SOD) were measured, by spectrophotometry methods, at 3 different time points: pre-test (TP1), post-test (TP2), and one hour after finishing the test (TP3). Significant (p < 0.05) group x time interactions were found for the three enzymes. Salivary POX, CAT and SOD increased in all groups from TP1 to TP2 and decreased from TP2 to TP3. Only the NSE showed a significant difference between TP1 to TP3 in POX and SOD by +0.011 ± 0.007 and +0.075 ± 0.020 (U/mL), respectively. The NSE showed significantly higher activity of POX, CAT and SOD in TP2 compared to the other groups. Furthermore, NSE and NSnE had higher activity of POX, CAT and SOD in TP1 and TP3 (p < 0.05) compared with SE and SnE. Only in the NSnE, were no differences observed in CAT compared with SE and SnE in TP3. These results showed that the antioxidant activity at rest and in the recovery time after the acute intense exercise was lower in SE and SnE compared with NSE and NSnE, suggesting that smoking habit may reduce the ameliorating effect of regular physical activity on acute exercise-induced oxidative stress.

Extracorporeal life support provides perfusion for patients with heart failure to allow time for recovery, function as a bridge for patients to heart transplantation, or serve as destination therapy for long term mechanical device support. Several biomarkers have been employed in attempt to predict these outcomes, but it remains to be determined which are suitable to guide clinical practice relevant to extracorporeal life support. Galectin-3 and soluble suppression of tumorigenicity-2 (sST2) are two of the more promising candidates with the greatest supporting evidence. In this review, we address the similarities and differences between galectin-3 and sST2 for prognostic prediction in adults and children with heart failure requiring extracorporeal life support and highlight the significant lack of progress in pediatric biomarker discovery and utilization.

Ulcerative colitis (UC), one of the two main types of inflammatory bowel disease, has no effective treatment. Rosmarinic acid (RA) is a polyphenol that, when administered orally, is metabolised in the small intestine, compromising its beneficial effects. We used chitosan/nutriose-coated niosomes loaded with RA to protect RA from gastric degradation and target the colon and evaluated their effect on acute colitis induced by 4% dextran sodium sulphate (DSS) for seven days in mice. RA-loaded nanovesicles (5, 10 and 20 mg/kg) or free RA (20 mg/kg) were orally administered from three days prior to colitis induction and during days 1, 3, 5 and 7 of DSS administration. RA-loaded nanovesicles improved body weight loss and disease activity index as well as increased mucus production and decreased myeloperoxidase activity and TNF-α production. Moreover, RA-loaded nanovesicles downregulated protein expression of inflammasome components such as NLR family pyrin domain-containing 3 (NLRP3), adaptor protein (ASC) and caspase-1, and the consequent reduction of IL-1β levels. Furthermore, nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) protein expression increased after the RA-loaded nanovesicles treatment However, these mechanistic changes were not detected with the RA-free treatment. Our findings suggest that the use of chitosan/nutriose-coated niosomes to increase RA local bioavailability could be a promising nutraceutical strategy for oral colon-targeted UC therapy.

Myotonic dystrophy type 1 (DM1), the most frequent inherited muscular dystrophy in adults, is caused by the CTG repeat expansion in the 3'UTR of the DMPK gene. Mutant DMPK RNA accumulates in nuclear foci altering diverse cellular functions including alternative splicing regulation. DM1 is a multisystemic condition, with debilitating central nervous system alterations. Although a defective neuroglia communication has been described as a contributor of the brain pathology in DM1, the specific cellular and molecular events potentially affected in glia cells have not been totally recognized. Thus, to study the effects of DM1 mutation on glial physiology, in this work, we have established an inducible DM1 model derived from the MIO-M1 cell line expressing 648 CUG repeats. This new model recreated the molecular hallmarks of DM1 elicited by a toxic RNA gain-of-function mechanism: accumulation of RNA foci colocalized with MBNL proteins and dysregulation of alternative splicing. By applying a microarray whole-transcriptome approach, we identified several gene changes associated with DM1 mutation in MIO-M1 cells, including the immune mediators CXCL10, CCL5, CXCL8, TNFAIP3, and TNFRSF9, as well as the microRNAs miR-222, miR-448, among others, as potential regulators. A gene ontology enrichment analyses revealed that inflammation and immune response emerged as major cellular deregulated processes in the MIO-M1 DM1 cells. Our findings indicate the involvement of an altered immune response in glia cells, opening new windows for the study of glia as potential contributor of the CNS symptoms in DM1.

The anticancer activities of Withaferin-A (Wi-A) and Withanone (Wi-N) from Ashwagandha and Caffeic Acid Phenethyl Ester (CAPE) from honeybee propolis have been well documented. Here, we examined the binding potential of these natural compounds to inhibit the constitutive phosphorylation of epidermal growth factor receptors (EGFRs). Exon 20 insertion mutants of EGFR, which show resistance to various FDA approved drugs and are linked to poor prognosis of lung cancer patients, were the primary focus of this study. Apart from exon 20 insertion mutants, the potential of natural compounds to serve as ATP competitive inhibitors of wildtype protein and other common mutants of EGFR, namely L858R and exon19del, were also examined. The potential of natural compounds was compared to the positive controls such as erlotinib, TAS6417 and poziotinib. Similar to known inhibitors, Wi-A and Wi-N could displace and binds at the ATP orthosteric site of exon19del, L858R and exon20, while CAPE was limited to wildtype EGFR and exon 20 insertion mutants only. Moreover, the binding free energy of the natural drugs against EGFRs was also comparable to the positive controls. This computational study suggests that Wi-A and Wi-N have potential against multiple mutated EGFRs, warranting further in vitro and in vivo experiments.

In vivo clinical applications of nanobodies (VHHs) require molecules that induce minimal immunoresponse and therefore possess sequences as similar as possible to the human VH domain. Although the relative sequence variability in llama nanobodies has been used to identify scaffolds with partially humanized signature, the transformation of the Camelidae hallmarks in the framework2 still represents a major problem. We assessed a set of mutants in silico and experimentally to elucidate what is the contribution of single residues to the VHH stability and how their combinations affect the mutant nanobody stability. We described at molecular level how the interaction among residues belonging to different structural elements enabled a model llama nanobody (C8WT, isolated from a naïve library) to be functional and maintain its stability, despite the analysis of its primary sequence would classify it as aggregation-prone. Five chimeras formed by grafting CDRs isolated from different nanobodies into C8WT scaffold were successfully expressed as soluble proteins and both tested clones preserved their antigen binding specificity. We identified a nanobody with human hallmarks that seems suitable for humanizing selected camelid VHHs by grafting heterologous CDRs in its scaffold and could serve for the preparation of a synthetic library of human-like single domains.

In contrast with some adult human organs, such as liver or skin, the adult human heart shows very limited self-regeneration capacity, attributed to the negligible presence of resident cardiac stem cells or cardiac progenitors [...].

GPR56 is required for the adipogenesis of preadipocytes, and the role of one of its ligands, type III collagen (ColIII), was investigated here. ColIII expression was examined by reverse transcription quantitative polymerase chain reaction, immunoblotting and immunostaining, and its function investigated by knockdown and genome editing in 3T3-L1 cells. Adipogenesis was assessed by oil red O staining of neutral cell lipids and production of established marker and regulator proteins. siRNA-mediated knockdown significantly reduced Col3a1 transcripts, ColIII protein and lipid accumulation in 3T3-L1 differentiating cells. Col3a1-/- 3T3-L1 genome-edited cell lines abolished adipogenesis, demonstrated by a dramatic reduction in adipogenic moderators: Pparγ2 (88%) and C/ebpα (96%) as well as markers aP2 (93%) and oil red O staining (80%). Col3a1-/- 3T3-L1 cells displayed reduced cell adhesion, sustained active β-catenin and deregulation of fibronectin (Fn) and collagen (Col4a1, Col6a1) extracellular matrix gene transcripts. Col3a1-/- 3T3-L1 cells also had dramatically reduced actin stress fibres. We conclude that ColIII is required for 3T3-L1 preadipocyte adipogenesis as well as the formation of actin stress fibres. The phenotype of Col3a1-/- 3T3-L1 cells is very similar to that of Gpr56-/- 3T3-L1 cells, suggesting a functional relationship between ColIII and Gpr56 in preadipocytes.

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Photodynamic therapy (PDT) is frequently used to treat non-muscle invasive bladder cancer due its low toxicity and high selectivity. Since recurrence often occurs, alternative approaches and/or designs of combined therapies to improve PDT effectiveness are needed. This work aimed to evaluate the cytotoxicity of 4,6,4'-trimethylangelicin (TMA) photoactivated by blue light (BL) on human bladder cancer T24 cells and investigate the mechanisms underlying its biological effects. TMA/BL exerted antiproliferative activity through the induction of apoptosis without genotoxicity, as demonstrated by the expression levels of phospho-H2AX, an indicator of DNA double-stranded breaks. It also modulated the Wnt canonical signal pathway by increasing the phospho-β-catenin and decreasing the nuclear levels of β-catenin. The inhibition of this pathway was due to the modulation of the GSK3β phosphorylation state (Tyr 216) that induces a proteasomal degradation of β-catenin. Indeed, a partial recovery of nuclear β-catenin expression and reduction of its phosphorylated form after treatment with LiCl were detected. As demonstrated by RT-PCR and cytofluorimetric analysis, TMA/BL also decreased the expression of CD44v6, a marker of cancer stem cells. Taken together, our data suggest that TMA photoactivated by BL may represent an interesting option for the photochemotherapy of noninvasive bladder carcinomas, since this treatment is able to inhibit key pathways for tumour growth and progression in the absence of genotoxic effects.

BYL719 (alpelisib) is a small molecule inhibitor of PI3K p110α developed for cancer therapy. Targeted suppression of PI3K has led to lifespan extension in rodents and model organisms. If PI3K inhibitors are to be considered as an aging therapeutic, it is important to understand the potential consequences of long-term exposure, and the most practical way to achieve this is through diet administration. Here, we investigated the pharmacokinetics of BYL719 delivered in diet and the efficacy of BYL719 to suppress insulin signaling when administered in the diet of 8-month-old male and female mice. Compared to oral gavage, diet incorporation resulted in a lower peak plasma BYL719 (3.6 vs. 9.2 μM) concentration but similar half-life (~1.5 h). Consuming BYL719 resulted in decreased insulin signaling in liver and muscle within 72 h, and mice still showed impaired glucose tolerance and insulin sensitivity following 6 weeks of access to a diet containing 0.3 g/kg BYL719. However, consuming BYL719 did not affect food intake, body mass, muscle function (rotarod and hang time performance) or cognitive behaviors. This provides evidence that BYL719 has long-term efficacy without major toxicity or side effects, and suggests that administering BYL719 in diet is suitable for studying the effect of pharmacological suppression of PI3K p110α on aging and metabolic function.

Cardiovascular diseases (CVDs) are seriously threatening to human life and health. Polyunsaturated fatty acids (PUFAs) are known for their role in preventing CVDs. It is beneficial to population health to promote the content of PUFAs in bovine milk. In recent years, limited research based on molecular mechanisms has focused on this field. The biological roles of numerous microRNAs (miRNAs) remain unknown. In this study, a promising and negatively correlated pair of the miRNA (miRNA-193a-5p) and a fatty acid desaturase 1 (FADS1) gene are identified and screened to explore whether they are potential factors of PUFAs' synthesis in bovine milk. The targeted relationship between miRNA-193a-5p and FADS1 in bovine mammary epithelial cells (BMECs) is demonstrated by dual luciferase reporter assays. qRT-PCR and western blot assays indicate that both the expression of mRNA and the protein FADS1 show a negative correlation with miRNA-193a-5p expression in BMECs. Also, miR-193a-5p expression is positively correlated with the expression of genes associated with milk fatty acid metabolism, including ELOVL fatty acid elongase 6 (ELOVL6) and diacylglycerol O-acyltransferase 2 (DGAT2). The expression of fatty acid desaturase 2 (FADS2) is negatively correlated with miR-193a-5p expression in BMECs. The contents of triglycerides (TAG), eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA) have a significant positive correlation with the expression of FADS1 and a significant negative correlation with the expression of miR-193a-5p in BMECs. For the first time, this study confirms that miRNA-193a-5p regulates PUFAs metabolism in BMECs by targeting FADS1, indicating that miRNA-193a-5p and FADS1 are underlying factors that improve PUFAs content in bovine milk.

Hair follicle outer root sheath (ORS) is a putative source of stem cells with therapeutic capacity. ORS contains several multipotent stem cell populations, primarily in the distal compartment of the bulge region. However, the bulge is routinely obtained using invasive isolation methods, which require human scalp tissue ex vivo. Non-invasive sampling has been standardized by means of the plucking procedure, enabling to reproducibly obtain the mid-ORS part. The mid-ORS shows potential for giving rise to multiple stem cell populations in vitro. To demonstrate the phenotypic features of distal, middle, and proximal ORS parts, gene and protein expression profiles were studied in physically separated portions. The mid-part of the ORS showed a comparable or higher NGFR, nestin/NES, CD34, CD73, CD44, CD133, CK5, PAX3, MITF, and PMEL expression on both protein and gene levels, when compared to the distal ORS part. Distinct subpopulations of cells exhibiting small and round morphology were characterized with flow cytometry as simultaneously expressing CD73/CD271, CD49f/CD105, nestin, and not CK10. Potentially, these distinct subpopulations can give rise to cultured neuroectodermal and mesenchymal stem cell populations in vitro. In conclusion, the mid part of the ORS holds the potential for yielding multiple stem cells, in particular mesenchymal stem cells.

The necrosome is a large-molecular-weight complex in which the terminal effector of the necroptotic pathway, Mixed Lineage Kinase Domain-Like protein (MLKL), is activated to induce necroptotic cell death. The precise mechanism of MLKL activation by the upstream kinase, Receptor Interacting Serine/Threonine Protein Kinase 3 (RIPK3) and the role of Receptor Interacting Serine/Threonine Protein Kinase 1 (RIPK1) in mediating MLKL activation remain incompletely understood. Here, we reconstituted human necrosome interactions in yeast by inducible expression of these necrosome effectors. Functional interactions were reflected by the detection of phosphorylated MLKL, plasma membrane permeabilization, and reduced proliferative potential. Following overexpression of human necrosome effectors in yeast, MLKL aggregated in the periphery of the cell, permeabilized the plasma membrane and compromised clonogenic potential. RIPK1 had little impact on RIPK3/MLKL-mediated yeast lethality; however, it exacerbated the toxicity provoked by co-expression of MLKL with a RIPK3 variant bearing a mutated RHIM-domain. Small molecule necroptotic inhibitors necrostatin-1 and TC13172, and viral inhibitors M45 (residues 1-90) and BAV_Rmil, abated the yeast toxicity triggered by the reconstituted necrosome. This yeast model provides a convenient tool to study necrosome protein interactions and to screen for and characterize potential necroptotic inhibitors.

Lipoxygenases are widespread enzymes that catalyze oxidation of polyunsaturated fatty acids (linoleic, linolenic, and arachidonic acid) to produce hydroperoxides. Lipoxygenase reactions can be desirable, but also lipoxygenases can react in undesirable ways. Most of the products of lipoxygenase reactions are aromatic compounds that can affect food properties, especially during long-term storage. Lipoxygenase action on unsaturated fatty acids could result in off-flavor/off-odor development, causing food spoilage. In addition, lipoxygenases are present in the human body and play an important role in stimulation of inflammatory reactions. Inflammation is linked to many diseases, such as cancer, stroke, and cardiovascular and neurodegenerative diseases. This review summarized recent research on plant families and species that can inhibit lipoxygenase activity.

Four decades of proteasome research have yielded extensive information on ubiquitin-dependent proteolysis. The archetype of proteasomes is a 20S barrel-shaped complex that does not rely on ubiquitin as a degradation signal but can degrade substrates with a considerable unstructured stretch. Since roughly half of all proteasomes in most eukaryotic cells are free 20S complexes, ubiquitin-independent protein degradation may coexist with ubiquitin-dependent degradation by the highly regulated 26S proteasome. This article reviews recent advances in our understanding of the biochemical and structural features that underlie the proteolytic mechanism of 20S proteasomes. The two outer α-rings of 20S proteasomes provide a number of potential docking sites for loosely folded polypeptides. The binding of a substrate can induce asymmetric conformational changes, trigger gate opening, and initiate its own degradation through a protease-driven translocation mechanism. Consequently, the substrate translocates through two additional narrow apertures augmented by the β-catalytic active sites. The overall pulling force through the two annuli results in a protease-like unfolding of the substrate and subsequent proteolysis in the catalytic chamber. Although both proteasomes contain identical β-catalytic active sites, the differential translocation mechanisms yield distinct peptide products. Nonoverlapping substrate repertoires and product outcomes rationalize cohabitation of both proteasome complexes in cells.

We report an extremely rare case of combined classical and periodontal Ehlers-Danlos syndrome (EDS) with early severe periodontitis and a generalized lack of attached gingiva. A German family with classical EDS was investigated by physical and dental evaluation and exome and Sanger sequencing. Due to the specific periodontal phenotype in the affected child, an additional diagnosis of periodontal EDS was suspected. Physical and genetic examination of two affected and three unaffected family members revealed a family diagnosis of classical EDS with a heterozygous mutation in COL5A1 (c.1502del; p.Pro501Leufs*57). Additional to the major clinical criteria for classical EDS-generalized joint hypermobility, hyperelastic skin, and atrophic scarring -the child aged four years presented with generalized alveolar bone loss up to 80%, early loss of two lower incisors, severe gingival recession, and generalized lack of attached gingiva. Due to these clinical findings, an additional diagnosis of periodontal EDS was suspected. Further genetic analysis revealed the novel missense mutation c.658T>G (p.Cys220Gly) in C1R in a heterozygous state. Early severe periodontitis in association with generalized lack of attached gingiva is pathognomonic for periodontal EDS and led to the right clinical and genetic diagnosis in the present case.

Nucleotides released by smooth muscle cells (SMCs) and by innervating nerve terminals activate specific P2 receptors and modulate bladder contraction. We hypothesized that cell surface enzymes regulate SMC contraction in mice bladder by controlling the concentration of nucleotides. We showed by immunohistochemistry, enzymatic histochemistry, and biochemical activities that nucleoside triphosphate diphosphohydrolase-1 (NTPDase1) and ecto-5'-nucleotidase were the major ectonucleotidases expressed by SMCs in the bladder. RT-qPCR revealed that, among the nucleotide receptors, there was higher expression of P2X1, P2Y1, and P2Y6 receptors. Ex vivo, nucleotides induced a more potent contraction of bladder strips isolated from NTPDase1 deficient (Entpd1-/-) mice compared to wild type controls. The strongest responses were obtained with uridine 5'-triphosphate (UTP) and uridine 5'-diphosphate (UDP), suggesting the involvement of P2Y6 receptors, which was confirmed with P2ry6-/- bladder strips. Interestingly, this response was reduced in female bladders. Our results also suggest the participation of P2X1, P2Y2 and/or P2Y4, and P2Y12 in these contractions. A reduced response to the thromboxane analogue U46619 was also observed in wild type, Entpd1-/-, and P2ry6-/- female bladders showing another difference due to sex. In summary, NTPDase1 modulates the activation of nucleotide receptors in mouse bladder SMCs, and contractions induced by P2Y6 receptor activation were weaker in female bladders.

The HPC-1/syntaxin 1A (Stx1a) gene, which is involved in synaptic transmission and neurodevelopmental disorders, is a TATA-less gene with several transcription start sites. It is activated by the binding of Sp1 and acetylated histone H3 to the -204 to +2 core promoter region (CPR) in neuronal cell/tissue. Furthermore, it is depressed by the association of class 1 histone deacetylases (HDACs) to Stx1a-CPR in non-neuronal cell/tissue. To further clarify the factors characterizing Stx1a gene silencing in non-neuronal cell/tissue not expressing Stx1a, we attempted to identify the promoter region forming DNA-protein complex only in non-neuronal cells. Electrophoresis mobility shift assays (EMSA) demonstrated that the -183 to -137 OL2 promoter region forms DNA-protein complex only in non-neuronal fetal rat skin keratinocyte (FRSK) cells which do not express Stx1a. Furthermore, the Yin-Yang 1 (YY1) transcription factor binds to the -183 to -137 promoter region of Stx1a in FRSK cells, as shown by competitive EMSA and supershift assay. Chromatin immunoprecipitation assay revealed that YY1 in vivo associates to Stx1a-CPR in cell/tissue not expressing Stx1a and that trichostatin A treatment in FRSK cells decreases the high-level association of YY1 to Stx1a-CPR in default. Reporter assay indicated that YY1 negatively regulates Stx1a transcription. Finally, mass spectrometry analysis showed that gene silencing factors, including HDAC1, associate onto the -183 to -137 promoter region together with YY1. The current study is the first to report that Stx1a transcription is negatively regulated in a cell/tissue-specific manner by YY1 transcription factor, which binds to the -183 to -137 promoter region together with gene silencing factors, including HDAC.

The bidirectional microbiota-gut-brain axis has raised increasing interest over the past years in the context of health and disease, but there is a lack of information on molecular mechanisms underlying this connection. We hypothesized that change in microbiota composition may affect brain epigenetics leading to long-lasting effects on specific brain gene regulation. To test this hypothesis, we used Zebrafish (Danio Rerio) as a model system. As previously shown, treatment with high doses of probiotics can modulate behavior in Zebrafish, causing significant changes in the expression of some brain-relevant genes, such as BDNF and Tph1A. Using an ultra-deep targeted analysis, we investigated the methylation state of the BDNF and Tph1A promoter region in the brain and gut of probiotic-treated and untreated Zebrafishes. Thanks to the high resolution power of our analysis, we evaluated cell-to-cell methylation differences. At this resolution level, we found slight DNA methylation changes in probiotic-treated samples, likely related to a subgroup of brain and gut cells, and that specific DNA methylation signatures significantly correlated with specific behavioral scores.

Hexavalent chromium [Cr(VI)] pollution is one of most serious heavy metal pollutants in the coastal area and posed serious threats to marine organisms and human beings. Many studies have been conducted on its toxicological effects on living organisms from morphological to physiological aspects. However, there are few studies about the transgenerational toxicological of Cr(VI). In this study, we exposed adult marine medaka fishes with Cr(VI) and their offspring with Cr(VI) to examine transgenerational effects of Cr(VI). We found that there were mechanisms such as changing reproduction modes in males to compensate for impacts on the reproduction. There were differences and similarities between the parental effect and the environmental effect, with the former one causing more serious adverse effects on the offspring of Cr(VI)-exposed fish. It was noteworthy that there was an interaction between the parental and offspring treatment which leads to the attenuation of the parental effects on offspring when the offspring also underwent the same treatment. In addition, physiological adaptation has also been observed in fish to improve their fitness. Overall, effects of Cr(VI) on fish and their offspring were studied to pave a way to study the of mechanisms of adaptation.

The binding of known odorant molecules to the human odorant-binding protein (hOBP) was evaluated in silico. Docking experiments elucidate the preferable binding site and binding affinity of odorant molecules to hOBP. The physicochemical properties molecular weight (MW), vapor pressure (Vp), hydrophobicity level (logP), number of double bonds (NºDB), degree of unsaturation (DoU) and the chemical classification, were selected for the study of odorant modulation. Here, these properties were analyzed concerning 30 pleasant and 30 unpleasant odorants, chosen to represent a wide variety of compounds and to determine their influence on the binding energy to hOBP. Our findings indicate that MW, logP and Vp are the most important odorant variables, directly correlated to odorant-binding energies (DGbinding) towards hOBP. Understanding how the odorants behave when complexed with the OBP in human olfaction opens new possibilities for the development of future biotechnological applications, including sensory devices, medical diagnosis, among others.

Endophytic fungi are widely present in internal plant tissues and provide different benefits to their host. Medicinal plants have unexplored diversity of functional fungal association; therefore, this study aimed to isolate endophytic fungi associated with leaves of medicinal plants Ephedra pachyclada and evaluate their plant growth-promoting properties. Fifteen isolated fungal endophytes belonging to Ascomycota, with three different genera, Penicillium, Alternaria, and Aspergillus, were obtained from healthy leaves of E. pachyclada. These fungal endophytes have varied antimicrobial activity against human pathogenic microbes and produce ammonia and indole acetic acid (IAA), in addition to their enzymatic activity. The results showed that Penicillium commune EP-5 had a maximum IAA productivity of 192.1 ± 4.04 µg mL-1 in the presence of 5 µg mL-1 tryptophan. The fungal isolates of Penicillium crustosum EP-2, Penicillium chrysogenum EP-3, and Aspergillus flavus EP-14 exhibited variable efficiency for solubilizing phosphate salts. Five representative fungal endophytes of Penicillium crustosum EP-2, Penicillium commune EP-5, Penicillium caseifulvum EP-11, Alternaria tenuissima EP-13, and Aspergillus flavus EP-14 and their consortium were selected and applied as bioinoculant to maize plants. The results showed that Penicillium commune EP-5 increased root lengths from 15.8 ± 0.8 to 22.1 ± 0.6. Moreover, the vegetative growth features of inoculated maize plants improved more than the uninoculated ones.

Parkinson's disease (PD) is a clinically heterogenic disorder characterized by distinct clinical entities. Most studies on motor deficits dichotomize PD into tremor dominant (TD) or non-tremor dominant (non-TD) with akinetic-rigid features (AR). Different pathophysiological mechanisms may affect the onset of motor manifestations. Recent studies have suggested that gut microbes may be involved in PD pathogenesis. The aim of this study was to investigate the gut microbiota and metabolome composition in PD patients in relation to TD and non-TD phenotypes. In order to address this issue, gut microbiota and the metabolome structure of PD patients were determined from faecal samples using 16S next generation sequencing and gas chromatography-mass spectrometry approaches. The results showed a reduction in the relative abundance of Lachnospiraceae, Blautia, Coprococcus, Lachnospira, and an increase in Enterobacteriaceae, Escherichia and Serratia linked to non-TD subtypes. Moreover, the levels of important molecules (i.e., nicotinic acid, cadaverine, glucuronic acid) were altered in relation to the severity of phenotype. We hypothesize that the microbiota/metabolome enterotypes associated to non-TD subtypes may favor the development of gut inflammatory environment and gastrointestinal dysfunctions and therefore a more severe α-synucleinopathy. This study adds important information to PD pathogenesis and emphasizes the potential pathophysiological link between gut microbiota/metabolites and PD motor subtypes.

CpG methylation in transposons, exons, introns and intergenic regions is important for long-term silencing, silencing of parasitic sequences and alternative promoters, regulating imprinted gene expression and determining X chromosome inactivation. Promoter CpG islands, although rich in CpG dinucleotides, are unmethylated and remain so during all phases of mammalian embryogenesis and development, except in specific cases. The biological mechanisms that contribute to the maintenance of the unmethylated state of CpG islands remain elusive, but the modification of established DNA methylation patterns is a common feature in all types of tumors and is considered as an event that intrinsically, or in association with genetic lesions, feeds carcinogenesis. In this review, we focus on the latest results describing the role that the levels of H3K4 trimethylation may have in determining the aberrant hypermethylation of CpG islands in tumors.

The mitochondrial citrate/isocitrate carrier, CIC, has been shown to play an important role in a growing list of human diseases. CIC belongs to a large family of nuclear-encoded mitochondrial transporters that serve the fundamental function of allowing the transit of ions and metabolites through the impermeable mitochondrial membrane. Citrate is central to mitochondrial metabolism and respiration and plays fundamental activities in the cytosol, serving as a metabolic substrate, an allosteric enzymatic regulator and, as the source of Acetyl-Coenzyme A, also as an epigenetic modifier. In this review, we highlight the complexity of the mechanisms of action of this transporter, describing its involvement in human diseases and the therapeutic opportunities for targeting its activity in several pathological conditions.

Urm1 (ubiquitin related modifier 1) is a molecular fossil in the class of ubiquitin-like proteins (UBLs). It encompasses characteristics of classical UBLs, such as ubiquitin or SUMO (small ubiquitin-related modifier), but also of bacterial sulfur-carrier proteins (SCP). Since its main function is to modify tRNA, Urm1 acts in a non-canonical manner. Uba4, the activating enzyme of Urm1, contains two domains: a classical E1-like domain (AD), which activates Urm1, and a rhodanese homology domain (RHD). This sulfurtransferase domain catalyzes the formation of a C-terminal thiocarboxylate on Urm1. Thiocarboxylated Urm1 is the sulfur donor for 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2U), a chemical nucleotide modification at the wobble position in tRNA. This thio-modification is conserved in all domains of life and optimizes translation. The absence of Urm1 increases stress sensitivity in yeast triggered by defects in protein homeostasis, a hallmark of neurological defects in higher organisms. In contrast, elevated levels of tRNA modifying enzymes promote the appearance of certain types of cancer and the formation of metastasis. Here, we summarize recent findings on the unique features that place Urm1 at the intersection of UBL and SCP and make Urm1 an excellent model for studying the evolution of protein conjugation and sulfur-carrier systems.

Recent evidence suggests that the reason Extra Virgin Olive Oil (EVOO) lowers blood pressure and reduces the risk of developing hypertension is partly due to minor components of EVOO, such as phenols. However, little is still known about the mechanism(s) through which EVOO phenols mediate anti-hypertensive effects. The aim of the present study was to investigate the mechanisms of action of EVOO phenols on mesenteric resistance arteries. A pressure myograph was used to test the effect of EVOO phenols on isolated mesenteric arteries in the presence of specific inhibitors of: 1) BKca channels (Paxillin, 10-5 M); 2) L-type calcium channels (Verapamil, 10-5 M); 3) Ryanodine receptor, RyR (Ryanodine, 10-5 M); 4) inositol 1,4,5-triphosphate receptor, IP3R, (2-Aminoethyl diphenylborinate, 2-APB, 3 × 10-3 M); 5) phospholipase C, PLC, (U73122, 10-5 M), and 6) GPCR-Gαi signaling, (Pertussis Toxin, 10-5 M). EVOO phenols induced vasodilation of mesenteric arteries in a dose-dependent manner, and this effect was reduced by pre-incubation with Paxillin, Verapamil, Ryanodine, 2-APB, U73122, and Pertussis Toxin. Our data suggest that EVOO phenol-mediated vasodilation requires activation of BKca channels potentially through a local increase of subcellular calcium microdomains, a pivotal mechanism on the base of artery vasodilation. These findings provide novel mechanistic insights for understanding the vasodilatory properties of EVOO phenols on resistance arteries.

In an era when cancer glycobiology research is exponentially growing, we are witnessing a progressive translation of the major scientific findings to the clinical practice with the overarching aim of improving cancer patients' management. Many mechanistic cell biology studies have demonstrated that heparan sulfate (HS) glycosaminoglycans are key molecules responsible for several molecular and biochemical processes, impacting extracellular matrix properties and cellular functions. HS can interact with a myriad of different ligands, and therefore, hold a pleiotropic role in regulating the activity of important cellular receptors and downstream signalling pathways. The aberrant expression of HS glycan chains in tumours determines main malignant features, such as cancer cell proliferation, angiogenesis, invasion and metastasis. In this review, we devote particular attention to HS biological activities, its expression profile and modulation in cancer. Moreover, we highlight HS clinical potential to improve both diagnosis and prognosis of cancer, either as HS-based biomarkers or as therapeutic targets.

The discovery and development of actinomycete secondary metabolites (ASMs) have played pivotal roles in the fields of human medicine and its related biotechnology sectors over the past several decades [...].

Autophagy, which is a conserved biological process and essential mechanism in maintaining homeostasis and metabolic balance, enables cells to degrade cytoplasmic constituents through lysosomes, recycle nutrients, and survive during starvation. Autophagy exerts an anticarcinogenic role in normal cells and inhibits the malignant transformation of cells. On the other hand, aberrations in autophagy are involved in gene derangements, cell metabolism, the process of tumor immune surveillance, invasion and metastasis, and tumor drug-resistance. Therefore, autophagy-targeted drugs may function as anti-tumor agents. Accumulating evidence suggests that flavonoids have anticarcinogenic properties, including those relating to cellular proliferation inhibition, the induction of apoptosis, autophagy, necrosis, cell cycle arrest, senescence, the impairment of cell migration, invasion, tumor angiogenesis, and the reduction of multidrug resistance in tumor cells. Flavonoids, which are a group of natural polyphenolic compounds characterized by multiple targets that participate in multiple pathways, have been widely studied in different models for autophagy modulation. However, flavonoid-induced autophagy commonly interacts with other mechanisms, comprehensively influencing the anticancer effect. Accordingly, targeted autophagy may become the core mechanism of flavonoids in the treatment of tumors. This paper reviews the flavonoid-induced autophagy of tumor cells and their interaction with other mechanisms, so as to provide a comprehensive and in-depth account on how flavonoids exert tumor-suppressive effects through autophagy.

For thousands of years, human beings have used natural products for the treatment of various types of pathologies [...].

While Next-Generation Sequencing (NGS) and technological advances have been useful in identifying genetic profiles of tumorigenesis, novel target proteins and various clinical biomarkers, cancer continues to be a major global health threat. DNA replication, DNA damage response (DDR) and repair, and cell cycle regulation continue to be essential systems in targeted cancer therapies. Although many genes involved in DDR are known to be tumor suppressor genes, cancer cells are often dependent and addicted to these genes, making them excellent therapeutic targets. In this review, genes implicated in DNA replication, DDR, DNA repair, cell cycle regulation are discussed with reference to peptide or small molecule inhibitors which may prove therapeutic in cancer patients. Additionally, the potential of utilizing novel synthetic lethal genes in these pathways is examined, providing possible new targets for future therapeutics. Specifically, we evaluate the potential of TONSL as a novel gene for targeted therapy. Although it is a scaffold protein with no known enzymatic activity, the strategy used for developing PCNA inhibitors can also be utilized to target TONSL. This review summarizes current knowledge on non-oncogene addiction, and the utilization of synthetic lethality for developing novel inhibitors targeting non-oncogenic addiction for cancer therapy.

Thermal interface materials (TIMs), typically composed of a polymer matrix with good wetting properties and thermally conductive fillers, are applied to the interfaces of mating components to reduce the interfacial thermal resistance. As a filler material, silver has been extensively studied because of its high intrinsic thermal conductivity. However, the high cost of silver and its toxicity has hindered the wide application of silver-based TIMs. Copper is an earth-abundant element and essential micronutrient for humans. In this paper, we present a copper-based multi-dimensional filler composed of three-dimensional microscale copper flakes, one-dimensional multi-walled carbon nanotubes (MWCNTs), and zero-dimensional copper nanoparticles (Cu NPs) to create a safe and low-cost TIM with a high thermal conductivity. Cu NPs synthesized by microwave irradiation of a precursor solution were bound to MWCNTs and mixed with copper flakes and polyimide matrix to obtain a TIM paste, which was stable even in a high-temperature environment. The cross-plane thermal conductivity of the copper-based TIM was 36 W/m/K. Owing to its high thermal conductivity and low cost, the copper-based TIM could be an industrially useful heat-dissipating material in the future.

Malignant melanoma favors spreading to bone, resulting in a weakened bone with a high fracture risk. Here, we revealed the disorganized alignment of apatite crystals in the bone matrix associated with the homing of cancer cells by developing an artificially controlled ex vivo melanoma bone metastasis model. The ex vivo metastasis model reflects the progressive melanoma cell activation in vivo, resulting in decreased bone mineral density and expression of MMP1-positive cells. Moreover, less organized intercellular connections were observed in the neighboring osteoblasts in metastasized bone, indicating the abnormal and randomized organization of bone matrix secreted by disconnected osteoblasts. Our study revealed that the deteriorated microstructure associated with disorganized osteoblast arrangement was a determinant of malignant melanoma-related bone dysfunction.

Progression from localized to metastatic disease requires cancer cells spreading to distant organs through the bloodstream. Only a small proportion of these circulating tumor cells (CTCs) survives dissemination due to anoikis, shear forces and elimination by the immune system. However, all metastases originate from CTCs capable of surviving and extravasating into distant tissue to re-initiate a tumor. Metastasis initiation is not always immediate as disseminated tumor cells (DTCs) may enter a non-dividing state of cell dormancy. Cancer dormancy is a reversible condition that can be maintained for many years without being clinically detectable. Subsequently, late disease relapses are thought to be due to cancer cells ultimately escaping from dormant state. Cancer dormancy is usually associated with minimal residual disease (MRD), where DTCs persist after intended curative therapy. Thus, MRD is commonly regarded as an indicator of poor prognosis in all cancers. In this review, we examine the current understanding of MRD and immunity during cancer progression to metastasis and discuss clinical perspectives for oncology.

So far, there is no effective disease-modifying therapies for Alzheimer's Disease (AD) in clinical practice. In this context, glycine-L-proline-L-glutamate (GPE) and its analogs may open the way for developing a novel molecule for treating neurodegenerative disorders, including AD. In turn, this study was aimed to investigate the neuroprotective potentials exerted by three novel GPE peptidomimetics (GPE1, GPE2, and GPE3) using an in vitro AD model. Anti-Alzheimer potentials were determined using a wide array of techniques, such as measurements of mitochondrial viability (MTT) and lactate dehydrogenase (LDH) release assays, determination of acetylcholinesterase (AChE), α-secretase and β-secretase activities, comparisons of total antioxidant capacity (TAC) and total oxidative status (TOS) levels, flow cytometric and microscopic detection of apoptotic and necrotic neuronal death, and investigating gene expression responses via PCR arrays involving 64 critical genes related to 10 different pathways. Our analysis showed that GPE peptidomimetics modulate oxidative stress, ACh depletion, α-secretase inactivation, apoptotic, and necrotic cell death. In vitro results suggested that treatments with novel GPE analogs might be promising therapeutic agents for treatment and/or or prevention of AD.

Primary myelofibrosis (PMF) is a myeloproliferative neoplasm characterized by hematopoietic stem-cell-derived clonal proliferation, leading to bone marrow (BM) fibrosis. Hematopoiesis alterations are closely associated with modifications of the BM microenvironment, characterized by defective interactions between vascular and endosteal niches. As such, neoangiogenesis, megakaryocytes hyperplasia and extensive bone marrow fibrosis, followed by osteosclerosis and bone damage, are the most relevant consequences of PMF. Moreover, bone tissue deposition, together with progressive fibrosis, represents crucial mechanisms of disabilities in patients. Although the underlying mechanisms of bone damage observed in PMF are still unclear, the involvement of cytokines, growth factors and bone marrow microenvironment resident cells have been linked to disease progression. Herein, we focused on the role of megakaryocytes and their alterations, associated with cytokines and chemokines release, in modulating functions of most of the bone marrow cell populations and in creating a complex network where impaired signaling strongly contributes to progression and disabilities.

Vascular endothelial growth factor A (VEGF-A or VEGF) is a highly conserved secreted signalling protein best known for its roles in vascular development and angiogenesis. Many non-endothelial roles for VEGF are now established, with the discovery that VEGF and its receptors VEGFR1 and VEGFR2 are expressed in many non-vascular cell-types, as well as various cancers. In addition to secreted VEGF binding to its receptors in the extracellular space at the cell membrane (i.e., in a paracrine or autocrine mode), intracellularly localised VEGF is emerging as an important signalling molecule regulating cell growth, survival, and metabolism. This intracellular mode of signalling has been termed "intracrine", and refers to the direct action of a signalling molecule within the cell without being secreted. In this review, we describe examples of intracrine VEGF signalling in regulating cell growth, differentiation and survival, both in normal cell homeostasis and development, as well as in cancer. We further discuss emerging evidence for the molecular mechanisms underpinning VEGF intracrine function, as well as the implications this intracellular mode of VEGF signalling may have for use and design of anti-VEGF cancer therapeutics.

Plants from the Rosacea family are rich in natural molecules with beneficial biological properties, and they are widely appreciated and used in the food industry, perfumery, and cosmetics. In this review, we are considering Rosa damascena Mill., Rosa alba L., Rosa centifolia L., and Rosa gallica L. as raw materials important for producing commercial products, analyzing and comparing the main biological activities of their essential oils, hydrolates, and extracts. A literature search was performed to find materials describing (i) botanical characteristics; (ii) the phytochemical profile; and (iii) biological properties of the essential oil sand extracts of these so called "old roses" that are cultivated in Bulgaria, Turkey, India, and the Middle East. The information used is from databases PubMed, Science Direct, and Google Scholar. Roses have beneficial healing properties due to their richness of beneficial components, the secondary metabolites as flavonoids (e.g., flavones, flavonols, anthocyanins), fragrant components (essential oils, e.g., monoterpenes, sesquiterpenes), and hydrolysable and condensed tannins. Rose essential oils and extracts with their therapeutic properties-as respiratory antiseptics, anti-inflammatories, mucolytics, expectorants, decongestants, and antioxidants-are able to act as symptomatic prophylactics and drugs, and in this way alleviate dramatic sufferings during severe diseases.

Cyclodextrins (CDs) are known to us for 130 years, yet they remain ever as new and as fascinating as in their early years, when Villiers marveled at the unexpected growth of "beautiful radiated crystals" in the alcoholic media of his experiments on bacterial fermentation of starches, or when Freudenberg struggled to solve the puzzle of their unusual shape and structure [...].

BACKGROUND: Breast cancer (BC) is the most common neoplasm in women. Many clinical and preclinical studies investigated the possible relationship between host metabolism and BC. Significant differences among BC subtypes have been reported for glucose metabolism. Insulin can promote tumorigenesis through a direct effect on epithelial tissues or indirectly by affecting the levels of other modulators, such as the insulin-like growth factor (IGF) family of receptors, sex hormones, and adipokines. The potential anti-cancer activity of metformin is based on two principal effects: first, its capacity for lowering circulating insulin levels with indirect endocrine effects that may impact on tumor cell proliferation; second, its direct influence on many pro-cancer signaling pathways that are key drivers of BC aggressiveness. METHODS: In the present review, the interaction between BC, host metabolism, and patients' prognosis has been reviewed across available literature evidence. CONCLUSIONS: Obesity, metabolic syndrome, and insulin resistance are all involved in BC growth and could have a relevant impact on prognosis. All these factors act through a pro-inflammatory state, mediated by cytokines originated in fat tissue, and seem to be related to a higher risk of BC development and worse prognosis.

Seasonal flu is an acute respiratory disease that exacts a massive toll on human populations, healthcare systems and economies. The disease is caused by an enveloped Influenza virus containing eight ribonucleoprotein (RNP) complexes. Each RNP incorporates multiple copies of nucleoprotein (NP), a fragment of the viral genome (vRNA), and a viral RNA-dependent RNA polymerase (POL), and is responsible for packaging the viral genome and performing critical functions including replication and transcription. A complete model of an Influenza RNP in atomic detail can elucidate the structural basis for viral genome functions, and identify potential targets for viral therapeutics. In this work we construct a model of a complete Influenza A RNP complex in atomic detail using multiple sources of structural and sequence information and a series of homology-modeling techniques, including a motif-matching fragment assembly method. Our final model provides a rationale for experimentally-observed changes to viral polymerase activity in numerous mutational assays. Further, our model reveals specific interactions between the three primary structural components of the RNP, including potential targets for blocking POL-binding to the NP-vRNA complex. The methods developed in this work open the possibility of elucidating other functionally-relevant atomic-scale interactions in additional RNP structures and other biomolecular complexes.

DNA in our cells is constantly modified by internal and external factors [...].

Herein we report the first proof for the application of type II 2'-deoxyribosyltransferase from Lactobacillus delbrueckii (LdNDT) in suicide gene therapy for cancer treatment. To this end, we first confirm the hydrolytic ability of LdNDT over the nucleoside-based prodrugs 2'-deoxy-5-fluorouridine (dFUrd), 2'-deoxy-2-fluoroadenosine (dFAdo), and 2'-deoxy-6-methylpurine riboside (d6MetPRib). Such activity was significantly increased (up to 30-fold) in the presence of an acceptor nucleobase. To shed light on the strong nucleobase dependence for enzymatic activity, different molecular dynamics simulations were carried out. Finally, as a proof of concept, we tested the LdNDT/dFAdo system in human cervical cancer (HeLa) cells. Interestingly, LdNDT/dFAdo showed a pronounced reduction in cellular viability with inhibitory concentrations in the low micromolar range. These results open up future opportunities for the clinical implementation of nucleoside 2'-deoxyribosyltransferases (NDTs) in cancer treatment.

A novel temperature stable alkaline protease yielding bacteria was isolated from the soils of Dachigam National Park, which is known to be inhabited by a wide variety of endemic plant and animal species of Western Himalaya. This high-potential protease producing isolate was characterized and identified as Bacillus amyloliquefaciens strain HM48 by morphological, Gram's staining and biochemical techniques followed by molecular characterization using 16S rRNA approach. The extracellular protease of B. amyloliquefaciens HM48 was purified by precipitating with ammonium sulfate (80%), followed by dialysis and Gel filtration chromatography increasing its purity by 5.8-fold. The SDS-PAGE analysis of the purified enzyme confirmed a molecular weight of about ≈25 kDa. The enzyme displayed exceptional activity in a broad temperature range (10-90 °C) at pH 8.0, retaining its maximum at 70 °C, being the highest reported for this proteolytic Bacillus sp., with KM and Vmax of 11.71 mg/mL and 357.14 µmol/mL/min, respectively. The enzyme exhibited remarkable activity and stability against various metal ions, surfactants, oxidizing agent (H2O2), organic solvents and displayed outstanding compatibility with widely used detergents. This protease showed effective wash performance by exemplifying complete blood and egg-yolk stains removal at 70 °C and efficiently disintegrated chicken feathers making it of vital importance for laundry purpose and waste management. For functional analysis, protease gene amplification of strain HM48 yielded a nucleotide sequence of about 700 bp, which, when checked against the available sequences in NCBI, displayed similarity with subtilisin-like serine protease of B. amyloliquefaciens. The structure of this protease and its highest-priority substrate β-casein was generated through protein modeling. These protein models were validated through futuristic algorithms following which protein-protein (protease from HM48 and β-casein) docking was performed. The interaction profile of these proteins in the docked state with each other was also generated, shedding light on their finer details. Such attributes make this thermally stable protease novel and suitable for high-temperature industrial and environmental applications.

Ginsenoside Rg3 exerts antiproliferation activity on cancer cells by regulating diverse noncoding RNAs. However, little is known about the role of long noncoding RNAs (lncRNAs) or their relationship with competitive endogenous RNA (ceRNA) in Rg3-treated cancer cells. Here, a lncRNA (ATXN8OS) was found to be downregulated via Rg3-mediated promoter hypermethylation in MCF-7 breast cancer cells. SiRNA-induced downregulation of ATXN8OS decreased cell proliferation but increased apoptosis, suggesting that the noncoding RNA possessed proproliferation activity. An in silico search for potential ATXN8OS-targeting microRNAs (miRs) identified a promising candidate (miR-424-5p) based on its high binding score. As expected, miR-424-5p suppressed proliferation and stimulated apoptosis of the MCF-7 cells. The in silico miR-target-gene prediction identified 200 potential target genes of miR-424-5p, which were subsequently narrowed down to seven that underwent hypermethylation at their promoter by Rg3. Among them, three genes (EYA1, DACH1, and CHRM3) were previously known oncogenes and were proven to be oppositely regulated by ATXN8OS and miR-424-5p. When taken together, Rg3 downregulated ATXN8OS that inhibited the tumor-suppressive miR-424-5p, leading to the downregulation of the oncogenic target genes.

Chronic heart failure (HF) is an important clinical, social, and economic problem. A key role in HF progression is played by oxidative stress. Free oxygen radicals, formed under the conditions of hypoxia and reperfusion, participate in myocardial stunning and other forms of post-reperfusion damage. HF patients also suffer from disorders connected with saliva secretion. However, still little is known about the mechanisms that impair the secretory function of salivary glands in these patients. In the presented study, we were the first to compare the antioxidant barrier, protein glycoxidation, and nitrosative/nitrative stress in non-stimulated (non-stimulated whole saliva (NWS)) and stimulated (SWS) saliva of HF patients. The study included 50 HF patients with normal saliva (NS) secretion (n = 27) and hyposalivation (HS) (n = 23), as well as an age- and gender-matched control group (n = 50). We demonstrated that, in NWS of HF patients with HS, the concentration of low-molecular-weight non-enzymatic antioxidants decreased (↓total polyphenols, ↓ascorbic acid, ↓reduced glutathione, ↓albumin) compared to HF patients with normal saliva (NS) secretion, as well as the control group (except albumin). We also observed increased content of protein glycoxidation products (↑dityrosine, ↑kynurenine, ↑glycophore) in NWS and SWS of HF patients with HS compared to healthy controls. Interestingly, the content of dityrosine, N-formylkynurenine, and glycophore in NWS was also significantly higher in HF patients with HS compared to those with NS secretion. The concentration of NO was considerably lower, while the levels of peroxynitrite and nitrotyrosine were significantly higher in NWS and SWS of HF subjects with HS compared to the controls. Salivary gland dysfunction occurs in patients with chronic HF with the submandibular salivary glands being the least efficient. Oxidative/nitrosative stress may be one of the mechanisms responsible for the impairment of salivary gland secretory function in HF patients.

Over the years, a wide variety of therapeutic antibodies has been successfully introduced in the auto-immunology clinic, and many more are on the way. Many of these treatments address either a pathogenic circulating molecule or a cell-bound molecule. Whereas addressing the former target results in neutralization of the soluble factor and binding to the latter target either inhibits cellular function or induces selective cell death. If this targeted molecule or cell is part of the immune system, this therapy evokes a state of immunodeficiency with infections as a possible consequence. Therefore, immune monitoring is needed to prevent such adverse side effects of immunotherapy. In this paper, different immunotherapies used in Sjögren's syndrome, as well as different approaches to monitoring the immune system, are discussed.

(1) Background: Retinal pigment epithelial cells (RPE) cells constitutively secrete vascular endothelial growth factor (VEGF) in the retina, protecting the neuronal cells and the choroid. Increased VEGF secretion, however, can result in neovascularization and edema. Many factors regulate VEGF secretion. In this study, we investigated the effect of external stimuli in relation to diurnal rhythm on constitutive VEGF secretion. (2) Methods: Single-eye RPE cell culture was prepared from porcine eyes. RPE cells were cultured in darkness, treated with daylight or room light, and treated with melatonin at different time frames, either respectively or in combination. Supernatants were collected and VEGF content evaluated using ELISA. Expression of the clock protein BMAL1 was evaluated with Western blot. (3) Results: VEGF secretion of the RPE shows a diurnal rhythm. While the rhythm is not influenced by either light or melatonin, the amount of secreted VEGF can be increased by nocturnal melatonin, especially in combination with morning daylight. These findings disclose another layer of VEGF regulation in the retina.

Human cartilage has relatively slow metabolism compared to other normal tissues. Cartilage damage is of great clinical consequence since cartilage has limited intrinsic healing potential. Cartilage tissue engineering is a rapidly emerging field that holds great promise for tissue function repair and artificial/engineered tissue substitutes. However, current clinical therapies for cartilage repair are less than satisfactory and rarely recover full function or return the diseased tissue to its native healthy state. Kartogenin (KGN), a small molecule, can promote chondrocyte differentiation both in vitro and in vivo. The purpose of this research is to optimize the chondrogenic process in mesenchymal stem cell (MSC)-based chondrogenic constructs with KGN for potential use in cartilage tissue engineering. In this study, we demonstrate that KGN treatment can promote MSC condensation and cell cluster formation within a tri-copolymer scaffold. Expression of Acan, Sox9, and Col2a1 was significantly up-regulated in three-dimensional (3D) culture conditions. The lacuna-like structure showed active deposition of type II collagen and aggrecan deposition. We expect these results will open new avenues for the use of small molecules in chondrogenic differentiation protocols in combination with scaffolds, which may yield better strategies for cartilage tissue engineering.

Reproducibility issues regarding in vitro cell culture experiments are related to genetic fluctuations and batch-wise variations of biological materials such as fetal calf serum (FCS). Genome sequencing may control the former, while the latter may remain unrecognized. Using a U937 macrophage model for cell differentiation and inflammation, we investigated whether the formation of effector molecules was dependent on the FCS batch used for cultivation. High resolution mass spectrometry (HRMS) was used to identify FCS constituents and to explore their effects on cultured cells evaluating secreted cytokines, eicosanoids, and other inflammatory mediators. Remarkably, the FCS eicosanoid composition showed more batch-dependent variations than the protein composition. Efficient uptake of fatty acids from the medium by U937 macrophages and inflammation-induced release thereof was evidenced using C13-labelled arachidonic acid, highlighting rapid lipid metabolism. For functional testing, FCS batch-dependent nanomolar concentration differences of two selected eicosanoids, 5-HETE and 15-HETE, were balanced out by spiking. Culturing U937 cells at these defined conditions indeed resulted in significant proteome alterations indicating HETE-induced PPARγ activation, independently corroborated by HETE-induced formation of peroxisomes observed by high-resolution microscopy. In conclusion, the present data demonstrate that FCS-contained eicosanoids, subject to substantial batch-wise variation, may modulate cellular effector functions in cell culture experiments.

The ubiquitin pathway is highly conserved across the eukaryotic domain of life and plays an essential role in a plethora of cellular processes. It is not surprising that many intracellular bacterial pathogens often target the essential host ubiquitin pathway. The intracellular bacterial pathogen Legionella pneumophila injects into the host cell cytosol multiple classes of classical and novel ubiquitin-modifying enzymes that modulate diverse ubiquitin-related processes in the host cell. Most of these pathogen-injected proteins, designated as effectors, mimic known E3-ubiquitin ligases through harboring F-box or U-box domains. The classical F-box effector, AnkB targets host proteins for K48-linked polyubiquitination, which leads to excessive proteasomal degradation that is required to generate adequate supplies of amino acids for metabolism of the pathogen. In contrast, the SidC and SdcA effectors share no structural similarity to known eukaryotic ligases despite having E3-ubiquitin ligase activity, suggesting that the number of E3-ligases in eukaryotes is under-represented. L. pneumophila also injects into the host many novel ubiquitin-modifying enzymes, which are the SidE family of effectors that catalyze phosphoribosyl-ubiquitination of serine residue of target proteins, independently of the canonical E1-2-3 enzymatic cascade. Interestingly, the environmental bacterium, L. pneumophila, has evolved within a diverse range of amoebal species, which serve as the natural hosts, while accidental transmission through contaminated aerosols can cause pneumonia in humans. Therefore, it is likely that the novel ubiquitin-modifying enzymes of L. pneumophila were acquired by the pathogen through interkingdom gene transfer from the diverse natural amoebal hosts. Furthermore, conservation of the ubiquitin pathway across eukaryotes has enabled these novel ubiquitin-modifying enzymes to function similarly in mammalian cells. Studies on the biological functions of these effectors are likely to reveal further novel ubiquitin biology and shed further lights on the evolution of ubiquitin.

Alzheimer's disease (AD) is generally recognized as a multifactorial neurodegenerative pathology with an increasing impact on society. Tenuazonic acid (TA) is a natural compound that was recently identified as a potential multitarget ligand with anti-cholinesterase, anti-amyloidogenic and antioxidant activities. Using its structure as a chemical scaffold, we synthesized and evaluated new derivatives (1-5), including tenuazonic-donepezil (TA-DNP) hybrids (4 and 5) due to the clinical importance of the anti-AD drug donepezil. These novel compounds all achieved activity in the micromolar range towards all selected targets and demonstrated to be potentially orally absorbed. Moreover, a selected compound (1) was further investigated as a chelating agent towards copper (II), zinc (II) and iron (III) and showed good chelating ability (pFe = 16.6, pCu = 11.6, pZn = 6.0 at pH 7.4). Therefore, the TA motif can be considered an interesting building block in the search for innovative multi-functional anti-neurodegenerative drugs, as exemplified by hybrid 5, a promising non-cytotoxic lead compound adequate for the early stages of AD, and capable of ameliorating the oxidative status of SH-SY5Y human neuroblastoma cells.

The aim of this study was to evaluate the mutual relationship among perivascular adipose tissue (PVAT) and endogenous and exogenous H2S in vasoactive responses of isolated arteries from adult normotensive (Wistar) rats and hypertriglyceridemic (HTG) rats, which are a nonobese model of metabolic syndrome. In HTG rats, mild hypertension was associated with glucose intolerance, dyslipidemia, increased amount of retroperitoneal fat, increased arterial contractility, and endothelial dysfunction associated with arterial wall injury, which was accompanied by decreased nitric oxide (NO)-synthase activity, increased expression of H2S producing enzyme, and an altered oxidative state. In HTG, endogenous H2S participated in the inhibition of endothelium-dependent vasorelaxation regardless of PVAT presence; on the other hand, aortas with preserved PVAT revealed a stronger anticontractile effect mediated at least partially by H2S. Although we observed a higher vasorelaxation induced by exogenous H2S donor in HTG rats than in Wistar rats, intact PVAT subtilized this effect. We demonstrate that, in HTG rats, endogenous H2S could manifest a dual effect depending on the type of triggered signaling pathway. H2S within the arterial wall contributes to endothelial dysfunction. On the other hand, PVAT of HTG is endowed with compensatory vasoactive mechanisms, which include stronger anti-contractile action of H2S. Nevertheless, the possible negative impact of PVAT during hypertriglyceridemia on the activity of exogenous H2S donors needs to be taken into consideration.

Protein degradation maintains cellular integrity by regulating virtually all biological processes, whereas impaired proteolysis perturbs protein quality control, and often leads to human disease. Two major proteolytic systems are responsible for protein breakdown in all cells: autophagy, which facilitates the loss of organelles, protein aggregates, and cell surface proteins; and the ubiquitin-proteasome system (UPS), which promotes degradation of mainly soluble proteins. Recent findings indicate that more complex protein structures, such as filamentous assemblies, which are not accessible to the catalytic core of the proteasome in vitro, can be efficiently degraded by this proteolytic machinery in systemic catabolic states in vivo. Mechanisms that loosen the filamentous structure seem to be activated first, hence increasing the accessibility of protein constituents to the UPS. In this review, we will discuss the mechanisms underlying the disassembly and loss of the intricate insoluble filamentous myofibrils, which are responsible for muscle contraction, and whose degradation by the UPS causes weakness and disability in aging and disease. Several lines of evidence indicate that myofibril breakdown occurs in a strictly ordered and controlled manner, and the function of AAA-ATPases is crucial for their disassembly and loss.

The use of 5-fluorouracil (5FU) is associated with multifaceted challenges and poor pharmacokinetics. Poly(lactic-co-glycolic acid)-lipid hybrid nanoparticles (PLNs)-based therapy has received attention as efficient carriers for a diversity of drugs. This study evaluated the in vivo chemotherapeutic and anti-proliferative efficacy of 5FU-loaded PLNs against 1,2-dimethylhydrazine (Di-MH) prompted colon dysplasia in mice compared to free 5FU. 5FU PLNs were prepared. Male Swiss albino mice were distributed to six experimental groups. Group 1: Saline group. All the other groups were injected weekly with Di-MH [20 mg/kg, s.c.]. Group 2: Di-MH induced colon dysplasia control group. Groups 3 and 4: Di-MH + free 5FU treated group [2.5 and 5 mg/kg]. Groups 5 and 6: Di-MH + 5FU-PLNs treated group [2.5 and 5 mg/kg]. Free 5FU and 5FU-PLNs doses were administered orally, twice weekly. Treatment with 5FU-PLNs induced a higher cytoprotective effect compared to free 5FU as indicated by lower mucosal histopathologic score and reduction in number of Ki-67 immunpositive proliferating nuclei. Additionally, there was significant upregulation of p53 and caspase 3 genes in colon specimens. Our results support the validity of utilizing the PLNs technique to improve the chemopreventive action of 5FU in treating colon cancer.

It has become impossible to review all the existing literature on Voltage-Dependent Anion selective Channel (VDAC) in a single article. A real Renaissance of studies brings this protein to the center of decisive knowledge both for cell physiology and therapeutic application. This review, after highlighting the similarities between the cellular context and the study methods of the solute carriers present in the inner membrane and VDAC in the outer membrane of the mitochondria, will focus on the isoforms of VDAC and their biochemical characteristics. In particular, the possible reasons for their evolutionary onset will be discussed. The variations in their post-translational modifications and the differences between the regulatory regions of their genes, probably the key to understanding the current presence of these genes, will be described. Finally, the situation in the higher eukaryotes will be compared to that of yeast, a unicellular eukaryote, where there is only one active isoform and the role of VDAC in energy metabolism is better understood.

Citrus hystrix DC. (CH) is found in many countries in Southeast Asia. This plant has been reported for anti-microbial, anti-cancer and anti-inflammatory bioactivities. However, the anti-inflammatory and anti-inflammasome properties of the leaves remain poorly understood. This study aimed to investigate the effect of CH leaves on NLRP3 and NF-κB signaling pathways. CH leaves were sequentially extracted using hexane, ethyl acetate and 95% ethanol to give three crude extracts. An active compound, lupeol was fractionated from the ethanolic extract using chromatographic techniques, and its structure was identified and confirmed by spectroscopic methods. Anti-inflammatory activities were observed on both lipopolysaccharide-stimulated and NLRP3 adenosine triphosphate-induced macrophages. The release of pro-inflammatory cytokines (IL-1β, IL-6 and TNF-α) was analyzed by Enzyme-Linked Immunosorbent Assay (ELISA). Real-time qRT-polymerase chain reaction (PCR) was used to measure inflammatory-associated gene expression. NF-κB protein expressions were investigated using the immunoblotting technique. The active fraction of ethanolic CH leaves and lupeol significantly reduced the release of pro-inflammatory cytokines and suppressed the expression of both inflammasome genes and NF-κB proteins. The ethanolic extract of CH leaves and lupeol showed potent anti-inflammatory activities by targeting NF-κB and NLRP3 signaling pathways.

Prion diseases are the hallmark protein folding neurodegenerative disease. Their transmissible nature has allowed for the development of many different cellular models of disease where prion propagation and sometimes pathology can be induced. This review examines the range of simple cell cultures to more complex neurospheres, organoid, and organotypic slice cultures that have been used to study prion disease pathogenesis and to test therapeutics. We highlight the advantages and disadvantages of each system, giving special consideration to the importance of strains when choosing a model and when interpreting results, as not all systems propagate all strains, and in some cases, the technique used, or treatment applied, can alter the very strain properties being studied.

Dopamine (DA), as one of the major neurotransmitters in the central nervous system (CNS) and periphery, exerts its actions through five types of receptors which belong to two major subfamilies such as D1-like (i.e., D1 and D5 receptors) and D2-like (i.e., D2, D3 and D4) receptors. Dopamine D3 receptor (D3R) was cloned 30 years ago, and its distribution in the CNS and in the periphery, molecular structure, cellular signaling mechanisms have been largely explored. Involvement of D3Rs has been recognized in several CNS functions such as movement control, cognition, learning, reward, emotional regulation and social behavior. D3Rs have become a promising target of drug research and great efforts have been made to obtain high affinity ligands (selective agonists, partial agonists and antagonists) in order to elucidate D3R functions. There has been a strong drive behind the efforts to find drug-like compounds with high affinity and selectivity and various functionality for D3Rs in the hope that they would have potential treatment options in CNS diseases such as schizophrenia, drug abuse, Parkinson's disease, depression, and restless leg syndrome. In this review, we provide an overview and update of the major aspects of research related to D3Rs: distribution in the CNS and periphery, signaling and molecular properties, the status of ligands available for D3R research (agonists, antagonists and partial agonists), behavioral functions of D3Rs, the role in neural networks, and we provide a summary on how the D3R-related drug research has been translated to human therapy.

Blueberries are consumed as healthy fruits that provide a variety of benefits to the nervous system. Scientists have found that blueberries can be used as a daily edible source for supplementation to prevent and minimize complexities of age-related diseases as well as to improve learning and memory in children. Anthocyanins are the most mentioned compounds among the components in blueberries, as they play a major role in providing the health benefits of this fruit. However, while they are highly active in impeding biological impairment in neuronal functions, they have poor bioavailability. This review focuses on neurological investigations of blueberries from in vitro cell studies to in vivo studies, including animal and human studies, with respect to their positive outcomes of neuroprotection and intervention in neurodegenerative conditions. Readers will also find information on the bioavailability of anthocyanins and the considerable factors affecting them so that they can make informed decisions regarding the daily consumption of blueberries. In this context, the ways in which blueberries or blueberry supplementation forms are consumed and which of these forms is best for maximizing the health benefits of blueberries should be considered important decision-making factors in the consumption of blueberries; all of these aspects are covered in this review. Finally, we discuss recent technologies that have been employed to improve the bioavailability of blueberry anthocyanins in the development of effective delivery vehicles supporting brain health.

Wnt/β-catenin signaling controls many biological processes for the generation and sustainability of proper tissue size, organization and function during development and homeostasis. Consequently, mutations in the Wnt pathway components and modulators cause diseases, including genetic disorders and cancers. Targeted treatment of pathway-associated diseases entails detailed understanding of the regulatory mechanisms that fine-tune Wnt signaling. Here, we identify the neurotrophin receptor-associated death domain (Nradd), a homolog of p75 neurotrophin receptor (p75NTR), as a negative regulator of Wnt/β-catenin signaling in zebrafish embryos and in mammalian cells. Nradd significantly suppresses Wnt8-mediated patterning of the mesoderm and neuroectoderm during zebrafish gastrulation. Nradd is localized at the plasma membrane, physically interacts with the Wnt receptor complex and enhances apoptosis in cooperation with Wnt/β-catenin signaling. Our functional analyses indicate that the N-glycosylated N-terminus and the death domain-containing C-terminus regions are necessary for both the inhibition of Wnt signaling and apoptosis. Finally, Nradd can induce apoptosis in mammalian cells. Thus, Nradd regulates cell death as a modifier of Wnt/β-catenin signaling during development.

As a multifunctional polypeptide, epidermal growth factor (EGF) increases growth performance or enhances resistance to diseases in commercial broilers under adverse conditions. In this study, a recombinant Lactococcus lactis was established to produce the secretary form of bioactive gEGF. The results of in vitro testing showed that gEGF promoted the proliferation of chicken embryo fibroblast cells. A total of 63 5-day-old broiler chickens were evenly divided into three groups and treated with either M17 medium (the control group), supernatant of LL-pNZ8149 fermentation product (the P-LL group), or supernatant of LL-pNZ8149-gEGF fermentation product (the gEGF group). In two weeks, many measurements of growth, immunity and the intestines were significantly higher in the gEGF group than those in the control and the P-LL groups. Our study showed that the bioactive gEGF could be expressed with Lactococcus lactis expression system with the potential to enhance growth performance, immune function, and intestinal development in broiler chickens.

The use of remedies based on medicinal plants continues to expand rapidly around the world, with many people now resorting to this type of product for the treatment and prevention of several pathologies [...].

Among seven homologs of cytochrome b561 in a model organism C. elegans, Cecytb-2 was confirmed to be expressed in digestive organs and was considered as a homolog of human Dcytb functioning as a ferric reductase. Cecytb-2 protein was expressed in Pichia pastoris cells, purified, and reconstituted into a phospholipid bilayer nanodisc. The reconstituted Cecytb-2 in nanodisc environments was extremely stable and more reducible with ascorbate than in a detergent-micelle state. We confirmed the ferric reductase activity of Cecytb-2 by analyzing the oxidation of ferrous heme upon addition of ferric substrate under anaerobic conditions, where clear and saturable dependencies on the substrate concentrations following the Michaelis-Menten equation were observed. Further, we confirmed that the ferric substrate was converted to a ferrous state by using a nitroso-PSAP assay. Importantly, we observed that the ferric reductase activity of Cecytb-2 became enhanced in the phospholipid bilayer nanodisc.

Lactobacillus strains dominate the vaginal habitat and they are associated with a lower risk of genital infections. In addition, they contribute to the conservation of the vaginal microbiota by producing postbiotic agents. Previous studies have shown that their predominance involves antimicrobial activity against urogenital pathologies. In this context, probiotics may improve treatment outcomes. The aim of this study was to evaluate the probiotic properties of lactobacilli strains of vaginal origin using a multidisciplinary approach. For this purpose, safety criteria, ability to resist at low pH and bile salts, antimicrobial activity, ability to produce biofilm, capacity to produce hydrogen peroxide and more importantly, auto-aggregation, co-aggregation (with Candida spp.) and adhesion to human cells were evaluated. The strains belonged to the species of L. crispatus, L. gasseri, L. rhamnosus and L. delbruckii. Among these, a strain of L. rhamnosus named AD3 showed the best probiotic properties. As probiotics are already in use in many clinical practice and there are no major safety concerns, L. rhamnosus AD3 showed promise in becoming a prevention and complementary treatment option for urogenital diseases. Indeed, these results suggest that strain L. rhamnosus AD3 is non-pathogenic and likely to be safe for human consumption. This study revealed the great amensalistic properties of a new L. rhamnosus strain which can aim to be used as probiotic in pharmaceutical applications.

Biliary tract cancer (BTC) compromises a heterogenous group of tumors with poor prognoses. Curative surgery remains the first choice for localized disease; however, most BTC patients have had unresectable or metastatic disease. The gold standard therapy for these patients is chemotherapy with gemcitabine and cisplatin. There are no consensus guidelines for standard treatment in a second-line setting, although the data of the ABC-06 trial showed a slight survival benefit from oxaliplatin and 5-fluorouracil combination chemotherapy. Recent progress in comprehensive genomic profiling for advanced BTC (ABTC) has helped to clarify tumorigenesis and facilitate the coming era of precision medicine. Generally, targeted agents fail to show significant clinical benefits in unselected populations. Only fibroblast growth factor receptor 2 (FGFR2) fusion and isocitrate dehydrogenase (IDH)- and BRAF mutation-enriched populations have survival benefits from the corresponding inhibitors. Several interesting targeted agents for monotherapies or combination therapies with other compounds are currently ongoing or recruiting. Here, we review the published data from clinical trials of second-line therapies after the failure of gemcitabine-based chemotherapy in ABTC. The results were stratified by different genetic alternations, as well as by chemotherapy, targeted therapy and immunotherapy.

Before replicating, Pospiviroidae viroids must move into the plant nucleus. However, the mechanisms of viroid nuclear import are not entirely understood. To study the nuclear import of viroids, we established a nuclear import assay system using onion cell strips and observed the import of Alexa Fluor-594-labeled citrus exocortis viroid (CEVd). To identify the plant factors involved in the nuclear import of viroids, we cloned the Viroid RNA-binding Protein 1 (VIRP1) gene from a tomato cultivar, Seokwang, and heterologously expressed and purified the VIRP1 protein. The newly prepared VIRP1 protein had alterations of amino acid residues at two points (H52R, A277G) compared with a reference VIRP1 protein (AJ249595). VIRP1 specifically bound to CEVd and promoted its nuclear import. However, it is still uncertain whether VIRP1 is the only factor required for the nuclear import of CEVd because CEVd entered the plant nuclei without VIRP1 in our assay system. The cause of the observed nuclear accumulation of CEVd in the absence of VIRP1 needs to be further clarified.

Articular cartilage is considered to have limited regenerative capacity, which has led to the search for therapies to limit or halt the progression of its destruction. Perlecan, a multifunctional heparan sulphate (HS) proteoglycan, promotes embryonic cartilage development and stabilises the mature tissue. We investigated the immunolocalisation of perlecan and collagen between donor-matched biopsies of human articular cartilage defects (n = 10 × 2) that were repaired either naturally or using autologous cell therapy, and with age-matched normal cartilage. We explored how the removal of HS from perlecan affects human chondrocytes in vitro. Immunohistochemistry showed both a pericellular and diffuse matrix staining pattern for perlecan in both natural and cell therapy repaired cartilage, which related to whether the morphology of the newly formed tissue was hyaline cartilage or fibrocartilage. Immunostaining for perlecan was significantly greater in both these repair tissues compared to normal age-matched controls. The immunolocalisation of collagens type III and VI was also dependent on tissue morphology. Heparanase treatment of chondrocytes in vitro resulted in significantly increased proliferation, while the expression of key chondrogenic surface and genetic markers was unaffected. Perlecan was more prominent in chondrocyte clusters than in individual cells after heparanase treatment. Heparanase treatment could be a means of increasing chondrocyte responsiveness to cartilage injury and perhaps to improve repair of defects.

Perinatal brain injuries, including encephalopathy related to fetal growth restriction, encephalopathy of prematurity, neonatal encephalopathy of the term neonate, and neonatal stroke, are a major cause of neurodevelopmental disorders. They trigger cellular and molecular cascades that lead in many cases to permanent motor, cognitive, and/or behavioral deficits. Damage includes neuronal degeneration, selective loss of subclasses of interneurons, blocked maturation of oligodendrocyte progenitor cells leading to dysmyelination, axonopathy and very likely synaptopathy, leading to impaired connectivity. The nature and severity of changes vary according to the type and severity of insult and maturation stage of the brain. Microglial activation has been demonstrated almost ubiquitously in perinatal brain injuries and these responses are key cell orchestrators of brain pathology but also attempts at repair. These divergent roles are facilitated by a diverse suite of transcriptional profiles and through a complex dialogue with other brain cell types. Adding to the complexity of understanding microglia and how to modulate them to protect the brain is that these cells have their own developmental stages, enabling them to be key participants in brain building. Of note, not only do microglia help build the brain and respond to brain injury, but they are a key cell in the transduction of systemic inflammation into neuroinflammation. Systemic inflammatory exposure is a key risk factor for poor neurodevelopmental outcomes in preterm born infants. Based on these observations, microglia appear as a key cell target for neuroprotection in perinatal brain injuries. Numerous strategies have been developed experimentally to modulate microglia and attenuate brain injury based on these strong supporting data and we will summarize these.

The occurrence and diversity of Lecanicillium and Sarocladium in maize seeds and their role in this cereal are poorly understood. Therefore, the present study aimed to investigate Sarocladium and Lecanicillium communities found in endosphere of maize seeds collected from fields in Poland and their potential to form selected bioactive substances. The sequencing of the internally transcribed spacer regions 1 (ITS 1) and 2 (ITS2) and the large-subunit (LSU, 28S) of the rRNA gene cluster resulted in the identification of 17 Sarocladium zeae strains, three Sarocladium strictum and five Lecanicillium lecanii isolates. The assay on solid substrate showed that S. zeae and S. strictum can synthesize bassianolide, vertilecanin A, vertilecanin A methyl ester, 2-decenedioic acid and 10-hydroxy-8-decenoic acid. This is also the first study revealing the ability of these two species to produce beauvericin and enniatin B1, respectively. Moreover, for the first time in the present investigation, pyrrocidine A and/or B have been annotated as metabolites of S. strictum and L. lecanii. The production of toxic, insecticidal and antibacterial compounds in cultures of S. strictum, S. zeae and L. lecanii suggests the requirement to revise the approach to study the biological role of fungi inhabiting maize seeds.

Recently, plant bioreactors have flourished into an exciting area of synthetic biology because of their product safety, inexpensive production cost, and easy scale-up. Duckweed is the smallest and fastest-growing aquatic plant, and has advantages including simple processing and the ability to grow high biomass in smaller areas. Therefore, duckweed could be used as a new potential bioreactor for biological products such as vaccines, antibodies, pharmaceutical proteins, and industrial enzymes. Duckweed has made a breakthrough in biosynthesis as a chassis plant and is being utilized for the production of plenty of biological products or bio-derivatives with multiple uses and high values. This review summarizes the latest progress on genetic background, genetic transformation system, and bioreactor development of duckweed, and provides insights for further exploration and application of duckweed.

The Human Genome Project, completed in 2003 by an international consortium, is considered one of the most important achievements for mankind in the 21st century [...].

Vascular endothelial growth factors (VEGFs) are vital regulators of angiogenesis that are expressed in response to soluble mediators, such as cytokines and growth factors. Their physiologic functions include blood vessel formation, regulation of vascular permeability, stem cell and monocyte/macrophage recruitment and maintenance of bone homeostasis and repair. In addition, angiogenesis plays a pivotal role in chronic pathologic conditions, such as tumorigenesis, inflammatory immune diseases and bone loss. According to their prevalence, morbidity and mortality, inflammatory diseases affecting periodontal tissues and oral cancer are relevant non-communicable diseases. Whereas oral squamous cell carcinoma (OSCC) is considered one of the most common cancers worldwide, destructive inflammatory periodontal diseases, on the other hand, are amongst the most prevalent chronic inflammatory conditions affecting humans and also represent the main cause of tooth loss in adults. In the recent years, while knowledge regarding the role of VEGF signaling in common oral diseases is expanding, new potential translational applications emerge. In the present narrative review we aim to explore the role of VEGF signaling in oral cancer and destructive periodontal inflammatory diseases, with emphasis in its translational applications as potential biomarkers and therapeutic targets.

Currently, food losses represent a serious imbalance in the dimensions of availability and accessibility in the global food system in the short term [...].

P14ARF (ARF; Alternative Reading Frame) is an extensively characterized tumor suppressor which, in response to oncogenic stimuli, mediates cell cycle arrest and apoptosis via p53-dependent and independent routes. ARF has been shown to be frequently lost through CpG island promoter methylation in a wide spectrum of human malignancies, such as colorectal, prostate, breast, and gastric cancers, while point mutations and deletions in the p14ARF locus have been linked with various forms of melanomas and glioblastomas. Although ARF has been mostly studied in the context of tumorigenesis, it has been also implicated in purely developmental processes, such as spermatogenesis, and mammary gland and ocular development, while it has been additionally involved in the regulation of angiogenesis. Moreover, ARF has been found to hold important roles in stem cell self-renewal and differentiation. As is often the case with tumor suppressors, ARF functions as a pleiotropic protein regulating a number of different mechanisms at the crossroad of development and tumorigenesis. Here, we provide an overview of the non-canonical functions of ARF in cancer and developmental biology, by dissecting the crosstalk of ARF signaling with key oncogenic and developmental pathways.

The recently identified severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus, the cause of coronavirus disease (COVID-19) and the associated ongoing pandemic, frequently leads to severe respiratory distress syndrome and pneumonia with fatal consequences. Although several factors of this infection and its consequences are not completely clear, the presence and involvement of specific chemokines is undoubtedly crucial for the development and progression of COVID-19. Cytokine storm and the often-resulting cytokine release syndrome (CRS) are pathophysiological hallmarks in COVID-19 infections related to its most severe and fatal cases. In this hyperinflammatory event, chemokines and other cytokines are highly upregulated and are therefore not fulfilling their beneficial function in the host response anymore but causing harmful effects. Here, we present the recent views on the involvement of chemokines and selected cytokines in COVID-19 and the therapeutics currently in clinical development targeting or interfering with them, discussing their potentials in the treatment of COVID-19 infections.

Meiosis is a specialized cell division process that mediates genetic information transfer to the next generation. Meiotic chromosomal segregation occurs when DNA replication is completed during the pre-meiotic S phase. Here, we show that Schizosaccharomyces pombe Pef1, an orthologue of mammalian cyclin-dependent kinase 5 (CDK5), is required to promote pre-meiotic DNA replication. We examined the efficiency of meiotic initiation using pat1-114 mutants and found that, meiotic nuclear divisions did not occur in the pef1Δ pat1-114 strain. Deletion of pef1 also suppressed the expression of DNA replication factors and the phosphorylation of Cdc2 Tyr-15. The double deletion of clg1 and psl1 arrested meiotic initiation in pat1-114 mutant cells, similar to that of pef1-deficient cells. Meiotic progression was also slightly delayed in the pas1-deficient strain. Our results reveal that Pef1 regulates cyclin-coordinated meiotic progression.

Plant-derived exosome-like nanovesicles (EPDENs) have recently been isolated and evaluated as potential bioactive nutraceutical biomolecules. It has been hypothesized that EPDENs may exert their activity on mammalian cells through their specific cargo. In this study, we isolated and purified EPDENs from the strawberry juice of Fragaria x ananassa (cv. Romina), a new cultivar characterized by a high content of anthocyanins, folic acid, flavonols, and vitamin C and an elevated antioxidant capacity. Fragaria-derived EPDENs were purified by a series of centrifugation and filtration steps. EPDENs showed size and morphology similar to mammalian extracellular nanovesicles. The internalization of Fragaria-derived EPDENs by human mesenchymal stromal cells (MSCs) did not negatively affect their viability, and the pretreatment of MSCs with Fragaria-derived EPDENs prevented oxidative stress in a dose-dependent manner. This is possibly due to the presence of vitamin C inside the nanovesicle membrane. The analysis of EPDEN cargo also revealed the presence of small RNAs and miRNAs. These findings suggest that Fragaria-derived EPDENs may be considered nanoshuttles contained in food, with potential health-promoting activity.

Inflammatory bowel disease (IBD) is a chronic inflammatory disorder of the gut that can lead to severe gastrointestinal symptoms, malnutrition, and complications such as fistulas and cancer. Mesenchymal stem/stromal cells (MSCs) are being investigated as a novel therapy for IBD and have been demonstrated to be safe and effective for perianal fistulizing Crohn's disease (PFCD). This systematic review aims to present the most recent studies on the safety and efficacy of MSC therapy in IBD. A detailed search strategy of clinical trials on MSCs and IBD was performed on PubMed, with 32 studies selected for inclusion in this review. The newest studies on local MSC injection for PFCD continue to support long-term efficacy while maintaining a favorable safety profile. The evidence for systemic MSC infusion in luminal IBD remains mixed due to marked methodological heterogeneity and unclear safety profiles. Although further studies are needed to better establish the role of this novel treatment modality, MSCs are proving to be a very exciting addition to the limited therapies available for IBD.

Quinpirole (QNP) sensitization is a well-established model of stereotypical checking relevant to obsessive-compulsive disorder. Previously, we found that QNP-treated rats display deficits in hippocampus-dependent tasks. The present study explores the expression of immediate early genes (IEG) during QNP-induced stereotypical checking in the hippocampus, anterior cingulate cortex (ACC), orbitofrontal cortex (OFC), and medial prefrontal cortex (mPFC). Adult male rats were injected with QNP (0.5 mg/mL/kg; n = 15) or saline (n = 14) daily for 10 days and exposed to an arena enriched with two objects. Visits to the objects and the corners of the arena were recorded. QNP-treated rats developed an idiosyncratic pattern of visits that persisted across experimental days. On day 11, rats were exposed to the arena twice for 5 min and sacrificed. The expression of IEGs Arc and Homer1a was determined using cellular compartment analysis of temporal activity by fluorescence in situ hybridization. IEG-positive nuclei were counted in the CA1 area of the hippocampus, ACC, OFC, and mPFC. We found significantly fewer IEG-positive nuclei in the CA1 in QNP-treated rats compared to controls. The overlap between IEG expressing neurons was comparable between the groups. We did not observe significant differences in IEG expression between QNP treated and control rats in ACC, OFC, and mPFC. In conclusion, treatment of rats with quinpirole decreases plasticity-related activity in the hippocampus during stereotypical checking.

Red palm weevil (Rhynchophorus ferrugineus Olivier, 1791, Coleoptera: Curculionidae) is a destructive pest of palms, rapidly extending its native geographical range and causing large economic losses worldwide. The present work describes isolation, identification, and bioinformatic analysis of antibacterial proteins and peptides from the immunized hemolymph of this beetle. In total, 17 different bactericidal or bacteriostatic compounds were isolated via a series of high-pressure liquid chromatography steps, and their partial amino acid sequences were determined by N-terminal sequencing or by mass spectrometry. The bioinformatic analysis of the results facilitated identification and description of corresponding nucleotide coding sequences for each peptide and protein, based on the recently published R. ferrugineus transcriptome database. The identified compounds are represented by several well-known bactericidal factors: two peptides similar to defensins, one cecropin-A1-like peptide, and one attacin-B-like protein. Interestingly, we have also identified some unexpected compounds comprising five isoforms of pheromone-binding proteins as well as seven isoforms of odorant-binding proteins. The particular role of these factors in insect response to bacterial infection needs further investigation.

Rheumatoid arthritis (RA) is a chronic, systemic, inflammatory autoimmune disease that affects about 1% of the global population, with a female-male ratio of 3:1. RA preferably affects the joints, with consequent joint swelling and deformities followed by ankylosis. However, evidence has accumulated showing that patients suffering from RA can also develop extra-articular manifestations, including cardiovascular disease states, neuropathies, and multiorgan dysfunction. In particular, peripheral nerve disorders showed a consistent impact in the course of the disease (prevalence about 20%) mostly associated to vasculitis of the nerve vessels leading to vascular ischemia, axonal degeneration, and neuronal demyelination. The pathophysiological basis of this RA-associated microvascular disease, which leads to impairment of assonal functionality, is still to be better clarified. However, endothelial dysfunction and alterations of the so-called brain-nerve barrier (BNB) seem to play a fundamental role. This review aims to assess the potential mechanisms underlying the impairment of endothelial cell functionality in the development of RA and to identify the role of dysfunctional endothelium as a causative mechanism of extra-articular manifestation of RA. On the other hand, the potential impact of lifestyle and nutritional interventions targeting the maintenance of endothelial cell integrity in patients with RA will be discussed as a potential option when approaching therapeutic solutions in the course of the disease.

Since the isolation and commercialization of insulin (a peptide composed of 51 amino acid residues) in the early 1920s, peptide drugs have reshaped the pharmaceutical industry [...].