Alzheimer's disease (AD) is the most common neurodegenerative disorders presenting with the pathological hallmarks of amyloid plaques and tau tangles. Over the past few years, great efforts have been made to explore reliable biomarkers of AD. High-throughput omics are a technology driven by multiple levels of unbiased data to detect the complex etiology of AD, and it provides us with new opportunities to better understand the pathophysiology of AD and thereby identify potential biomarkers. Through revealing the interaction networks between different molecular levels, the ultimate goal of multi-omics is to improve the diagnosis and treatment of AD. In this review, based on the current AD pathology and the current status of AD diagnostic biomarkers, we summarize how genomics, transcriptomics, proteomics and metabolomics are all conducing to the discovery of reliable AD biomarkers that could be developed and used in clinical AD management.

A recent explosion of methods to produce human trophoblast and stem cells (hTSCs) is fuelling a renewed interest in this tissue. The trophoblast is critical to reproduction by facilitating implantation, maternal physiological adaptations to pregnancy and the growth of the fetus through transport of nutrients between the mother and fetus. More broadly, the trophoblast has phenotypic properties that make it of interest to other fields. Its angiogenic and invasive properties are similar to tumours and could identify novel drug targets, and its ability to regulate immunological tolerance of the allogenic fetus could lead to improvements in transplantations. Within this review, we integrate and assess transcriptomic data of cell-based models of hTSC alongside in vivo samples to identify the utility and applicability of these models. We also integrate single-cell RNA sequencing data sets of human blastoids, stem cells and embryos to identify how these models may recapitulate early trophoblast development.

The non-classical human leukocyte antigen (HLA)-G exerts immune-suppressive properties modulating both NK and T cell responses. While it is physiologically expressed at the maternal-fetal interface and in immune-privileged organs, HLA-G expression is found in tumors and in virus-infected cells. So far, there exists little information about the role of HLA-G and its interplay with immune cells in biopsies, surgical specimen or autopsy tissues of lung, kidney and/or heart muscle from SARS-CoV-2-infected patients compared to control tissues. Heterogeneous, but higher HLA-G protein expression levels were detected in lung alveolar epithelial cells of SARS-CoV-2-infected patients compared to lung epithelial cells from influenza-infected patients, but not in other organs or lung epithelia from non-viral-infected patients, which was not accompanied by high levels of SARS-CoV-2 nucleocapsid antigen and spike protein, but inversely correlated to the HLA-G-specific miRNA expression. High HLA-G expression levels not only in SARS-CoV-2-, but also in influenza-infected lung tissues were associated with a high frequency of tissue-infiltrating immune cells, but low numbers of CD8+ cells and an altered expression of hyperactivation and exhaustion markers in the lung epithelia combined with changes in the spatial distribution of macrophages and T cells. Thus, our data provide evidence for an involvement of HLA-G and HLA-G-specific miRNAs in immune escape and as suitable therapeutic targets for the treatment of SARS-CoV-2 infections.

Eph receptors and their ligands, Ephrins, are involved in the thymocyte-thymic epithelial cell (TEC) interactions, key for the functional maturation of both thymocytes and thymic epithelium. Several years ago, we reported that the lack of EphA4, a Eph of the subfamily A, coursed with reduced proportions of double positive (DP) thymocytes apparently due to an altered thymic epithelial stroma [Munoz et al. in J Immunol 177:804-813, 2006]. In the present study, we reevaluate the lymphoid, epithelial, and extracellular matrix (ECM) phenotype of EphA4-/- mice grouped into three categories with respect to their proportions of DP thymocytes. Our results demonstrate a profound hypocellularity, specific alterations of T cell differentiation that affected not only DP thymocytes, but also double negative and single positive T cell subsets, as well as the proportions of positively and negatively selected thymocytes. In correlation, thymic histological organization changed markedly, especially in the cortex, as well as the proportions of both Ly51+UEA-1- cortical TECs and Ly51-UEA-1+ medullary TECs. The alterations observed in the expression of ECM components (Fibronectin, Laminin, Collagen IV), integrin receptors (VLA-4, VLA-6), chemokines (CXCL12, CCL25, CCL21) and their receptors (CXCR4, CCR7, CCR9) and in vitro transwell assays on the capacity of migration of WT and mutant thymocytes suggest that the lack of EphA4 alters T-cell differentiation by presumably affecting cell adhesion between TECs and T-TEC interactions rather than by thymocyte migration.

Repair-supportive mesenchymal cells (RSMCs) have been recently reported in the context of naphthalene (NA)-induced airway injury and regeneration. These cells transiently express smooth muscle actin (Acta2) and are enriched with platelet-derived growth factor receptor alpha (Pdgfra) and fibroblast growth factor 10 (Fgf10) expression. Genetic deletion of Ctnnb1 (gene coding for beta catenin) or Fgf10 in these cells using the Acta2-Cre-ERT2 driver line after injury (defined as NA-Tam condition; Tam refers to tamoxifen) led to impaired repair of the airway epithelium. In this study, we demonstrate that RSMCs are mostly captured using the Acta2-Cre-ERT2 driver when labeling occurs after (NA-Tam condition) rather than before injury (Tam-NA condition), and that their expansion occurs mostly between days 3 and 7 following NA treatment. Previous studies have shown that lineage-traced peribronchial GLI1+ cells are transiently amplified after NA injury. Here, we report that Gli1 expression is enriched in RSMCs. Using lineage tracing with Gli1Cre-ERT2 mice combined with genetic inactivation of Fgf10, we show that GLI1+ cells with Fgf10 deletion fail to amplify around the injured airways, thus resulting in impaired airway epithelial repair. Interestingly, Fgf10 expression is not upregulated in GLI1+ cells following NA treatment, suggesting that epithelial repair is mostly due to the increased number of Fgf10-expressing GLI1+ cells. Co-culture of SCGB1A1+ cells with GLI1+ cells isolated from non-injured or injured lungs showed that GLI1+ cells from these two conditions are similarly capable of supporting bronchiolar organoid (or bronchiolosphere) formation. Single-cell RNA sequencing on sorted lineage-labeled cells showed that the RSMC signature resembles that of alveolar fibroblasts. Altogether, our study provides strong evidence for the involvement of mesenchymal progenitors in airway epithelial regeneration and highlights the critical role played by Fgf10-expressing GLI1+ cells in this context.

Deficiency of decidual NK (dNK) cell number and function has been widely regarded as an important cause of spontaneous abortion. However, the metabolic mechanism underlying the crosstalk between dNK cells and embryonic trophoblasts during early pregnancy remains largely unknown. Here, we observed that enriched glutamine and activated glutaminolysis in dNK cells contribute to trophoblast invasion and embryo growth by insulin-like growth factor-1 (IGF-1) and growth differentiation factor-15 (GDF-15) secretion. Mechanistically, these processes are dependent on the downregulation of EGLN1-HIF-1α mediated by α-ketoglutarate (α-KG). Blocking glutaminolysis with the GLS inhibitor BPTES or the glutamate dehydrogenase inhibitor EGCG leads to early embryo implantation failure, spontaneous abortion and/or fetal growth restriction in pregnant mice with impaired trophoblast invasion. Additionally, α-KG supplementation significantly alleviated pregnancy loss mediated by defective glutaminolysis in vivo, suggesting that inactivated glutamine/α-ketoglutarate metabolism in dNK cells impaired trophoblast invasion and induced pregnancy loss.

The mitochondrial quality control of lung epithelial cells is disturbed during sepsis, which contributes to abnormal mitochondrial function and acute lung injury. Melatonin is one of the primary hormones secreted by the pineal gland, displaying favorable antioxidative actions in sepsis and cardiopulmonary disease. However, the potential roles and molecular basis of melatonin in lipopolysaccharide (LPS)-treated lung epithelial cells have not been explored and reported. Herein, we investigated whether melatonin could protect against sepsis-induced acute lung injury (ALI) and LPS-treated lung epithelial cells through the mitochondrial quality control as well as its possible molecular targets. Wild type and Sirt3 knockout mice were intratracheally instilled with LPS for 12 h to construct an in vivo acute lung injury model. Both A549 lung epithelial cells and primary alveolar type II (AT-II) cells were used to explore the possible roles of melatonin in vitro by incubating with small interfering RNA against Sirt3. To determine the involvement of the melatonin receptor, cells and mice were treated with si Mtnr1b and luzindole. Melatonin pretreatment significantly inhibited pathological injury, inflammatory response, oxidative stress, and apoptosis in LPS-treated lung tissues and LPS-treated lung epithelial cells. Furthermore, melatonin also shifted the dynamic course of mitochondria from fission to fusion, inhibited mitophagy and fatty acid oxidation in LPS-treated lung epithelial cells in vitro and in vivo. However, SIRT3 inhibition abolished the protective roles of melatonin in acute lung injury. Mechanistically, we found that melatonin increased the activity and expression of SIRT3, which further promoted the deacetylation of SOD2 at K122 and K68. More importantly, melatonin exerted pulmonary protection by activating MTNR1B but not MTNR1A during ALI. Collectively, melatonin could preserve the mitochondrial quality control of lung epithelial cells through the deacetylation of SOD2 in a SIRT3-dependent manner, which eventually alleviated sepsis-induced injury, inflammation, oxidative stress, and apoptosis. Thus, melatonin may serve as a promising candidate against ALI in the future.

High temperature-induced crop failures are prominent nowadays in major staples, including rice, wheat, and maize; however, crops such as foxtail millet (Setaria italica) are resilient to temperature stress. In this study, a novel small heat shock protein of foxtail millet, SisHSP21.9, is identified and characterized for its role in conferring tolerance to high-temperature stress. SisHSP21.9 is a panicoid-specific gene, which is highly upregulated during high-temperature in leaves, and the protein is localized in the chloroplast. Its expression is directly regulated by heat shock factor, SiHSFA2e, during temperature stress. Further, overexpression of SiHSP21.9 in rice enhanced the survival of transgenics during high-temperature stress (> 80% survival frequency), and the transgenic lines showed improved plant architecture and overall grain yield. Compared to WT plants, transgenic lines maintained optimal photosynthesis rates with higher photosystem efficiencies at high temperatures, and this is conferred through protecting the components of photosystems, chlorophyll-binding proteins, and chloroplast-localized functional proteins by SisHSP21.9. Prolonged high-temperature stress showed minimal damage to chloroplast proteins resulting in comparatively lower yield loss (35-37%) in transgenic lines. Altogether, the study suggests that SisHSP21.9 is a potential candidate for designing thermotolerant crops for climate-resilient agriculture; however, further research is needed because tolerance to abiotic stresses is polygenic.

The specification, characterization, and fate of alveolar type 1 and type 2 (AT1 and AT2) progenitors during embryonic lung development are poorly defined. Current models of distal epithelial lineage formation fail to capture the heterogeneity and dynamic contribution of progenitor pools present during early development. Furthermore, few studies explore the pathways involved in alveolar progenitor specification and fate. In this paper, we build upon our previously published work on the regulation of airway epithelial progenitors by fibroblast growth factor receptor 2b (FGFR2b) signalling during early (E12.5) and mid (E14.5) pseudoglandular stage lung development. Our results suggest that a significant proportion of AT2 and AT1 progenitors are lineage-flexible during late pseudoglandular stage development, and that lineage commitment is regulated in part by FGFR2b signalling. We have characterized a set of direct FGFR2b targets at E16.5 which are likely involved in alveolar lineage formation. These signature genes converge on a subpopulation of AT2 cells later in development and are downregulated in AT2 cells transitioning to the AT1 lineage during repair after injury in adults. Our findings highlight the extensive heterogeneity of pneumocytes by elucidating the role of FGFR2b signalling in these cells during early airway epithelial lineage formation, as well as during repair after injury.

Hydrogen sulfide (H2S) has been known for years as a poisoning gas and until recently evoked mostly negative associations. However, the discovery of its gasotransmitter functions suggested its contribution to various physiological and pathological processes. Although H2S has been found to exert cytoprotective effects through modulation of antioxidant, anti-inflammatory, anti-apoptotic, and pro-angiogenic responses in a variety of conditions, its role in the pathophysiology of skeletal muscles has not been broadly elucidated so far. The classical example of muscle-related disorders is Duchenne muscular dystrophy (DMD), the most common and severe type of muscular dystrophy. Mutations in the DMD gene that encodes dystrophin, a cytoskeletal protein that protects muscle fibers from contraction-induced damage, lead to prominent dysfunctions in the structure and functions of the skeletal muscle. However, the main cause of death is associated with cardiorespiratory failure, and DMD remains an incurable disease. Taking into account a wide range of physiological functions of H2S and recent literature data on its possible protective role in DMD, we focused on the description of the 'old' and 'new' functions of H2S, especially in muscle pathophysiology. Although the number of studies showing its essential regulatory action in dystrophic muscles is still limited, we propose that H2S-based therapy has the potential to attenuate the progression of DMD and other muscle-related disorders.

The viral epidemics and pandemics have stimulated the development of known and the discovery of novel antiviral agents. About a hundred mono- and combination antiviral drugs have been already approved, whereas thousands are in development. Here, we briefly reviewed 7 classes of antiviral agents: neutralizing antibodies, neutralizing recombinant soluble human receptors, antiviral CRISPR/Cas systems, interferons, antiviral peptides, antiviral nucleic acid polymers, and antiviral small molecules. Interferons and some small molecules alone or in combinations possess broad-spectrum antiviral activity, which could be beneficial for treatment of emerging and re-emerging viral infections.

Colorectal cancer (CRC) is a leading cause of cancer-related death worldwide, largely due to the development of colorectal liver metastases (CRLM). For the establishment of CRLM, CRC cells must remodel their tumor-microenvironment (TME), avoid the immune system, invade the underlying stroma, survive the hostile environment of the circulation, extravasate into the liver, reprogram the hepatic microenvironment into a permissive pre-metastatic niche, and finally, awake from a dormant state to grow out into clinically detectable CRLM. These steps form part of the invasion-metastasis cascade that relies on reciprocal interactions between the tumor and its ever-changing microenvironment. Such interplay provides a strong rational for therapeutically targeting the TME. In fact, several TME constituents, such as VEGF, TGF-β coreceptor endoglin, and CXCR4, are already targeted in clinical trials. It is, however, of utmost importance to fully understand the complex interactions in the invasion-metastasis cascade to identify novel potential therapeutic targets and prevent the establishment of CRLM, which may ultimately greatly improve patient outcome.

Lactate dehydrogenase 5 (LDH5) is overexpressed in many cancers and is a potential target for anticancer therapy due to its role in aerobic glycolysis. Small-molecule drugs have been developed as competitive inhibitors to bind substrate/cofactor sites of LDH5, but none reached the clinic to date. Recently, we designed the first LDH5 non-competitive inhibitor, cGmC9, a peptide that inhibits protein-protein interactions required for LDH5 enzymatic activity. Peptides are gaining a large interest as anticancer agents to modulate intracellular protein-protein interactions not targetable by small molecules; however, delivery of these peptides to the cytosol, where LDH5 and other anticancer targets are located, remains a challenge for this class of therapeutics. In this study, we focused on the cellular internalisation of cGmC9 to achieve LDH5 inhibition in the cytosol. We designed cGmC9 analogues and compared them for LDH5 inhibition, cellular uptake, toxicity, and antiproliferation against a panel of cancer cell lines. The lead analogue, [R/r]cGmC9, specifically impairs proliferation of cancer cell lines with high glycolytic profiles. Proteomics analysis showed expected metabolic changes in response to decreased glycolysis. This is the first report of a peptide-based LDH5 inhibitor able to modulate cancer metabolism and kill cancer cells that are glycolytic. The current study demonstrates the potential of using peptides as inhibitors of intracellular protein-protein interactions relevant for cancer pathways and shows that active peptides can be rationally designed to improve their cell permeation.

Trophoblasts are specialized epithelial cells that perform critical functions during blastocyst implantation and mediate maternal-fetal communication during pregnancy. However, our understanding of human trophoblast biology remains limited since access to first-trimester placental tissue is scarce, especially between the first and fourth weeks of development. Moreover, animal models inadequately recapitulate unique aspects of human placental physiology. In the mouse system, the isolation of self-renewing trophoblast stem cells has provided a valuable in vitro model system of placental development, but the derivation of analogous human trophoblast stem cells (hTSCs) has remained elusive until recently. Building on a landmark study reporting the isolation of bona fide hTSCs from blastocysts and first-trimester placental tissues in 2018, several groups have developed methods to derive hTSCs from pluripotent and somatic cell sources. Here we review the biological and molecular properties that define authentic hTSCs, the trophoblast potential of distinct pluripotent states, and methods for inducing hTSCs in somatic cells by direct reprogramming. The generation of hTSCs from pluripotent and somatic cells presents exciting opportunities to elucidate the molecular mechanisms of human placental development and the etiology of pregnancy-related diseases.

Experimental autoimmune-orchitis (EAO), a rodent model of chronic testicular inflammation and fibrosis, replicates pathogenic changes seen in some cases of human spermatogenic disturbances. During EAO, increased levels of pro-inflammatory and pro-fibrotic mediators such as TNF, CCL2, and activin A are accompanied by infiltration of leukocytes into the testicular parenchyma. Activin A levels correlate with EAO severity, while elevated CCL2 acting through its receptor CCR2 mediates leukocyte trafficking and recruits macrophages. CCR2 + CXCR4 + macrophages producing extracellular matrix proteins contribute widely to fibrogenesis. Furthermore, testicular macrophages (TMs) play a critical role in organ homeostasis. Therefore, we aimed to investigate the role of the activin A/CCL2-CCR2/macrophage axis in the development of testicular fibrosis. Following EAO induction, we observed lower levels of organ damage, collagen deposition, and leukocyte infiltration (including fibronectin+, collagen I+ and CXCR4+ TMs) in Ccr2-/- mice than in WT mice. Furthermore, levels of Il-10, Ccl2, and the activin A subunit Inhba mRNAs were lower in Ccr2-/- EAO testes. Notably, fibronectin+ TMs were also present in biopsies from patients with impaired spermatogenesis and fibrotic alterations. Overexpression of the activin A antagonist follistatin reduced tissue damage and collagen I+ TM accumulation in WT EAO testes, while treating macrophages with activin A in vitro increased the expression of Ccr2, Fn1, Cxcr4, and Mmp2 and enhanced migration along a CCL2 gradient; these effects were abolished by follistatin. Taken together, our data indicate that CCR2 and activin A promote fibrosis during testicular inflammation by regulating macrophage function. Inhibition of CCR2 or activin A protects against damage progression, offering a promising avenue for therapeutic intervention.

Ataxia telangiectasia mutated (ATM) is a serine-threonine protein kinase and important regulator of the DNA damage response (DDR). One critical ATM target is the structural subunit A (PR65-S401) of protein phosphatase 2A (PP2A), known to regulate diverse cellular processes such as mitosis and cell growth as well as dephosphorylating many proteins during the recovery from the DDR. We generated mouse embryonic fibroblasts expressing PR65-WT, -S401A (cannot be phosphorylated), and -S401D (phospho-mimetic) transgenes. Significantly, S401 mutants exhibited extensive chromosomal aberrations, impaired DNA double-strand break (DSB) repair and underwent increased mitotic catastrophe after radiation. Both S401A and the S401D cells showed impaired DSB repair (nonhomologous end joining and homologous recombination repair) and exhibited delayed DNA damage recovery, which was reflected in reduced radiation survival. Furthermore, S401D cells displayed increased ERK and AKT signaling resulting in enhanced growth rate further underscoring the multiple roles ATM-PP2A signaling plays in regulating prosurvival responses. Time-lapse video and cellular localization experiments showed that PR65 was exported to the cytoplasm after radiation by CRM1, a nuclear export protein, in line with the very rapid pleiotropic effects observed. A putative nuclear export sequence (NES) close to S401 was identified and when mutated resulted in aberrant PR65 shuttling. Our study demonstrates that the phosphorylation of a single, critical PR65 amino acid (S401) by ATM fundamentally controls the DDR, and balances DSB repair quality, cell survival and growth by spatiotemporal PR65 nuclear-cytoplasmic shuttling mediated by the nuclear export receptor CRM1.

Ataxia telangiectasia is a rare neurodegenerative disease caused by biallelic mutations in the ataxia telangiectasia mutated gene. No cure is currently available for these patients but positive effects on neurologic features in AT patients have been achieved by dexamethasone administration through autologous erythrocytes (EryDex) in phase II and phase III clinical trials, leading us to explore the molecular mechanisms behind the drug action. During these investigations, new ATM variants, which originated from alternative splicing of ATM messenger, were discovered, and detected in vivo in the blood of AT patients treated with EryDex. Some of the new ATM variants, alongside an in silico designed one, were characterized and examined in AT fibroblast cell lines. ATM variants were capable of rescuing ATM activity in AT cells, particularly in the nuclear role of DNA DSBs recognition and repair, and in the cytoplasmic role of modulating autophagy, antioxidant capacity and mitochondria functionality, all of the features that are compromised in AT but essential for neuron survival. These outcomes are triggered by the kinase and further functional domains of the tested ATM variants, that are useful for restoring cellular functionality. The in silico designed ATM variant eliciting most of the functionality recover may be exploited in gene therapy or gene delivery for the treatment of AT patients.

BACKGROUND: Parkinson's disease (PD) is characterized by selective and progressive dopamine (DA) neuron loss in the substantia nigra and other brain regions, with the presence of Lewy body formation. Most PD cases are sporadic, whereas monogenic forms of PD have been linked to multiple genes, including Leucine kinase repeat 2 (LRRK2) and PTEN-induced kinase 1 (PINK1), two protein kinase genes involved in multiple signaling pathways. There is increasing evidence to suggest that endogenous DA and DA-dependent neurodegeneration have a pathophysiologic role in sporadic and familial PD. METHODS: We generated patient-derived dopaminergic neurons and human midbrain-like organoids (hMLOs), transgenic (TG) mouse and Drosophila models, expressing both mutant and wild-type (WT) LRRK2 and PINK1. Using these models, we examined the effect of LRRK2 and PINK1 on tyrosine hydroxylase (TH)-DA pathway. RESULTS: We demonstrated that PD-linked LRRK2 mutations were able to modulate TH-DA pathway, resulting in up-regulation of DA early in the disease which subsequently led to neurodegeneration. The LRRK2-induced DA toxicity and degeneration were abrogated by wild-type (WT) PINK1 (but not PINK1 mutations), and early treatment with a clinical-grade drug, α-methyl-L-tyrosine (α-MT), a TH inhibitor, was able to reverse the pathologies in human neurons and TG Drosophila models. We also identified opposing effects between LRRK2 and PINK1 on TH expression, suggesting that functional balance between these two genes may regulate the TH-DA pathway. CONCLUSIONS: Our findings highlight the vital role of the TH-DA pathway in PD pathogenesis. LRRK2 and PINK1 have opposing effects on the TH-DA pathway, and its balance affects DA neuron survival. LRRK2 or PINK1 mutations can disrupt this balance, promoting DA neuron demise. Our findings provide support for potential clinical trials using TH-DA pathway inhibitors in early or prodromic PD.

Synapsin I (SynI) is a synaptic vesicle (SV)-associated phosphoprotein that modulates neurotransmission by controlling SV trafficking. The SynI C-domain contains a highly conserved ATP binding site mediating SynI oligomerization and SV clustering and an adjacent main Ca2+ binding site, whose physiological role is unexplored. Molecular dynamics simulations revealed that the E373K point mutation irreversibly deletes Ca2+ binding to SynI, still allowing ATP binding, but inducing a destabilization of the SynI oligomerization interface. Here, we analyzed the effects of this mutation on neurotransmitter release and short-term plasticity in excitatory and inhibitory synapses from primary hippocampal neurons. Patch-clamp recordings showed an increase in the frequency of miniature excitatory postsynaptic currents (EPSCs) that was totally occluded by exogenous Ca2+ chelators and associated with a constitutive increase in resting terminal Ca2+ concentrations. Evoked EPSC amplitude was also reduced, due to a decreased readily releasable pool (RRP) size. Moreover, in both excitatory and inhibitory synapses, we observed a marked impaired recovery from synaptic depression, associated with impaired RRP refilling and depletion of the recycling pool of SVs. Our study identifies SynI as a novel Ca2+ buffer in excitatory terminals. Blocking Ca2+ binding to SynI results in higher constitutive Ca2+ levels that increase the probability of spontaneous release and disperse SVs. This causes a decreased size of the RRP and an impaired recovery from depression due to the failure of SV reclustering after sustained high-frequency stimulation. The results indicate a physiological role of Ca2+ binding to SynI in the regulation of SV clustering and trafficking in nerve terminals.

Signaling from the Rho family small GTPases controls a wide range of signaling outcomes. Key among the downstream effectors for many of the Rho GTPases are the p21-activated kinases, or PAK group. The PAK family comprises two types, the type I PAKs (PAK1, 2 and 3) and the type II PAKs (PAK4, 5 and 6), which have distinct structures and mechanisms of regulation. In this review, we discuss signal transduction from Rho GTPases with a focus on the type II PAKs. We discuss the role of PAKs in signal transduction pathways and selectivity of Rho GTPases for PAK family members. We consider the less well studied of the Rho GTPases and their PAK-related signaling. We then discuss the molecular basis for kinase domain recognition of substrates and for regulation of signaling. We conclude with a discussion of the role of PAKs in cross talk between Rho family small GTPases and the roles of PAKs in disease.

Cervical cancer is the fourth most frequently diagnosed and fatal gynecological cancer. 15-61% of all cases metastasize and develop chemoresistance, reducing the 5-year survival of cervical cancer patients to as low as 17%. Therefore, unraveling the mechanisms contributing to metastasis is critical in developing better-targeted therapies against it. Here, we have identified a novel mechanism where nuclear Caspase-8 directly interacts with and inhibits the activity of CDK9, thereby modulating RNAPII-mediated global transcription, including those of cell-migration- and cell-invasion-associated genes. Crucially, low Caspase-8 expression in cervical cancer patients leads to poor prognosis, higher CDK9 phosphorylation at Thr186, and increased RNAPII activity in cervical cancer cell lines and patient biopsies. Caspase-8 knock-out cells were also more resistant to the small-molecule CDK9 inhibitor BAY1251152 in both 2D- and 3D-culture conditions. Combining BAY1251152 with Cisplatin synergistically overcame chemoresistance of Caspase-8-deficient cervical cancer cells. Therefore, Caspase-8 expression could be a marker in chemoresistant cervical tumors, suggesting CDK9 inhibitor treatment for their sensitization to Cisplatin-based chemotherapy.

The study aimed to investigate the potential role of lysine-specific demethylase 5A (KDM5A) in cisplatin-induced ototoxicity. The effect of the KDM5A inhibitor CPI-455 was assessed by apoptosis assay, immunofluorescence, flow cytometry, seahorse respirometry assay, and auditory brainstem response test. RNA sequencing, qRT-PCR, and CUT&Tag assays were used to explore the mechanism underlying CPI-455-induced protection. Our results demonstrated that the expression of KDM5A was increased in cisplatin-injured cochlear hair cells compared with controls. CPI-455 treatment markedly declined KDM5A and elevated H3K4 trimethylation levels in cisplatin-injured cochlear hair cells. Moreover, CPI-455 effectively prevented the death of hair cells and spiral ganglion neurons and increased the number of ribbon synapses in a cisplatin-induced ototoxicity mouse model both in vitro and in vivo. In HEI-OC1 cells, KDM5A knockdown reduced reactive oxygen species accumulation and improved mitochondrial membrane potential and oxidative phosphorylation under cisplatin-induced stress. Mechanistically, through transcriptomics and epigenomics analyses, a set of apoptosis-related genes, including Sos1, Sos2, and Map3k3, were regulated by CPI-455. Altogether, our findings indicate that inhibition of KDM5A may represent an effective epigenetic therapeutic target for preventing cisplatin-induced hearing loss.

Fibrosis is a relentlessly progressive and irreversible cause of organ damage, as in chronic kidney disease (CKD), but its underlying mechanisms remain elusive. We found that a circular RNA, circPTPN14, is highly expressed in human kidneys with biopsy-proved chronic interstitial fibrosis, mouse kidneys subjected to ischemia/reperfusion (IR) or unilateral ureteral obstruction (UUO), and TGFβ1-stimulated renal tubule epithelial cells (TECs). The intrarenal injection of circPTPN14 shRNA alleviated the progression of fibrosis in kidneys subjected to IR or UUO. Knockdown of circPTPN14 in TECs inhibited TGFβ1-induced expression of profibrotic genes, whereas overexpressing circPTPN14 increased the profibrotic effect of TGFβ1. The profibrotic action of circPTPN14 was ascribed to an increase in MYC transcription. The binding of circPTPN14 to the KH3 and KH4 domains of far upstream element (FUSE) binding protein 1 (FUBP1) enhanced the interaction between FUBP1 and FUSE domain, which was required for the initiation of MYC transcription. In human kidneys (n = 30) with biopsy-proved chronic interstitial fibrosis, the expression of circPTPN14 positively correlated with MYC expression. Taken together these studies show a novel mechanism in the pathogenesis of renal fibrosis, mediated by circPTPN14, which can be a target in the diagnosis and treatment of CKD.

Class I phosphoinositide 3-kinases (PI3Ks) are a family of lipid kinases. They are super elevated in many human cancer types and exert their main cellular functions by activating Akt to trigger an array of distinct responses, affecting metabolism and cell polarity. The signal equally plays important roles in cardiovascular pathophysiology. PI3K is required for cardiogenesis and regulation of cardiac structure and function. Overexpression of PI3K governs the development of cardiac pressure overload adaptation and compensatory hypertrophy. Therefore, inhibition of PI3K shortens life span, enhances cardiac dysfunction and pathological hypertrophy. The inverse inhibition effect, however, desirably destroys many cancer cells by blocking several aspects of the tumorigenesis phenotype. Given the contrasting effects in cardio-oncology; the best therapeutic strategy to target PI3K in cancer, while maintaining or rather increasing cardiac safety is under intense investigational scrutiny. To improve our molecular understanding towards identifying cardiac safety signalling of PI3K and/or better therapeutic strategy for cancer treatment, this article reviews PI3K signalling in cardio-oncology. PI3K signalling at the interface of metabolism, inflammation and immunity, and autonomic innervation networks were examined. Examples were then given of cardiovascular drugs that target the networks, being repurposed for cancer treatment. This was followed by an intersection scheme of the networks that can be functionalised with machine learning for safety and risk prediction, diagnoses, and defining new novel encouraging leads and targets for clinical translation. This will hopefully overcome the challenges of the one-signalling-one-health-outcome alliance, and expand our knowledge of the totality of PI3K signalling in cardio-oncology.

The cystic fibrosis transmembrane conductance regulator (CFTR) is an anion channel that is crucial for fluid homeodynamics throughout the male reproductive tract. Previous evidence shed light on a potential molecular partnership between this channel and aquaporins (AQPs). Herein, we explore the role of CFTR on AQPs-mediated glycerol permeability in mouse Sertoli cells (mSCs). We were able to identify the expression of CFTR, AQP3, AQP7, and AQP9 in mSCs by RT-PCR, Western blot, and immunofluorescence techniques. Cells were then treated with CFTRinh-172, a specific CFTR inhibitor, and its glycerol permeability was evaluated by stopped-flow light scattering. We observed that CFTR inhibition decreased glycerol permeability in mSCs by 30.6% when compared to the control group. A DUOLINK proximity ligation assay was used to evaluate the endogenous protein-protein interactions between CFTR and the various aquaglyceroporins we identified. We positively detected that CFTR is in close proximity with AQP3, AQP7, and AQP9 and that, through a possible physical interaction, CFTR can modulate AQP-mediated glycerol permeability in mSCs. As glycerol is essential for the control of the blood-testis barrier and elevated concentration in testis results in the disruption of spermatogenesis, we suggest that the malfunction of CFTR and the consequent alteration in glycerol permeability is a potential link between male infertility and cystic fibrosis.

Endothelial cells (EC) in vivo buffer and regulate the transfer of plasma fatty acid (FA) to the underlying tissues. We hypothesize that inflammation could alter the functionality of the EC, i.e., their capacity and uptake of different FA. The aim of this work is to verify the functionality of inflamed cells by analyzing their ability to uptake and accumulate exogenous saturated FA. Control and inflammatory human microvascular endothelial cells stimulated in vitro with two deuterium-labeled saturated FA (D-FA), i.e., palmitic (D31-PA) and myristic (D27-MA) acids. Cells were measured both by spontaneous and stimulated Raman imaging to extract detailed information about uptaken FA, whereas coherent anti-Stokes Raman scattering and fluorescence imaging showed the global content of FA in cells. Additionally, we employed atomic force microscopy to obtain a morphological image of the cells. The results indicate that the uptake of D-FA in inflamed cells is dependent on their concentration and type. Cells accumulated D-FA when treated with a low concentration, and the effect was more pronounced for D27-MA, in normal cells, but even more so, in inflamed cells. In the case of D31-PA, a slightly increased uptake was observed for inflamed cells when administered at higher concentration. The results provide a better understanding of the EC inflammation and indicate the impact of the pathological state of the EC on their capacity to buffer fat. All the microscopic methods used showed complementarity in the analysis of FA uptake by EC, but each method recognized this process from a different perspective.

Epstein-Barr virus (EBV), a human oncogenic herpesvirus with a typical life cycle consisting of latent phase and lytic phase, is associated with many human diseases. EBV can express a variety of proteins that enable the virus to affect host cell processes and evade host immunity. Additionally, these proteins provide a basis for the maintenance of viral infection, contribute to the formation of tumors, and influence the occurrence and development of related diseases. Posttranslational modifications (PTMs) are chemical modifications of proteins after translation and are very important to guarantee the proper biological functions of these proteins. Studies in the past have intensely investigated PTMs of EBV-encoded proteins. EBV regulates the progression of the latent phase and lytic phase by affecting the PTMs of its encoded proteins, which are critical for the development of EBV-associated human diseases. In this review, we summarize the PTMs of EBV-encoded proteins that have been discovered and studied thus far with focus on their effects on the viral life cycle.

Mechanoreceptors are implicated as functional afferents within mucosa of the airways and the recent discovery of mechanosensitive channels Piezo1 and Piezo2 has proved essential for cells of various mechanically sensitive tissues. However, the role for Piezo1/2 in vocal fold (VF) mucosal epithelia, a cell that withstands excessive biomechanical insult, remains unknown. The purpose of this study was to test the hypothesis that Piezo1 is required for VF mucosal repair pathways of epithelial cell injury. Utilizing a sonic hedgehog (shh) Cre line for epithelial-specific ablation of Piezo1/2 mechanoreceptors, we investigated 6wk adult VF mucosa following naphthalene exposure for repair strategies at 1, 3, 7 and 14 days post-injury (dpi). PIEZO1 localized to differentiated apical epithelia and was paramount for epithelial remodeling events. Injury to wildtype epithelium was most appreciated at 3 dpi. Shhcre/+; Piezo1loxP/loxP, Piezo2 loxP/+ mutant epithelium exhibited severe cell/nuclear defects compared to injured controls. Conditional ablation of Piezo1 and/or Piezo2 to uninjured VF epithelium did not result in abnormal phenotypes across P0, P15 and 6wk postnatal stages compared to heterozygote and control tissue. Results demonstrate a role for Piezo1-expressing VF epithelia in regulating self-renewal via effects on p63 transcription and YAP subcellular translocation-altering cytokeratin differentiation.

BACKGROUND: Allergic disorders are common all over the world. The pathogenesis of allergy is unclear. Therapies for allergic disorders require improvement. Endoplasmic reticulum (ER) stress is one of the factors influencing immune response. The purpose of this study is to improve the effectiveness of immunotherapy for experimental respiratory allergy by targeting the ER stress signal pathway. METHODS: Committed CD4+ T cells were isolated from blood samples collected from patients with allergic rhinitis (AR) and TCR ovalbumin transgenic mice. The effects of TCR engagement and 3-methyl-4-nitrophenol (MNP) on inducing ER stress in committed CD4+ T cells were evaluated. RESULTS: ER stress was detected in antigen-specific CD4+ T cells (sCD4+ T cells) of AR patients. The environmental pollutant MNP increased the expression of the X-binding protein-1 (XBP1) in the committed CD4+ T cells during the TCR engagement. XBP1 mediated the effects of MNP on inhibiting regulatory T cell (Treg) generation. The effects of MNP on induction of protein 20 (Rnf20) in CD4+ T cells were mediated by XBP1. Inhibition of Rnf20 rescued the Treg development from MNP-primed sCD4+ T cells. The ablation of Rnf20 improved the immunotherapy of AR through the restoration of the Treg generation. CONCLUSIONS: ER stress can be detected in CD4+ T cells in TCR engagement. Exposure to MNP exacerbates ER stress in committed CD4+ T cells. Regulation of the ER stress-related Rnf20 expression can restore the generation of Treg from CD4+ T cells of subjects with allergic diseases.

Autism spectrum disorders (ASD) are neurodevelopmental disorders. Genetic factors, along with non-genetic triggers, have been shown to play a causative role. Despite the various causes, a triad of common symptoms defines individuals with ASD; pervasive social impairments, impaired social communication, and repeated sensory-motor behaviors. Therefore, it can be hypothesized that different genetic and environmental factors converge on a single hypothetical neurobiological process that determines these behaviors. However, the cellular and subcellular signature of this process is, so far, not well understood. Here, we performed a comparative study using "omics" approaches to identify altered proteins and, thereby, biological processes affected in ASD. In this study, we mined publicly available repositories for genetic mouse model data sets, identifying six that were suitable, and compared them with in-house derived proteomics data from prenatal zinc (Zn)-deficient mice, a non-genetic mouse model with ASD-like behavior. Findings derived from these comparisons were further validated using in vitro neuronal cell culture models for ASD. We could show that a protein network, centered on VAMP2, STX1A, RAB3A, CPLX2, and AKAP5, is a key convergence point mediating synaptic vesicle release and recycling, a process affected across all analyzed models. Moreover, we demonstrated that Zn availability has predictable functional effects on synaptic vesicle release in line with the alteration of proteins in this network. In addition, drugs that target kinases, reported to regulate key proteins in this network, similarly impacted the proteins' levels and distribution. We conclude that altered synaptic stability and plasticity through abnormal synaptic vesicle dynamics and function may be the common neurobiological denominator of the shared behavioral abnormalities in ASD and, therefore, a prime drug target for developing therapeutic strategies.

Gastrulation and neurulation are successive morphogenetic processes that play key roles in shaping the basic embryonic body plan. Importantly, they operate through common cellular and molecular mechanisms to set up the three spatially organized germ layers and to close the neural tube. During gastrulation and neurulation, convergent extension movements driven by cell intercalation and oriented cell division generate major forces to narrow the germ layers along the mediolateral axis and elongate the embryo in the anteroposterior direction. Apical constriction also makes an important contribution to promote the formation of the blastopore and the bending of the neural plate. Planar cell polarity proteins are major regulators of asymmetric cell behaviors and critically involved in a wide variety of developmental processes, from gastrulation and neurulation to organogenesis. Mutations of planar cell polarity genes can lead to general defects in the morphogenesis of different organs and the co-existence of distinct congenital diseases, such as spina bifida, hearing deficits, kidney diseases, and limb elongation defects. This review outlines our current understanding of non-canonical Wnt signaling, commonly known as Wnt/planar cell polarity signaling, in regulating morphogenetic movements of gastrulation and neural tube closure during development and disease. It also attempts to identify unanswered questions that deserve further investigations.

Type 1 diabetes (T1D) is characterized by an immune-mediated progressive destruction of the insulin-producing β-cells. Proinflammatory cytokines trigger endoplasmic reticulum (ER) stress and subsequent insulin secretory deficiency in cultured β-cells, mimicking the islet microenvironment in T1D. β-cells undergo physiologic ER stress due to the high rate of insulin production and secretion under stimulated conditions. Severe and uncompensated ER stress in β-cells is induced by several pathological mechanisms before onset and during T1D. We previously described that the small drug Compound A (CpdA), a selective glucocorticoid receptor (GR/NR3C1, nuclear receptor subfamily 3, group C, member 1) ligand with demonstrated inflammation-suppressive activity in vivo, is an effective modulator of effector T and dendritic cells and of macrophages, yet, in a GR-independent manner. Here, we focus on CpdA's therapeutic potential in T1D cellular and animal models. We demonstrate that CpdA improves the unfolded protein response (UPR) by attenuating ER stress and favoring the survival and function of β-cells exposed to an environment of proinflammatory cytokines. CpdA administration to NODscid mice adoptively transferred with diabetogenic splenocytes (from diabetic NOD mice) led to a delay of disease onset and reduction of diabetes incidence. Histological analysis of the pancreas showed a reduction in islet leukocyte infiltration (insulitis) and preservation of insulin expression in CpdA-treated normoglycemic mice in comparison with control group. These new findings together with our previous reports justify further studies on the administration of this small molecule as a novel therapeutic strategy with dual targets (effector immune and β-cells) during autoimmune diabetes.

Spaceflight entails a variety of environmental and psychological stressors that may have long-term physiological and genomic consequences. Metabolomics, an approach that investigates the terminal metabolic outputs of complex physiological alterations, considers the dynamic state of the human body and allows the identification and quantification of down-stream metabolites linked to up-stream physiological and genomic regulation by stress. Employing a metabolomics-based approach, this study investigated longitudinal metabolic perturbations of male (n = 40) and female (n = 11) astronauts on 4-6-month missions to the International Space Station (ISS). Proton nuclear magnetic resonance (1H-NMR) spectroscopy followed by univariate, multivariate and machine learning analyses were used on blood serum to examine sex-specific metabolic changes at various time points throughout the astronauts' missions, and the metabolic effects of long-duration space travel. Space travel resulted in sex-specific changes in energy metabolism, bone mineral and muscle regulation, immunity, as well as macromolecule maintenance and synthesis. Additionally, metabolic signatures suggest differential metabolic responses-especially during the recovery period-with females requiring more time to adjust to return to Earth. These findings provide insight into the perturbations in glucose and amino acid metabolism and macromolecule biosynthesis that result from the stressors of long-duration spaceflight. Metabolomic biomarkers may provide a viable approach to predicting and diagnosing health risks associated with prolonged space travel and other physiological challenges on Earth.

Sulforaphane has been investigated in human pathologies and preclinical models of airway diseases. To provide further mechanistic insights, we explored L-sulforaphane (LSF) in the ovalbumin (OVA)-induced chronic allergic airways murine model, with key hallmarks of asthma. Histological analysis indicated that LSF prevented or reversed OVA-induced epithelial thickening, collagen deposition, goblet cell metaplasia, and inflammation. Well-known antioxidant and anti-inflammatory mechanisms contribute to the beneficial effects of LSF. Fourier transform infrared microspectroscopy revealed altered composition of macromolecules, following OVA sensitization, which were restored by LSF. RNA sequencing in human peripheral blood mononuclear cells highlighted the anti-inflammatory signature of LSF. Findings indicated that LSF may alter gene expression via an epigenetic mechanism which involves regulation of protein acetylation status. LSF resulted in histone and α-tubulin hyperacetylation in vivo, and cellular and enzymatic assays indicated decreased expression and modest histone deacetylase (HDAC) inhibition activity, in comparison with the well-known pan-HDAC inhibitor suberoylanilide hydroxamic acid (SAHA). Molecular modeling confirmed interaction of LSF and LSF metabolites with the catalytic domain of metal-dependent HDAC enzymes. More generally, this study confirmed known mechanisms and identified potential epigenetic pathways accounting for the protective effects and provide support for the potential clinical utility of LSF in allergic airways disease.