Search Results (297)

Intracranial aneurysms (IAs) represent a major clinical concern due to their risk of rupture and the resulting morbidity and mortality. Both environmental and genetic factors contribute to IA susceptibility, yet the genetic causes of IA remain poorly understood. We previously identified several single nucleotide variants (SNVs) in collagen XXII (COL22A1) in affected individuals with IA. However, the functional impact of these variants has not been determined, and it remains unclear whether and how they increase IA susceptibility. Here, we tested the functional effect of these variants in a zebrafish embryo model. Inducible overexpression of six human COL22A1 SNVs increased the incidence of cranial hemorrhage in zebrafish embryos, while overexpression of wild-type COL22A1 had no significant effect. Overexpression of DNA construct encoding COL22A1 P989L variant disrupted intracranial vascular architecture, leading to reduced vessel length, altered vascular surface parameters, and abnormal arterial patterning. Overexpression of the P989L SNV also caused pronounced vascular leakage, reduced pericyte number, and decreased expression of the tight junction proteins Claudin-5 and ZO-1. P989L SNV overexpression was also associated with increased expression of the endoplasmic reticulum stress marker hspa5. In silico modeling suggested that the P989L variant likely perturbs triple-helix formation in COL22A1, thereby causing protein misfolding and compromising its function. Together, these findings demonstrate the deleterious effects of IA-associated COL22A1 variants on vascular function and stability and suggest that these variants may increase the incidence of IA in humans. Full article

The purpose of this study was to investigate the adverse effects of 1.5 μg·L⁻¹ environmentally relevant chlorantraniliprole (CAP) on oxidase biomarkers (juvenile, 2.5 g) for 2, 4, and 8 h and transcriptomic response (adult, 254.8 g) for 96 and 192 h in American shad Alosa sapidissima (Wilson, 1981). American shad is sensitive to pollutants and has become an important economic fish in China, especially for recirculating the aquaculture system and photovoltaic farming. For juvenile shad under short-time CAP exposure, acid phosphatase (ACP) and aryl hydrocarbon receptase (AHR) at the protein level significantly increased at 2 h, and for longer-time exposure, alkaline phosphatase (AKP), polyphenol oxidase enzyme (PPO), and tumor necrosis factor alpha (TNFα) at the protein level significantly decreased; ryanodine receptase (RYR) at the protein level was significantly increased at 8 h. Interestingly, malondialdehyde (MDA) contents, biomarkers of oxidative stress, were significantly decreased for depletion at 2 h and 4 h, while they increased for eliminating free radicals at 8 h via longer-time CAP exposure duration. With the same CAP exposure for adult shad, the number of congested and dilated sinuses of the liver changed, with fine granular brown pigmentation and vacuolization of hepatocytes at 96 h, while the sinuses and central veins were dilated and edematous degeneration occurred at 192 h for longer-time exposure. The detected enzymatic activities, except for adenosine 5′-monophosphate (AMP)-activated protein kinase (AMPK), significantly decreased, and MDA contents significantly increased in adult shad at 96 and 192 h. Ribosome, proteasome, spliceosome, protein processing in endoplasmic reticulum, oxidative phosphorylation, glycerophospholipid metabolism, biosynthesis of amino acids, ferroptosis, peroxisome, apoptosis, necroptosis, and mTOR signaling pathways were the most significantly enriched pathways. For qPCR verification, the genes ppa2, pla1a, psmb13a, pkz and stat1b were significantly upregulated, while hspa8b, capn2, tram2, asns, bcl2l1, diablo, and prkcb were downregulated in adult shad. The results reveal elevated oxidative stress causing time-dependent hepatic damage via 1.5 μg·L⁻¹ CAP exposure both in juvenile and adult shad. Full article

Background: Low-dose aspirin (LDA) reduces preeclampsia (PE) risk by up to 40%, yet its molecular effects on chorion trophoblast cells (CTCs), a fetal membrane lineage at the feto-maternal interface, remain obscure. CTCs form a structural and immunoregulatory barrier whose dysfunction drives inflammation-associated membrane pathology in PE. Extracellular vesicles (EVs) released by CTCs may encode cellular stress and adaptation states, offering a molecular window into aspirin’s timing-dependent effects on PE risk modification. Methods: Human CTCs were challenged with cigarette smoke extract (CSE) to model oxidative stress-driven PE pathology. Two paradigms were tested: (1) prophylactic aspirin (4 and 40 µg/mL) before and/or flanking the CSE, and (2) therapeutic aspirin after the CSE challenge. The EVs were isolated via ultracentrifugation and size-exclusion chromatography, characterized by nanoparticle tracking and immunoblotting, and profiled by quantitative mass spectrometry. A network pathway analysis and machine learning biomarker selection defined the EV-encoded molecular states. Results: The CTC-derived EVs from the CSE-exposed cells carried a PE-like proteomic signature marked by suppressed VEGF/ECM remodeling, activated TNF-p53 apoptotic signaling, and heightened inflammation. Prophylactic low-dose aspirin shifted the EV cargo toward an EV-encoded signature consistent with preserved angiogenic potential (enrichment of VEGFA, COL1A1, and MMP14) and predicted attenuation of apoptotic and NF-κB pathway activity by an Ingenuity Pathway Analysis. High-dose aspirin produced broad transcriptional suppression without an accompanying pro-angiogenic EV signature. Therapeutic (post-injury) aspirin partially attenuated the injury-associated EV cargo but did not restore the angiogenic EV signature. An exploratory machine learning analysis of EV proteomes identified a candidate prophylactic biomarker panel anchored by HSPA8, SERPINF2, COL4A1, and PLOD1, mapped to the predicted angiogenic recovery and redox-balance pathways. These EV cargo readouts represent the predicted molecular states and require functional validation before clinical interpretation. Conclusions: The CTC-derived EV proteomic signatures capture the dose- and timing-dependent aspirin effects in this in vitro CTC model, positioning the chorion as a candidate pharmacological “secondary responder” favoring cellular resilience over classical anti-inflammatory suppression. As an exploratory hypothesis-generating study, EV-based molecular profiling could provide a foundation for future investigations aimed at stratifying aspirin responders from non-responders, although clinical validation in maternal plasma cohorts will be required before any translational application. Full article

Fetal growth restriction (FGR) is frequently defined as the failure of the fetus to reach its genetically predetermined growth potential. Heat shock proteins (HSPs) are extreme-temperature-resistant molecules that help proteostasis. The aim of this prospective case–control immunohistochemistry study is to evaluate the expression of HSP90 and HSP70 in the placentas of pregnancies complicated with FGR and compare their levels with the control placentas of normal-growth pregnancies. A prospective case–control study was conducted including people undergoing singleton pregnancies who gave birth in a tertiary university hospital in Central Greece. Participants were divided into two equal groups: an FGR pregnancy group and a control group with normal growth. Immunohistochemistry of placental samples was assessed using anti-HSP90 alpha/beta antibody (clone F-8, Santa Cruz Biotechnology, Dallas, TX, USA) and anti-HSC70/HSP70 antibody (clone W27, sc-24, Santa Cruz Biotechnology, Dallas, TX, USA). A scoring system was created to quantify the expression of HSP90 and HSP70 in each sample, and the grade of staining was measured at four points. A total of 80 pregnant people were prospectively enrolled in our study, with 40 in each group. Both constitutive (HSP90β and HSC70/HSPA8) and stress-inducible (HSP90α and HSP70/HSPA1A/B) isoforms were analyzed. When comparing the total score of HSP expression, a statistically significant difference was observed for both HSP90 and HSP70. For HSP90 expression, only the Hofbauer cell’s stain was identified as a statistically significant independent factor, meaning that its positive expression was observed in Hofbauer cells. For HSP70 expression, only the staining of syncytiotrophoblasts was identified as an independent factor. FGR is a common pregnancy complication and a leading cause of stillbirth, neonatal mortality, and short- and long-term neonatal morbidity worldwide. Based on our findings, the lower expression levels of both HSP90 and HSP70 are associated with FGR, revealing a possible association with stress response in FGR pathophysiology. However, more robust data from larger-scale prospective studies are needed to elucidate the possible role of HSPs as potential FGR biomarkers. Full article

Heat stress (HS) is among the most economically consequential environmental challenges to global dairy production, causing progressive declines in milk yield, compositional quality, and mammary cellular integrity. The temperature–humidity index (THI) is the primary thermal load metric, with performance-impairment thresholds typically beginning at THI 68 in Holstein cattle, with severe impacts manifesting beyond THI 72; breed-specific thresholds for Jersey, Brown Swiss, and Simmental cows differ owing to their lower metabolic heat load and greater inherent thermotolerance. At the molecular level, HS activates heat shock protein networks—notably HSPA1A, HSP90B1, and HSPH1—through HSF1/HSF4 transcriptional activation, while simultaneously suppressing casein genes (CSN1S1, CSN2, CSN3), lipogenic genes (FASN, SCD, CD36), amino acid transporters (SLC7A5, SLC38A2), and mTOR-AKT-STAT5 translational machinery, collectively impairing milk biosynthetic capacity. Pro-apoptotic signaling (BAX, CASP3 upregulation; BCL2 downregulation) and mitochondrial dysfunction further compromise mammary epithelial viability. Post-transcriptional regulation through miRNA, circRNA, and lncRNA competing endogenous RNA networks, alongside epitranscriptomic m6A modifications, adds further regulatory complexity. Genome-wide association studies have identified SNPs in HSP70A1A, HSPA4, TLR4, and PRLR as thermotolerance candidates compatible with sustained milk production. Nutritional supplementation with methionine, arginine, and taurine partially restores cellular synthetic capacity. Integrating multi-trait genomic selection with Bos indicus introgression, precision cooling, and targeted nutrition offers the most viable path toward climate-resilient, high-producing dairy cattle. Full article

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by immune dysregulation and multi-organ damage. Abnormal B cell activation and autoantibody production constitute the core pathological mechanism of SLE. However, the proportion, BCR pairing types, clonal evolution patterns, and transcriptomic features of dual BCR B cells in SLE remain incompletely elucidated. In this study, we employed single-cell RNA sequencing (scRNA-seq) combined with single-cell B cell receptor repertoire sequencing (scBCR-seq) to preliminarily analyze the proportion and characteristics of dual BCR B cells in SLE model mice (MRL/Lpr and SLE.Yaa) as well as in peripheral blood from SLE patients. The results showed: (1) Compared with control groups, the proportion of dual BCR B cells in SLE model mice and patients exhibited a decreasing trend, whereas the diversity of the CDR3 repertoire decreased and clonality increased. Increased clonal sharing was observed between single BCR B cells and dual BCR B cells. The main pairing types of dual BCR B cells were H + κ1 + κ2, H1 + H2 + κ, and H1 + H2 + κ + λ, with preferential utilization of autoimmunity-associated V gene families such as IGHV4-34, and high expression of IGHG subtypes. (2) Tracking analysis of B cell receptor clonality and effector molecule expression revealed that in SLE, dual BCR B cells tend to enrich in IFN-α/γ responses, TNF-NFκB inflammation, and complement pathways, and highly express interferon-related genes such as Ly6a, Isg15, MX1, and IFI6. (3) In both single BCR B and dual BCR B cells from SLE patients, the proportion of the naïve B cell subset decreased, whereas the proportions of plasma and Breg subsets increased and exhibited clonal expansion. SLE dual BCR Breg cells highly expressed IL10, HSPA1A, and others. This study is the first to reveal, at the high-throughput single-B-cell level, that the proportion, subset origin distribution, CDR3 repertoire composition, and effector molecule expression of dual BCR B cells display unique characteristics in SLE model mice and patients, providing baseline comparative data and novel research perspectives for further investigation into B cell effector functions and mechanisms in SLE patients. Full article

Protein–protein interactions (PPIs) play a central role in regulating cellular processes. However, the identification and characterization of senescence-associated PPIs remain challenging. In this study, we developed and evaluated an integrative methodology based on selective proteomics to identify PPIs associated with cellular senescence induced by TMPRSS11a. We started from isolated proteins that co-immunoprecipitated with TMPRSS11a and were subsequently identified by mass spectrometry. Building on this dataset, we implemented a workflow combining selective proteomics, structural bioinformatics, and experimental validation. Using this approach, we investigated the interaction between the transmembrane serine protease TMPRSS11a and the chaperone HSPA8. Structural bioinformatics analyses were performed to identify potential residues involved in the interaction interface, and the proximity of the TMPRSS11a–HSPA8 complex was evaluated using an in vitro proximity ligation assay. Our results provide evidence for an interaction between TMPRSS11a and HSPA8 and suggest its association with an enhanced senescence response. Overall, this study presents a workflow to investigate senescence-associated PPIs from proteomics-derived candidate proteins. Full article

Coronavirus disease 2019 continues to pose global health challenges, with the pandemic significantly burdening several economies, healthcare systems, and the social lives of individuals. Furthermore, new cases continue to be reported, underscoring the need for therapeutic strategies targeting conserved regions and host–virus interactions. Building on earlier virtual screening for small molecules, all-atom molecular dynamics simulations and binding-free-energy calculations were performed to elucidate how the two previously identified small molecules (NSC36398 and NSC281245) may affect the dynamic behaviour of the interaction between heat shock 70 kDa protein 8 (HSPA8) and the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike glycoprotein. Post-MD analyses refined prior docking predictions, where NSC281245 was found to bind tightly to the complex with limited perturbations at the HSPA8-spike protein interaction surface, whereas NSC36398 appeared to induce allosteric-like domain-level destabilisation effects while maintaining stable polar contacts with the protein. Our findings demonstrate the potential of NSC36398 as a promising modulator for disrupting the HSPA8-spike protein complex, which may serve as a structural lead for designing next-generation inhibitors of host–virus interactions. Full article

Photoaging is an extrinsic skin aging process caused by chronic ultraviolet (UV) radiation. A core pathological feature of photoaging is excessive oxidative stress, which can further induce ferroptosis. The HSP70 family plays a critical role in this stress response by protecting the key antioxidant enzyme GPX4. In this study, we established UV-induced photoaging models in cultured cells and 3D skin organoids. UPLC-MS/MS analysis of Chenpi transdermal permeate (prepared by in vitro transdermal penetration of Chenpi extract through mouse skin) identified hesperidin as the primary bioactive compound of Chenpi (dried peel of the plant Citrus reticulata Blanco after the aging process). The efficacy of hesperidin was validated in human keratinocytes (HaCaTs), fibroblasts (HSFs), and skin organoids. Mechanistically, transcriptomic and metabolomics analysis indicated that ferroptosis is a key pathway through which hesperidin ameliorates photoaging. Limited proteolysis mass spectrometry (LiP-MS), transcriptomics, and molecular dynamics simulation results demonstrated that hesperidin directly binds to the molecular chaperone HSPA1L. By upregulating HSPA1L expression, hesperidin enhanced the stability of GPX4 and suppressed UV-triggered ferroptosis. Our findings identify the HSPA1L/GPX4 axis as a critical redox regulatory pathway targeted by hesperidin, providing a mechanistic foundation for anti-photoaging therapies. Full article

With the rapid development of cancer treatment, immunotherapy has revolutionized renal cell carcinoma (RCC) treatment, yet patient responses remain heterogeneous. Here, a computational pipeline was constructed by integrating single-cell and bulk RNA sequencing data to identify immune-related candidate driver genes and characterize their impact on RCC immunotherapy. Based on gene regulatory networks (GRN), 25 immune-related candidate driver genes were identified, leading to the stratification of patients into three clusters (C1–C3). Compared to the C2/C3 cluster, the C1 cluster exhibited elevated immune infiltration, tumor mutation burden and checkpoint expression, which may represent immunotherapy responders. Dynamic analysis of GRNs revealed the critical role of candidate driver genes in predicting the efficacy of immunotherapy. IRF1, IRF9 and STAT1 in lymphoid cells of C1 participated in anti-tumor immune response by impacting target genes CD8A, HLA-A/E, TAP1 and PD-1. JUN, FOS, STAT3, JUND and NR2F1 were up-regulated in clusters C2 and C3, leading to tumor progression and immune evasion by influencing target genes HSPA1A, CXCL9 and PDGFR. In conclusion, integration of the transcriptome with molecular networks provided a network-based framework to uncover immune-related candidate driver genes for stratifying RCC patients, thereby serving as potential therapeutic targets to improve the outcome of RCC immunotherapy. Full article

Background: Metabolic-dysfunction-associated steatotic liver disease (MASLD) is a growing global health burden, yet no approved pharmacological therapy currently exists. Purpose: The purpose of this study is to investigate the prophylactic and therapeutic potential of Rebamipide, a mucosal-protective and anti-inflammatory drug, in a high-fat diet (MHFD)-induced MASLD rat model, integrating quantitative liver proteomics, network analysis, and histopathology. Methods: Male Wistar rats were fed MHFD for 16 weeks and treated with Rebamipide either prophylactically (Reb T1, co-administered with diet) or therapeutically (Reb T2, administered post-NASH onset). Label-free LC-MS/MS proteomics combined with principal component analysis (PCA), partial squares discriminant analysis (PLS-DA), and enrichment analyses (including Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), Reactome via g: Profiler, network mapping, and Rat Genome Database (RGD) mining) revealed that MHFD had the following impacts: it induced the profound suppression of mitochondrial chaperones (Hspa9), microsomal triglyceride transfer protein (Mttp), and cytochrome P450 isoforms (Cyp2c6); it disrupted lipid trafficking, oxidative stress defense, and xenobiotic metabolism. Results: Rebamipide prophylaxis preserved lipid-handling proteins, prevented glycogen loss, and maintained antioxidant defenses. In contrast, therapeutic administration reversed established steatosis and remodeled metabolic pathways, enhancing fatty acid β-oxidation, detoxification, and mitochondrial protein import. Nine shared proteins across all comparisons, including MTTP and multiple Stress-70 mitochondrial isoforms, mapped to three core genes (Mttp, Cyp2c6, Hspa9) central to lipid transport, protein import, and metabolic stress adaptation. KEGG and Reactome analyses highlighted Rebamipide’s modulation of bile acid synthesis, ceramide and phosphatidylcholine metabolism, lipoprotein remodeling, and MAPK signaling. Histopathological evaluation confirmed Rebamipide’s efficacy, showing reduced steatosis and the normalization of the hepatocyte structure, with near-complete restoration in the therapeutic (Reb T2) group compared to partial protection in the Reb T1 group. Conclusions: These findings demonstrate Rebamipide’s dual-phase, multi-targeted mechanism: early protection against diet-induced metabolic injury and robust reversal of established MASLD pathology. The identified protein triad (Mttp, Cyp2c6, Hspa9) and associated pathways provide novel biomarker candidates and mechanistic insight supporting Rebamipide’s repurposing as a therapeutic for metabolic liver disease. Full article

Endogenous heat shock cognate 73 kDa protein (HSC70) plays a role in early embryonic development and cellular stress regulation. This study evaluated the effects of exogenous recombinant HSC70 supplementation on bovine embryo development and the expression of apoptosis-related genes under in vitro conditions. Expression analyses of HSPA1A, HSPA8, BCL-2, and BAX were performed on Day 7 bovine embryos produced in vivo and in vitro. In vitro embryos exhibited higher basal expressions of HSPA8, BAX and BCL-2 compared with in vivo embryos (p ≤ 0.001). Supplementation with 500 or 1000 ng/mL HSC70 was associated with increased expression of HSPA1A, HSPA8, BCL-2, and BAX relative to control embryos (p ≤ 0.01). The 1000 ng/mL group showed significantly higher HSPA8 expression compared with both the control and 500 ng/mL groups. Morphological evaluation indicated that embryos supplemented with 500 ng/mL were associated with improved blastocyst yield and quality compared with control and 1000 ng/mL groups.In conclusion, supplementation with 500 ng/mL recombinant HSC70 was associated with modulation of apoptosis-related gene expression and improved morphological developmental parameters under in vitro conditions. These findings indicate dose-dependent regulatory effects of HSC70 on apoptosis-related signaling pathways; however, as apoptosis was assessed at the transcriptional level, the results should be interpreted as molecular associations rather than direct confirmation of altered apoptotic activity. Full article

To explore the regulatory aspects of mRNAs and miRNAs in suicide, we integrated transcriptomic data from GEO datasets. The analysis of mRNA expression in the prefrontal cortex of suicide victims with major depressive disorder revealed a differential profile with 27 downregulated mRNAs, including HSPA1A, HSPA1B, DNAJB1, NR4A1, and GADD45B, which are involved in proteostasis, transcriptional regulation, and apoptosis. Functional enrichment analysis using KEGG and Gene Ontology (GO) revealed significant associations with synaptic plasticity, neuronal survival, and signaling pathways, including MAPK, TGF-β, Wnt, p53, and neurotrophins. Subsequently, using the GSE34120 GEO dataset of miRNAs from the frontal cortex of suicide victims, 105 dysregulated miRNAs were identified. The networks revealed compact regulatory modules with hsa-miR-576-3p, hsa-miR-493, and hsa-miR-550, as well as highly connected central nodes such as hsa-miR-30b, hsa-miR-16a-5p, hsa-miR-181a-5p, and hsa-miR-184. The integration of both profiles allowed the elaboration of miRNA–mRNA regulatory networks in which TP53, FOXO3, RELA, and FOS interact with multiple dysregulated miRNAs. These findings support the notion that suicide involves complex post-transcriptional dysregulation, particularly related to astrocytic function and neurotrophic signaling, with potential diagnostic and therapeutic applications. Full article

Wuweiganlu (WGL) is a traditional formulation widely applied in the treatment of rheumatoid arthritis (RA), yet the identity of its bioactive constituents remains inadequately defined. In this study, ultra-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS) and untargeted serum metabolomics were employed to characterize the active components of WGL. Fraxin was identified as a principal compound from WGL. To investigate its therapeutic mechanism in RA, a series of in silico and experimental approaches were conducted. Network pharmacology analysis and RNA sequencing identified heat shock protein family member 8 (HSPA8) as a potential molecular target of Fraxin, which was further validated by molecular docking studies. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses indicated that Fraxin exerts its effects primarily by modulating cell apoptosis through the PI3K signaling pathway. In vitro experiments demonstrated that Fraxin significantly reduced inflammatory responses and downregulated HSPA8 expression in lipopolysaccharide (LPS)-stimulated fibroblast-like synoviocytes (FLs) and macrophages. In vivo, Fraxin administration markedly reduced paw swelling, alleviated bone deformities, and improved bone volume fraction (BV/TV) in male IL1RA-deficient mice exhibiting spontaneous arthritis. Histological analysis confirmed that Fraxin attenuated joint inflammation by modulating the inflammatory microenvironment. Additionally, Fraxin inhibited synovial hyperplasia by regulating mitochondrial membrane potential collapse in FLs. Functional assays revealed that this regulation occurred via the inhibition of HSPA8/PI3K/AKT signaling axis, thereby suppressing aberrant FLS proliferation and contributing to the attenuation of RA progression. Full article

Immune thrombocytopenia (ITP) is a hematological disorder commonly found in individuals of any gender, race, or age. Patients with ITP will present with thrombocytopenia either in a primary form or because of an infection or a dysfunction in the immune system. The severity of ITP is linked to diminished production of platelets due to the blockage of production in the bone marrow niche and increased destruction of platelets, which confirms the diagnosis of the disorder. The investigation of the pathogenesis of ITP is of critical importance as it can give an important indication of the state of the patient, guiding us through risk assessment and treatment. Proteomics can provide tools to explore the protein profile of ITP. In this review, we aimed to uncover different biomarkers, both diagnostic and prognostic, that have been investigated with proteomic methodologies and that might help in understanding the pathogenesis of ITP and providing personalized treatment to patients. Several differentially abundant proteins were identified, including haptoglobin isoforms, heat shock proteins (HSPA6, HSPA8), integrin β3 (ITGB3), 14-3-3 protein eta (YWHAH), vitamin D-binding protein, fibrinogen chains, MYH9, and FETUB, which are involved in key signaling pathways, such as PI3K/akt, TNF-a, and mTOR, and they demonstrate potential as diagnostic and prognostic biomarkers. Collectively, current data support the value of proteomics for uncovering the molecular landscape of ITP and guiding the development of precision diagnostics and personalized therapeutic strategies. Full article