Search Results (650)

Background Plasma membrane proteins (PMPs) play key roles in cell signalling, adhesion, and trafficking, and are attractive therapeutic targets in cancer due to their surface accessibility. However, their typically low abundance limits detection by conventional proteomic approaches. Methods: To improve PMP detection, we employed a surface proteomics workflow combining cell surface biotinylation and affinity purification prior to LC-MS/MS analysis in cervical (SiHa) and bladder (UMUC3) cancer cell lines cultured under normoxic (21% O₂) or hypoxic (0.1% O₂) conditions. Results: In SiHa cells, 43 hypoxia-upregulated proteins were identified exclusively in the biotin-enriched fraction, including ITGB2, ITGA7, AXL, MET, JAG2, and CAV1/CAV2. In UMUC3 cells, 32 unique upregulated PMPs were detected, including CD55, ADGRB1, SLC9A1, NECTIN3, and ACTG1. These proteins were not observed in corresponding whole-cell lysates and are associated with extracellular matrix remodelling, immune modulation, and ion transport. Biotinylation enhanced the detection of membrane-associated pathways such as ECM organisation, integrin signalling, and PI3K–Akt activation. Protein–protein interaction analysis revealed links between membrane receptors and intracellular stress regulators, including mitochondrial proteins. Conclusions: These findings demonstrate that surface biotinylation improves the sensitivity and selectivity of plasma membrane proteomics under hypoxia, revealing hypoxia-responsive proteins and pathways not captured by standard whole-cell analysis. Full article

Heat stress in dairy cows, caused by high temperature and humidity during summer, has led to significant declines in milk production and severe economic losses for farms. Exosomes—extracellular vesicles carrying bioactive molecules—are critical for intercellular communication and immunity but remain understudied in heat-stressed Holstein cows. In this study, we extracted exosomes from three heat-stressed (HS) cows and three non-heat-stressed (Ctr) cows and employed proteomics to analyze plasma exosomes. We identified a total of 28 upregulated and 18 downregulated proteins in the HS group compared to the control group. Notably, we observed a significant upregulation of key protein groups, including cytoskeletal regulators, signaling mediators, and coagulation factors, alongside the downregulation of HP-25_1. These differentially expressed proteins demonstrate strong potential as heat stress biomarkers. GO and KEGG analyses linked the differentially expressed proteins to actin cytoskeleton regulation and endoplasmic reticulum pathways. Additionally, protein–protein interaction (PPI) analysis revealed the PI3K-Akt signaling pathway as a central node in the cellular response to heat stress. These findings establish plasma exosomes as valuable biospecimens, provide valuable insights into the molecular mechanisms of heat stress response, and may contribute to the development of precision breeding strategies for enhanced thermal resilience in dairy herds. Full article

Cancer is a systemic disease rather than a localized pathology and is characterized by widespread effects, including whole-body exhaustion and chronic inflammation. A thorough understanding of cancer pathophysiology requires a systemic approach that accounts for the complex interactions between cancer cells and host tissues. To explore these dynamics, we employed a comprehensive metabolomic analysis of plasma samples from patients with either esophageal or head and neck squamous cell carcinoma (SCC). Plasma samples from 149 patients were metabolically profiled and correlated with clinical data. Among the metabolites identified, lysophosphatidylcholine (LPC) emerged as the sole biomarker strongly correlated with prognosis. A significant reduction in plasma LPC levels was linked to poorer overall survival. Plasma LPC levels demonstrated minimal correlation with patient-specific factors, such as tumor size and general condition, but showed significant association with the response to immune checkpoint inhibitor therapy. Proteomic and cytokine analyses revealed that low plasma LPC levels reflected systemic chronic inflammation, characterized by high levels of inflammatory proteins, the cytokines interleukin-6 and tumor necrosis factor-α, and coagulation-related proteins. These findings indicate that plasma LPC levels may be used as reliable biomarkers for predicting prognosis and evaluating the efficacy of immunotherapy in patients with SCC. Full article

Background: Rheumatoid arthritis (RA) is an autoimmune disease that remains incurable. An increasing number of proteomic genome-wide association studies (GWASs) are emerging, offering immense potential for identifying novel therapeutic targets for diseases. This study aims to identify potential therapeutic targets for RA based on human plasma proteome. Methods: Protein quantitative trait loci were extracted and integrated from eight large-scale proteomic GWASs. Proteome-wide Mendelian randomization (Pro-MR) was performed to prioritize proteins causally associated with RA. Further validation of the reliability and stratification of prioritized proteins was performed using MR meta-analysis, colocalization, and transcriptome-wide summary-data-based MR. Subsequently, prioritized proteins were characterized through protein–protein interaction and enrichment analyses, pleiotropy assessment, genetically engineered mouse models, cell-type-specific expression analysis, and druggability evaluation. Phenotypic expansion analyses were also conducted to explore the effects of the prioritized proteins on phenotypes such as endocrine disorders, cardiovascular diseases, and other immune-related diseases. Results: Pro-MR prioritized 32 unique proteins associated with RA risk. After validation, prioritized proteins were stratified into four reliability tiers. Prioritized proteins showed interactions with established RA drug targets and were enriched in an immune-related functional profile. Four trans-associated proteins exhibited vertical or horizontal pleiotropy with specific genes or proteins. Genetically engineered mouse models for 18 prioritized protein-coding genes displayed abnormal immune phenotypes. Single-cell RNA sequencing data were used to validate the enriched expression of several prioritized proteins in specific synovial cell types. Nine prioritized proteins were identified as targets of existing drugs in clinical trials or were already approved. Further phenome-wide MR and mediation analyses revealed the effects and potential mediating roles of some prioritized proteins on other phenotypes. Conclusions: This study identified 32 plasma proteins as potential therapeutic targets for RA, expanding the prospects for drug discovery and deepening insights into RA pathogenesis. Full article

Background: The endogenous opioid system plays a pivotal role in numerous physiological processes and is implicated in a range of diseases, including atherosclerosis, a condition contributing to nearly 50% of deaths in Western societies. Objectives: This study investigates the effects of opioid receptor blockade, using naloxone, on the plasma lipid profile and atherosclerosis progression. Methods: ApoE^−/− mice with advanced atherosclerosis were treated with naloxone for seven days, and the effects on atherosclerotic plaque development and liver steatosis were evaluated. Results: A proteomic analysis of liver samples post-treatment identified 38 proteins with altered abundance. The results revealed that naloxone treatment led to an increase in HDL cholesterol, a lipid fraction associated with protective cardiovascular effects. Furthermore, naloxone did not influence the progression of atherosclerotic plaques or the development of liver steatosis. Conclusions: In conclusion, while short-term naloxone treatment in mice with advanced atherosclerosis does not alter overall atherosclerotic plaque progression or liver steatosis, the observed elevation in HDL cholesterol and the extensive changes in liver protein abundance underscore the complex and multifaceted role of the opioid system in lipid metabolism and cardiovascular health. These findings provide a foundation for further exploration of opioid receptor antagonists as modulators of lipid profiles and potential contributors to cardiovascular therapy. Full article

Rhabdomyosarcoma (RMS), the most common pediatric soft tissue sarcoma, comprises embryonal (ERMS) and alveolar (ARMS) subtypes with distinct histopathological features, clinical outcomes, and therapeutic responses. To better characterize their molecular distinctions, we performed untargeted plasma proteomics and metabolomics profiling in children with ERMS (n = 18), ARMS (n = 17), and matched healthy controls (n = 18). Differential expression, functional enrichment (GO, KEGG, RaMP-DB), co-expression network analysis (WGCNA/WMCNA), and multi-omics integration (DIABLO, MOFA) revealed distinct molecular signatures for each subtype. ARMS displayed elevated oncogenic and stemness-associated proteins (e.g., cyclin E1, FAP, myotrophin) and metabolites involved in lipid transport, fatty acid metabolism, and polyamine biosynthesis. In contrast, ERMS was enriched in immune-related and myogenic proteins (e.g., myosin-9, SAA2, S100A11) and metabolites linked to glutamate/glycine metabolism and redox homeostasis. Pathway analyses highlighted subtype-specific activation of PI3K-Akt and Hippo signaling in ARMS and immune and coagulation pathways in ERMS. Additionally, the proteomics and metabolomics datasets showed association with clinical parameters, including disease stage, lymph node involvement, and age, demonstrating clear molecular discrimination consistent with clinical observation. Co-expression networks and integrative analyses further reinforced these distinctions, uncovering coordinated protein–metabolite modules. Our findings reveal novel, subtype-specific molecular programs in RMS and propose candidate biomarkers and pathways that may guide precision diagnostics and therapeutic targeting in pediatric sarcomas. Full article

Artificial insemination (AI) is a key breeding technique in the swine industry; however, the lack of reliable biomarkers for semen quality limits its effectiveness. Seminal plasma (SP) contains extracellular vesicles (EVs) that present a promising, non-invasive biomarker for semen quality. This study explores the biochemical profiles of boar SP to assess semen quality through near-infrared spectroscopy (NIRS) and proteomics of SP-EVs. Fresh semen from mature Duroc boars was evaluated based on sperm motility, classifying samples as Passed (≥70%) or Failed (<70%). NIRS analysis identified distinct variations in water structures at specific wavelengths (C1, C5, C12 nm), achieving high accuracy (92.2%), sensitivity (94.2%), and specificity (90.3%) through PCA-LDA. Proteomic analysis of SP-EVs revealed 218 proteins in Passed and 238 in Failed samples. Nexin-1 and seminal plasma protein pB1 were upregulated in Passed samples, while LGALS3BP was downregulated. The functional analysis highlighted pathways associated with single fertilization, filament organization, and glutathione metabolism in Passed samples. Integrating NIRS with SP-EV proteomics provides a robust approach to non-invasive assessment of semen quality. These findings suggest that SP-EVs could serve as effective biosensors for rapid semen quality assessment, enabling better boar semen selection and enhancing AI practices in swine breeding. Full article

Background: Immune checkpoint inhibitors (ICIs) have transformed cancer treatment, yet severe immune-related adverse events (irAEs) often necessitate immunotherapy discontinuation and cause life-threatening complications. Circulating plasma proteins, dynamically accessible and functionally linked to immunity, may predict and offer novel targets for irAEs. Methods: Leveraging multi-omics integration, we conducted bidirectional two-sample Mendelian randomization (MR) using protein quantitative trait loci (pQTLs) from 4998 plasma proteins and genome-wide association data of irAE phenotypes. A causal inference framework combining colocalization analysis, multivariable MR (MVMR) adjusting for body mass index (BMI) confounding, and mediation MR elucidated BMI-independent pathways. Systems biology approaches including tissue-specific expression profiling, pathway enrichment, and protein interaction network analysis revealed spatial and functional drivers of irAE pathogenesis. Results: Proteome-wide MR mapping identified eight plasma proteins (CCL20, CSF1, CXCL9, CD40, TGFβ1, CLSTN2, TNFSF12, TGFα) causally associated with all-grade irAEs, and five (CCL20, CCL25, CXCL10, ADA, TGFα) with high-grade irAEs. Colocalization prioritized CD40/TNFSF12 (all-grade) and ADA/CCL25 (high-grade) as therapeutic targets (PPH4 > 0.7). CXCL9/TNFSF12 (all-grade) and CCL25 (high-grade) exerted BMI-independent effects, suggesting intrinsic immune dysregulation mechanisms. Tissue-specific gene expression patterns, CSF1, TGFβ1 in lung, TNFSF12 in the ileum may explain organ-specific irAE vulnerabilities. High-grade irAEs correlated with compartmentalized immune dysregulation and IL-17/immunodeficiency pathway activation. Conclusions: This study establishes the causal atlas of plasma proteins in irAE pathogenesis, bridging biomarker discovery with actionable therapeutic targets. These advances align with next-generation immunotherapy goals: maximizing efficacy while taming the immune storm. Full article

Sperm freezability exhibits marked individual variability, yet the mechanisms remain unclear. Using bulls as the experimental model, we integrated proteomic (sperm) and metabolomic (seminal plasma) analyses of high-freezability (HF) and control (CF) bulls to identify key biomarkers associated with sperm freezability. Post-thaw motility and membrane integrity were significantly higher in HF bulls (p < 0.05). Sperm proteome analysis revealed upregulated antioxidant proteins (PRDX2, GSTM4), heat shock proteins (HSP70, HSP90), and key enzymes in arginine and proline metabolism (PRODH, LAP3). Seminal plasma metabolomics revealed elevated spermine in HF bulls. Meanwhile, we found that spermine abundance was positively correlated with post-thaw motility, as well as with the expression levels of both PRODH and LAP3 (r > 0.6, p < 0.05). Functional validation demonstrated that 200 μM spermine supplementation in cryopreservation extenders enhanced post-thaw motility, kinematic parameters (VAP, VSL, VCL), membrane integrity, and acrosome integrity (p < 0.05). Concurrently, spermine enhanced antioxidant enzyme (SOD, CAT, GSH-Px) activity and reduced ROS and MDA levels (p < 0.05). Our study reveals a spermine-driven antioxidant network coordinating sperm–seminal plasma synergy during cryopreservation, offering novel strategies for semen freezing optimization. Full article

Background/Objectives: Cell–cell communication (CCC) is a critical process within the tumor microenvironment, governing regulatory interactions between cancer cells and other cellular subpopulations. Aiming to improve the accuracy and completeness of intercellular gene-regulatory network inference, we constructed a novel spatial-resolved gene-regulatory network framework (spGRN). Methods: Firstly, the spatial multi-omics data of colorectal cancer (CRC) patients were analyzed. We precisely located the tumor boundaries and then systematically constructed the spGRN framework to study the network regulation. Subsequently, the key signaling molecules obtained by the spGRN were identified and further validated by the spatial-proteomics dataset. Results: Through the constructed spatial gene regulatory network, we found that in the communication with malignant cells, the highly expressed ligands LIF and LGALS3BP and receptors IL6ST and ITGB1 in fibroblasts can promote tumor proliferation, and the highly expressed ligands S100A8/S100A9 in plasma cells play an important role in regulating inflammatory responses. Further, validation of the key signaling molecules by the spatial-proteomics dataset highlighted the role of these genes in mediating the regulation of boundary-related cells. Furthermore, we applied the spGRN to publicly available single-cell and spatial-transcriptomics datasets from three other cancer types. The results demonstrate that ITGB1 and its target genes FOS/JUN were commonly expressed in all four cancer types, indicating their potential as pan-cancer therapeutic targets. Conclusion: the spGRN was proven to be a useful tool to select signal molecules as potential biomarkers or valuable therapeutic targets. Full article

Background: The reproductive ability of equine species is a critical component of equine breeding programs, with sperm quality serving as a primary determinant of reproductive success. In this study, we perform an integrative analysis of proteomics and metabolomics in seminal plasma to identify proteins and metabolites associated with sperm quality and reproductive ability in equine species. Methods: We utilized the CEROS instrument to assess the morphology and motility of sperm samples from three horses and three donkeys. Additionally, we statistically analyzed the mating frequency and pregnancy rates in both species. Meanwhile, the 4D-DIA high-throughput proteomic and metabolomic profiling of seminal plasma samples from horses and donkeys revealed a complex landscape of proteins and metabolites. Results: Our findings reveal a certain degree of correlation between seminal plasma proteins and metabolites and sperm quality, as well as overall fertility. Notably, we found that the proteins B3GAT3, XYLT2, CHST14, HS2ST1, GLCE, and HSPG2 in the glycosaminoglycan biosynthesis signaling pathway; the metabolites D-glucose, 4-phosphopantetheine, and 4-hydroxyphenylpyruvic acid in the tyrosine metabolism, starch, and source metabolisms; and pantothenate CoA biosynthesis metabolism present unique characteristics in the seminal plasma of equine species. Conclusions: This comprehensive approach provides new insights into the molecular mechanisms underlying sperm quality and has identified potential proteins and metabolites that could be used to indicate reproduction ability. The findings from this study could be instrumental in developing novel strategies to enhance equine breeding practices and reproductive management. Future research will focus on exploring their potential for clinical application in the equine industry. Full article

Background: Rheumatoid arthritis (RA) is a chronic autoimmune disorder that predominantly affects synovial joints, leading to inflammation, pain, and progressive joint damage. Despite therapeutic advancements, the molecular basis of established RA remains poorly defined. Methods: In this study, we conducted an untargeted plasma proteomic analysis using two-dimensional differential gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) in samples from RA patients and healthy controls in the discovery phase. Results: Significantly (ANOVA, p ≤ 0.05, fold change > 1.5) differentially abundant proteins (DAPs) were identified. Notably, upregulated proteins included mitochondrial dicarboxylate carrier, hemopexin, and 28S ribosomal protein S18c, while CCDC124, osteocalcin, apolipoproteins A-I and A-IV, and haptoglobin were downregulated. Receiver operating characteristic (ROC) analysis identified CCDC124, osteocalcin, and metallothionein-2 with high diagnostic potential (AUC = 0.98). Proteins with the highest selected frequency were quantitatively verified by multiple reaction monitoring (MRM) analysis in the validation cohort. Bioinformatic analysis using Ingenuity Pathway Analysis (IPA) revealed the underlying molecular pathways and key interaction networks involved STAT1, TNF, and CD40. These central nodes were associated with immune regulation, cell-to-cell signaling, and hematological system development. Conclusions: Our combined proteomic and bioinformatic approaches underscore the involvement of dysregulated immune pathways in RA pathogenesis and highlight potential diagnostic biomarkers. The utility of these markers needs to be evaluated in further studies and in a larger cohort of patients. Full article

Glioblastoma is a highly aggressive, infiltrative brain tumor of the central nervous system (CNS). Its extensive molecular and biochemical heterogenicity hinders the identification of reliable biomarkers and therapeutic targets, thereby making prognosis and existing therapy ineffective. In recent years, breakthroughs in the use of proteomics on a range of biological samples, such as plasma, cerebrospinal fluid (CSF), tissues, brain cells, and exosomes, represent a potential improvement to GBM investigations. Mass spectrometry-based approaches represent an important technique in the characterization of the tumoral proteome, for the identification of differentially expressed proteins, and for studying altered molecular pathways involved in tumor stages. Proteomics studies advance our knowledge about GBM pathogenesis, the discovery of reliable diagnostic and prognostic markers, and therapeutic approaches, also. In this context, for the effective application of proteomics on GBM, it is mandatory to develop a translational network by integrating hospitals, biobanks, and research institutions into a single network, to enable a collaborative approach across disciplines, thereby enabling rapid translation to clinical application of new proteomic insights. Today, high-quality biobanks play a key role in enabling collaborative, ethically compliant research, supporting the effective application of proteomics in glioblastoma studies and the translation of discoveries into clinical practice. This review explores current trends in proteomics and GBM research, highlighting how leveraging biobank infrastructure and fostering institutional cooperation can drive the development of targeted pilot projects to enhance the impact and effectiveness of glioblastoma research. Full article

Cardiovascular diseases are a leading cause of mortality, driving the search for alternative preventive strategies. This study investigates the antioxidant effects, among others, of a mixture of four bioactive peptides (BPs) derived from dry-cured pork ham on endothelial cells from healthy (C-HUVECs) and gestational diabetes (GD-HUVECs) pregnancies, as well as human plasma, using an integrative omics approach. Human umbilical vein endothelial cells (HUVECs) were treated with 300 μM purified BP, followed by transcriptomic and proteomic analyses. The results revealed significant alterations in mitochondrial gene expression and downregulation of genes associated with inflammation and oxidative stress in healthy HUVECs. Furthermore, BP treatment modulated key signalling pathways, including Ras and MAPK, leading to changes in the phosphorylation of ERK, AKT, and NF-κB, suggesting potential cardioprotective effects. The effects of BP were compared to those of the antioxidant hydroxytyrosol, highlighting their relative efficacy in vascular protection. The proteomic analysis of human plasma demonstrated BP-induced modulation of lipid metabolism, inflammation, and oxidative stress with notable changes in proteins such as APOA1 and MMP-8. These natural compounds demonstrate significant preventive potential in vascular health, highlighting their promise as effective tools for reducing cardiovascular risk before the progression of the pathology. These findings emphasize the importance of integrative omics in understanding the mechanisms behind BP’s effects and suggest promising applications for nutraceuticals aimed at cardiovascular protection. Full article

Sperm cryopreservation is a fundamental reproductive biotechnology, enabling the long-term storage of genetic material and supporting assisted reproduction programs. Despite its widespread application, the process is associated with significant limitations due to the cryo-induced cellular damage that occurs during freezing and thawing. These injuries primarily affect the plasma membrane, nuclear DNA, and motility, thereby compromising the fertilizing potential of spermatozoa. Furthermore, interspecies variability in terms of cryo-sensitivity poses a major challenge to the development of standardized cryopreservation protocols. Recent advances have focused on mitigating cryodamage through the use of various strategies. The inclusion of antioxidants in cryopreservation media has proven effective in reducing oxidative stress, thereby enhancing cellular protection. Similarly, the addition of lipid-based supplements contributes to membrane stabilization, improving post-thaw sperm viability and functionality. Moreover, the application of omics technologies, such as transcriptomics and proteomics, has facilitated a deeper understanding of molecular damage and protective responses, paving the way for the development of tailored, species-specific protocols. These integrated approaches optimize cryopreservation conditions, maximizing post-thaw survival and the fertilizing capacity of sperm. Enhancing cryopreservation techniques not only improves the outcomes of assisted reproductive technologies, but also plays a crucial role in the conservation of genetically valuable livestock species. In conclusion, the integration of biotechnological and molecular tools holds significant promise for overcoming the current limitations and advancing the efficacy of sperm cryopreservation. Full article