Search Results (218)

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

Recurrent pregnancy loss (RPL) is defined as the occurrence of two or more pregnancy losses before 20 weeks of gestation. RPL is a common medical condition among reproductive-age women, with approximately 23 million cases reported annually worldwide. Up to 5% of pregnant women may experience two or more consecutive pregnancy losses. Previous studies have investigated risk factors for RPL, including maternal age, uterine pathology, genetic anomalies, infectious agents, endocrine disorders, thrombophilia, and immune dysfunction. However, RPL is a disease caused by a complex interaction of genetic factors, environmental factors (e.g., diet, lifestyle, and stress), epigenetic factors, and the immune system. In addition, due to the lack of research on genetics research related to RPL, the etiology remains unclear in up to 50% of cases. Platelets play a critical role in pregnancy maintenance. This study examined the associations of platelet receptor and ligand gene variants, including integrin subunit beta 3 (ITGB3) rs2317676 A > G, rs3809865 A > T; fibrinogen gamma chain (FGG) rs1049636 T > C, rs2066865 T > C; glycoprotein 1b subunit alpha (GP1BA) rs2243093 T > C, rs6065 C > T; platelet endothelial cell adhesion molecule 1 (PECAM1) rs2812 C > T; and platelet endothelial aggregation receptor 1 (PEAR1) rs822442 C > A, rs12137505 G > A, with RPL prevalence. In total, 389 RPL patients and 375 healthy controls (all Korean women) were enrolled. Genotyping of each single nucleotide polymorphism was performed using polymerase chain reaction–restriction fragment length polymorphism and the TaqMan genotyping assay. All samples were collected with approval from the Institutional Review Board at Bundang CHA Medical Center. The ITGB3 rs3809865 A > T genotype was strongly associated with RPL prevalence (pregnancy loss [PL] ≥ 2: adjusted odds ratio [AOR] = 2.505, 95% confidence interval [CI] = 1.262–4.969, p = 0.009; PL ≥ 3: AOR = 3.255, 95% CI = 1.551–6.830, p = 0.002; PL ≥ 4: AOR = 3.613, 95% CI = 1.403–9.307, p = 0.008). The FGG rs1049636 T > C polymorphism was associated with a decreased risk in women who had three or more pregnancy losses (PL ≥ 3: AOR = 0.673, 95% CI = 0.460–0.987, p = 0.043; PL ≥ 4: AOR = 0.556, 95% CI = 0.310–0.997, p = 0.049). These findings indicate significant associations of the ITGB3 rs3809865 A > T and FGG rs1049636 T > C polymorphisms with RPL, suggesting that platelet function influences RPL in Korean women. Full article

Inherited epidermolysis bullosa (EB) is a heterogeneous clinical entity that includes over 30 phenotypically and/or genotypically distinct inherited disorders, characterized by mechanical skin fragility and bullae formation. Junctional EB (JEB) is an autosomal recessive disease characterized by an intermediated cleavage level within the skin layers, commonly at the “lamina lucida”. Laryngo-onycho-cutaneous syndrome (LOC) is an extremely rare variant of JEB, characterized by granulation tissue formation in specific body sites (skin, larynx, and nails). Although most cases of JEB are caused by pathogenic variants occurring in the genes encoding for classical components of the lamina lucida, such as laminin 332 (LAMA3, LAMB3, LAMC2), integrin α6β4 (ITGA6, ITGB4), and collagen XVII (COL17A1), other variants have also been described. We report the case of a 4-month-old male infant who presented with recurrent bullous and erosive lesions from the first month of life. At the first dermatological evaluation, the patient was agitated and exhibited hoarse breathing, a clinical sign suggestive of laryngeal involvement. Multiple polygonal skin erosions were observed on the cheeks, along with similar isolated, roundish lesions on the scalp and legs. Notably, nail dystrophy and near-complete anonychia were evident on the left first and fifth toes. Due to the coexistence of skin erosions and nail dystrophy in such a young infant, a congenital bullous disorder was suspected, prompting molecular analysis of all potentially involved genes. In the patient’s DNA, clinical exome sequencing (CES) identified a pathogenic variant, apparently in homozygosity, in the exon 1 of the LAMA3 gene (18q11.2; NM_000227.6): c.47G > A;p.Trp16*. The presence of this variant was confirmed, in heterozygosity, in the genomic DNA of the patient’s mother, while it was absent in the father’s DNA. Subsequently, trio-based SNP array analysis was performed, revealing a paternally derived pathogenic microdeletion encompassing the LAMA3 locus (18q11.2). To our knowledge, this is the first reported case of JEB with a LOC-like phenotype caused by a maternally inherited monoallelic nonsense mutation in LAMA3, unmasked by an almost complete deletion of the paternal allele. The combined use of exome sequencing and SNP array is proving essential for elucidating autosomal recessive diseases with a discordant segregation. This is pivotal for providing accurate genetic counseling to parents regarding future pregnancies. Full article

The discovery of bioactive natural compounds from microbes holds promise for regenerative medicine. In this study, we identified and characterized a steroid-like compound, 3,12-dioxochola-4,6-dien-24-oic acid (DOCDA), from a crude extract of Rhodococcus sp. DOCDA significantly promoted wound healing by enhancing HaCaT cell invasion and migration. It upregulated key growth factors (EGF, VEGF-A, IGF, TGF-β, and HGF), indicating the activation of regenerative signaling. Additionally, DOCDA increased the expression of genes related to focal adhesion and cytoskeletal regulation (ITGB1, ITGA4, FAK, SRC, RHOA, CDC42, RAC1, and paxillin), supporting enhanced cellular motility and remodeling. Notably, DOCDA promoted stem-like properties in HaCaT cells, as shown by increased spheroid formation and elevated levels of the stemness markers ALDH1 and CD44. Target prediction and molecular docking identified the glucocorticoid receptor (GR) as the primary target of DOCDA, with a docking score of −7.7 kcal/mol. Network and pathway enrichment analysis revealed that GR-linked pathways were significantly associated with wound healing, including steroid hormone signaling, inflammation, immune responses, and cell migration. In vivo, the topical application of DOCDA led to over 70% wound closure in mice by day 5. These findings suggest that DOCDA is a steroid-like compound that accelerates wound healing and may serve as a potential agent in regenerative therapy. 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

In sheep production, due to the limitations of breeding conditions, the uteri of ewes are often infected with bacteria, resulting in the failure of embryo implantation or loss, causing huge losses to the sheep industry. Therefore, in this study, by using RT-qPCR, Western blot, and immunofluorescence, we investigated the effects of LPS infusion on the immune microenvironment and endometrial receptivity, which play an important role in the process of embryo implantation in ruminants, during the three critical periods of embryo implantation in sheep. The results showed that LPS infusion at day 12, day 16, and day 20 significantly increased the expression of Th1 cytokines (TNF-α, IL-1β, IL-8, IL-6), while significantly decreasing the expression of Th2 cytokines (IL-4 and IL-10) and disrupting the expression of implantation factors, such as ITGB3, ITGB5, VEGF, and LIF, in the endometrial tissues of sheep. Additionally, the protein expression level of TLR4 and the phosphorylation level of ERK were significantly elevated at day 12, day 16, and day 20 after LPS infusion, suggesting that LPS may impair endometrial receptivity through the TLR4/ERK pathway. Validation was conducted in a receptive model of sEECs using TLR4 and ERK phosphorylation inhibitors. Compared with the LPS group, TLR4 and ERK phosphorylation inhibitors significantly reduced the expression of TLR4 and p-ERK, down-regulated Th1 cytokines, up-regulated Th2 cytokines, and alleviated the disruption of genes for attachment. Treatment with 50 μM PTE can significantly alleviate the abnormal expression of implantation genes caused by LPS, and its mechanism may be related to the regulation of the ERK signaling pathway. Full article

The Tibetan goat (Capra hircus) exhibits remarkable adaptations to high-altitude hypoxia, yet the molecular mechanisms remain unclear. This study integrates RNA-seq, WGCNA, and machine learning to explore gene-environment interactions (G × E) in hypoxia adaptation. Fibroblasts from the Tibetan goat and Yunling goat were cultured under hypoxic (1% O₂) and normoxic (21% O₂) conditions, respectively. This identified 68 breed-specific (G), 100 oxygen-responsive (E), and 620 interaction-driven (I) Differentially Expressed Genes (DEGs). The notably higher number of interaction-driven DEGs compared to other effects highlights transcriptional plasticity. We defined two gene sets: Environmental Stress Genes (n = 632, E ∪ I) and Genetic Adaptation Genes (n = 659, G ∪ I). The former were significantly enriched in pathways related to oxidative stress defense and metabolic adaptation, while the latter showed prominent enrichment in pathways associated with vascular remodeling and transcriptional regulation. CTNNB1 emerged as a key regulatory factor in both gene sets, interacting with CASP3 and MMP2 to form the core of the protein–protein interaction (PPI) network. Machine learning identified MAP3K5, TGFBR2, RSPO1 and ITGB5 as critical genes. WGCNA identified key modules in hypoxia adaptation, where FOXO3, HEXIM1, and PPARD promote the stabilization of HIF-1α and metabolic adaptation through the HIF-1 signaling pathway and glycolysis. These findings underscore the pivotal role of gene–environment interactions in hypoxic adaptation, offering novel perspectives for both livestock breeding programs and biomedical research initiatives. Full article

Calpains, calcium-dependent cytosolic cysteine proteases, perform controlled proteolysis of their substrates for various cellular and physiological activities. In different cancers, missense mutations accumulate in the genes coding for the calpain cleavage sites in various calpain substrates termed as the calpain cleavage-related mutations (CCRMs). However, the impact of such CCRMs on the calpain–substrate interaction is yet to be explored. This study focuses on the interaction of wild-type and mutant β-integrins with calpain-1 and 2 in uterine corpus endometrial carcinoma (UCEC). A total of 48 calpain substrates with 176 CCRMs were retrieved from different datasets and shortlisted on the basis of their involvement in cancer pathways. Finally, three calpain substrates, ITGB1, ITGB3, and ITGB7, were selected to assess the structural changes due to CCRMs. These CCRMs were observed towards the C-terminal of the cytoplasmic domain within the calpain cleavage site. The wild-type and mutant proteins were docked with calpain-1 and 2, followed by molecular simulation. The interaction between mutant substrates and calpains showcased variations compared to their respective wild-type counterparts. This may be attributed to mutations in the calpain cleavage sites, highlighting the importance of the cytoplasmic domain of β-integrins in the interactions with calpains and subsequent cellular signaling. Highlights: 1. Calpain cleavage-related mutations (CCRMs) can alter cellular signaling. 2. CCRMs impact the structure of C-domains of human integrin-β subunits. 3. Altered structure influences the cleavability of human integrin-β subunits by human calpains. 4. Altered cleavability impacts the cell signaling mediated through calpain–integrin-β axis. 5. Presence of CCRMS may influence the progression of uterine corpus endometrial carcinoma (UCEC). Full article

Background: The role of extracellular matrix adhesion components in modulation of the treatment sensitivity of ovarian cancer (OC) cells is not well understood. Methods and Results: Analysis of ovarian cancer TCGA gene expression datasets revealed an inverse correlation between genes involved in cell-cycle progression and extracellular matrix interactions, including laminin-binding receptor integrin β4, a major component of extracellular matrix adhesion. Gene ontology analysis also showed that in patient populations with low ITGB4 expression, cell cycle-related programs were activated, while in populations with high expression of ITGB4, the activation of these cell cycle programs was lower. Suppression of proliferation with the CDK4/6 inhibitor Palbociclib stimulated integrin β4 expression and induced protection against Cisplatin in cells naturally expressing low levels of integrin β4. Additionally, ovarian cancer patient-derived organoids showed reduced Cisplatin sensitivity when pretreated with Palbociclib. Our data also showed that integrin β4 overexpression decreased ovarian cancer cell proliferation and at the same time, attenuated Cisplatin response. Conclusions: In summary, our investigations support the idea that integrin β4, and likely its matrix ligands, play critical roles in the regulation of cellular growth and the chemoresistance of ovarian cancer cells. Full article

Necrotizing enterocolitis (NEC) is an acute intestine dysfunction intestinal disorder characterized by inflammation and cell death, including pyroptosis. Previous studies have implicated pyroptosis, particularly via NLRP3 inflammatory activation, and contribute to the development of NEC. However, the genetic and molecular mechanisms underlying pyroptosis in NEC pathogenesis and sequelae remain unclear. Our study aimed to identify the pyroptosis-related cell populations and genes and explore potential therapeutic targets. Single-cell RNA sequencing (scRNA-seq) data were analyzed to identify the cell populations related to NEC and pyroptosis. Weighted gene correlation network analysis (WGCNA) of bulk RNA-seq was performed to identify gene modules associate with pyroptosis. Cell–cell communication was employed to investigate intercellular signaling networks. Gene Set Enrichment Analysis (GSEA) was conducted to compare the pathways enriched in the high and low TREM1-expressing subgroups. Immunofluorescence staining was performed to detect the TREM1⁺CD163⁺ macrophages in the intestines. PCR and Western blot were performed to detect the expression of mRNA and proteins in the intestine tissues and cells. scRNA-seq analysis revealed increased macrophage abundance in NEC, with one macrophage cluster (cluster 4) exhibiting a markedly elevated pyroptosis score. WGCNA identified a gene module (MEbrown) that positively correlated with pyroptosis. Five genes (TREM1, TLN1, NOTCH2, MPZL1, and ADA) within this module were identified as potential diagnostic markers of pyroptosis. Furthermore, we identified a novel macrophage subpopulation, TREM1⁺CD163⁺, in NEC. Cell–cell communication analysis suggested that TREM1⁺CD163⁺ macrophages interact with other cells primarily through the NAMPT/ITGA5/ITGB1 and CCL3/CCR1 pathways. GSEA revealed a significant association between high TREM1 expression and pathways related to pyroptosis, cell proliferation, and inflammation. In vivo and in vitro experiments confirmed an increase in TREM1⁺CD163⁺ macrophages in NEC-affected intestines. TREM1 inhibition in THP-1 cells significantly reduced the expression of pro-inflammatory cytokines and pyroptosis-related genes and proteins. We identified the TREM1⁺CD163+ macrophage population that plays a crucial role in pyroptosis during NEC progression. Our findings elucidate the biological functions and molecular mechanisms of TREM1, demonstrating its upregulation in vivo and pro-pyroptosis effects in vitro. These insights advance our understanding of the role of pyroptosis in NEC pathogenesis and suggest TREM1 is a potential therapeutic target for NEC. Full article

Migraine is a chronic neurovascular disease with unclear pathophysiological mechanisms. In this study, its pathogenic mechanisms were investigated through bioinformatics analysis of migraine-related pathways and key genes. Female Sprague Dawley rats were divided into control and migraine model groups. The control group received saline, while the migraine model group received nitroglycerin (NTG) to induce migraines over four weeks. Migraine-like behaviors were assessed within two hours following the final NTG injection. Genes of hypothalamus were identified using DESeq2. Gene ontology enrichment and KEGG pathway analyses were conducted, followed by the identification of hub genes based on protein interaction networks by using algorithms such as Closeness, Degree, and Maximum Neighborhood Component. Rats with NTG-induced migraine showed increased head scratching and cage climbing and a reduced sucrose preference. Transcriptome analysis revealed 1564 differentially expressed genes, with 1233 upregulated and 331 downregulated. Pathways linked to inflammation, PI3K–Akt signaling, and cytokine–cytokine receptor interactions were found to have enriched expression of several genes. Further protein interaction network analysis identified nine hub genes: Alb, Tgfb1, Cd4, Ptprc, Itgb1, Icam1, Col1a1, Pxdn, and Itgad. These findings suggest that migraine involves PI3K–Akt signaling and cytokine–cytokine receptor interactions, providing insights into molecular mechanisms and potential therapeutic targets. However, the study was limited by a small sample size and reliance on a single experimental model, which may constrain the clinical applicability of the findings. Full article

Background: Hodgkin lymphoma (HL) is a B-cell-derived malignancy and one of the most frequent types of lymphoma. The tumour cells typically exhibit multiple genomic alterations together with aberrantly activated signalling pathways, driven by paracrine and/or autocrine modes. SPP1 (alias osteopontin) is a cytokine acting as a signalling activator and has been connected with relapse in HL patients. To understand its pathogenic role, here, we investigated the mechanisms and function of deregulated SPP1 in HL. Methods: We screened public patient datasets and cell lines for aberrant SPP1 expression. HL cell lines were stimulated with SPP1 and subjected to siRNA-mediated knockdown. Gene and protein activities were analyzed by RQ-PCR, ELISA, Western blot, and immuno-cytology. Results: SPP1 expression was detected in 8.3% of classic HL patients and in HL cell line SUP-HD1, chosen to serve as an experimental model. The gene encoding SPP1 is located at chromosomal position 4q22 and is genomically amplified in SUP-HD1. Transcription factor binding site analysis revealed TALE and HOX factors as potential regulators. Consistent with this finding, we showed that aberrantly expressed PBX1 and HOXB9 mediate the transcriptional activation of SPP1. RNA-seq data and knockdown experiments indicated that SPP1 signals via integrin ITGB1 in SUP-HD1. Accordingly, SPP1 activated NFkB in addition to MAPK/ERK which in turn mediated the nuclear import of ETS2, activating oncogenic JUNB expression. Conclusions: SPP1 is aberrantly activated in HL cell line SUP-HD1 via genomic copy number gain and by homeodomain transcription factors PBX1 and HOXB9. SPP1-activated NFkB and MAPK merit further investigation as potential therapeutic targets in affected HL patients. Full article

Objectives: The identification of methods to improve the endometrial receptivity (ER) is increasingly of interest. The effect of the electromagnetic field associated with Quantum Molecular Resonance (QMR) on ER was investigated here. Methods: Ishikawa cells were used to evaluate the effects of QMR both on the expression of a group of genes involved in ER, i.e., HOXA10, HOXA11, LIF, ITGB3, and ITGAV, and on cell toxicity. Endometrial samples were obtained from six patients during routine diagnostic procedures, four of which were subsequently used to assess the transcriptional response to QMR through microarray. Results: Compared to unexposed controls, a single exposure of Ishikawa cells to QMR for 20 min was associated with a significant and power-dependent up-regulation of all the selected ER-related genes up to 8 power units (PU). Repeated exposure to QMR, up to three consecutive days, showed a significant up-regulation of all the selected genes at power values of 4 PU, from day two onwards. Negligible cytotoxicity was observed. Gene set enrichment analysis, on microarray data of endometrial biopsies stimulated for three consecutive days at 4 PU, showed a significant enrichment of specific gene sets, related to the proteasome system, the cell adhesion, the glucocorticoid receptor, and cell cycle pathways. Conclusions: Our results suggest a possible favorable impact of QMR on ER. Full article

Integrin β4 (ITGB4) mediates lung endothelial cell (EC) inflammation attenuated by simvastatin, an HMG CoA-reductase inhibitor. The cytoplasmic domain of ITGB4 is predicted to bind kindlin-2. Kindlin-2 expression is mediated by SMURF1, an E3 ubiquitin ligase that promotes kindlin-2 ubiquitination and degradation. We hypothesized that increased kindlin-2 expression via the inhibition of SMURF1 mediates EC inflammatory responses relevant to acute lung injury (ALI). To investigate this, human lung ECs were treated with simvastatin (5 µM, 16 h) prior to the immunoprecipitation of kindlin-2 and Western blotting for ITGB4. Next, ECs were treated with a SMURF1 inhibitor, A01, and increased kindlin-2 expression was confirmed. In assays of barrier function, kindlin-2 was silenced (siRNA) in ECs prior to thrombin and measurements of transendothelial resistance (TER) and FITC-dextran transwell flux. Repeat assessments of barrier function were performed in A01-treated ECs. Finally, mice were pretreated with A01 prior to LPS; bronchoalveolar lavage (BAL) fluid was collected, and their lungs were used for histology. Simvastatin increased ITGB4:kindlin-2 association, while A01 increased kindlin-2 expression. Thrombin-induced EC barrier disruption was both increased after kindlin-2 silencing and decreased by A01. Finally, murine ALI was significantly attenuated by A01. Our findings suggest that the augmentation of kindlin-2 may serve as a novel ALI therapeutic strategy. Full article

The Inner Mongolia cashmere goat is a local breed valued for both its cashmere and meat production. Early growth traits include birth weight and weaning weight. Generally, cashmere goats with higher birth and weaning weights tend to exhibit a higher carcass weight at slaughter. Consequently, exploring the genetic variation associated with these early growth traits is crucial in enhancing the meat production performance of Inner Mongolia cashmere goats. For this study, we performed genome-wide association analysis and haplotype analysis on the early growth traits of 212 Inner Mongolia cashmere goats (72 rams, 140 ewes) to identify molecular markers and haplotypes significantly associated with birth and weaning weights. Through genome-wide association analysis and gene annotation, we identified 21 SNPs and 117 candidate genes linked to early growth traits. Notably, genes such as RUNX1T1, ERBIN, MYO15B, NT5C, GRB2, ITGB4, and GALK emerged as significant factors influencing the early growth of Inner Mongolia cashmere goats. Additionally, nine haplotype blocks related to early growth traits were constructed, resulting in eight haplotype combinations. In conclusion, the haplotype combinations A1A1, C1C1, and D1D1 were found to be beneficial for the genetic improvement of early growth traits in Inner Mongolia cashmere goats. Full article