👤 Shan Mou

Cutaneous squamous cell carcinoma (cSCC) involves complex immune interactions. This study aimed to identify a T cell-related gene signature to characterize the immune landscape and aid in molecular diagnosis. We integrated single-cell RNA sequencing (scRNA-seq) and five bulk microarray datasets, utilizing an independent RNA-seq cohort for external validation. Feature genes were identified from the intersection of scRNA-seq-defined T cell-related genes (TRGs) and bulk differentially expressed genes using machine learning. A diagnostic nomogram was constructed, and its performance was assessed via ROC curves. In addition, immune infiltration, immunofluorescence staining, drug interactions, and clinical expression (qRT-PCR) were evaluated. Screening yielded 28 T cell-related DEGs enriched in extracellular matrix functions. machine learning selected a core signature: APOE, CYBA, and S100A2. The diagnostic model demonstrated high diagnostic performance in the studied cohorts (AUC > 0.9) across training and external validation cohorts. Clinically, qRT-PCR supported significant upregulation of CYBA and S100A2. APOE exhibited distinct immunomodulatory connectivity, correlating positively with Th17 cells and negatively with Tregs, whereas CYBA and S100A2 were associated with Treg infiltration. Immunofluorescence results revealed significantly elevated levels of S100A2 and Foxp3 in cSCC tissues compared to the control group. Pharmacogenetic analysis highlights the association of these genes, particularly the APOE gene, with drug response. This T cell-associated signature highlights the potential link between molecular diagnosis and immune characterization. Specifically, CYBA and S100A2 are identified as promising diagnostic candidate signatures, while APOE may reflect immunomodulatory heterogeneity. These findings offer insights for developing diagnostic strategies and targeted immunotherapies in cSCC. Show less

Purinergic receptor P2X7 has been considered as a potential new target for detecting and treating high-risk plaque. Nanobodies are the smallest antibody fragments with high antigen binding ability and specificity, which are well-suited for radionuclide imaging. The present study aimed to develop a novel P2X7-targeted nanobody SPECT tracer and to investigate its potential for identification of atherosclerotic plaque (AP). The anti-P2X7 nanobody 1c81 was site-specifically conjugated with [ Show less

Glioma, characterized by its cellular and molecular heterogeneity, presents formidable challenges in treatment strategy and prognostic assessment. The tumor microenvironment (TME) profoundly influences tumor behavior and treatment response, with tumor-associated neutrophils (TANs) playing a complex but understudied role. This study aimed to investigate the heterogeneity and role of TANs in glioma and to develop a prognostic model. Analysis of scRNA-seq data identified cellular subpopulations and differentially expressed neutrophil-related genes (DE-NRGs). Bulk RNA-seq was obtained from four independent datasets. Molecular subtypes of glioma samples were determined by consensus clustering. WGCNA was conducted to elucidate the association between gene modules and subtypes. We developed a risk score model. Expression of selected genes was confirmed using immunohistochemistry (IHC). In vitro experiments were also performed for functional verification, including CCK8, EdU, Transwell, and TUNEL assays. A total of 108 DE-NRGs for TANs were identified based on scRNA-seq data. Two molecular subtypes were characterized, showing significant differences in prognosis and clinical features. Immune-related analyses demonstrated varied immunological characteristics between subtypes. The risk score model was constructed with 7 genes, including AEBP1, CAVIN1, DCTD, DEPP1, DUSP6, FKBP9, and UGCG. It showed significant prognostic value and was validated across three external datasets. The mutation landscape highlighted higher IDH mutation prevalence in low-risk groups. Drug sensitivity analysis indicated TMZ resistance in high-risk groups. In vitro experiments showed that UGCG could promote glioma cell proliferation, migration, and invasion, while decreasing apoptosis. This study explored the heterogeneity of TANs and developed a prognostic model, providing insights for prognostic prediction and guiding personalized treatment strategies in glioma. Declaration of Generative AI in Scientific Writing: The authors declare nonuse of generative AI and AI-assisted technologies in the writing process. Show less

The efficacy of immune checkpoint inhibitors in the treatment of hepatocellular carcinoma (HCC) remains limited, highlighting the need for further investigation into the mechanisms underlying treatment resistance. Accumulating evidence indicates that tumor-associated macrophages (TAM) within the tumor microenvironment demonstrate a key role in immune evasion and treatment resistance. This study explored the role of TAMs in the HCC tumor microenvironment. Our findings reveal that TAMs expressing CX3C motif chemokine receptor 1 (CX3CR1) induced T-cell exhaustion through IL27 secretion in orthotopic models of HCC following treatment with anti-PD1. Moreover, we identified prostaglandin E2 (PGE2), released by immune-attacked tumor cells, as a key regulator of TAM transition to a CX3CR1+ phenotype. To augment the therapeutic response to anti-PD1 therapy, we propose targeting CX3CR1+ TAMs in addition to anti-PD1 therapy. Our study contributes to the understanding of the role of TAMs in cancer immunotherapy and highlights potential clinical implications for HCC treatment. The combination of targeting CX3CR1+ TAMs with anti-PD1 therapy holds promise for enhancing the efficacy of immunotherapeutic interventions in patients with HCC. Show less

Ovarian cancer (OC) is the most fatal gynecologic cancer. The branched-chain First, the expression level of BCKDK in OC cell lines or tissues was determined using tissue microarray- (TMA-) based immunohistochemistry or western blotting. Then, growth curve analysis, anchorage-independent cell transformation assays, wound healing assays, cell migration assays, and tumor xenografts were used to test whether BCKDK could promote cell transformation or metastasis. Finally, the signaling pathways involved in this process were investigated by western blotting or immunoprecipitation. We found that the expression of BCKDK was upregulated in OC tissues and the high expression of BCKDK was correlated with an advanced pathological grade in patients. The ectopic overexpression of BCKDK promoted the proliferation and migration of OC cells, and the knockdown of BCKDK with shRNAs inhibited the proliferation and migration of OC ex vivo and Our results demonstrate that BCKDK promotes OC proliferation and migration by activating the MEK/ERK signaling pathway. Targeting the BCKDK-MEK axis may provide a new therapeutic strategy for treating patients with OC. Show less

Hyperlipidemia is one of the key risk factors causing many chronic diseases, and lowering blood lipid levels can prevent many diseases. In this paper, a hyperlipidemic cell model of oleic acid (OA) induced hepatocellular carcinoma cells (HepG2) was established using polyphenols extracted from Shanxi-aged vinegar (SAVEP). The effects of SAVEP on nuclear damage, mitochondrial membrane potential, apoptosis, cellular lipid deposition, and lipid metabolism protein expression in HepG2 hyperlipidemic cells were examined to investigate the lipid-lowering mechanism of SAVEP at the cellular level. The results showed that SAVEP could reduce the content of TC/TG index, repair the nuclear damage, reduce lipid accumulation and finally decrease the rate of apoptosis by up-regulating the expression of key proteins such as PPARα, LXRα, and ABCA1 in the process of lipid metabolism. PRACTICAL APPLICATIONS: In this thesis, the hypolipidemic activity of polyphenol extracts from Shanxi-aged vinegar was analyzed on the level of HepG2 cells. The hypolipidemic mechanism of oxidative stress, lipid metabolism and inflammatory stress was also elucidated. It provided a theoretical basis for the in-depth understanding of the hypolipidemic health effects of Shanxi-aged vinegar. Show less

Glucose-dependent insulinotropic polypeptide receptor (GIPR) has been identified as a contributor to obesity, and GIPR knockout mice are protected against diet-induced obesity (DIO). Therefore, we developed the anti-GIPR antagonistic monoclonal antibody (mAb) alone and in combination with DPP-4 inhibitor as potential therapeutic strategy for treating obesity and dyslipidemia based on this genetic evidence. Fully neutralized GIPR activity of GIPR-monoclonal antibody (mAb) was assessed by regulating the in vitro production of cAMP in the mouse GIPR stably expressing cells. Chronic efficacies of GIPR-mAb alone and in combination with DPP-4 inhibitor Sitagliptin in diabetic or DIO mice were both investigated. Multiple metabolic parameters including body weight, glucose level, fat mass, lipid metabolism-related indicators as well as H&E staining and immunohistochemical analysis were performed. Role of GIPR in pancreatic cells on regulating fat metabolism was explored in GIPR β-cell knockout mouse model. Chronic treatment of GIPR-mAb improved body weight control, glucose metabolism, and was associated with reduced fat mass, enhanced pancreatic function and exchange ratio of the resting respiratory in diabetic mice. In addition, further study of anti-GIPR mAb combined with Sitagliptin in DIO mice demonstrated significantly improved weight loss compare to the both monomer treatment. Furthermore, we demonstrated important role of GIPR in β-cell in regulating the fat mass and response to antagonistic GIPR-mAb in a conditional GIPR-knockout mouse. Chronic treatment with anti-GIPR mAb alone and combined with DPP-4 inhibitor provide preclinical therapeutic approaches to treat obesity. Show less

Skeletal muscle has a remarkable ability to regenerate owing to its resident stem cells (also called satellite cells, SCs). SCs are normally quiescent; when stimulated by damage, they activate and expand to form new fibers. The mechanisms underlying SC proliferative progression remain poorly understood. Here we show that DHX36, a helicase that unwinds RNA G-quadruplex (rG4) structures, is essential for muscle regeneration by regulating SC expansion. DHX36 (initially named RHAU) is barely expressed at quiescence but is highly induced during SC activation and proliferation. Inducible deletion of Dhx36 in adult SCs causes defective proliferation and muscle regeneration after damage. System-wide mapping in proliferating SCs reveals DHX36 binding predominantly to rG4 structures at various regions of mRNAs, while integrated polysome profiling shows that DHX36 promotes mRNA translation via 5'-untranslated region (UTR) rG4 binding. Furthermore, we demonstrate that DHX36 specifically regulates the translation of Gnai2 mRNA by unwinding its 5' UTR rG4 structures and identify GNAI2 as a downstream effector of DHX36 for SC expansion. Altogether, our findings uncover DHX36 as an indispensable post-transcriptional regulator of SC function and muscle regeneration acting through binding and unwinding rG4 structures at 5' UTR of target mRNAs. Show less

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes the Coronavirus disease 2019 (COVID-19), which was declared a pandemic by the World Health Organization (WHO) in March 2020. The disease has caused more than 5.1 million deaths worldwide. While cells in the respiratory system are frequently the initial target for SARS-CoV-2, clinical studies suggest that COVID-19 can become a multi-organ disease in the most severe cases. Still, the direct affinity of SARS-CoV-2 for cells in other organs such as the kidneys, which are often affected in severe COVID-19, remains poorly understood. In this study, we employed a human induced pluripotent stem (iPS) cell-derived model to investigate the affinity of SARS-CoV-2 for kidney glomerular podocytes. We studied uptake of the live SARS-CoV-2 virus as well as pseudotyped viral particles by human iPS cell derived podocytes using qPCR, western blot, and immunofluorescence. Global gene expression and qPCR analyses revealed that human iPS cell-derived podocytes express many host factor genes (including ACE2, BSG/CD147, PLS3, ACTR3, DOCK7, TMPRSS2, CTSL CD209, and CD33) associated with SARS-CoV-2 binding and viral processing. Infection of podocytes with live SARS-CoV-2 or spike-pseudotyped lentiviral particles revealed viral uptake by the cells at low Multiplicity of Infection (MOI of 0.01) as confirmed by RNA quantification and immunofluorescence studies. Our results also indicate that direct infection of human iPS cell-derived podocytes by SARS-CoV-2 virus can cause cell death and podocyte foot process retraction, a hallmark of podocytopathies and progressive glomerular diseases including collapsing glomerulopathy observed in patients with severe COVID-19 disease. Additionally, antibody blocking experiments identified BSG/CD147 and ACE2 receptors as key mediators of spike binding activity in human iPS cell-derived podocytes. These results show that SARS-CoV-2 can infect kidney glomerular podocytes Many patients with COVID19 disease exhibit multiorgan complications, suggesting that SARS-CoV-2 infection can extend beyond the respiratory system. Acute kidney injury is a common COVID-19 complication contributing to increased morbidity and mortality. Still, SARS-Cov-2 affinity for specialized kidney cells remain less clear. By leveraging our protocol for stem cell differentiation, we show that SARS-CoV-2 can directly infect kidney glomerular podocytes by using multiple Spike-binding proteins including ACE2 and BSG/CD147. Our results also indicate that infection by SARS-CoV-2 virus can cause podocyte cell death and foot process effacement, a hallmark of podocytopathies including collapsing glomerulopathy observed in patients with severe COVID-19 disease. This stem cell-derived model is potentially useful for kidney-specific antiviral drug screening and mechanistic studies of COVID-19 organotropism. Show less

Manipulation of neural stem and progenitor cells (NSPCs) is critical for the successful treatment of spinal cord injury (SCI) by NSPC transplantation, since their differentiation into neurons and oligodendrocytes can be inhibited by factors present in inflamed myelin. In this study, we examined the effects of LINGO-1 on spinal cord-derived NSPC (sp-NSPC) differentiation, the underlying mechanisms of action, and the functional recovery of mice after transplantation of manipulated cells. sp-NSPCs were harvested from female adult C57/BL6 mice after SCI induced with an NYU impactor. These cells were infected with lentiviral vectors containing LINGO-1 shRNA sequence or a scrambled control and transplanted into SCI mice. Tuj-1- and GFAP-positive cells were assessed by immunofluorescence staining. Wnt5a, p-JNK, JNK, and β-catenin expression was determined by Western blot and RT-qPCR. miRNAs were sequenced to detect changes in miRNA expression. Motor function was evaluated 0-35 days post-surgery by means of the Basso Mouse Scale (BMS) and by the rotarod performance test. We discovered that LINGO-1 shRNA increased neuronal differentiation of sp-NSPCs while decreasing astrocyte differentiation. These effects were accompanied by elevated Wnt5a protein expression, but unexpectedly, no changes in Wnt5a mRNA levels. miRNA-sequence analysis demonstrated that miR-15b-3p was a downstream mediator of LINGO-1 which suppressed Wnt5a expression. Transplantation of LINGO-1 shRNA-treated sp-NSPCs into SCI mice promoted neural differentiation, wound compaction, and motor function recovery. LINGO-1 shRNA promotes neural differentiation of sp-NSPCs and Wnt5a expression, probably by downregulating miR-15b-3p. Transplantation of LINGO-1 shRNA-treated NSPCs promotes recovery of motor function after SCI, highlighting its potential as a target for SCI treatment. Show less

Podocytes are terminally differentiated cells with little proliferative capacity. The high expression levels of cell cycle inhibitory proteins, including p21, p27, and p57, play an important role in maintaining the low level of proliferation of mature podocytes. In the present study, we aimed to explore the role of yes-associated protein (YAP) signalling in adriamycin-induced podocyte re-entry into the cell cycle and dedifferentiation. Proliferating cell nuclear antigen (PCNA)-, cyclin-dependent kinase 4 (CDK4)-, and Cyclin D1-positive podocytes were found in mice with adriamycin-induced nephropathy. In vitro, adriamycin administration increased the percentage of cells in S phase and the upregulation of mesenchymal-related marker proteins. CDK4 and cyclin D1 were significantly up-regulated after incubation with adriamycin. Overexpression of YAP in podocytes promoted their entry into the cell cycle; up-regulated cyclin D1, desmin, and snail2 expression and down-regulated Wilms' tumour 1 (WT1) and nephrin production. Recombinant murine FGF-basic induced podocytes to re-enter the cell cycle, inhibited WT1 and nephrin, and increased desmin and snail2 expression. Pretreating podocytes with verteporfin, an inhibitor of YAP/ TEA domain transcription factor (TEAD), decreased the adriamycin-induced overexpression of cyclin D1 and reduced the ratio of S-phase podocytes. This result was further verified by knocking down YAP expression using RNA interference. In conclusion, adriamycin induced podocytes to re-enter the cell cycle via upregulation of CDK4 and cyclin D1 expression, which was at least partly mediated by YAP signalling. Re-entry into the cell cycle induced the over-expression of mesenchymal markers in podocytes. Show less

Non-obstructive azoospermia (NOA), a severe form of male infertility, is often suspected to be linked to currently undefined genetic abnormalities. To explore the genetic basis of this condition, we successfully sequenced ~650 infertility-related genes in 757 NOA patients and 709 fertile males. We evaluated the contributions of rare variants to the etiology of NOA by identifying individual genes showing nominal associations and testing the genetic burden of a given biological process as a whole. We found a significant excess of rare, non-silent variants in genes that are key epigenetic regulators of spermatogenesis, such as BRWD1, DNMT1, DNMT3B, RNF17, UBR2, USP1 and USP26, in NOA patients (P = 5.5 × 10(-7)), corresponding to a carrier frequency of 22.5% of patients and 13.7% of controls (P = 1.4 × 10(-5)). An accumulation of low-frequency variants was also identified in additional epigenetic genes (BRDT and MTHFR). Our study suggested the potential associations of genetic defects in genes that are epigenetic regulators with spermatogenic failure in human. Show less