-
Frontiers in Physiology 2022We aim to explore the detailed molecular mechanisms of membrane nephropathy (MN) related genes by bioinformatics analysis. Two microarray datasets (GSE108109 and...
We aim to explore the detailed molecular mechanisms of membrane nephropathy (MN) related genes by bioinformatics analysis. Two microarray datasets (GSE108109 and GSE104948) with glomerular gene expression data from 65 MN patients and 9 healthy donors were obtained from the Gene Expression Omnibus (GEO) database. After processing the raw data, DEGs screening was conducted using the LIMMA (linear model for microarray data) package and Gene set enrichment analysis (GSEA) was performed with GSEA software (v. 3.0), followed by gene ontology (GO) enrichment and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment. The protein-protein interaction (PPI) network analysis was carried out to determine the hub genes, by applying the maximal clique centrality (MCC) method, which was visualized by Cytoscape. Finally, utilizing the Nephroseq v5 online platform, we analyzed subgroups associated with hub genes. The findings were further validated by immunohistochemistry (IHC) staining in renal tissues from MN or control patients. A sum of 370 DEGs (188 up-regulated genes, 182 down-regulated genes) and 20 hub genes were ascertained. GO and KEGG enrichment analysis demonstrated that DEGs of MN were preponderantly associated with cell damage and complement cascade-related immune responses. Combined with literature data and hub gene-related MN subset analysis, CTSS, ITGB2, and HCK may play important roles in the pathological process of MN. This study identified novel hub genes in MN using bioinformatics. We found that some hub genes such as CTSS, ITGB2, and HCK might contribute to MN immunopathological process, providing new insights for further study of the molecular mechanisms underlying glomerular injury of MN.
PubMed: 35812314
DOI: 10.3389/fphys.2022.914382 -
BMC Genomics Dec 2023Pulmonary arterial hypertension (PAH) is a devastating chronic cardiopulmonary disease without an effective therapeutic approach. The underlying molecular mechanism of...
BACKGROUND
Pulmonary arterial hypertension (PAH) is a devastating chronic cardiopulmonary disease without an effective therapeutic approach. The underlying molecular mechanism of PAH remains largely unexplored at single-cell resolution.
METHODS
Single-cell RNA sequencing (scRNA-seq) data from the Gene Expression Omnibus (GEO) database (GSE210248) was included and analyzed comprehensively. Additionally, microarray transcriptome data including 15 lung tissue from PAH patients and 11 normal samples (GSE113439) was also obtained. Seurat R package was applied to process scRNA-seq data. Uniform manifold approximation and projection (UMAP) was utilized for dimensionality reduction and cluster identification, and the SingleR package was performed for cell annotation. FindAllMarkers analysis and ClusterProfiler package were applied to identify differentially expressed genes (DEGs) for each cluster in GSE210248 and GSE113439, respectively. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genome (KEGG) were used for functional enrichment analysis of DEGs. Microenvironment Cell Populations counter (MCP counter) was applied to evaluate the immune cell infiltration. STRING was used to construct a protein-protein interaction (PPI) network of DEGs, followed by hub genes selection through Cytoscape software and Veen Diagram.
RESULTS
Nineteen thousand five hundred seventy-six cells from 3 donors and 21,896 cells from 3 PAH patients remained for subsequent analysis after filtration. A total of 42 cell clusters were identified through UMAP and annotated by the SingleR package. 10 cell clusters with the top 10 cell amounts were selected for consequent analysis. Compared with the control group, the proportion of adipocytes and fibroblasts was significantly reduced, while CD8+ T cells and macrophages were notably increased in the PAH group. MCP counter revealed decreased distribution of CD8+ T cells, cytotoxic lymphocytes, and NK cells, as well as increased infiltration of monocytic lineage in PAH lung samples. Among 997 DEGs in GSE113439, module 1 with 68 critical genes was screened out through the MCODE plug-in in Cytoscape software. The top 20 DEGs in each cluster of GSE210248 were filtered out by the Cytohubba plug-in using the MCC method. Eventually, WDR43 and GNL2 were found significantly increased in PAH and identified as the hub genes after overlapping these DEGs from GSE210248 and GSE113439.
CONCLUSION
WDR43 and GNL2 might provide novel insight into revealing the new molecular mechanisms and potential therapeutic targets for PAH.
Topics: Humans; Pulmonary Arterial Hypertension; Transcriptome; Adipocytes; CD8-Positive T-Lymphocytes; Databases, Factual; Computational Biology; Gene Expression Profiling
PubMed: 38110868
DOI: 10.1186/s12864-023-09892-3 -
Clinical Cancer Research : An Official... May 2021Merkel cell carcinoma (MCC) is an aggressive cutaneous neuroendocrine carcinoma that can be divided into two classes: virus-positive (VP) MCC, associated with oncogenic...
PURPOSE
Merkel cell carcinoma (MCC) is an aggressive cutaneous neuroendocrine carcinoma that can be divided into two classes: virus-positive (VP) MCC, associated with oncogenic Merkel cell polyomavirus (MCPyV); and virus-negative (VN) MCC, associated with photodamage.
EXPERIMENTAL DESIGN
We classified 346 MCC tumors from 300 patients for MCPyV using a combination of IHC, ISH, and qPCR assays. In a subset of tumors, we profiled mutation status and expression of cancer-relevant genes. MCPyV and molecular profiling results were correlated with disease-specific outcomes. Potential prognostic biomarkers were further validated by IHC.
RESULTS
A total of 177 tumors were classified as VP-MCC, 151 tumors were VN-MCC, and 17 tumors were indeterminate. MCPyV positivity in primary tumors was associated with longer disease-specific and recurrence-free survival in univariate analysis, and in multivariate analysis incorporating age, sex, immune status, and stage at presentation. Prioritized oncogene or tumor suppressor mutations were frequent in VN-MCC but rare in VP-MCC. mutation developed with recurrence in one VP-MCC case. Importantly, for the first time we find that VP-MCC and VN-MCC display distinct sets of prognostic molecular biomarkers. For VP-MCC, shorter survival was associated with decreased expression of immune markers including granzyme and IDO1. For VN-MCC, shorter survival correlated with high expression of several genes including .
CONCLUSIONS
MCPyV status is an independent prognostic factor for MCC. Features of the tumor genome, transcriptome, and microenvironment may modify prognosis in a manner specific to viral status. MCPyV status has clinicopathologic significance and allows for identification of additional prognostic subgroups.
Topics: Aged; Aged, 80 and over; Biomarkers, Tumor; Carcinoma, Merkel Cell; Cell Transformation, Viral; DNA Copy Number Variations; Disease Susceptibility; Female; Gene Expression Profiling; High-Throughput Nucleotide Sequencing; Humans; Immunohistochemistry; Kaplan-Meier Estimate; Male; Merkel cell polyomavirus; Middle Aged; Mutation; Neoplasm Staging; Oncogenes; Polyomavirus Infections; Prognosis; Tumor Microenvironment
PubMed: 33547200
DOI: 10.1158/1078-0432.CCR-20-0864 -
Frontiers in Immunology 2022Acute rejection (AR) in kidney transplantation is an established risk factor that reduces the survival rate of allografts. Despite standard immunosuppression, molecules...
BACKGROUND
Acute rejection (AR) in kidney transplantation is an established risk factor that reduces the survival rate of allografts. Despite standard immunosuppression, molecules with regulatory control in the immune pathway of AR can be used as important targets for therapeutic operations to prevent rejection.
METHODS
We downloaded the microarray data of 15 AR patients and 37 non-acute rejection (NAR) patients from Gene Expression Omnibus (GEO). Gene network was constructed, and genes were classified into different modules using weighted gene co-expression network analysis (WGCNA). Kyoto Encyclopedia of Genes and Genomes (KEGG) and Cytoscape were applied for the hub genes in the most related module to AR. Different cell types were explored by xCell online database and single-cell RNA sequencing. We also validated the SLAMF8 and TLR4 levels in Raw264.7 and human kidney tissues of TCMR.
RESULTS
A total of 1,561 differentially expressed genes were filtered. WGCNA was constructed, and genes were classified into 12 modules. Among them, the green module was most closely associated with AR. These genes were significantly enriched in 20 pathway terms, such as cytokine-cytokine receptor interaction, chemokine signaling pathway, and other important regulatory processes. Intersection with GS > 0.4, MM > 0.9, the top 10 MCC values and DEGs in the green module, and six hub genes (DOCK2, NCKAP1L, IL2RG, SLAMF8, CD180, and PTPRE) were identified. Their expression levels were all confirmed to be significantly elevated in AR patients in GEO, Nephroseq, and quantitative real-time PCR (qRT-PCR). Single-cell RNA sequencing showed that AR patient had a higher percentage of native T, CD1C+_B DC, NKT, NK, and monocytes in peripheral blood mononuclear cells (PBMCs). Xcell enrichment scores of 20 cell types were significantly different (p<0.01), mostly immune cells, such as B cells, CD4+ Tem, CD8+ T cells, CD8+ Tcm, macrophages, M1, and monocytes. GSEA suggests that highly expressed six hub genes are correlated with allograft rejection, interferon γ response, interferon α response, and inflammatory response. In addition, SLAMF8 is highly expressed in human kidney tissues of TCMR and in M1 phenotype macrophages of Raw264.7 cell line WGCNA accompanied by high expression of TLR4.
CONCLUSION
This study demonstrates six hub genes and functionally enriched pathways related to AR. SLAMF8 is involved in the M1 macrophages TLR4, which contributed to AR process.
Topics: Gene Regulatory Networks; Humans; Kidney Transplantation; Leukocytes, Mononuclear; Macrophages; Membrane Proteins; Signaling Lymphocytic Activation Molecule Family; Toll-Like Receptor 4
PubMed: 35432371
DOI: 10.3389/fimmu.2022.846695 -
Journal of Healthcare Engineering 2022The aim of this study was to compare changes in the metabolite levels of ex-smokers and nonsmokers using a metabolomics approach, accounting for the weight gain in...
The aim of this study was to compare changes in the metabolite levels of ex-smokers and nonsmokers using a metabolomics approach, accounting for the weight gain in ex-smokers. Volunteer ex-smokers and nonsmokers were recruited from two cohorts Shijingshan (174) and Xishan (78), respectively, at a 1 : 1 ratio for age and sex. Nontargeted metabolomics was performed on the volunteers' blood samples using liquid chromatography-mass spectrometry, and multivariate statistical analysis was performed using principal component analysis and orthogonal partial least squares discriminant analysis. Enrichment analysis was used to identify Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways associated with differential metabolites and weighted gene co-expression network analysis and maximal correlation coefficient (MCC) algorithms were used to identify key metabolites. The results revealed no significant differences between the distribution of blood metabolite levels in the ex-smokers and nonsmokers. The biosynthesis of valine, leucine, and isoleucine was determined to be associated with differential metabolites, and five key metabolites were identified. Further analysis revealed differences in weight gain and regained metabolite levels in ex-smokers, and 10 differential metabolites were identified that may be associated with weight gain in ex-smokers. These findings suggest that quitting smoking restores metabolites to almost normal levels and results in weight gain. The identified key metabolites and metabolic pathways may also provide a basis for clinical studies.
Topics: Ex-Smokers; Humans; Metabolic Networks and Pathways; Metabolomics; Non-Smokers; Weight Gain
PubMed: 35469229
DOI: 10.1155/2022/6480749 -
Gerontology 2023Aging, an inevitable physiological process, leads to morphological and histological degenerative changes in the mandibular condylar cartilage (MCC); however, the...
INTRODUCTION
Aging, an inevitable physiological process, leads to morphological and histological degenerative changes in the mandibular condylar cartilage (MCC); however, the molecular mechanism has not yet been elucidated, and little information is available on age-related factors. Therefore, this study was designed to identify age-related factors by investigating the age-related differentially expressed genes (DEGs) and localization of their translated protein expression in the mandibular condyle.
METHODS
Mandibular condyles were collected from 10- and 50-week-old mice. Total RNA was extracted from the samples and then analyzed using cap analysis of gene expression (CAGE) to identify age-related DEGs. Gene ontology (GO) enrichment analysis was performed to determine which biological processes were most affected by aging in terms of gene expression using Metascape. The mandibular condyle samples were processed for histology to investigate morphological changes caused by aging and for immunohistochemistry to localize the protein expression encoded by age-related genes identified with CAGE. Semi-quantitative immunohistochemistry was performed to assess age-related extracellular matrix (ECM) protein levels in the MCC. The histological sections were also used for Alcian blue histochemistry to detect glycosaminoglycans (GAGs).
RESULTS
GO enrichment analysis revealed that the genes related to "extracellular matrix organization," including Acan, Col1a1, Col1a2, Col2a1, Mmp3, Mmp9, and Mmp13, were most differentially expressed in the aged mandibular condyle. Among these seven genes, Mmp3 was upregulated, and the others were downregulated with aging. Histological examination showed the age-related morphological and histological changes in the MCC. Immunohistochemical investigation showed the localization of matrix metalloproteinases (MMPs)-3, -9, and -13 and their substrate proteins, aggrecan, type I collagen, and type II collagen, in the mandibular condyle at 10 and 50 weeks, indicating different localizations between the young and the aged. In the aged MCC, semi-quantitative immunohistochemistry showed a significant decrease in the aggrecan protein level, and Alcian blue histochemistry showed a decrease in GAGs.
CONCLUSION
MMP-3, MMP-9, and MMP-13 contribute to the remodeling of the ECM of the MCC and subchondral bone during aging by degrading ECM proteins at specific times and sites under the regulation of their production and secretion.
Topics: Mice; Animals; Matrix Metalloproteinase 3; Mandibular Condyle; Immunohistochemistry; Aggrecans; Alcian Blue; Gene Expression
PubMed: 37769633
DOI: 10.1159/000533921 -
Chest Nov 2021Although mucus plugging is a well-reported feature of asthma, whether asthma and type 2 inflammation affect mucociliary clearance (MCC) is unknown.
BACKGROUND
Although mucus plugging is a well-reported feature of asthma, whether asthma and type 2 inflammation affect mucociliary clearance (MCC) is unknown.
RESEARCH QUESTION
Does type 2 inflammation influence mucus clearance rates in patients with mild asthma who are not receiving corticosteroids?
STUDY DESIGN AND METHODS
The clearance rates of inhaled radiolabeled particles were compared between patients with mild asthma with low (n = 17) and high (n = 18) levels of T2 inflammation. Fraction exhaled nitric oxide (Feno) was used to prospectively segregate subjects into T2 Lo (Feno < 25 ppb) and T2 Hi (Feno > 35 ppb) cohorts. Bronchial brush samples were collected with fiber-optic bronchoscopy, and quantitative polymerase chain reaction was performed to measure expression of genes associated with T2 asthma. MCC rate comparisons were also made with a historical group of healthy control subjects (HCs, n = 12).
RESULTS
The T2 Lo cohort demonstrated increased MCC when compared with both T2 Hi and historic HCs. MCC within the T2 Hi group varied significantly, with some subjects having low or zero clearance. MCC decreased with increasing expression of several markers of T2 airway inflammation (CCL26, NOS2, and POSTN) and with Feno. MUC5AC and FOXJ1 expression was similar between the T2Lo and T2Hi cohorts.
INTERPRETATION
Increasing T2 inflammation was associated with decreasing MCC. High rates of MCC in T2 Lo subjects may indicate a compensatory mechanism present in mild disease but lost with high levels of inflammation. Future studies are required to better understand mechanisms and whether impairments in MCC in more severe asthma drive worse clinical outcomes.
Topics: Adult; Asthma; Bronchial Provocation Tests; Bronchoscopy; Cell Adhesion Molecules; Chemokine CCL26; Correlation of Data; Cross-Sectional Studies; Female; Gene Expression Profiling; Humans; Inflammation; Male; Mucociliary Clearance; Mucus; Nitric Oxide Synthase Type II; Radiopharmaceuticals; Respiratory Function Tests; Respiratory Tract Absorption; Severity of Illness Index
PubMed: 34029561
DOI: 10.1016/j.chest.2021.05.013 -
Biomedicine & Pharmacotherapy =... Oct 2021MicroRNAs play an important role in health and disease. TGF-β signaling, upregulated by HIV Tat, and in chronic airway diseases and smokers upregulates miR-145-5p to...
BACKGROUND
MicroRNAs play an important role in health and disease. TGF-β signaling, upregulated by HIV Tat, and in chronic airway diseases and smokers upregulates miR-145-5p to suppress cystic fibrosis transmembrane conductance regulator (CFTR). CFTR suppression in chronic airway diseases like Cystic Fibrosis, COPD and smokers has been associated with suppressed MCC and recurrent lung infections and inflammation. This can explain the emergence of recurrent lung infections and inflammation in people living with HIV.
METHODS
Tat-induced aberrant microRNAome was identified by miRNA expression analysis. microRNA mimics and antagomirs were used to validate the identified miRNAs involved in Tat mediated CFTR mRNA suppression. CRISPR-based editing of the miRNA target sites in CFTR 3'UTR was used to determine rescue of CFTR mRNA and function in airway epithelial cell lines and in primary human bronchial epithelial cells exposed to TGF-β and Tat.
FINDINGS
HIV Tat upregulates miR-145-5p and miR-509-3p. The two miRNAs demonstrate co-operative effects in suppressing CFTR. CRISPR-based editing of the miRNA target site preserves CFTR mRNA and function in airway epithelial cells INTERPRETATION: Given the important roles of TGF-β signaling and the multitude of genes regulated by miRNAs, we demonstrate that CRISPR-based gene-specific microRNA antagonism approach can preserve CFTR mRNA and function in the context of HIV Tat and TGF-β signaling without suppressing expression of other genes regulated by miR-145-5p.
Topics: 3' Untranslated Regions; Bronchi; CRISPR-Cas Systems; Cell Line; Cells, Cultured; Cystic Fibrosis Transmembrane Conductance Regulator; Epithelial Cells; Gene Editing; Humans; MicroRNAs; RNA, Messenger; Signal Transduction; Transforming Growth Factor beta; Up-Regulation; tat Gene Products, Human Immunodeficiency Virus
PubMed: 34463266
DOI: 10.1016/j.biopha.2021.112090 -
Organotypic Epithelial Raft Cultures as a Three-Dimensional In Vitro Model of Merkel Cell Carcinoma.Cancers Feb 2022Merkel cell carcinoma (MCC) is a rare type of skin cancer for which an in vitro model is still lacking. MCC tumorigenesis is associated either with the integration of...
Merkel cell carcinoma (MCC) is a rare type of skin cancer for which an in vitro model is still lacking. MCC tumorigenesis is associated either with the integration of Merkel cell polyomavirus into the host genome, or with the accumulation of somatic mutations upon chronic exposure to UV light. Transgenic animals expressing the viral oncoproteins, which are constitutively expressed in virus-related MCC, do not fully recapitulate MCC. Although cell-line-derived xenografts have been established for the two subtypes of MCC, they still present certain limitations. Here, we generated organotypic epithelial raft cultures (OERCs) of MCC by using primary human keratinocytes and both virus-positive and virus-negative MCC cell lines. The primary human keratinocytes and the tumor cells were grown on top of a dermal equivalent. Histological and immunohistochemical examination of the rafts confirmed the growth of MCC cells. Furthermore, gene expression analysis revealed differences in the expression profiles of the distinct tumor cells and the keratinocytes at the transcriptional level. In summary, considering the limited availability of patient samples, OERCs of MCC may constitute a suitable model for evaluating the efficacy and selectivity of new drug candidates against MCC; moreover, they are a potential tool to study the oncogenic mechanisms of this malignancy.
PubMed: 35205840
DOI: 10.3390/cancers14041091 -
Journal of Cell Science Nov 2022The gene mutated in colorectal cancer (MCC) encodes a coiled-coil protein implicated, as its name suggests, in the pathogenesis of hereditary human colon cancer. To...
The gene mutated in colorectal cancer (MCC) encodes a coiled-coil protein implicated, as its name suggests, in the pathogenesis of hereditary human colon cancer. To date, however, the contributions of MCC to intestinal homeostasis and disease remain unclear. Here, we examine the subcellular localization of MCC, both at the mRNA and protein levels, in the adult intestinal epithelium. Our findings reveal that Mcc transcripts are restricted to proliferating crypt cells, including Lgr5+ stem cells, where the Mcc protein is distinctly associated with the centrosome. Upon intestinal cellular differentiation, Mcc is redeployed to the apical domain of polarized villus cells where non-centrosomal microtubule organizing centers (ncMTOCs) are positioned. Using intestinal organoids, we show that the shuttling of the Mcc protein depends on phosphorylation by casein kinases 1δ and ε, which are critical modulators of WNT signaling. Together, our findings support a role for MCC in establishing and maintaining the cellular architecture of the intestinal epithelium as a component of both the centrosome and ncMTOC.
Topics: Humans; Microtubule-Organizing Center; Centrosome; Intestines; Cell Differentiation; Proteins; Intestinal Mucosa
PubMed: 36217793
DOI: 10.1242/jcs.259272