NOD2- and disease-specific gene expression profiles of peripheral blood mononuclear cells from Crohn’s disease patients
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ABSTRACT: Employing microarray assays, a total of 267 genes were identified that were significantly up- or downregulated in PBMCs of WT-NOD2 patients, compared to healthy donors after challenge with vitamin D (+/-D) and/or a combination (+/-LP) of LPS (lipopolysaccharide) and PGN (peptidoglycan) (p < 0.05; threshold: ≥ 2-fold change). For further analysis by real-time PCR, 12 genes with known impact on inflammation and immunity were selected that fulfilled predefined expression criteria. In a larger cohort of patients and controls, a disease-associated expression pattern, with higher transcript levels in vitamin D-treated PBMCs from 5 patients, was observed for three of these genes, CLEC5A (p < 0.030), lysozyme (LYZ; p < 0.047) and TREM1 (p < 0.023). Six genes were found to be expressed in a NOD2- dependent manner (CD101, p < 0.002; CLEC5A, p < 0.020; CXCL5, p < 0.009; IL-24, p < 0.044; ITGB2, p < 0.041; LYZ, p < 0.042). Interestingly, the highest transcript levels were observed in patients with heterozygous NOD2 mutations.
Project description:Methods: This study analyzed lung tissue sequencing data from early-stage COPD patients (GSE47460) and smoke-exposed mice. We employed Weighted Gene Co-Expression Network Analysis (WGCNA) and machine learning to identify potential pathogenic genes. Further analyses included single-cell sequencing from mice and COPD patients to identify gene expression in specific cell subgroups. Cell-chat and pseudo-temporal analyses were conducted, with findings validated in smoke-exposed mice. Additionally, Mendelian randomization (MR) was utilized to verify the relationship between candidate genes and lung function/COPD. Lastly, functional validation was performed in vitro in cell cultures. Results: "Machine learning analysis of 30 differentially expressed genes pinpointed 8 key genes, with CLEC5A identified as a potential pathogenic factor in early COPD. Bioinformatics suggested CLEC5A's role in macrophage-mediated inflammation in COPD. Two-sample Mendelian randomization linked CLEC5A SNPs with FEV1, FEV1/FVC, and emphysema/chronic bronchitis. In vitro, CLEC5A knockdown reduced inflammatory markers in macrophages. Conclusion: Our study identifies CLEC5A as a pivotal gene in early-stage COPD, contributing to its pathogenesis through pro-inflammatory mechanisms. This discovery offers insights for early diagnosis and treatment strategies in COPD, highlighting CLEC5A as a target for further research.
Project description:Recurrent mutations in ASXL1 are found in various hematological malignancies and are associated with poor prognosis. In particular, ASXL1 mutations are frequently found in patients with hematological malignancies associated with myelodysplasia including myelodysplastic syndromes (MDS), and chronic myelomonocytic leukemia. Although loss-of-function ASXL1 mutations promote myeloid transformation, a large subset of ASXL1 mutations is thought to result in stable truncation of ASXL1. Here we demonstrate that C-terminal truncating ASXL1 mutations (ASXL1-MT) inhibit myeloid differentiation and induce MDS-like disease in mice, displaying all the features of human MDS including multi-lineage myelodysplasia, pancytopenia and occasional progression to overt leukemia. Concerning the molecular mechanisms, ASXL1-MT derepressed expression of Hoxa9 and miR-125a through inhibiting PRC2-mediated methylation of H3K27. miR-125a targeted expression of a surface receptor Clec5a, which was found to supports for myeloid differentiation. In addition, HOXA9 expression was high in MDS patients with ASXL1 mutations while Clec5a expression was generally low in MDS patients. Thus, ASXL1-MT induced MDS-like disease in mice via derepression of Hoxa9 and miR-125a, and Clec5a downregulation. Our data provide evidence for a novel axis of MDS pathogenesis (ASXL1 mutations-upregulation of HoxA9 and miR-125a-downregulation of Clec5a) and implicate both ASXL1 mutants and miR-125a as therapeutic targets in MDS.
Project description:Recurrent mutations in ASXL1 are found in various hematological malignancies and are associated with poor prognosis. In particular, ASXL1 mutations are frequently found in patients with hematological malignancies associated with myelodysplasia including myelodysplastic syndromes (MDS), and chronic myelomonocytic leukemia. Although loss-of-function ASXL1 mutations promote myeloid transformation, a large subset of ASXL1 mutations is thought to result in stable truncation of ASXL1. Here we demonstrate that C-terminal truncating ASXL1 mutations (ASXL1-MT) inhibit myeloid differentiation and induce MDS-like disease in mice, displaying all the features of human MDS including multi-lineage myelodysplasia, pancytopenia and occasional progression to overt leukemia. Concerning the molecular mechanisms, ASXL1-MT derepressed expression of Hoxa9 and miR-125a through inhibiting PRC2-mediated methylation of H3K27. miR-125a targeted expression of a surface receptor Clec5a, which was found to supports for myeloid differentiation. In addition, HOXA9 expression was high in MDS patients with ASXL1 mutations while Clec5a expression was generally low in MDS patients. Thus, ASXL1-MT induced MDS-like disease in mice via derepression of Hoxa9 and miR-125a, and Clec5a downregulation. Our data provide evidence for a novel axis of MDS pathogenesis (ASXL1 mutations-upregulation of HoxA9 and miR-125a-downregulation of Clec5a) and implicate both ASXL1 mutants and miR-125a as therapeutic targets in MDS.
Project description:Background: Survivin was described as strongly expressed in human cancer. To date, little is known about the association between Survivin splice variants and the other apoptosis-related genes. In this study, we analyzed the apoptosis gene signature of Survivin and its variant expression in breast cancer. Methods: Expression of the 5 transcripts was measured by RT-PCR in 135 breast carcinomas. Human Apoptosis Gene Arrays were used to screen the genes that could be associated with Survivin variants. Cox survival analysis was analyzed according to the breast cancer patient outcome. Results: Significant associations between Survivin transcript and the apoptotic gene array were found. Interestingly, Survivin-3B variant showed major inverse correlations with the pro-apoptotic gene array. Overexpression of Survivin-3B strongly inhibits 5-fluorouracil/epirubicin/Cyclophosphamide induced-apoptosis in breast tumor cell lines. In breast carcinomas, uni- and multivariate analysis showed patients with high level of survivin-3B expression had a shorter overall (p=0.030 and p=0.042, respectively) and disease-free (p=0.024 and p=0.009) survival. Conclusion: Our data indicate Survivin-3B could have an anti-apoptotic activity and its expression level might be used as a prognostic marker in breast carcinoma. The expression of 96 apoptosis genes and was analyzed in 41 breast carcinomas and 1 pool of normal mammary tissues. In parallel, every survivin isoform expression was quantified by real-time quantitative PCR. Data obtained with the macro-array were correlated with survivin isoform levels detected by RT-PCR.
Project description:Background: Survivin was described as strongly expressed in human cancer. To date, little is known about the association between Survivin splice variants and the other apoptosis-related genes. In this study, we analyzed the apoptosis gene signature of Survivin and its variant expression in breast cancer. Methods: Expression of the 5 transcripts was measured by RT-PCR in 135 breast carcinomas. Human Apoptosis Gene Arrays were used to screen the genes that could be associated with Survivin variants. Cox survival analysis was analyzed according to the breast cancer patient outcome. Results: Significant associations between Survivin transcript and the apoptotic gene array were found. Interestingly, Survivin-3B variant showed major inverse correlations with the pro-apoptotic gene array. Overexpression of Survivin-3B strongly inhibits 5-fluorouracil/epirubicin/Cyclophosphamide induced-apoptosis in breast tumor cell lines. In breast carcinomas, uni- and multivariate analysis showed patients with high level of survivin-3B expression had a shorter overall (p=0.030 and p=0.042, respectively) and disease-free (p=0.024 and p=0.009) survival. Conclusion: Our data indicate Survivin-3B could have an anti-apoptotic activity and its expression level might be used as a prognostic marker in breast carcinoma.
Project description:Profiling of ileal mucosal gene expression in Chron's disease patients with and without NOD2 risk alleles; through microarray analyses it was discovered that 18 genes related to lymphocyte and phagocyte activation recuitment were upregulated in affected ileum in NOD2R patients as compared to ileum in NOD2s (no risk alleles) patients, thus supporting the concept that NOD2 risk alleles contribute to impaired regulation of inflammation in the ileum.
Project description:To explore the differentially expressed (DE) circular (circ)RNAs, competing endogenous RNAs networks, and potential biomarkers in distal cholangiocarcinoma (dCCA), using microarray analysis, a comprehensive circRNA expression profiling was constructed with six paired dCCA tumor and adjacent normal samples. A total of 171 DE circRNAs were identified. Three upregulated (hsa_circ_0060144, hsa_circ_0008274, and hsa_circ_0000673) and two downregulated circRNAs (hsa_circ_0139402 and hsa_circ_0001714) were validated with RT-qPCR in 40 dCCA patients. Among them, hsa_circ_0000673 had an idea prognostic efficiency (AUC=0.85, p<0.01) and high level of hsa_circ_0000673 was associated with late tumor stage (p<0.01), poor histodifferentiation (p=0.041), and residual tumor (p=0.044). Gene ontology analysis of the host genes of 171 DE circRNAs showed that “response to nutrient”, “adherens junction”, and “cell adhesion molecule binding” were mainly enriched in terms of biological process, cellular component, and molecular functions respectively. Nine target micro(mi)RNAs of the five DE circRNAs were screened out and enriched in cancer associated pathways. Thereafter, 188 DE mRNAs were predicted as target genes of the nine miRNAs. Finally, protein-protein interaction networks were constructed and 20 hubgenes were identified. This research provided a landscape of DE circRNAs in dCCA, constructed circRNA-miRNA-mRNA regulating networks, and identified hsa_circ_0000673 as a potential biomarker and target for treating dCCA.
Project description:Crohn’s disease (CD) is a chronic inflammatory intestinal disease, often characterized by aberrant healing and stricturing complications. Mechanisms underlying NOD2-pathogenicity and salvage pathways in anti-TNF and refractory patients remain largely uncharacterized. Here we show that loss of NOD2 function leads to aberrant activated fibroblast and macrophage homeostasis through the upregulation of a pathogenic signature, and propose new precision therapeutic approaches involving gp130 blockade for select CD patients, to potentially supplement anti-TNF therapy
Project description:In this study, we analyse the in-vivo modulation of the transcriptome of human PBMCs by a bolus of vitamin D3 (80,000 IU) after 24 hours.
Project description:Three independent repeats of FAIRE-seq from PBMCs obtained before and 1 and 2 days after an oral dose of 2,000 µg (80,000 IU) of vitamin D3