Antibody-based gene expression profiling of peripheral blood from patients with idiopathic inflammatory myopathy / 中华风湿病学杂志

ABSTRACT

Objective:To elucidate the pathophysiological mechanisms of idiopathic inflammatory myopathy subtypes by analyzing the gene expression profiles of peripheral blood mononuclear cells (PBMCs) from anti-MDA5 antibody-positive and anti-Jo-1 antibody-positive myositis patients.Methods:Gene expression profiling screening and analysis of PBMCs from 12 anti-MDA5 positive, 16 anti-Jo-1 positive myositis patients and 43 healthy controls were performed using Illumina HT-12 v4 expression profiling microarrays. Applying the unpaired t test with Benjamini-Hochberg correction, the genes with the absolute value of fold change (FC) in gene expression signal ≥2 and adjusted P<0.05 were selected as differentially expressed genes. Differential gene sets were subjected to Gene Ontology (GO) functional enrichment analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, with P<0.05 as the threshold for being significantly enriched. Validation of differentially expressed genes by real time-PCR. The Kolmogorov-Smirnov test was used to test the normality of continuous variables. If the distribution was normal and the variance was homogeneous, analysis of variance (one-way ANOVA) was used.If the distribution was not normal, Kruskal-Wallis test was used, and P<0.05 was regarded as statistically significant difference. Results:Analysis of gene expression profiles of PBMCs from patients with positive anti-MDA5 and anti-Jo-1 antibody revealed significant differences in gene expression of PBMCs from patients with the two myositis subtypes. The number of differentially expressed genes that specifically up-regulated in anti-MDA5 antibody positive patients was 407, and the GO functional enrichment analysis was mainly enriched in biological processes such as innate immune response ( P<0.001), response to virus ( P<0.001) and type Ⅰ interferon signaling pathway ( P<0.001), and the KEGG pathway enrichment analysis was mainly enriched in the viral infection-associated pathway ( P<0.001), RIG-Ⅰ like receptor signaling pathway ( P<0.001) and Toll-like receptor signaling pathway ( P=0.002), etc. The 259 differential genes specifically down-regulated in the anti-MDA5 antibody positive group were mainly enriched in biological processes such as immune response ( P=0.006), TGF-β receptor signaling pathway ( P=0.010) and natural killer cell mediated immunity ( P=0.015) in GO functional enrichment analysis. There were 162 differentially expressed genes up-regulated specifically in anti-Jo-1 antibody positive patients, and GO functional enrichment analysis was mainly enriched in biological processes such as nucleosome assembly ( P<0.001), negative regulation of cell growth ( P=0.001), negative regulation of apoptotic process P=0.004), and innate immune response in mucosa ( P=0.012), and the KEGG pathway enrichment analysis mainly enriched in metabolic-related signaling pathways ( P<0.001) and immune-related pathways ( P<0.001), etc. Real-time PCR confirmed that IFIH1 ( P=0.037), ISG15 ( P=0.003), and DDX58 ( P=0.032) in the RIG-Ⅰ-like receptor pathway as well as chemokines MCP-1 ( P=0.003), MCP-2 ( P<0.001), and transcription factor BATF2 ( P=0.002), and inflammatory signaling pathway-associated MYD88 ( P<0.001) were highly expressed in PBMCs from anti-MDA5 antibody-positive myositis patients. Conclusion:The gene expression profile of PBMCs in anti-MDA5 antibody-positive patients suggests that the pathogenesis of patients with anti-MDA 5 antibody positive is closely related to biological processes such as innate immune response, viral infection, and interferon response.