PURPOSE Clostridioides difficile toxin B (TcdB) plays a critical role in C. difficile infection (CDI), a common and costly healthcare-associated disease. The aim of the current study was to explore the intracellular and potent systemic effects of TcdB on human colon epithelial cells utilizing Gene Expression Omnibus and bioinformatic methods. METHODS Two datasets (GSE63880 and GSE29008) were collected to extract data components of mRNA of TcdB-treated human colon epithelial cells; "limma" package of "R" software was used to screen the differential genes, and "pheatmap" package was applied to construct heat maps for the differential genes; Metascape website was utilized for protein-protein interaction network and Molecular Complex Detection analysis, and Genome Ontology (GO) was used to analyze the selected differential genes. Quantitative real-time PCR (qRT-PCR) and Western blot were performed to validate the expression of hub genes. RESULTS GO terms involved in DNA replication and cell cycle were identified significantly enriched in TcdB-treated human colon epithelial cells. Moreover, the decreased expression of DNA replication-related genes, MCM complex, and CDC45 in C. difficile (TcdA-/TcdB+)-infected Caco-2 cells were validated via qRT-PCR and Western blot assays. CONCLUSIONS In conclusion, the integrated analysis of different gene expression datasets allowed us to identify a set of genes and GO terms underlying the mechanisms of CDI induced by TcdB. It would aid in understanding of the molecular mechanisms underlying TcdB-exposed colon epithelial cells and provide the basis for developing diagnosis biomarkers, treatment, and prevention strategies.

中文翻译：

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基于综合的生物信息学分析鉴定毒素B在艰难梭菌毒力中的作用。

目的艰难梭菌毒素B（TcdB）在艰难梭菌感染（CDI）中起着关键作用，艰难梭菌感染是一种常见且代价高昂的医疗保健相关疾病。本研究的目的是利用基因表达综合技术和生物信息学方法，探索TcdB对人结肠上皮细胞的细胞内和强力全身作用。方法收集两个数据集（GSE63880和GSE29008）以提取经TcdB处理的人结肠上皮细胞mRNA的数据。使用“ R”软件的“ limma”软件包筛选差异基因，并使用“ pheatmap”软件包构建差异基因的热图。利用Metascape网站进行蛋白质-蛋白质相互作用网络和分子复合物检测分析，然后使用Genome Ontology（GO）分析选定的差异基因。进行实时定量PCR（qRT-PCR）和蛋白质印迹以验证毂基因的表达。结果确定了参与DNA复制和细胞周期的GO术语在TcdB处理的人结肠上皮细胞中明显富集。此外，通过qRT-PCR和Western印迹试验验证了感染艰难梭菌（TcdA- / TcdB +）的Caco-2细胞中DNA复制相关基因，MCM复合物和CDC45的表达降低。结论总之，对不同基因表达数据集的综合分析使我们能够鉴定出TcdB诱导CDI机制的一组基因和GO术语。