Clostridium perfringens (C. perfringens), a toxin-producing enteric pathogen, causes a variety of intestinal infections in humans and animals. C. perfringens beta2 (CPB2) toxin has been considered to be a strong virulence factor for C. perfringens infectious enteric diseases (CPED). Altered levels and functions of microRNA-21-5p (miR-21-5p) have been associated with apoptosis and inflammation response in pathological processes. However, little is known about its functional mechanism in CPED. Here, we found that miR-21-5p expressed in multiple tissues of pig, had a highest level in jejunum, and significantly upregulated in intestinal porcine epithelial cells (IPEC-J2) exposed to CPB2 toxin. Noteworthily, transfection of CPB2-treated IPEC-J2 cells with miR-21-5p mimic increased cell viability and Bcl2 expression, as well as reduced cytotoxicity, apoptosis rates and Bax level. Moreover, overexpression of miR-21-5p significantly suppressed the levels of interleukin (IL)-6, IL-8, TNF-α, IL-1β and nuclear factor-kappa B (NF-κB p65) activity induced by CPB2 toxin, whereas that of the IL-10 was increased in IPEC-J2 cells. On the contrary, transfection of miR-21-5p inhibitor promoted CPB2-induced cell apoptosis and inflammation. Furthermore, we validated that programmed cell death 4 (PDCD4) was strikingly downregulated in CPB2-treated IPEC-J2 cells. PDCD4 exhibited opposing effects to those of miR-21-5p mimic on IPEC-J2 cells, and restoration of PDCD4 expression counteracted the suppressive effect of miR-21-5p on CPB2-induced apoptosis and inflammatory response. Collectively, our findings demonstrated that miR-21-5p was involved in regulating the immune response triggered by CPB2 toxin and contributed to protective effects in CPB2-induced CPED cell model by targeting PDCD4.

产气荚膜梭菌( C. perfringens ) 是一种产生毒素的肠道病原体，可引起人类和动物的多种肠道感染。产气荚膜梭菌β2（CPB2）毒素被认为是产气荚膜梭菌传染性肠道疾病（CPED）的强毒力因子。 microRNA-21-5p (miR-21-5p) 水平和功能的改变与病理过程中的细胞凋亡和炎症反应有关。然而，对其在 CPED 中的作用机制知之甚少。在这里，我们发现miR-21-5p在猪的多个组织中表达，在空肠中水平最高，并且在暴露于CPB2毒素的猪肠上皮细胞（IPEC-J2）中显着上调。值得注意的是，用 miR-21-5p 模拟物转染 CPB2 处理的 IPEC-J2 细胞可增加细胞活力和 Bcl2 表达，并降低细胞毒性、凋亡率和 Bax 水平。此外，miR-21-5p的过表达显着抑制CPB2毒素诱导的白细胞介素（IL）-6 、 IL-8 、 TNF-α 、 IL-1β和核因子-κB（NF-κB p65）活性水平。而 IPEC-J2 细胞中IL-10的含量增加。相反，转染miR-21-5p抑制剂可促进CPB2诱导的细胞凋亡和炎症。此外，我们验证了程序性细胞死亡 4 (PDCD4) 在 CPB2 处理的 IPEC-J2 细胞中显着下调。 PDCD4 对 IPEC-J2 细胞表现出与 miR-21-5p 模拟物相反的作用，PDCD4 表达的恢复抵消了 miR-21-5p 对 CPB2 诱导的细胞凋亡和炎症反应的抑制作用。 总的来说，我们的研究结果表明，miR-21-5p 参与调节 CPB2 毒素触发的免疫反应，并通过靶向PDCD4在 CPB2 诱导的 CPED 细胞模型中发挥保护作用。