College of Veterinary Medicine, Shanxi Agricultural University, Taigu, 030801, China; College of Animal Science and Technology, Henan Institute of Science and Technology, Xinxiang, 453003, China.
College of Veterinary Medicine, Shanxi Agricultural University, Taigu, 030801, China.
Vet Parasitol. 2021 Feb;290:109351. doi: 10.1016/j.vetpar.2021.109351. Epub 2021 Jan 7.
The present study aimed to investigate the effects of death receptor adapter proteins, namely, TNF receptor-associated death domain (TRADD) and Fas-associated death domain (FADD) proteins, on Eimeria tenella-induced host cell apoptosis. Gene silencing, culture technique for primary chick embryo cecal epithelial cells, enzyme-linked immunosorbent assay, Hoechst-Annexin V/PI apoptosis staining, fluorescence quantitative PCR, and flow cytometry were used to detect the E. tenella host cell apoptotic rate, RIP1 and FADD protein expression levels, and caspase-8 activity of the TRADD siRNA-treated and FADD siRNA-treated groups. Results showed that the apoptotic rate in the TRADD siRNA group was significantly higher than that in the NC siRNA group at 4 h post-infection with E. tenella (P < 0.05). The RIP1 protein expression level in the TRADD siRNA group was significantly lower than that in the NC siRNA group at 4-24 h (P < 0.05). The FADD expression and apoptotic rates in the TRADD siRNA group were significantly lower than those in the NC siRNA group at 24-120 h (P < 0.05). The caspase-8 activity and apoptotic rates in the FADD siRNA group were significantly lower than those in the NC siRNA group (P < 0.05) at 24-120 h. These findings indicated that E. tenella inhibited the host cell apoptosis through the TRADD-RIP1 pathway at the early developmental stage and promoted host cell apoptosis via the TRADD-FADD-caspase-8 apoptotic pathway at the middle and late developmental stages.
本研究旨在探讨死亡受体衔接蛋白，即 TNF 受体相关死亡域（TRADD）和 Fas 相关死亡域（FADD）蛋白，对柔嫩艾美耳球虫诱导的宿主细胞凋亡的影响。采用基因沉默、原代鸡胚盲肠上皮细胞培养技术、酶联免疫吸附试验、Hoechst-Annexin V/PI 凋亡染色、荧光定量 PCR 和流式细胞术检测 TRADD siRNA 处理组和 FADD siRNA 处理组的柔嫩艾美耳球虫感染后宿主细胞凋亡率、RIP1 和 FADD 蛋白表达水平以及 caspase-8 活性。结果显示，感染柔嫩艾美耳球虫后 4 h，TRADD siRNA 组的凋亡率明显高于 NC siRNA 组（P<0.05）。TRADD siRNA 组的 RIP1 蛋白表达水平在 4-24 h 时明显低于 NC siRNA 组（P<0.05）。TRADD siRNA 组的 FADD 表达和凋亡率在 24-120 h 时明显低于 NC siRNA 组（P<0.05）。FADD siRNA 组的 caspase-8 活性和凋亡率在 24-120 h 时明显低于 NC siRNA 组（P<0.05）。这些发现表明，柔嫩艾美耳球虫在早期发育阶段通过 TRADD-RIP1 途径抑制宿主细胞凋亡，在中晚期发育阶段通过 TRADD-FADD-caspase-8 凋亡途径促进宿主细胞凋亡。
