College of Veterinary Medicine, Henan Agricultural University, Zhengzhou, 450046, China.
College of Animal Science, Tarim University, Alar, 843300, Xinjiang, China.
Parasit Vectors. 2022 Aug 16;15(1):291. doi: 10.1186/s13071-022-05415-3.
MicroRNAs (miRNAs) are involved in the regulation of both the innate and adaptive immune response to Cryptosporidium parvum infection. We previously reported that C. parvum upregulated miR‑942‑5p expression in HCT‑8 cells via TLR2/TLR4‑NF‑κB signaling. In the present study, the role of miRNA-942-5p in the regulation of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated HCT-8 cell apoptosis induced by C. parvum was investigated.
Quantitative real-time polymerase chain reaction, western blotting, flow cytometry, and immunofluorescence were used for analysis.
Forced expression of miRNA-942-5p resulted in decreased apoptosis and an increased C. parvum burden in HCT-8 cells. The opposite results were observed using the suppressed expression of miRNA-942-5p. The miRNA-942-5p led to the translational suppression of IFI27 gene through targeting the 3'-untranslated region of the IFI27 gene. Moreover, overexpression of the IFI27 gene produced a high apoptotic ratio and low C. parvum burden. In contrast, a low apoptotic ratio and a high C. parvum burden were observed following downregulation of the IFI27 gene. Both miR-942-5p and the IFI27 gene influenced TRAIL and caspase-8 expression induced by C. parvum in HCT-8 cells. Moreover, TRAIL promoted HCT-8 cell apoptosis in a concentration-dependent manner.
These data suggested that C. parvum induced the downregulation of IFI27 via relief of miR-942-5p-mediated translational suppression. IFI27 downregulation was affected the burden of C. parvum by regulating HCT-8 cell apoptosis through TRAIL-dependent pathways. Future studies should determine the mechanisms by which C. parvum infection increases miR-942-5p expression and the role of miR-942-5p in hosts' anti-C. parvum immunity in vivo.
微小 RNA（miRNA）参与了对隐孢子虫感染的先天和适应性免疫反应的调节。我们之前报道过，隐孢子虫通过 TLR2/TLR4-NF-κB 信号通路上调 HCT-8 细胞中 miR-942-5p 的表达。在本研究中，研究了 miRNA-942-5p 在调节隐孢子虫诱导的 HCT-8 细胞肿瘤坏死因子相关凋亡诱导配体（TRAIL）介导的细胞凋亡中的作用。
采用定量实时聚合酶链反应、western blot、流式细胞术和免疫荧光法进行分析。
miR-942-5p 的强制表达导致 HCT-8 细胞中的凋亡减少和隐孢子虫负担增加。使用 miR-942-5p 的抑制表达观察到相反的结果。miR-942-5p 通过靶向 IFI27 基因的 3'-非翻译区导致 IFI27 基因的翻译抑制。此外，IFI27 基因的过表达产生高凋亡率和低隐孢子虫负担。相反，下调 IFI27 基因观察到低凋亡率和高隐孢子虫负担。miR-942-5p 和 IFI27 基因均影响隐孢子虫诱导的 HCT-8 细胞中 TRAIL 和 caspase-8 的表达。此外，TRAIL 以浓度依赖的方式促进 HCT-8 细胞凋亡。
这些数据表明，隐孢子虫通过解除 miR-942-5p 介导的翻译抑制诱导 IFI27 的下调。IFI27 的下调通过 TRAIL 依赖途径调节 HCT-8 细胞凋亡，从而影响隐孢子虫的负担。未来的研究应确定隐孢子虫感染增加 miR-942-5p 表达的机制以及 miR-942-5p 在宿主抗隐孢子虫免疫中的作用。
