Cryptosporidium parvum upregulates miR-942-5p expression in HCT-8 cells via TLR2/TLR4-NF-κB signaling

Abstract

Background: Micro (mi)RNAs are small noncoding RNA molecules that function in RNA silencing and post-transcriptional regulation of gene expression. This study investigated host miRNA activity in the innate immune response to Cryptosporidium parvum infection. Methods: In vitro infection model adopts HCT-8 human ileocecal adenocarcinoma cells infected with C. parvum. The expression of miR-942-5p was estimated using quantitative real-time polymerase chain reaction (qPCR). The TLRs-NF-κB signaling was confirmed by qPCR, western blotting, TLR4-and TLR2-specific short-interfering (si)RNA, and NF-κB inhibition. Results: HCT-8 cells express all known toll-like receptors (TLRs). Cryptosporidium parvum infection of cultured HCT-8 cells upregulated TLR2 and TLR4, and downstream TLR effectors, including NF-κB and suppressed IκBα (nuclear factor of kappa light polypeptide gene enhancer in B cells inhibitor, alpha). The expression of miR-942-5p was significantly upregulated at 4, 8, 12 and 24 h post-infection, and especially at 8 hpi. The results of TLR4-and TLR2-specific siRNA and NF-κB inhibition showed that upregulation of miR-942-5p was promoted by p65 subunit-dependent TLR2/TLR4-NF-κB pathway signaling. Conclusions: miR-942-5p of HCT-8 cells was significantly upregulated after C. parvum infection, especially at 8 hpi, in response to a p65-dependent TLR2/TLR4-NF-κB signaling. TLR4 appeared to play a dominant role.

Is Cryptosporidium a hijacker able to drive cancer cell proliferation?

“…For instance, the activation of NF-κB signaling, coupled with the induction of B7-H1 by downregulation of miR-513 in Cryptosporidium infected cells may contribute to the apoptotic resistance of infected cells ( Gong et al, 2010 ). Consistently, a genome-wide miRNA expression analysis showed significant modulation of miRNA expression profile of miR-942 in C. parvum gastro-intestinal epithelial cells through the activation of the TLR4/NF-κB pathway ( Zhang et al, 2020a , Zhang et al, 2020b ). On the other hand, as explained above, the expression of chemokines as CCL20 can be downregulated by the infection.…”

“…( Kumar et al, 2018 ), 17. ( Zhang et al, 2020a , Zhang et al, 2020b ), 18. ( Zhou et al, 2009 ), 19.…”

MiR-942-5p targeting the IFI27 gene regulates HCT-8 cell apoptosis via a TRAIL-dependent pathway during the early phase of Cryptosporidium parvum infection

“…To test this hypothesis, we transfected siRNA-IFI27 into HCT-8 cells and infected the cells with C. parvum. The infected cells were treated with various concentrations of recombinant TRAIL (10,25, and 50 ng/ ml). After 6 h, the cell apoptosis ratio was assessed in all cultures.…”

“…Our previous study of miRNA expression in HCT-8 cells infected with C. parvum found that miR-942-5p, miR-181d, miR-3976, miR-18b-3p, miR-34b-5p, and miR-3591-3p may regulate apoptosis during the early phase of infection [ 24 ]. In a more recent study, we showed that C. parvum upregulated miR-942-5p expression in HCT-8 cells via the Toll-like receptor (TLR) 2/TLR4-NF-κB signaling pathway [ 25 ]. The present study investigated the mechanism of miR-942-5p in regulating HCT-8 cell apoptosis triggered by C. parvum infection, namely that the upregulation of miR-942-5p decreased HCT-8 cell apoptosis by targeting the IFI27 gene via the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) pathway and protected C. parvum from being cleared during the early stage of infection.…”

Response to comments on our article (Yin YL et al., Parasit Vectors, 10.1186/s13071-021-04739-w) by Yuqing Wang and colleagues

“…Cryptosporidiosis, caused by Cryptosporidium spp., is an important zoonotic parasitic disease, seriously threatening the health of humans and many animals [ 1 , 2 ]. Considering that the severity of cryptosporidiosis is closely associated with host status, especially immunity, understanding the host response to infection is critical to effectively develop well-directed control strategies against cryptosporidiosis [ 3 ].…”