Giardia duodenalis induces extrinsic pathway of apoptosis in intestinal epithelial cells through activation of TNFR1 and K63 de-ubiquitination of RIP1 in vitro.

Giardia duodenalis is one of main causative agents of diarrhea that affects the health of millions of people on a global scale per year. It has been clear that attachment of G. duodenalis trophozoites to intestinal epithelium cells (IECs) can induce cell death, while the underlying cellular and molecular mechanisms remain to be explored. It was shown in this study that treatment of Caco-2 cells with Giardia trophozoites could result in reduced cell viability. RNA sequencing analysis demonstrated that expressions of many apoptosis-related genes and some deubiquitinase genes displayed marked changes in trophozoite-treated cells. Trophozoites activated the death-signaling receptor TNFR1 on the IEC surface and caspase-3/8 (CASP3/8) signaling pathways in Caco-2 cells. K63 ubiquitination level of RIP1 was reduced upon stimulation with trophozoites, in parallel, the expressions of deubiquitinases CYLD and A20 were increased. The caspase inhibitor Q-VD-OPH could rescue trophozoite-induced cell apoptosis. Likewise, TNFR1, CYLD, and A20 silencing decreased the levels of cleaved CASP3/8 in trophozoite-treated cells and reversed the pro-apoptosis induction effect of trophozoites. These data suggest that Giardia trophozoite stimulation can activate CASP3/8 signaling pathways via activation of TNFR1 and K63 de-ubiquitination of RIP1 caused by up-regulated expressions of CYLD and A20, and promote Caco-2 cell apoptosis. The present study deepens our understanding of the mechanism of interaction between Giardia and IECs.