Inhibition of microRNA-297 alleviates THLE-2 cell injury induced by hypoxia/reoxygenation by inhibiting NLRP3 inflammasome activation via sirtuin 3.

Abstract

It has been acknowledged that microRNAs (miRNAs/miRs) assume a critical role in hypoxia/reoxygenation (H/R) - induced hepatocyte injury. Therefore, cell experiments were performed in this study to investigate the mechanism of miR-297 in H/R-induced hepatocyte injury with the involvement of sirtuin 3 (SIRT3) and NOD-like receptor pyrin domain containing 3 (NLRP3). Initially, transformed human liver epithelial-2 (THLE-2) cells were utilized for H/R challenge. After miR-297 antagomir and NLRP3 adenovirus vector delivery, THLE-2 cell proliferation and apoptosis were measured by MTT, EdU, and TUNEL assays, respectively. Enzyme-linked immunosorbent assay was conducted to evaluate the levels of apoptosis-related indicators (Bax and Bcl-2) and inflammation-related indicators (interleukin 6 (IL-6) and IL-10), Western blot analysis to detect NLRP3, and cleaved caspase-1 expression. The binding relation between miR-297 and SIRT3 was examined using dual-luciferase assay. The results showed that miR-297 antagomir repressed the apoptosis and inflammation induced by H/R treatment in THLE-2 cells. Mechanistically, miR-297 antagomir diminished the extent of IκBα and nuclear factor-kappa B (NF-κB) phosphorylation and NLRP3 activation in H/R-induced THLE-2 cells by targeting SIRT3. Furthermore, NLRP3 overexpression normalized the promoting effects of miR-297 antagomir on proliferation and its inhibitory effects on apoptosis and inflammation in H/R-induced THLE-2 cells. In summary, our results elucidated that miR-297 antagomir repressed H/R-induced THLE-2 cell injury via SIRT3 promotion and NLRP3 inactivation.