Hepatitis B virus X mediates podocyte pyroptosis by regulating the ROS/NLRP۳ signaling pathway in hepatitis B virus-associated glomerulonephritis

Objective(s): This study was designed to investigate whether HBx-induced podocyte injury is related to the nucleotide-binding oligomerization domain-like receptor protein ۳ (NLRP۳) inflammasome and the specific mechanism of the oxidative stress pathway in hepatitis B virus-associated glomerulonephritis (HBV-GN).Materials and Methods: The HBx gene was overexpressed in renal podocytes to mimic HBV-GN. Podocyte morphology was observed under a scanning electron microscope. Reactive oxygen species (ROS) generation was detected by dichlorodihydrofluorescein diacetate (DCFH-DA) assay. The podocytes in each group were treated with Hoechst ۳۳۳۴۲ and subjected to immunofluorescence staining. Caspase-۱ activity and LDH levels were assessed with a Caspase-۱ Activity Assay Kit and an LDH ELISA Kit, respectively. The expression of all pyroptosis-related proteins was examined by Western blot analysis.Results: Pyroptosis-related proteins, including NLRP۳, apoptosis-associated speck-like protein containing card (ASC), caspase-۱, IL-۱β, and IL-۱۸ (P<۰.۰۵), were up-regulated upon HBx overexpression, and caspase-۱ enzyme activity and LDH and Desmin expression were also enhanced (P<۰.۰۵). NLRP۳ knockdown attenuated the increased expression of pyroptosis-related proteins upon HBx overexpression (P<۰.۰۵), which was also achieved by the addition of an ROS inhibitor (P<۰.۰۵).Conclusion: HBx regulates podocyte pyroptosis in HBV-GN by targeting the NLRP۳ inflammasome, and mitochondrial oxidative stress plays an important role in this process.

Keywords:

Glomerulonephritis , HBx , NLRP۳ , Pyroptosis , ROS