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Hepatitis B virus X mediates podocyte pyroptosis by regulating the ROS/NLRP3 signaling pathway in hepatitis B
Yani Yu1,2, Hui Dong3,2, Jingyi Sun1
1Department of Nephrology, The Affiliated Hospital of Qingdao University, Qingdao 266003, Shandong, China.
Objectives:
This study was designed to investigate whether HBx-induced podocyte injury is related to the nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) 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 33342 and subjected to immunofluorescence staining. Caspase-1 activity and LDH levels were assessed with a Caspase-1 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 NLRP3, apoptosis-associated speck-like protein containing card (ASC), caspase-1, IL-1β, and IL-18 (P<0.05), were up-regulated upon HBx overexpression, and caspase-1 enzyme activity and LDH and Desmin expression were also enhanced (P<0.05). NLRP3 knockdown attenuated the increased expression of pyroptosis-related proteins upon HBx overexpression (P<0.05), which was also achieved by the addition of an ROS inhibitor (P<0.05).
Conclusion:
HBx regulates podocyte pyroptosis in HBV-GN by targeting the NLRP3 inflammasome, and mitochondrial oxidative stress plays an important role in this process.
