HBx-induced PLA₂R overexpression mediates podocyte pyroptosis through the ROS-NLRP3 signaling pathway

Abstract

Introduction

Hepatitis B virus-associated glomerulonephritis (HBV-GN) is one of the main types of secondary glomerular diseases, and podocyte injury is an important pathogenic mechanism of HBV-GN, participating in the occurrence and development of HBV-GN. However, the specific mechanism of podocyte injury remains to be studied.

Methods

Human renal podocytes cultured in vitro were divided into six groups. The podocyte morphology was observed under a transmission electron microscope, and the expression of M-type phospholipase A₂ receptor (M-PLA₂R) on the podocyte membrane was observed by indirect immunofluorescence staining under a fluorescence microscope. The pyroptosis rate and reactive oxygen species (ROS) of podocytes were assessed by FLICA/PI double staining and flow cytometry. Western blot (WB) and quantitative real-time PCR (qPCR) were used to determine the expression of PLA₂R, nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing card (ASC), caspase-1, IL-1β, and IL-18.

Results

Hepatitis B virus X (HBx) transfected into human renal podocytes in vitro induced the overexpression of PLA₂R. Moreover, the overexpressed PLA₂R combined with secretory phospholipase A₂ group IB (sPLA₂-IB) aggravated podocyte injury and increased the pyroptosis rate. In addition, the expression of ROS, the NLRP3 inflammasome and downstream inflammatory factors was increased. In contrast, after inhibiting the expression of PLA₂R and ROS, podocyte damage was alleviated, and the pyroptosis rate and the expression of genes related to the ROS-NLRP3 signaling pathway were decreased.

Conclusion

HBx-induced PLA₂R overexpression on the podocyte membrane can significantly upregulate the ROS-NLRP3 signaling pathway, thereby mediating podocyte pyroptosis.

Keywords:

• HBx
• M-type phospholipase A₂ receptor
• reactive oxygen species
• nucleotide-binding oligomerization domain-like receptor protein 3
• pyroptosis

Author contributions

Moxuan Feng and Yani Yu designed the experimental idea and wrote the main manuscript. Yueqi Chen and Xiaoqian Yang performed the experiments and collected the relevant data. Baoshuang Li finished the formal analysis and investigation. Wei Jiang prepared Figures 1–5 and approved the final version of the manuscript. All authors reviewed the manuscript.

Disclosure statement

No potential conflict of interest was reported by the author(s).

Data availability statement

All data generated or analyzed during this study are included in this article. Further enquiries can be directed to the corresponding author.

Additional information

Funding

This work was supported by the National Natural Science Foundation of China [grant number NSFC 81870494], the Chinese Society of Nephrology [grant number 20010080800], and the Qingdao Outstanding Health Professional Development Fund [grant number 2020-2022].