Downregulation of USP46 alleviates hypoxia/reoxygenation-induced pyroptosis in renal tubular epithelial cells by inhibiting NLRP3 deubiquitination
10.12464/j.issn.1674-7445.2025072
- VernacularTitle:下调USP46通过抑制NLRP3去泛素化减轻细胞焦亡改善肾小管上皮细胞缺氧/复氧损伤
- Author:
Shiqi GU
1
;
Haiqiang NI
1
;
Xuan PENG
1
;
Xiaotian CAO
1
;
Nianqiao GONG
1
Author Information
1. Institute of Organ Transplantation, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Key Laboratory of Organ Transplantation of Ministry of Education, Key Laboratory of Organ Transplantation of National Health Commission of China, Key Laboratory of Organ Transplantation of Chinese Academy of Medical Sciences, Wuhan 430030, China.
- Publication Type:OriginalArticle
- Keywords:
Kidney transplantation;
Ischemia-reperfusion injury;
Pyroptosis;
Ubiquitin-specific protease 46;
NOD-like receptor protein 3;
Inflammasome;
Deubiquitination;
Oxidative stress
- From:
Organ Transplantation
2025;16(3):416-424
- CountryChina
- Language:Chinese
-
Abstract:
Objective To investigate the role and mechanism of ubiquitin-specific protease 46 (USP46) in hypoxia/reoxygenation (H/R)-induced pyroptosis of renal tubular epithelial cells. Methods Renal tubular epithelial cells were divided into negative control siRNA group (si-CTL group), USP46 knockdown group (si-USP46 group), negative control siRNA + H/R treatment group (si-CTL+H/R group), and USP46 knockdown + H/R treatment group (si-USP46+H/R group). Flow cytometry was used to detect cell apoptosis in each group. Real-time quantitative polymerase chain reaction (RT-qPCR) was employed to measure the messenger RNA (mRNA) expression of USP46, NOD-like receptor protein 3 (NLRP3), gasdermin D (GSDMD), interleukin (IL)-18, and IL-1β. Western blotting was used to detect the protein expression of USP46, NLRP3, GSDMD, and cleaved cysteinyl aspartate specific proteinase (C-Caspase)-1. The levels of inflammatory factors and lactate dehydrogenase (LDH) in the cell supernatants were detected, and the levels of reactive oxygen species (ROS) and malondialdehyde (MDA) in the cells were detected. Co-immunoprecipitation was used to verify the interaction between USP46 and NLRP3. Results Compared with the si-CTL group, the si-CTL+H/R group exhibited increased cell apoptosis, elevated protein expression of USP46, NLRP3, GSDMD-N and C-Caspase-1, increased mRNA expression of USP46, NLRP3, GSDMD, IL-18 and IL-1β, higher levels of IL-18, IL-1β, TNF-α and LDH, and increased ROS and MDA levels (all P < 0.05). Compared with the si-CTL+H/R group, the si-USP46+H/R group showed decreased cell apoptosis, reduced protein expression of USP46, NLRP3, GSDMD-N and C-Caspase-1, decreased mRNA expression of USP46, GSDMD and IL-18, lower levels of IL-18, IL-1β, TNF-α and LDH, and decreased ROS and MDA levels (all P < 0.05). Co-immunoprecipitation results indicated that USP46 could bind to NLRP3. Conclusions Downregulation of USP46 alleviates H/R-induced pyroptosis in renal tubular epithelial cells, possibly by inhibiting USP46-dependent NLRP3 deubiquitination and promoting NLRP3 ubiquitination and degradation.
