NLRP3炎性小体在乙型肝炎病毒相关性肾炎足细胞焦亡中的作用
The Role of NLRP3 Inflammasome Activation in Podocyte Pyroptosis in Hepatitis B Virus Associated-Glomerulonephritis
DOI: 10.12677/ACM.2022.1281010, PDF,    国家自然科学基金支持
作者: 孙静宜, 蒋 伟：青岛大学附属医院肾内科，山东 青岛
关键词: 乙型肝炎病毒相关性肾炎；NLRP3炎性小体；足细胞；焦亡；HBV-GN； NLRP3 Inflammasome； Podocyte； Pyroptosis
摘要: 目的：本研究旨在探讨NLRP3炎性小体在乙型肝炎病毒相关性肾炎(Hepatitis B virus associat-ed-Glomerulonephritis, HBV-GN)足细胞焦亡中的作用。方法：通过将携带HBx基因的载体及NLRP3 siRNA载体转染至人肾足细胞中，使足细胞株分为空白对照组、空转染组、HBx转染组和HBx+NLRP3 siRNA转染组后，完成以下实验。通过流式细胞术及Hoechst 33342染色，分别检测各组足细胞焦亡率及细胞核的数量及形态学变化。通过实时荧光定量逆转录聚合酶链式反应(Quantitative real-time reverse transcription polymerase chain reaction, qRT-RCR)、蛋白质免疫印迹(Western blot, WB)以及酶联免疫吸附实验(Enzyme linked immunosorbent assay, ELISA)检测4组足细胞的NLRP3、凋亡相关斑点样蛋白(apoptosis-associated speek-like protein containing CARD, ASC)、caspase-1、白细胞介素(Interleukin, IL)-1β、IL-18的表达。通过ELISA试剂盒检测乳酸脱氢酶(lactate dehydrogenase, LDH)水平。通过免疫荧光染色检测各组足细胞中Nephrin蛋白、Desmin蛋白的表达。结果：HBx过表达使NLRP3炎性小体、Caspase-1、IL-1β、IL-18、LDH、Desmin表达均升高，同时导致Nephrin表达下降，最终引起了足细胞焦亡(P < 0.05)。然而，抑制NLRP3炎症小体的生成可以减轻足细胞焦亡，减少了焦亡相关蛋白的表达(P < 0.05)。结论：NLRP3炎性小体介导的炎症信号通路在HBx过表达引起的足细胞焦亡中发挥了重要作用，NLRP3炎性小体有望成为HBV-GN的潜在治疗靶点之一。
Abstract: Objective: This study was designed to investigate the role of NLRP3 inflammasome in podocyte py-roptosis in Hepatitis B virus associated-Glomerulonephritis (HBV-GN). Methods: The vectors carry-ing HBx gene and NLRP3 siRNA were transfected into human renal podocytes. The podocyte lines were divided into blank control group, empty plasmid group, HBx group and HBx+NLRP3 siRNA group. The podocyte pyroptosis rate and the numerical and morphological changes of nuclei were detected by flow cytometry and Hoechst 33342 staining, respectively. NLRP3, apoptosis associated speek-like protein containing CARD (ASC), caspase-1, IL-1β and IL-18 were detected by RT-PCR, Western blot and ELISA. The level of lactate dehydrogenase (LDH) was detected by ELISA kit. The expressions of nephrin protein and desmin protein in podocytes of each group were detected by immunofluorescence staining. Results: The overexpression of HBx increased the expression of NLRP3, caspase-1, IL-1β, IL-18, LDH and desmin, which led to the decrease of nephrin and finally mediated podocyte pyroptosis (P < 0.05). Moreover, the addition of NLRP3 siRNA decreased podo-cyte pyroptosis as well as the expression of pyrosis-related proteins compared with the HBx group (P < 0.05). Conclusion: NLRP3 inflammasome plays an important role in HBx-induced podocyte py-roptosis, suggesting that NLRP3 inflammasome is a potential therapeutic target for alleviating HBV-GN inflammation.
文章引用：孙静宜, 蒋伟. NLRP3炎性小体在乙型肝炎病毒相关性肾炎足细胞焦亡中的作用[J]. 临床医学进展, 2022, 12(8): 7012-7025. https://doi.org/10.12677/ACM.2022.1281010

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