Deng Junhui, Tan Wei, Luo Qinglin, Lin Lirong, Zheng Luquan, Yang Jurong
The Third Affiliated Hospital of Chongqing Medical University, Chongqing, China.
Front Physiol. 2021 Apr 29;12:663216. doi: 10.3389/fphys.2021.663216. eCollection 2021.
Acute kidney injury (AKI) is a complication of sepsis. Pyroptosis of gasdermin D (GSDMD)-mediated tubular epithelial cells (TECs) play important roles in pathogenesis of sepsis-associated AKI. Long non-coding RNA (lncRNA) maternally expressed gene 3 (), an imprinted gene involved in tumorigenesis, is implicated in pyroptosis occurring in multiple organs. Herein, we investigated the role and mechanisms of in regulation of TEC pyroptosis in lipopolysaccharide (LPS)-induced AKI. Male C57BL/6 mice and primary human TECs were treated with LPS for 24 h to establish the animal and cell models, respectively, of sepsis-induced AKI. Renal function was assessed by evaluation of serum creatinine and urea levels. Renal tubule injury score was assessed by Periodic acid-Schiff staining. Renal pyroptosis was assessed by evaluating expression of caspase-1, GSDMD, and inflammatory factors IL-1β and IL-18. Cellular pyroptosis was assessed by analyzing the release rate of LDH, expression of IL-1β, IL-18, caspase-1, and GSDMD, and using EtBr and EthD2 staining. expression in renal tissues and cells was detected using RT-qPCR. The molecular mechanisms of in LPS-induced AKI were assessed through bioinformatics analysis, RNA-binding protein immunoprecipitation, dual luciferase reporter gene assays, and a rescue experiment. Pyroptosis was detected in both LPS-induced animal and cell models, and the expression of in these models was significantly up-regulated. -knockdown TECs treated with LPS showed a decreased number of pyroptotic cells, down-regulated secretion of LDH, IL-1β, and IL-18, and decreased expression of GSDMD, compared with those of controls; however, there was no difference in the expression of caspase-1 between knockdown cells and controls. Bioinformatics analysis screened out miR-18a-3P, and further experiments demonstrated that controls GSDMD expression by acting as a ceRNA for miR-18a-3P to promote TECs pyroptosis. Our study demonstrates that lncRNA promoted renal tubular epithelial pyroptosis by regulating the miR-18a-3p/GSDMD pathway in LPS-induced AKI.
急性肾损伤（AKI）是脓毒症的一种并发症。gasdermin D（GSDMD）介导的肾小管上皮细胞（TECs）焦亡在脓毒症相关AKI的发病机制中起重要作用。长链非编码RNA（lncRNA）母系表达基因3（）是一个参与肿瘤发生的印记基因，与多个器官发生的焦亡有关。在此，我们研究了在脂多糖（LPS）诱导的AKI中调控TECs焦亡的作用及机制。分别用LPS处理雄性C57BL/6小鼠和原代人TECs 24小时，以建立脓毒症诱导的AKI动物模型和细胞模型。通过评估血清肌酐和尿素水平来评估肾功能。通过高碘酸-希夫染色评估肾小管损伤评分。通过评估半胱天冬酶-1、GSDMD以及炎性因子IL-1β和IL-18的表达来评估肾脏焦亡。通过分析乳酸脱氢酶（LDH）释放率、IL-1β、IL-18、半胱天冬酶-1和GSDMD的表达，并使用溴化乙锭（EtBr）和乙锭二聚体（EthD2）染色来评估细胞焦亡。使用逆转录定量聚合酶链反应（RT-qPCR）检测肾脏组织和细胞中的表达。通过生物信息学分析、RNA结合蛋白免疫沉淀、双荧光素酶报告基因检测和挽救实验评估在LPS诱导的AKI中的分子机制。在LPS诱导的动物模型和细胞模型中均检测到焦亡，且这些模型中的表达显著上调。与对照组相比，用LPS处理的敲低TECs显示焦亡细胞数量减少、LDH、IL-1β和IL-18分泌下调以及GSDMD表达降低；然而，敲低细胞与对照组之间半胱天冬酶-1的表达没有差异。生物信息学分析筛选出miR-18a-3P，进一步实验表明通过作为miR-18a-3P的竞争性内源RNA（ceRNA）来调控GSDMD表达，从而促进TECs焦亡。我们的研究表明，lncRNA通过在LPS诱导的AKI中调控miR-18a-3p/GSDMD途径促进肾小管上皮细胞焦亡。
