Xu Ke, Mo Xiaojun, Wang Yijun, Zeng Zhenhua, Xu Ziqiang, Yue Dongyou, Li Guicheng, Li Tao, Liu Junhong, Yuan Jiemin
Department of Critical Care Medicine, First People's Hospital of Chenzhou, Chenzhou, Hunan, China.
Department of Emergency, First People's Hospital of Chenzhou, Chenzhou, Hunan, China.
Histol Histopathol. 2023 Apr;38(4):443-452. doi: 10.14670/HH-18-531. Epub 2022 Oct 6.
Sepsis-induced acute kidney injury (AKI) is known to result from the inflammatory responses. MiRNAs participate in the development of sepsis-induced AKI. Nevertheless, the function of miR-527 in sepsis-induced AKI remains unclear.
Cell viability was evaluated by CCK8 assay, and TUNEL staining was applied to assess cell apoptosis. Pro-inflammatory cytokine (TNF-α, IL-6 and IL-1β) levels were evaluated by ELISA. Meanwhile, the relation among miR-527 and Beclin1 was detected by dual luciferase report assay. Western blot and RT-qPCR were used to examine the protein and mRNA levels, respectively. Furthermore, an in vivo model was constructed to assess the function of miR-527 in sepsis-induced AKI.
MiR-527 downregulation significantly alleviated the symptoms of sepsis-induced AKI in mice. MiR-527 level in HK-2 cells was significantly upregulated by LPS, and downregulation of miR-527 notably reversed LPS-induced inhibition of HK-2 cell viability by inhibiting apoptosis. In addition, LPS greatly increased TNF-α, IL-6 and IL-1β levels in supernatant of HK-2 cells, while miR-527 inhibitor partially restored this phenomenon. Meanwhile, Beclin1 was found to be the downstream mRNA of miR-527, and miR-527 inhibitor notably upregulated the level of LC3. MiR-527 downregulation reversed LPS-induced HK-2 cell injury through suppression of TGF-β pathway.
Downregulation of miR-527 alleviated sepsis-induced AKI via targeting Beclin1. Thus, miR-527 might act as a vital mediator in sepsis-induced AKI.
脓毒症诱导的急性肾损伤（AKI）已知是由炎症反应引起的。微小RNA（miRNA）参与脓毒症诱导的AKI的发展。然而，miR-527在脓毒症诱导的AKI中的作用仍不清楚。
通过CCK8法评估细胞活力，并应用TUNEL染色评估细胞凋亡。通过酶联免疫吸附测定（ELISA）评估促炎细胞因子（TNF-α、IL-6和IL-1β）水平。同时，通过双荧光素酶报告基因测定检测miR-527与Beclin1之间的关系。分别使用蛋白质免疫印迹法（Western blot）和逆转录定量聚合酶链反应（RT-qPCR）检测蛋白质和mRNA水平。此外，构建体内模型以评估miR-527在脓毒症诱导的AKI中的作用。
miR-527下调显著减轻了小鼠脓毒症诱导的AKI症状。脂多糖（LPS）显著上调HK-2细胞中miR-527的水平，而miR-527的下调通过抑制凋亡显著逆转了LPS诱导的HK-2细胞活力抑制。此外，LPS显著增加了HK-2细胞上清液中TNF-α、IL-6和IL-1β的水平，而miR-527抑制剂部分恢复了这一现象。同时，发现Beclin1是miR-527的下游mRNA，miR-527抑制剂显著上调了LC3的水平。miR-527下调通过抑制转化生长因子-β（TGF-β）途径逆转了LPS诱导的HK-2细胞损伤。
miR-527的下调通过靶向Beclin1减轻了脓毒症诱导的AKI。因此，miR-527可能是脓毒症诱导的AKI中的重要介质。
