Department of Anesthesiology, The Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, P.R. China.
Zhejiang Provincial Engineering Institute on Eye Diseases, Zhejiang Provincial Key Laboratory of Ophthalmology, Zhejiang Provincial Clinical Research Center for Eye Diseases, Eye Center Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, P.R. China.
Ren Fail. 2023;45(2):2261552. doi: 10.1080/0886022X.2023.2261552. Epub 2023 Oct 2.
Circular RNAs (circRNAs) are promising biomarkers and therapeutic targets for acute kidney injury (AKI). In this study, we investigated the mechanism by which circRNA itchy E3 ubiquitin protein ligase (circ-ITCH) regulates sepsis-induced AKI.
A sepsis-induced AKI mouse model was created using LPS induction and circ-ITCH overexpression. Circ-ITCH levels were confirmed RT-qPCR. Kidney tissue changes were examined through various stains and TUNEL. Enzyme-linked immunosorbent assay (ELISA) gauged oxidative stress and inflammation. Mitochondrial features were studied with electron microscopy. RT-qPCR and western blotting assessed mitochondrial function parameters. Using starBase, binding sites between circ-ITCH and miR-214-3p, as well as miR-214-3p and ABCA1, were predicted. Regulatory connections were proven by dual-luciferase assay, RT-qPCR, and western blotting.
Circ-ITCH expression was downregulated in LPS-induced sepsis mice. Overexpression of circ-ITCH ameliorates indicators of renal function (serum creatinine [SCr], blood urea nitrogen [BUN], neutrophil gelatinase-associated lipocalin [NGAL], and kidney injury molecule-1 [Kim-1]), reduces renal cell apoptosis, mitigates oxidative stress markers (reactive oxygen species [ROS] and malondialdehyde [MDA]), and diminishes inflammatory markers (interleukin [IL]-1β, IL-6, and tumor necrosis factor [TNF-α]). Moreover, circ-ITCH overexpression alleviated mitochondrial damage and dysfunction. Furthermore, circ-ITCH acts as a sponge for miR-214-3p, thereby upregulating ABCA1 expression. In addition, the miR-214-3p inhibitor repressed oxidative stress, inflammation, and mitochondrial dysfunction, which was reversed by circ-ITCH knockdown. Further cellular analysis in HK-2 cells supported these findings, highlighting the protective role of circ-ITCH against sepsis-induced AKI, particularly through the miR-214-3p/ABCA1 axis.
The novel circ-ITCH/miR-214-3p/ABCA1 pathway plays an essential role in the regulation of oxidative stress and mitochondrial dysfunction in sepsis-induced AKI.
环状 RNA（circRNA）是急性肾损伤（AKI）有前途的生物标志物和治疗靶点。在这项研究中，我们研究了环状 RNA 痒 E3 泛素蛋白连接酶（circ-ITCH）调节脓毒症诱导的 AKI 的机制。
使用 LPS 诱导和 circ-ITCH 过表达创建了脓毒症诱导的 AKI 小鼠模型。通过 RT-qPCR 确认 circ-ITCH 水平。通过各种染色和 TUNEL 检查肾组织变化。酶联免疫吸附测定（ELISA）测量氧化应激和炎症。电子显微镜研究线粒体特征。RT-qPCR 和 Western blot 评估线粒体功能参数。使用 starBase 预测 circ-ITCH 与 miR-214-3p 以及 miR-214-3p 与 ABCA1 之间的结合位点。通过双荧光素酶测定、RT-qPCR 和 Western blot 证明调节关系。
LPS 诱导的脓毒症小鼠中 circ-ITCH 的表达下调。circ-ITCH 的过表达改善了肾功能指标（血清肌酐[SCr]、血尿素氮[BUN]、中性粒细胞明胶酶相关脂质运载蛋白[NGAL]和肾损伤分子-1[Kim-1]），减少了肾细胞凋亡，减轻了氧化应激标志物（活性氧[ROS]和丙二醛[MDA]），并减少了炎症标志物（白细胞介素[IL]-1β、IL-6 和肿瘤坏死因子[TNF-α]）。此外，circ-ITCH 的过表达减轻了线粒体损伤和功能障碍。此外，circ-ITCH 作为 miR-214-3p 的海绵，从而上调 ABCA1 的表达。此外，miR-214-3p 抑制剂抑制氧化应激、炎症和线粒体功能障碍，而 circ-ITCH 敲低则逆转了这些作用。在 HK-2 细胞中的进一步细胞分析支持了这些发现，强调了 circ-ITCH 对脓毒症诱导的 AKI 的保护作用，特别是通过 miR-214-3p/ABCA1 轴。
新型 circ-ITCH/miR-214-3p/ABCA1 通路在调节脓毒症诱导的 AKI 中的氧化应激和线粒体功能障碍中发挥重要作用。
