Center of Organ Transplantation, The Second Xiangya Hospital of Central South University, Changsha, 410011, China.
Department of Nephrology, The Second Xiangya Hospital of Central South University, No 139 Renmin Road, Changsha, 410011, China.
Sci Rep. 2022 Jul 14;12(1):11994. doi: 10.1038/s41598-022-15607-7.
Our previous research demonstrated that NOD-like receptor family CARD domain-containing protein 4 (NLRC4) inflammasome was overexpressed in renal tissues of patients with diabetic nephropathy (DN). This study further investigated the effect of circRNAs-miRNAs interaction on NLRC4 and their potential mechanisms. DN mice models were first established using STZ. Then, pyroptosis related marker expression was detected using qPCR, western blot (WB), and immunohistochemistry analysis. After that, differentially expressed circRNAs, miRNAs, and mRNAs were investigated using next-generation sequencing. Additionally, the function and potential mechanism of circ_0000181 and miR-667-5p on pyroptosis were measured in vitro DN cell model using MTS, WB, and Enzyme-linked immunosorbent assay. There was an apparent elevation of NLRC4, Caspase1, IL-1β, and IL-18 levels in DN mice. The next-generation sequencing results revealed that there were 947 circRNAs and 390 miRNAs significantly different between the DN and sham kidney tissue, of which circ_0000181 and miR-667-5p had potential targeting effects with NLRC4. Dual-luciferase and functional rescue experiments demonstrated that circ_0000181 promoted NLRC4 inflammasome activation via competitive sponge of miR-667-5p, promoted the release of IL-1β and IL-18, and caused pyroptosis. Altogether, circ_0000181 regulates miR-667-5p/NLRC4 axis to promote pyroptosis progression in DN.
我们之前的研究表明，核苷酸结合寡聚化结构域样受体家族 C 型富含半胱氨酸域蛋白 4（NLRC4）炎症小体在糖尿病肾病（DN）患者的肾组织中过度表达。本研究进一步探讨了 circRNAs-miRNAs 相互作用对 NLRC4 的影响及其潜在机制。首先使用 STZ 建立 DN 小鼠模型。然后，通过 qPCR、western blot（WB）和免疫组织化学分析检测细胞焦亡相关标志物的表达。之后，使用下一代测序技术研究差异表达的 circRNAs、miRNAs 和 mRNAs。此外，在体外 DN 细胞模型中使用 MTS、WB 和酶联免疫吸附测定法测量 circ_0000181 和 miR-667-5p 对细胞焦亡的功能和潜在机制。在 DN 小鼠中，NLRC4、Caspase1、IL-1β 和 IL-18 水平明显升高。下一代测序结果显示，DN 肾组织和假手术肾组织之间有 947 个 circRNAs 和 390 个 miRNAs 存在显著差异，其中 circ_0000181 和 miR-667-5p 与 NLRC4 具有潜在的靶向作用。双荧光素酶和功能拯救实验表明，circ_0000181 通过竞争性海绵吸附 miR-667-5p 促进 NLRC4 炎症小体的激活，促进 IL-1β 和 IL-18 的释放，并引起细胞焦亡。总之，circ_0000181 通过调节 miR-667-5p/NLRC4 轴促进 DN 中细胞焦亡的进展。
