摘要： 目的 探讨miR-494-3p对高糖（HG）诱导的小鼠肾足细胞（MPC5）损伤的影响及其对钙调磷酸酶调节蛋白1 （RCAN1）的调控作用。方法 HG诱导MPC5细胞建立细胞损伤模型，随机分为NC组、HG组、HG+anti-miR-NC组、HG+anti-miR-494-3p组、HG+pcDNA组、HG+RCAN1组、HG+anti-miR-494-3p+si-NC组、HG+anti-miR-494-3p+si-RCAN1组。采用实时PCR （qRT-PCR）检测miR-494-3p、RCAN1 mRNA表达量；流式细胞术检测细胞凋亡； ELISA法检测白细胞介素（IL）-1β、肿瘤坏死因子-α （TNF-α）水平；双荧光素酶报告实验检测miR-494-3p mimic对Wt-RCAN1荧光素酶活性的影响； Western blotting检测RCAN1蛋白表达量。结果 与NC组比较，HG组miR-494-3p表达量升高，凋亡率、IL-1β、TNF-α水平升高，RCAN1 mRNA及蛋白水平降低（P < 0.05）；与HG+anti-miR-NC组比较，HG+anti-miR-494-3p组凋亡率、IL-1β、TNF-α水平降低（P < 0.05）； miR-494-3p mimic可降低Wt-RCAN1荧光素酶活性（P < 0.05）；与HG+pcDNA组比较，HG+RCAN1组凋亡率、IL-1β、TNF-α水平降低（P < 0.05）；与HG+anti-miR-NC+si-NC组比较，HG+anti-miR-494-3p+si-RCAN1组凋亡率、IL-1β、TNF-α水平升高（P < 0.05）。结论 沉默miR-494-3p可通过促进RCAN1表达抑制MPC5细胞凋亡、炎症反应，从而减轻HG诱导的MPC5细胞损伤。

miR-494-3p靶向调控RCAN1对高糖诱导的足细胞损伤的影响

郭晓莉¹, 杜燕², 李莎莎², 袁二磊³

1. 西安市第三医院肾脏内科, 西安 710018;
2. 西安医学院第一附属医院肾脏内科, 西安 710077;
3. 陕西双博中医肝肾病医院肾脏内科, 西安 710016

收稿日期:2022-11-08出版日期:2023-08-30发布日期:2023-08-07
通讯作者:杜燕E-mail:evcco57@163.com
作者简介:郭晓莉(1978-),女,主治医师,硕士.
基金资助:陕西省重点研发计划（2022SF-132）


关键词: miR-494-3p, 钙调磷酸酶调节蛋白1, 足细胞, 炎症, 细胞凋亡
Abstract: Objective To investigate the effect of miR-494-3p on high glucose (HG) -induced mouse renal podocyte (MPC5) injury and RCAN1 regulation. Methods A cell damage model was established using HG to induce injury in MPC5 cells,which were subsequently divided into the following groups:NC group,HG group,HG+anti-miR-NC group,HG+anti-miR-494-3p group,HG+pcDNA group,HG+RCAN1 group,HG+ anti-miR-494-3p+si-NC group,and HG+anti-miR-494-3p+si-RCAN1 group. qRT-PCR was performed to determine the mRNA expression levels of miR-494-3p and RCAN1,apoptosis was evaluated using flow cytometry,and the levels of IL-1β and TNF-α were determined by ELISA. Additionally,dual-luciferase reporter assays were conducted to assess the effect of miR-494-3p mimics on Wt-RCAN1 luciferase activity,and western blotting was performed to detect RCAN1,Bcl-2,and Bax protein expression. Results miR-494-3p expression,apoptosis rate,and Bax protein,IL-1β,and TNF-α levels were increased,and RCAN1 mRNA and protein levels were decreased in the HG group compared with the NC group (P < 0.05). Additionally,the apoptosis rate and IL-1β and TNF-α levels were decreased in the HG+anti-miR-494-3p group compared with the HG+anti-miR-NC group (P < 0.05). miR-494-3p mimics were found to reduce Wt-RCAN1 luciferase activity (P < 0.05). The apoptosis rate and IL-1β and TNF-α levels were decreased in the HG+RCAN1 group compared with the HG+pcDNA group (P < 0.05). Moreover,the apoptosis rate and IL-1β and TNF-α levels were increased in the HG+anti-miR-494-3p+si-RCAN1 group compared with the HG+anti-miR-NC+si-NC group (P < 0.05). Conclusion Silencing miR-494-3p inhibited MPC5 cell apoptosis and inflammatory response by promoting RCAN1 expression,thereby reducing HG-induced MPC5 cell damage.
Key words: miR-494-3p, RCAN1, podocyte, inflammation, apoptosis
PDF全文下载地址:

https://journal.cmu.edu.cn/CN/article/downloadArticleFile.do?attachType=PDF&id=3265