Department of Cardiovascular Surgery, General Hospital of Ningxia Medical University, Yinchuan, China.
Eur Rev Med Pharmacol Sci. 2020 Feb;24(4):2070-2076. doi: 10.26355/eurrev_202002_20385.
The aim of this study was to explore the potential effect of miRNA-1297 on myocardial fibrosis (MF) and its underlying mechanism.
MF model was established by cardiac perfusion of Angiotensin II (Ang-II) in mice. The primary myocardial fibroblasts were extracted from MF mice (Ang-II infusion group) and controls (sham group), respectively. The relative levels of miRNA-1297 and ULK1 in the in vivo and in vitro MF models were determined by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). Meanwhile, the protein expressions of fibrosis-related genes in MF mice and primary myocardial fibroblasts were determined by Western Blot. Subsequently, the Dual-Luciferase Reporter Gene Assay was applied to verify the downstream gene of miRNA-1297. In addition, a series of rescue experiments were conducted to elucidate the role of miRNA-1297/ULK1 in regulating MF.
Masson staining showed plenty of micro-vessels around myocardial tissues and significantly increased contents of intercellular collagen in Ang-II infusion group when compared with those in the sham group. Western blot results revealed that the protein expressions of Col1a1 and α-SMA were significantly upregulated in myocardial tissues of MF mice. QRT-PCR data illustrated that miRNA-1297 was remarkably downregulated in MF model. ULK1 was verified as the target gene of miRNA-1297, which was upregulated in the MF model. The overexpression of miRNA-1297 or the knockdown of ULK1 could downregulate the protein levels of Col1a1 and α-SMA in primary myocardial fibroblasts extracted from MF mice. Notably, ULK1 overexpression could reverse the regulatory effect of miRNA-1297 on MF.
MiRNA-1297 suppresses myocardial fibrosis via down-regulating ULK1.
本研究旨在探讨 miRNA-1297 对心肌纤维化（MF）的潜在影响及其作用机制。
通过向小鼠心脏灌注血管紧张素 II（Ang-II）建立 MF 模型。分别从 MF 小鼠（Ang-II 输注组）和对照组（假手术组）提取原代心肌成纤维细胞。通过定量实时聚合酶链反应（qRT-PCR）测定体内和体外 MF 模型中 miRNA-1297 和 ULK1 的相对水平。同时，通过 Western Blot 测定 MF 小鼠和原代心肌成纤维细胞中纤维化相关基因的蛋白表达。随后，应用双荧光素酶报告基因检测法验证 miRNA-1297 的下游基因。此外，进行了一系列的挽救实验，以阐明 miRNA-1297/ULK1 在调节 MF 中的作用。
Masson 染色显示 Ang-II 输注组心肌组织周围有大量微血管，细胞间胶原含量明显增加，与假手术组相比。Western blot 结果显示，MF 小鼠心肌组织中 Col1a1 和α-SMA 的蛋白表达明显上调。qRT-PCR 数据表明，MF 模型中 miRNA-1297 显著下调。ULK1 被验证为 miRNA-1297 的靶基因，在 MF 模型中上调。miRNA-1297 的过表达或 ULK1 的敲低可下调从 MF 小鼠提取的原代心肌成纤维细胞中 Col1a1 和α-SMA 的蛋白水平。值得注意的是，ULK1 的过表达可逆转 miRNA-1297 对 MF 的调节作用。
miRNA-1297 通过下调 ULK1 抑制心肌纤维化。
