Department of Biochemistry and Molecular Biology, Shanxi Medical University, Taiyuan, Shanxi Province, P. R. China.
Shanxi Key Laboratory of Birth Defect and Cell Regeneration, Shanxi Medical University, Taiyuan, Shanxi Province, P. R. China.
J Cell Physiol. 2019 Aug;234(8):13452-13463. doi: 10.1002/jcp.28025. Epub 2019 Jan 30.
Cardiac microvascular endothelial cells (CMECs) play a critical role in the physiological regulation of coronary blood flow and its dysfunction is associated with myocardium ischemic injury. This study was performed to clarify the effect of microRNA-128 (miR-128) on the CMEC injury in coronary heart disease (CHD) by binding to insulin receptor substrate 1 (IRS1).
The rat CMECs were cultured by explant culture method and identified by CD31 immunofluorescence assay. CMECs were treated with homocysteine (Hcy), which underwent stress of CHD, followed by treatment of miR-128 mimics/inhibitors or IRS1 siRNA. Expression of miR-128, IRS1, and vascular endothelial growth factor (VEGF) was determined. The viability, apoptosis, migration ability, and tube formation ability of CMECs were evaluated. The superoxide dismutase (SOD), malondialdehyde (MDA), and reactive oxygen species (ROS) of CMECs were evaluated, respectively.
In rat CMECs, miR-128 was poorly expressed and IRS1 was highly expressed. Notably, miR-128 targeted and negatively regulated IRS1. Additionally, the treatment with Hcy in CMECs led to reduced viability, migration ability, tube formation, VEGF expression, SOD activity as well as increased cell apoptosis, MDA and ROS levels. The experimental results demonstrated that miR-128 mimics and IRS1 siRNA in rat CMECs promoted viability, migration ability, tube formation, VEGF expression, SOD activity, while repressing cell apoptosis, MDA and ROS levels. MiR-128 inhibitors could reverse the tendencies.
Collectively, our study provides evidence that miR-128 targeted and negatively regulated IRS1 expression, whereby the functional injury of CMECs induced by Hcy was ameliorated. Furthermore, protection of miR-128 was stimulated by reducing oxidative stress.
心肌微血管内皮细胞（CMECs）在调节冠状动脉血流的生理功能中起着关键作用，其功能障碍与心肌缺血损伤有关。本研究旨在通过与胰岛素受体底物 1（IRS1）结合，阐明 microRNA-128（miR-128）对冠心病（CHD）CMEC 损伤的影响。
采用组织块培养法培养大鼠 CMECs，并通过 CD31 免疫荧光法进行鉴定。用同型半胱氨酸（Hcy）处理 CMECs，使其发生 CHD 应激，然后用 miR-128 模拟物/抑制剂或 IRS1 siRNA 处理。测定 miR-128、IRS1 和血管内皮生长因子（VEGF）的表达。评估 CMEC 的活力、凋亡、迁移和管形成能力。分别评估 CMEC 的超氧化物歧化酶（SOD）、丙二醛（MDA）和活性氧（ROS）水平。
在大鼠 CMECs 中，miR-128 表达水平较低，IRS1 表达水平较高。值得注意的是，miR-128 靶向并负调控 IRS1。此外，在 CMECs 中用 Hcy 处理会导致活力、迁移能力、管形成、VEGF 表达、SOD 活性降低以及细胞凋亡、MDA 和 ROS 水平增加。实验结果表明，miR-128 模拟物和 IRS1 siRNA 在大鼠 CMECs 中促进活力、迁移能力、管形成、VEGF 表达、SOD 活性，同时抑制细胞凋亡、MDA 和 ROS 水平。miR-128 抑制剂可以逆转这些趋势。
综上所述，我们的研究提供了证据表明，miR-128 靶向并负调控 IRS1 的表达，从而减轻 Hcy 诱导的 CMEC 功能损伤。此外，通过降低氧化应激来刺激 miR-128 的保护作用。
