Department of Cardiology, Heze Municipal Hospital, No. 2888, Caozhou West Road, Heze, 274000, China.
Department of Internal Medicine, Licun Township Health Center, Heze, 274038, China.
Mol Biotechnol. 2021 Sep;63(9):840-848. doi: 10.1007/s12033-021-00341-8. Epub 2021 Jun 7.
Atherosclerosis (AS) is a dynamic and multi-stage process that involves various cells types, such as vascular smooth muscle cells (VSMCs) and molecules such as microRNAs. In this study, we investigated how miR-338-3p works in the process of AS. To determine how miR-338-3p was expressed in AS, an AS rat model was established and primary rat VSMCs were cultured. Real-time polymerase chain reaction was performed to detect miR-338-3p expression. Markers of different VSMC phenotypes were tested by Western blot. Immunofluorescent staining was employed to observe the morphologic changes of VSMCs transfected with miR-338-3p mimics. A dual luciferase reporter assay system was used to verify that desmin was a target of miR-338-3p. To further identify the role of miR-338-3p in the development of AS, VSMC proliferation and migration were evaluated by EdU incorporation assay, MTT assay, and wound healing assay. miR-338-3p expression was upregulated in the aortic tissues of an AS rat model and in primary rat VSMCs from a later passage. The transfection of miR-338-3p mimics in VSMCs promoted the synthetic cell phenotype. Bioinformatics analysis proposed desmin as a candidate target for miR-338-3p and the dual luciferase reporter assay confirmed in vivo that desmin was a direct target of miR-338-3p. The MTT and EdU incorporation assay revealed increased cell viability when miR-338-3p mimics were transfected. The increased expression of PCNA was a consistent observation, although a positive result was not obtained with respect to VSMC mobility. In AS, miR-338-3p expression was elevated. Elevated miR-338-3p inhibited the expression of desmin, thus promoting the contractile-to-synthetic VSMC phenotypic transition. In addition to morphologic changes, miR-338-3p enhanced the proliferative but not mobile ability of VSMCs. In summary, miR-338-3p promotes the development of AS.
动脉粥样硬化（AS）是一个动态的多阶段过程，涉及多种细胞类型，如血管平滑肌细胞（VSMCs）和分子如 microRNAs。在这项研究中，我们研究了 miR-338-3p 在 AS 中的作用机制。为了确定 miR-338-3p 在 AS 中的表达情况，建立了 AS 大鼠模型并培养了原代大鼠 VSMCs。通过实时聚合酶链反应检测 miR-338-3p 的表达。通过 Western blot 检测不同 VSMC 表型标志物。采用免疫荧光染色观察转染 miR-338-3p 模拟物的 VSMCs 的形态变化。采用双荧光素酶报告基因检测系统验证结蛋白是 miR-338-3p 的靶标。为了进一步确定 miR-338-3p 在 AS 发展中的作用，通过 EdU 掺入实验、MTT 实验和划痕愈合实验评估 VSMC 的增殖和迁移。AS 大鼠模型主动脉组织和传代后期的原代大鼠 VSMCs 中 miR-338-3p 的表达上调。VSMCs 转染 miR-338-3p 模拟物可促进合成细胞表型。生物信息学分析提出结蛋白是 miR-338-3p 的候选靶标，双荧光素酶报告基因检测系统证实结蛋白是 miR-338-3p 的直接靶标。MTT 和 EdU 掺入实验显示转染 miR-338-3p 模拟物可提高细胞活力。PCNA 表达增加是一个一致的观察结果，尽管 VSMC 迁移方面没有得到阳性结果。在 AS 中，miR-338-3p 的表达上调。miR-338-3p 升高抑制结蛋白的表达，从而促进收缩型至合成型 VSMC 表型转变。除了形态变化外，miR-338-3p 还增强了 VSMCs 的增殖能力，但不影响其迁移能力。综上所述，miR-338-3p 促进 AS 的发展。
