Chu Heng, Fan XingLi, Zhang Zhe, Han Lin
Department of Thoracic Surgery, Qingdao Hospital, University of Health and Rehabilitation Sciences (Qingdao Municipal Hospital), Qingdao, Shandong, 266000, China.
Department of Cardiovascular Surgery, Changhai Hospital Affiliated to Naval Medical University, Shanghai 200433, China.
Open Med (Wars). 2023 Aug 31;18(1):20230777. doi: 10.1515/med-2023-0777. eCollection 2023.
Calcific aortic valve disease (CAVD) is an important cause of disease burden among aging populations. Excessive active endoplasmic reticulum stress (ERS) was demonstrated to promote CAVD. The expression level of miR-199a-5p in patients with CAVD was reported to be downregulated. In this article, we aimed to investigate the function and mechanism of miR-199a-5p in CAVD. The expression level of miR-199a-5p and ERS markers was identified in calcific aortic valve samples and osteogenic induction by real-time quantitative polymerase chain reaction (RT-qPCR), immunohistochemistry, and western blotting (WB). Alizarin red staining, RT-qPCR, and WB were used for the verification of the function of miR-199a-5p. The dual luciferase reporter assay and rescue experiment were conducted to illuminate the mechanism of miR-199a-5p. In our study, the expression level of miR-199a-5p was significantly decreased in calcified aortic valves and valve interstitial cells' (VICs) osteogenic induction model, accompanying with the upregulation of ERS markers. Overexpression of miR-199a-5p suppressed the osteogenic differentiation of VICs, while downregulation of miR-199a-5p promoted this function. 78 kDa glucose-regulated protein (GRP78) and activating transcription factor 6 (ATF6), both of which were pivotal modulators in ERS, were potential targets of miR-199a-5p. miR-199a-5p directly targeted GRP78 and ATF6 to modulate osteoblastic differentiation of VICs. miR-199a-5p inhibits osteogenic differentiation of VICs by regulating ERS via targeting GRP78 and ATF6.
钙化性主动脉瓣疾病（CAVD）是老年人群疾病负担的一个重要原因。研究表明，过度的内质网应激（ERS）会促进CAVD的发展。据报道，CAVD患者中miR-199a-5p的表达水平下调。在本文中，我们旨在研究miR-199a-5p在CAVD中的作用及其机制。通过实时定量聚合酶链反应（RT-qPCR）、免疫组织化学和蛋白质免疫印迹法（WB），检测钙化主动脉瓣样本和成骨诱导中的miR-199a-5p及ERS标志物的表达水平。茜素红染色、RT-qPCR和WB用于验证miR-199a-5p的功能。采用双荧光素酶报告基因检测和挽救实验来阐明miR-199a-5p的作用机制。在我们的研究中，钙化主动脉瓣和瓣膜间质细胞（VICs）成骨诱导模型中miR-199a-5p的表达水平显著降低，同时ERS标志物上调。miR-199a-5p过表达抑制了VICs的成骨分化，而miR-199a-5p下调则促进了这一功能。78 kDa葡萄糖调节蛋白（GRP78）和激活转录因子6（ATF6）是ERS中的关键调节因子，均为miR-199a-5p的潜在靶点。miR-199a-5p直接靶向GRP78和ATF6，从而调节VICs的成骨细胞分化。miR-199a-5p通过靶向GRP78和ATF6调节ERS，进而抑制VICs的成骨分化。
