As a basic member of the Class III histone deacetylases, SIRT1 has been implicated in the occurrence and progression of diabetic retinopathy (DR). The current study aimed to investigate the roles of SIRT1/miR-20a/YAP/HIF1α/VEGFA in DR. The expression of SIRT1 was initially determined through RT-qPCR and western blot analysis following the successful establishment of a DR mouse model, followed by detection of SIRT1 catalytic activity. Retinal microvascular endothelial cells (RMECs) were cultured in media supplemented with normal glucose (NG) or high glucose (HG). Thereafter, SIRT1 was either silenced or overexpressed in RMECs, after which EdU staining and Matrigel-based tube formation assay were performed to assess cell proliferation and tube formation. The binding relationship between YAP, HIF1α, and VEGFA was further illustrated using dual-luciferase reporter assay. Pre-retinal neovascular cell number was tallied with the IB4 positive vascular endothelial cells determined by immunofluorescence. SIRT1 was poorly expressed in mice with DR and HG-treated RMECs with low catalytic activity. The proliferation and tube formation capabilities of RMECs were elevated under HG conditions, which could be reversed following overexpression of SIRT1. SIRT1 was identified to positively regulate the expression of miR-20a with YAP detected as the key target gene of miR-20a. Our data suggested that YAP could upregulate VEGFA via induction of HIF1α. Moreover, SIRT1 overexpression strongly repressed RMEC proliferation and angiogenesis, which could be reversed via restoration of YAP/HIF1α/VEGFA expression. Taken together, the key findings of our study suggest that upregulation of SIRT1 inhibits the development of DR via miR-20a-induced downregulation of YAP/HIF1α/VEGFA.

中文翻译：

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组蛋白脱乙酰基酶SIRT1的过表达通过miR-20a升高和YAP /HIF1α/ VEGFA耗竭在糖尿病性视网膜病中发挥抗血管生成作用。

作为III类组蛋白脱乙酰基酶的基本成员，SIRT1与糖尿病性视网膜病（DR）的发生和发展有关。目前的研究旨在调查SIRT1 / miR-20a / YAP /HIF1α/ VEGFA在DR中的作用。成功建立DR小鼠模型后，首先通过RT-qPCR和Western blot分析确定SIRT1的表达，然后检测SIRT1的催化活性。视网膜微血管内皮细胞（RMEC）在补充了正常葡萄糖（NG）或高葡萄糖（HG）的培养基中培养。此后，沉默SIRT1或在RMEC中过表达，然后进行EdU染色和基于Matrigel的管形成试验以评估细胞增殖和管形成。YAP，HIF1α，VEGFA用双荧光素酶报告基因检测法进一步说明。用免疫荧光测定的IB4阳性血管内皮细胞计数视网膜前新血管细胞数目。SIRT1在具有低催化活性的DR和HG处理的RMEC的小鼠中表达较差。在HG条件下，RMEC的增殖和管形成能力有所提高，在SIRT1过表达后可以逆转。通过检测到YAP作为miR-20a的关键靶基因，可以确定SIRT1可以正向调节miR-20a的表达。我们的数据表明，YAP可以通过诱导HIF1α上调VEGFA。此外，SIRT1的过表达强烈抑制RMEC增殖和血管生成，这可以通过恢复YAP /HIF1α/ VEGFA表达来逆转。在一起