基于GAS5/miR-455-5p/DDAH1信号途径探讨高良姜素抗动脉粥样硬化作用及机制研究

Alpinia officinarum hance shows great cardiovascular protection effects. Multiple studies have proved that galangin, the main active component from alpinia officinarum hance, has potent activity of anti-atherosclerosis. But the mechanism is still not fully illustrated. The pre-experiment found that galangin can resist high-fat diet-induced vascular endothelial damage in ApoE-/- mice and regulate the expression of GAS5 and miR-455-5p in endothelial cells. Since numerous studies have shown that asymmetric dimethylarginine (ADMA) can result in atherosclerosis, based on our pre-studys, we found that the gene of DDAH1 coding the key hydrolytic enzyme of ADMA was miR-455-5p’s target gene, and GAS5 was also predicted to bind to miR-455-5p by bioinformatics analysis. We further proved that the expression of miR-455-5p was significantly increased, and expression of GAS5 and DDAH1 were both significantly decreased in human atherosclerotic plaque tissue. In summary, we speculated that galangin may affect the level of ADMA through the GAS5/miR-455-5p/DDAH1 pathway against AS endothelial injury. This study intends to use knockout mice and endothelial cell models to elucidate the new mechanisms of galangin reversing endothelial dysfunction. Our study will illustrate a new mechanism of galangin’s anti-atherosclerosis effect, and will provide evidence of developing drugs from galangin to prevent and treat atherosclerosis.

高良姜具有心血管保护功效，国内外研究已证实高良姜素是其抗动脉粥样硬化（AS）的主要活性成分，但机制尚不明确。本项目预实验发现高良姜素可抵御高脂饮食诱导的ApoE-/-小鼠血管内皮损伤，调控内皮细胞中GAS5、miR-455-5p表达。现有研究已证实非对称性二甲基精氨酸（ADMA）可导致AS，我们前期的研究发现ADMA的水解酶DDAH1是miR-455-5p的靶基因，并预测GAS5也可能与miR-455-5p结合，且证实人AS斑块组织中GAS5及DDAH1表达降低，miR-455-5p表达升高。综上，我们推测高良姜素可能通过作用GAS5调控miR-455-5p/DDAH1通路影响ADMA水平，抵御AS内皮损伤。本研究拟使用基因敲除小鼠和内皮细胞模型，通过RNA干扰、报告基因及miRNA pull-down等技术，阐明高良姜素对内皮损伤保护的新机制，为发现心血管疾病新的防治靶点提供依据。

本项目从中药心血管药理学出发，利用动物和细胞生物学相结合的手段，查明了高良姜素抗动脉粥样硬化（AS）作用并进行初步机制探索，并从内皮炎症、巨噬细胞表型转化等多角度挖掘高良姜素抗炎作用的可能，同时收集临床样本探究与AS高危因素高血压的发病风险相关的药物基因组生物标志物。项目发现高良姜素可通过调控AS小鼠体内DDAH1/p-eNOS/ADMA水平，影响NO产生；影响AS小鼠脂质累积和肝功能；同时通过过表达miR-455-5p AS小鼠模型证实miR-455-5p在主动脉内膜中的过表达能逆转高良姜素的抗AS作用；高良姜素可抑制AS小鼠外周血中单个核细胞向促炎症表型M1型分化，促进其向抗炎症表型M2型分化。通过细胞实验发现，高良姜素抗内皮炎症的作用经NF-kB通路介导；同时其可以抑制原代小鼠巨噬细胞向M1型分化，促进其向M2型分化。项目还从内皮炎症和AS相关基因出发，收集中国南北方多个中心临床样本，进行系统的药物基因组学研究，发现PVT1 rs10956390和rs80177647、SOCS3 rs8064821多态性位点与高血压发病风险相关。项目还应用公共数据库，分析SOCS3 rs8064821基因型与SOCS3表达水平相关性，并在收集的样本中进行验证，为寻找高血压生物标志物，挖掘AS发病关键基因提供方向。

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