S-腺苷同型半胱氨酸对miR-128-转录因子EB通路介导的巨噬泡沫细胞胆固醇外流影响的研究

Elevated levels of S-adenosylhomocysteine (SAH) are positively associated with development and progression of atherosclerosis. Inhibition of macrophage cholesterol efflux is one of the important mechanisms promoting atherosclerosis (AS) development. However, the role of SAH in macrophage cholesterol efflux is unclear. Our recent findings revealed that there was a negative correlation between plasma SAH levels and cholesterol efflux capacity among patients with coronary artery disease. SAH intervention in macrophage foam cells significantly impairs autophagy, ABCA1 expression and cholesterol efflux, which are associated with miR-128 up-regulation and transcription factor EB (TFEB) down-regulation. Based on the above findings, we would firstly elucidate that miR-128 can specifically regulate TFEB and then inhibit autophagy and ABCA1 expression which mediate the inhibitory effect of SAH on cholesterol efflux from macrophage foam cells. Next, we would use ApoE-/- mouse model to confirm that whether the inhibitory effect of SAH on macrophage cholesterol efflux is through miR-128-TFEB-autophagy/ABCA1 pathway. Furthermore, we intend to use of betaine to reduce SAH levels for verifying that SAH plays a critical role in the development of AS by inhibiting cholesterol efflux. Our study provides a novel molecular insight into mechanisms of SAH-associated inhibition of macrophage cholesterol efflux that may contribute to the development of AS, and as a new therapeutic target to AS caused by disordered methionine metabolism.

S-腺苷同型半胱氨酸（SAH）水平升高与动脉粥样硬化（AS）的发生发展呈正相关，然而SAH对AS发展的重要机制—巨噬细胞胆固醇外流的影响尚不清楚。我们初步研究发现冠心病患者血浆SAH水平与胆固醇外流能力呈负相关；在巨噬泡沫细胞中，SAH降低自噬和ABCA1表达，抑制胆固醇外流，其效应与上调miR-128和下调转录因子EB（TFEB）的表达相关。在此基础上，本项目拟采用巨噬泡沫细胞模型，明确miR-128可以特异性调控TFEB进而抑制自噬和ABCA1表达介导了SAH抑制胆固醇外流作用；接着通过ApoE-/-小鼠模型验证SAH抑制巨噬细胞胆固醇外流的作用机制涉及miR-128-TFEB-自噬/ABCA1通路；并采用甜菜碱干预降低SAH水平来验证SAH通过抑制胆固醇外流促AS发展的机制。本研究将从脂质代谢角度阐明SAH促AS的作用机制，为蛋氨酸代谢紊乱所致AS的防治提供理论基础。

动脉粥样硬化性心血管疾病（AS）是中老年人群的常见疾病，位居我国居民疾病死因的首位，多项研究表明S-腺苷同型半胱氨酸（SAH）水平升高与动脉粥样硬化的发生发展呈正相关，然而SAH对AS发展的重要机制—巨噬细胞胆固醇外流的影响尚不明确。本项目主要从人群大样本前瞻性队列、巨噬泡沫细胞模型和ApoE-/-小鼠模型研究了SAH水平与胆固醇外流能力的关系和SAH调控巨噬泡沫细胞胆固醇外流的具体机制。人群研究结果表明冠心病患者血浆SAH水平与全因及心血管死亡风险呈显著的正相关关系，而与胆固醇外流能力成负相关关系，表明高浓度SAH可能通过抑制巨噬细胞胆固醇外流来促进AS的发展。在巨噬泡沫细胞模型中，研究发现SAH显著上调miRNA-128的表达，进而调控TFEB的活性来抑制巨噬泡沫细胞内的自噬和ABCA1表达，进而抑制巨噬泡沫细胞的胆固醇外流，miR-128在SAH抑制巨噬泡沫细胞胆固醇外流能力中具有特异性调控作用。在ApoE-/-小鼠模型中，从整体水平上研究了SAH对小鼠腹腔巨噬细胞胆固醇外流的影响，结果表明SAH通过激活小鼠腹腔巨噬泡沫细胞的miR-128-TFEB-自噬/ABCA1-胆固醇外流通路调控巨噬细胞的胆固醇外流。研究还进一步在高SAH的巨噬泡沫细胞模型和ApoE-/-小鼠动物模型中采用甜菜碱干预降低SAH水平，结果表明甜菜碱干预降低SAH水平后，改善了巨噬泡沫细胞胆固醇外流能力和AS，且与调控TFEB-自噬/ABCA1-胆固醇外流通路有关，证实了SAH与胆固醇外流的因果关系，表明能够以SAH为干预靶点防治蛋氨酸代谢紊乱引起的AS。本项目研究揭示了SAH抑制巨噬泡沫细胞胆固醇外流的具体机制，为防治蛋氨酸代谢紊乱引起的AS提供新的理论依据和实践基础。