ANGPTL-4/Profilin-1通路在糖尿病视网膜病变代谢记忆分子机制中的作用

Diabetic retinopathy (DR) is the primary leading cause of blindness in working-age people. The effect of metabolic memory of hyperglycemia, which not only affects the pathogenesis but also the therapeutic effect and prognosis of DR, prolongs DR progression after good control of hyperglycemia. Hitherto, relative research focusing on metabolic memory effect is still at its preliminary stage. Recently, we have provided proofs that the expression levels of angiopoietin-like protein 4 (ANGPTL-4), a novel adipocytokine, and profilin-1, are both significantly elevated and correlated under high glucose (HG) conditions of DR; furthers, ANGPTL-4 correlates with serum lipids and vascular endothelial growth factor, and profilin-1 mediates critical pathology of DR, such as inflammation, apoptosis and angiogenesis, etc. However, the mechanism of ANGPTL-4 affects profilin-1 remains unknown. To clarify this, we adopt in vivo and in vitro models of hyperglycemia metabolic memory to demonstrate that HG-activated ANGPTL-4 mediates inflammation, apoptosis and angiogenesis via up-regulating profilin-1, favoring DR vascular inflammatory dysfunction progression. Then, we reveal the therapy-targeting role of ANGPTL-4/profilin-1 pathway for DR via small interfering RNA. The prospection of this research would help elucidate the role of ANGPTL-4/profilin-1 pathway in DR metabolic memory and provide new thoughts for clinical prevention and therapy of DR.

糖尿病视网膜病变（DR）是工作年龄人群中首位致盲性病变，高血糖独特的“代谢记忆”效应造成DR患者在血糖控制正常稳定后，病情仍持续进展，故研究其具体机制非常必要。申请人前期研究发现，ANGPTL-4和Profilin-1在DR高糖环境中均高表达且具有相关性，ANGPTL-4与血脂和VEGF关系密切，而Profilin-1在DR血管炎性病变等过程中发挥重要作用；但ANGPTL-4对Profilin-1的具体调控机制尚不明确。为阐明其机制，我们利用高糖代谢记忆细胞和动物模型，验证高糖活化ANGPTL-4，进而上调Profilin-1，可能通过激活细胞凋亡、炎症及血管生成因子通路，最终引起视网膜血管炎性损伤；并进行体内干预，检测ANGPTL-4/Profilin-1通路是否为DR治疗的有效靶点。本课题有望阐明ANGPTL-4/Profilin-1通路在代谢记忆中的作用，为DR的临床防治提供新思路。

糖尿病视网膜病变（DR）是一种常见的糖尿病慢性并发症。本研究通过探索高糖高脂条件下血管内皮细胞生物特性的改变，进而揭示DR发生发展机制及寻找新的DR治疗靶点。首先，在高糖代谢记忆HRMECs细胞模型中，验证ANGPTL-4激活Profilin-1的上下游关系；其次，探讨ANGPTL-4通过LDL/ApoB激活Profilin-1，活化下游炎症、凋亡及血管生成因子通路，进而促进DR高糖代谢记忆的作用机制；第三，在高糖代谢记忆大鼠模型中，探讨通过干预ANGPTL-4/ Profilin-1通路，逆转高糖代谢记忆效应，进而治疗DR的可能性。为此，我们采用了体内和体外实验研究方法，利用WB、RT-PCR与ELISA等检测细胞和上清中各蛋白表达及活性；利用流式细胞术、TUNEL试剂盒与CLSM等检测各组细胞的凋亡情况；利用透射电镜观察视网膜细胞基底膜厚度等超微结构的改变；利用FITC-dextran与鼠尾静脉注射Evan Blue检测细胞与视网膜血管的渗透性情况；利用96孔板检测细胞增殖，Transwell小室检测细胞侵袭，划痕实验检测细胞迁徙，Matrigel生物胶检测管腔形成。我们发现，高糖引起视网膜血管内皮细胞ANGPTL-4表达水平升高，活化的ANGPTL-4通过上调LDL/ApoB，进而促进Profilin-1高表达，激活炎症、凋亡和血管生成因子通路，最终促进DR代谢记忆的形成。ANGPTL-4/Profilin-1通路不仅是高糖代谢记忆中的一个关键信号通路，也可能是糖代谢与脂代谢的一个连接点，在糖尿病合并异常脂代谢最终导致DR发病过程中可能发挥重要作用。