晚期糖基化终产物对心肌微血管内皮细胞管样结构形成和崩解的影响及机制

目的研究晚期糖基化终产物(AGEs)对大鼠心肌微血管内皮细胞(CMECs)管样结构形成和崩解的影响及其初步机制。方法分离、培养SD大鼠CMECs,将AGEs-BSA与大鼠CMECs共同孵育,实验分为对照组(只加DMEM培养液)、BSA组(100 μg/ml BSA)、AGEs-BSA组(100μg/ml AGEs-BSA)。分别在孵育第1、3天时,用管样结构形成实验观察CMECs管样结构的长度;流式细胞仪检测CMECs的凋亡;ELISA法测定CMECs半胱天冬酶3(caspase-3)的活性。孵育1天采用Western blot法检测CMECs血管内皮细胞生长因子(VEGF)蛋白的表达。结果第1天时,与对照组和BSA组比较,AGEs-BSA组CMECs管样结构的长度明显增加,VEGF蛋白表达显著增加,差异均有统计学意义(P<0.01);而CMECs早期凋亡率及caspase-3的活性,差异无统计学意义(P>0.05)。第3天时,与对照组和BSA组比较,AGEs-BSA组CMECs的管样结构长度明显降低,CMECs早期凋亡率及caspase-3的活性显著增加,差异有统计学意义(P<0.01)。结论 AGEs-BSA在早期可能通过自分泌VEGF促进CMECs管样结构的形成,晚期可能通过促进CMECs的凋亡,加速管样结构的崩解。

Effects of advanced glycation end products on tubule-like structure formation and collapse in cardiac microvascular endothelial cells and their mechanisms

Objective To investigate the effects of advanced glycation end products(AGEs) on tubule-like structure formation (TLS) and collapse in cardiac microvascular endothelial cells (CMECs) and their possible mechanisms. Methods The CMECs from male SD rats were isolated and cultured by enzymatic digestion method. The TLS formation and collapse in CMECs were induced on collagen Ⅰ,and the length of TLS was quantified on day 1 and day 3. The expression of vascular endothelial growth factor(VEGF) protein was tested by Western blot on day 1. The rate of apoptosis was measured using flow cytometry on day 1 and day 3. The activity of caspase-3 was measured by ELISA on day 1 and day 3. Results the AGEs-BSA increased the TLS formation on day 1,but sharply accelerated the collapse of the TLS on day 3 (P<0.01). The AGEs-BSA enhanced the expression of VEGF (P<0.01) and had no effects on the rate of apoptosis and the activity of caspase-3 (P>0.05) on day 1,but increased the rate of apoptosis and the activity of caspase-3 dramatically on day3 (P<0.01).Conclusions The present study suggests that AGEs-BSA may increase the TLS formation of CMEC via autocrine of VEGF in the early stage, but accelerated the collapse of TLS in the CMECs via apoptosis in the late stage.