目的 探讨高糖对血管内皮细胞分泌高迁移率族蛋白B1(HMGB1)的影响,以及高糖条件培养液对血管内皮细胞氧化应激水平的影响.方法 1.采用不同糖浓度的培养基(5.6mmol/L,20mmol/L,33mmol/L)体外培养人脐静脉血管内皮细胞株(Ea.hy926),台盼兰排斥法检测48h以内细胞的存活率,使用Western blot检测培养液上清中HMGB1的表达水平.2.分别用33mmol/L高糖培养基、混合液(33mmol/L组上清液与33mmol/l高糖培养基混合),刺激血管内皮细胞,采用分光光度比色法检测血管内皮细胞内丙二醛(MDA)和超氧化物岐化酶(SOD)的水平.结果 1.与正常对照组相比,高糖组培养液上清中HMGB1表达均明显增加(P<0.05).台盼蓝排斥法检测不同糖浓度48h内对细胞生存率影响较小.2.与单纯高糖培养基刺激组相比,混合液刺激组的血管内皮细胞内MDA、SOD水平有显著差异(P<0.05).结论 高糖可诱导体外血管内皮细胞表达分泌HMGB1.高糖环境下,机体可能通过增加分泌HMGB1,加强氧化应激反应而促进脑血管病的发生.%Objective To observe the effects of high glucose on the expression of HMGB1 in cultured vascular endothelial cells( VECs ) and the effects of the culture fluid with high glucose condition on oxidative stress. Methods 1. The human umbilical veins endothelial line ( Ea. hy926 ) were cultured in vitro. VECs were cultured with glucose in different concentrations( 5. 6 mmol/L,20 mmol/L and 33mmol/L ) for 48h. The levels of HMGB1 in the supernatant were measured by Western blot. The cell viahility within 48h was examined by trypan blue exclusion test. 2.VECs were divided into two groups: high glucose group ( 33mmol/l ) and the mixed group ( the supernatant of 33mmol/L group plus 33mmol/L high glucose medium ). 24 hours later, the levels of MDA and SOD were measured by UV-Spectrometer. Results 1. Compared with normal control, the expression of HMGB1 was significantly increased in the supernatant of high glucose group( P ＜0. 05 ). Within 48h, no differences in cell viability were found compared with control group by Trypan hlue exclusion test. 2. The level of MDA was significantly higher in the mixed group than that in 33mmo/L high glucose group. And the activity of SOD was lower in the mixed group than that in high glucose group ( P ＜ 0. 05 ). Conclusion High glucose could significantly induce the expression of HMCB1 in cultured VESs. In high glucose condition,the level of HMCB1 increases, which may play a role in oxidative stress and promote the occurrence of cerebrovascular disease.
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