Bcl-2过度表达与糖尿病孕鼠模型血管早期病变的相关性研究

The relationship between overexpression of Bcl-2 and the early vascular lesions in pregnancy diabetic model rat

目的探讨糖尿病孕鼠模型是否存在早期血管病变及抗凋亡因子Bcl-2在血管病变中的作用。方法以链脲菌素诱导Wistar大鼠致妊娠期糖尿病,并于孕21天处死孕鼠,取孕鼠颈动脉作为研究血管,对血管组织HE染色,以KS400图像分析系统观察血管中段横截面内中膜厚度(intima-middle thickness,IMT),免疫组化SP法检测Bcl-2表达;对颈动脉血管平滑肌细胞(vascular smooth muscle cell,VSMC)进行原代培养,以磷脂酰丝氨酸外翻分析及流式细胞仪检测细胞凋亡;采用实时定量PCR法及蛋白印迹法检测细胞内Bcl-2的表达。结果①同正常孕鼠组相比,妊娠期糖尿病孕鼠颈动脉血管壁显著增厚[(12.56±2.62)μm和(8.45±1.25)μm,P<0.001];②正常孕鼠颈动脉平滑肌细胞加入高糖培养后,细胞凋亡率显著降低(P<0.001);③与正常孕鼠相比,妊娠期糖尿病孕鼠血管平滑肌细胞及血管组织内Bcl-2mRNA及蛋白表达增加。结论糖尿病孕鼠模型血管已发生早期病变,可能与高糖环境导致细胞内Bcl-2过度表达、抑制细胞凋亡有关。

Objective To study whether the diabetic pregnant rat model has early vascular disease,and the role of anti-apoptotic factor Bcl-2 in vascular disease.Methods Wistar diabetic rats were induced by streptozotocin during pregnancy and were sacrificed on 21 days gestation.Carotid vessels were taken as the study materies.Intima-middle thickness（IMT）of were studied by HE staining using the KS400 image analysis system.Expression of Bcl-2 on the carotid artery and smooth muscle cells were determinged by immunohistochemical method.The apoptosis of primary cultured vascular smooth muscle cell were analysed by phosphatidylserine and flow cytometry.Real-time PCR and western blotting methods were used to detect intracellular Bcl-2 expression.Results ①Compared with normal pregnant rats,carotid artery wall in pregnant rats with gestational diabetes were significantly thicker[（32.56±2.62）μm vs（21.45±1.25）μm,P=0.0001];②The apoptosis of carotid artery smooth muscle cells in normal pregnant rat after cultured in high glucose was significantly reduced（P=0.0001）;③Compared with normal pregnant rats,Bcl-2 RNA and protein expression of vascular smooth muscle cells in gestational diabetes rats were significantly increased（P=0.0001）.Conclusions Early vascular lesions in diabetic pregnant rat model have occurred and may result in overexpression of Bcl-2 and thus inhibit apoptosis.