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目的:探讨获取高纯度的原代SD大鼠脑动脉内皮细胞(cerebral artery endothelial cell,CAEC)的培养方法。方法:选取5~6周SD雄性大鼠6只,体重110~140 g,取大脑,取脑动脉,剪碎,接种于明胶包被的直径35 mm培养皿中,内皮细胞培养基培养;采用倒置显微镜观察内皮细胞生长状况及其形态;免疫荧光法检测动脉内皮标志物Ⅷ因子相关抗原的表达。结果:培养60 h后可见细胞从贴壁的血管片段周围长出,细胞呈梭形;培养7 d后细胞可融合成片,融合后的脑动脉细胞呈典型的“鹅卵石”样外观;免疫荧光法检测显示动脉内皮标志物Ⅷ因子相关抗原表达阳性,内皮细胞纯度90%以上。结论:该方法能够成功获得纯度较高的原代SD大鼠脑动脉内皮细胞。
Objective: To explore a method of culturing high purity primary cerebral artery endothelial cells (CAEC) of SD rats. Methods: Six male Sprague-Dawley rats, weighing 110-140 g, were enrolled in this study. Sixty male Sprague-Dawley rats aged 5-6 weeks were enrolled in this study. The brains were removed from the cerebral arteries and cut into pieces. The gelatin-coated 35 mm diameter petri dishes were cultured in endothelial cell culture medium. Inverted microscope was used to observe the growth and morphology of endothelial cells. The expression of factor Ⅷ related antigen was detected by immunofluorescence. RESULTS: After cultured for 60 h, the cells grew out from the adherent vascular segments, and the cells were fusiform. After 7 days of culture, the cells could be integrated into pieces. The fused cerebral arteries showed the typical appearance of “pebbles” Immunofluorescence assay showed that endothelial marker Ⅷ factor-related antigen expression was positive, and the endothelial cell purity was over 90%. Conclusion: This method can successfully obtain primary SD rat cerebral artery endothelial cells with high purity.