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目的 探讨端粒酶催化亚单位转染血管内皮细胞后,血管内皮细胞增生活性的变化。方法 应用胶原酶消化法培养脐静脉血管内皮细胞(UE),ABC法检测内皮细胞CD34表达。脂质体介导内皮细胞转染,MTT法测定细胞代谢活性并测定细胞生长曲线。基于逆转录聚合酶链反应的端粒重复末端扩增分析法检测端粒酶活性表达。结果 培养的细胞贴壁后为单层呈铺路石状排列。细胞爬片CD34染色呈阳性,pBABE-HYGRO-hERT转染内皮细胞测定端粒酶活性的吸光度为0.889。MTT实验表明:在各个时间点转染阳性细胞(HC)的吸光值都高于脐静脉血管内皮细胞(UE)(P<0.05)。UE与HC的细胞生长曲线形态相似,8 d内HC数量增长了约4倍。在各个时间点:HC较UE细胞数多(P<0.05)。结论 pBABE-HYGRO-hTERT转染内皮细胞后,提高了内皮细胞端粒酶的活性和增殖能力。
Objective To investigate the changes of vascular endothelial cell proliferation after transfection of vascular endothelial cells with telomerase catalytic subunit. Methods Human umbilical vein endothelial cells (UE) were cultured with collagenase digestion method, and CD34 expression in endothelial cells was detected by ABC method. Liposome-mediated endothelial cell transfection, MTT assay of cell metabolic activity and determination of cell growth curve. Telomerase activity was detected by telomere repeat end amplification assay based on reverse transcription polymerase chain reaction. Results The cultured cells were monolayer arranged in paving way. CD34 staining was positive for cell slide, and the absorbance of telomerase activity was 0.889 in pBABE-HYGRO-hERT transfected endothelial cells. MTT assay showed that the absorbance of transfected cells (HC) at all time points was higher than that of umbilical vein endothelial cells (UE) (P <0.05). The morphology of cell growth curve between UE and HC was similar, and the number of HC increased about 4 times within 8 days. At each time point: HC was more than that of UE cells (P <0.05). Conclusion After transfection of endothelial cells with pBABE-HYGRO-hTERT, the activity of telomerase and proliferation of endothelial cells were increased.