MDM2基因抑制LPS诱导的人脐静脉内皮细胞增殖率下降和凋亡率升高

目的:研究MDM2基因对LPS诱导人脐静脉内皮细胞(HUVECs)增殖率下降和凋亡率升高的影响。方法:分别用不同浓度的LPS作用于体外培养至对数期的HUVEC细胞,24 h后用MTT法检测LPS对HUVEC细胞增殖的抑制作用。用3 μg/ml LPS处理HUVEC细胞后检测细胞增殖率和凋亡率;利用RT-PCR和Western blot分析HUVEC细胞中MDM2 mRNA和蛋白表达水平。利用Lipofectamine TM 2000转染重组质粒pcDNA3.1-MDM2和pMDM2-shRNA以实现基因的过表达和敲除,RT-PCR和Western blot用于检测转染效果;转染后检测不同处理组HUVEC细胞的增殖率和凋亡率。结果:不同浓度LPS作用24 h后,HUVEC细胞的增殖受到抑制,且具有浓度依赖性。与0 μg/ml处理相比,经3 μg/ml LPS处理后,HUVEC细胞增殖率降低,凋亡率升高( P <0.05);同时,MDM2 mRNA和蛋白表达水平均明显下调( P <0.05)。与pcDNA3.1组相比,MDM2基因过表达后,HUVEC细胞增殖率明显升高,凋亡率显著降低( P <0.05)。与shNC组相比,MDM2基因敲除后,HUVEC细胞增殖率明显降低,凋亡率显著升高( P <0.05)。MDM2基因过表达抑制LPS诱导的HUVEC细胞增殖率下降、凋亡率升高( P <0.05)。结论: MDM2基因过表达抑制LPS诱导的HUVEC细胞增殖率下降和凋亡率升高。

MDM2 inhibited decreased proliferation and increased apoptosis of human umbilical vein endothelial cells induced by LPS

Objective: To study the effect of MDM2 gene on the decreased proliferation and increased apoptosis of human umbilical vein endothelial cells induced by LPS. Methods: LPS at different concentrations was applied to human umbilical vein endothelial cells(HUVECs) cultured to the logarithmic phase in vitro,respectively.The inhibition of LPS on the proliferation of HUVECs was detected by MTT assay after 24 h.The proliferation rate and apoptosis rate of HUVECs treated with 3 μg/ml LPS were detected,and the mRNA and protein expression of MDM2 in HUVECs were analyzed by RT-PCR and Western blot respectively.Lipofectamine TM 2000 was used to transfect recombinant plasmid pcDNA3.1-MDM2 and pMDM2-shRNA into HUVECs to achieve gene over-expression and knock-out.To detect transfection efficiency,the expression of MDM2 was measured by RT-PCR and Western blot.After transfection,the proliferation rate,apoptosis rate of HUVECs and the mRNA and protein expression of MDM2 in different treatment groups were determined separately. Results: The proliferation of HUVECs was inhibited after being treated by LPS at different concentrations for 24 h in concentration-dependent manner.Compared with 0 μg/ml,the proliferation rate of HUVECs treated with 3 μg/ml LPS reduced and apoptosis rate was increased( P <0.05).And both mRNA and protein levels of MDM2 in HUVECs decreased( P <0.05).Compared with pcDNA3.1 group,the proliferation rate of HUVECs obviously increased,and apoptosis rate sharply decreased after overexpression of MDM2 gene( P <0.05).Compared with shNC group,the proliferation rate of HUVECs distinctly increased,and apoptosis rate clearly decreased after MDM2 knock-out( P <0.05).Overexpression of MDM2 inhibited decreased proliferation of HUVECs and increased apoptosis rate induced by LPS( P <0.05). Conclusion: Overexpression of MDM2 gene inhibits decreased proliferation and increased apoptosis of HUVECs induced by LPS.