肾上腺素对U251细胞增殖及凋亡的影响

Effects of epinephrine on proliferation and apoptosis of U251 cells

目的:探讨肾上腺素对人胶质瘤细胞系U251细胞增殖、凋亡、周期的作用。方法:应用RT-PCR方法检测U251细胞中β-肾上腺素能受体(β-AR) mRNA的表达。MTT法检测肾上腺素对U251细胞增殖的影响;克隆形成实验检测肾上腺素对U251细胞克隆形成的影响;Annexin V-FITC/PI检测肾上腺素对U251细胞凋亡的影响;流式细胞术检测肾上腺素对U251细胞周期的影响;Western Blot检测周期、凋亡和自噬相关蛋白表达变化。结果:人胶质瘤细胞U251表达β-AR;肾上腺素以浓度依赖性的方式抑制U251细胞增殖(P <0. 05);肾上腺素抑制U251细胞的克隆形成(P <0. 05);流式结果显示,随着肾上腺素浓度的增加,细胞的凋亡率明显增加(P <0. 05),G0/G1期细胞比例显著增加(P <0. 05),G2/M期细胞比例下降(P <0. 05);肾上腺素处理U251细胞48 h,细胞周期蛋白Cyclin D表达下调,促凋亡蛋白Bax表达上调,自噬相关蛋白LC3-II/LC3-I比值升高。结论:肾上腺素可能通过激活细胞自噬抑制人胶质瘤细胞的增殖,使细胞阻滞在G0/G1期。

Objective:To explore the effects of epinephrine on cell proliferation,apoptosis and cell cycle of human glioma cell line U251.Methods:The mRNA expression ofβ-adrenergic receptor(β-AR)in human glioma cell line U251 was detected by reverse transcription-polymerase chain reaction(RT-PCR).The effects of epinephrine on U251 cell proliferation were investigated by MTT assay.The effect of epinephrine on U251 cell colony formation was examined by colony formation assay.Annexin V-FITC/PI was performed to examine cell apoptosis.Flow cytometry was performed to examine cell cycle.Western Blot was used to detect the changes of proteins related with cell cycle,apoptosis and autophagy.Results:The mRNA ofβ-AR was expressed in U251 cells.Epinephrine inhibited the proliferation of U251 cells in a dose-dependent manner(P<0.05).Epinephrine prevented U251 cells colony formation(P<0.05).Moreover,application of epinephrine raised U251 cell apoptosis ratio and the proportion at G0/G1 phase(P<0.05),and reduced the proportion of G2/M phase(P<0.05).Cyclin D was down-regulated,Bax and the ratio of LC3-II/LC3-I were up-regulated after treatment of epinephrine for 48 h.Conclusion:These observations together suggest that epinephrine inhibits human glioma cell proliferation via promoting cell apoptosis and resting cell at G0/G1 phase by activation of autophagy process.