Ghrelin对3T3-L1前脂肪细胞增殖和凋亡作用的研究

Effect of ghrelin on the proliferation and apoptosis of 3T3-L1 preadipocytes and its possible mechanism

目的研究ghrelin对3T3-L1前脂肪细胞增殖和凋亡的影响及其分子机制。方法体外培养3T3-L1前脂肪细胞,四甲基偶氮唑盐法(MTT法)检测ghrelin对3T3-L1前脂肪细胞增殖影响的剂量-时间效应关系;Annexin Ⅴ-FITC/PI流式细胞术分析ghrelin对3T3-L1前脂肪细胞凋亡率的影响;Western blot方法检测ghrelin干预后p-ERK1/2和p-Akt蛋白表达的变化。结果Ghrelin时间依赖性促进3T3-L1前脂肪细胞增殖,ghrelin10-7～10-15mol/L作用24h明显促进3T3-L1前脂肪细胞增殖,与对照组相比差异有统计学意义(P<0.05),其中ghrelin10-11mol/L促增殖效应最明显(P<0.01);10-11、10-13mol/L ghrelin干预后前脂肪细胞凋亡率较对照组显著降低(P<0.05),其中10-11mol/L时抑凋亡效应最明显;10-11mol/L ghrelin干预后前脂肪细胞和成熟脂肪细胞中p-ERK1/2和p-Akt蛋白表达量较对照组显著增加(P<0.01)。结论Ghrelin促进3T3-L1前脂肪细胞增殖并且抑制其凋亡,ERK1/2和PI3K/Akt信号通路可能参与了ghrelin调节细胞增殖和凋亡的过程。

Objective To explore the effect of ghrelin on the proliferation and apoptosis of 3T3-L1 preadipocytes and its molecular mechanism. Methods 3T3-L1 preadipocytes were cultured in vitro. MTT method was used to detect the dose-time response on the proliferation of 3T3-L1 preadipocytes by ghrelin. The influence of ghrelin on the apoptotic rate of 3T3-L1 preadipocytes was analyzed by flow cytometry （FCM）. The protein expression of p-ERK1/2 and p-Akt after intervened by ghrelin was determined by Western blot. Results Ghrelin promoted the proliferation of 3T3-L1 preadipocytes and the effect was time-dependent. Compared with the control group, in the concentrations of ghrelin from 10^-7 mol/L to 10^-15 mol/L for 24 hours, the proliferation of 3T3-L1 preadipocytes was significantly increased （P 〈 0.05）, and the most pronounced effect was at 10^-11 mol/L （P 〈 0.01）. After the intervention of ghrelin （10^-11, 10^-13 mol/L）, the apoptotic rate of preadipocytes decreased significantly when compared with control group （P 〈 0.05 ） , and the most pronounced effect was also at 10^-11 mol/L. The protein expression levels of p-ERK1/2 and p-Akt in preadipocytes and terminally differentiated adipocytes were significantly enhanced by the intervention of ghrelin at 10^-11 mol/L （P 〈 0.01 ）. Conclusions Ghrelin could efficiently promote the proliferation of 3T3-L1 preadipocytes and inhibit its apop＇tosis, and the signal pathway of ERK1/2 and PI3K/Akt may be involved in the regulation of cell proliferation and apoptosis process.