代综方对3T3-L1脂肪细胞分化及相关基因表达的影响

Effects of Dai Zong Fang on 3T3-L1 adipocyte differentiation and related gene expression

目的:研究代综方(Dai Zong Fang,DZF)对3T3-L1脂肪细胞分化及分化过程中相关基因表达的影响。方法:体外培养3T3-L1前脂肪细胞,鸡尾酒式诱导脂肪细胞分化,给予不同浓度DZF干预6 d。BODIPY493/503与DAPI双荧光染色观察脂肪细胞分化情况;甘油磷酸氧化酶-过氧化物酶法测定细胞内甘油三酯(triglyceride,TG)含量;RT-PCR检测CCAAT增强子结合蛋白-α(CCAAT enhancer binding protein-α,C/EBP-α)、过氧化物酶体增殖物激活受体-γ(peroxisome proliferator activated receptor-γ,PPAR-γ)、乙酰辅酶A羧化酶(acetyl-Co A carboxylase,ACC)、脂肪酸合成酶(fatty acid synthetase,FAS)的m RNA表达;Western blot法检测PPAR-γ、C/EBP-α的蛋白表达。结果:DZF抑制脂肪细胞分化,降低细胞内TG含量,并呈剂量依赖效应;与对照组比较,DZF大剂量组明显下调ACC的m RNA表达(P<0.05),DZF小中大剂量均明显下调FAS的m RNA表达(P<0.01);与对照组比较,DZF中、大剂量组明显下调C/EBPα、PPAR-γ的m RNA表达(P<0.01),Western蛋白表达结果与m RNA结果一致。结论:DZF能够明显抑制3T3-L1脂肪细胞的分化程度,其机制可能与下调C/EBPα、PPAR-γ、ACC、FAS基因的表达有关。

Objectives：To study the effect of Dai Zong Fang（DZF） on 3T3-L1 preadipocytes differentiation and changes in expression of differentiation associated genes. Methods：3T3-L1 preadipocytes cells were induced into mature adipocytes by Cocktail（1-methyl-3-isobutylxanthine,dexamethasone and insulin）,treated by different concentrations of DZF from 2 d to 8 d of differentiation. The lipid accumulation of adipocyte was observed by BODIPY493/503 and DAPI double fluorescent staining. The intracellular triglyceride（TG）was detected by glycerol phosphate oxidase and peroxidase. RT-PCR was used to detect m RNA expression of CCAAT enhancer binding protein-α（C/EBP-α）,peroxisome proliferators-activated receptors-γ（PPAR-γ）,acetyl-Co A carboxylase（ACC）,fatty acid synthetase（FAS）. Western blot was used to detect protein expression of PPAR-γ and C/EBP-α. Results：DZF could inhibit adipocyte differentiation and reduce intracellular TG content in a dose-dependent manner. ACC m RNA expression was significantly downregulated in large-dose DZF group than in control group（P0.05）. FAS m RNA expression was significantly down-regulated in small,medium,large-dose DZF groups than in control group（P0.01）. C/EBPα and PPAR-γ m RNA expression was significantly downregulated in medium,large-dose DZF groups than in control group（P0.01）. The result of Western blot was consistent with that of m RNA results. Conclusion ：DZF could significantly inhibit the differentiation of adipocyte,the mechanisms might be related to the down-regulation of C/EBPα,PPAR-γ,ACC,FAS gene expression.