Apelin-13对3T3-L1前体脂肪细胞增殖与分化的影响

Effects of Apelin-13 on proliferation and differentiation of 3T3-L1 preadipocytes

目的探讨Apelin-13对3T3-L1前体脂肪细胞增殖与分化的影响及其可能的作用机制.方法体外培养3T3-L1前体脂肪细胞.取对数生长期细胞,分别予以不同浓度Apelin-13进行干预,四甲基偶氮唑盐(MTT)法检测Apelin-13对3T3-L1前体脂肪细胞增殖的影响."经典鸡尾酒法"诱导3 T3-L1前体脂肪细胞分化.将对数生长期细胞分为实验组和对照组.实验组分别于诱导分化第2、4、6、8天更换培养液的同时,予以细胞存活率抑制作用最强浓度的Apelin-13进行干预.对照组不做处理.于诱导分化第8天,根据油红O染色、脂质和甘油三酯含量检测细胞分化程度.于诱导分化第2、4、6、8天,分别采用RT-PCR和Western印迹检测过氧化物酶体增殖物活化受体γ(PPARγ)mRNA和蛋白表达的变化.结果 Apelin-13浓度越高,干预时间越长,3T3-L1前体脂肪细胞存活率越低.以100μmol/L Apelin-13干预96 h后,3T3-L1前体脂肪细胞存活率最低.经Apelin-13干预的3T3-L1前体脂肪细胞脂滴、脂质和甘油三酯含量均明显少于对照组(t=4.526、5.353、4.827,P均<0.05).与对照组相比,诱导分化第6、8天,经Apelin-13干预的3T3-L1前体脂肪细胞PPARγmRNA和蛋白表达量显著下降(t=4.962、5.416、4.734、5.627,P均<0.05).结论 Apelin-13呈浓度和时间依赖性抑制3 T3-L1前体脂肪细胞的增殖,并抑制3 T3-L1前体脂肪细胞分化过程中脂滴的形成,减少脂质聚集和甘油三酯含量.其作用机制可能与通过抑制PPARγ的表达有关.

Objective To explore the effects of Apelin-13 on the proliferation and differentiation of 3T3-L1 preadipocytes and its possible mechanisms. Methods 3T3-L1 preadipocytes were cultured in vitro. The logarithmic growth cells were treated with different concentrations of Apelin-13. The effects of Apelin-13 on the proliferation of 3T3-L1 preadipocytes were detected by methyl thiazolyl tetrazolium (MTT) assay. The"classical cocktail" method was used to induce the differentiation of 3T3-L1 preadipocytes. The logarithmic growth cells were divided into experimental group and control group. In experimental group, the culture me-diums were replaced on the 2nd, 4th, 6th and 8th day of differentiation induction, respectively;at the same time cells were intervened with Apelin-13 in a concentration with the strongest inhibitory effect on cell liva-bility. Cells in control group received no intervention. On the 8th day of differentiation induction, the degree of differentiation was detected by Oil red O staining, lipid and triglyceride assessment. Peroxisome prolifera-tor activated receptor γ( PPARγ) mRNA and protein expression were detected by RT-PCR and Western blotting on the 2nd, 4th, 6th and 8th day of differentiation induction, respectively. Results The cell liva-bility of 3T3-L1 preadipocytes was decreased along with the increase of the concentration as well as the inter-vention time of Apelin-13. The lowest livability of 3T3-L1 preadipocytes was observed after 96 hours of inter-vention with 100 μmol/L Apelin-13. Compared with control group, the lipid droplets, lipid and triglyceride content of 3T3-L1 preadipocytes treated with Apelin-13 were significantly lower (t=4. 526, 5. 353, 4. 827, all P<0. 05). The expression of PPARγmRNA and protein in 3T3-L1 preadipocytes treated with Apelin-13 were decreased significantly on the 6th and 8th day of differentiation (t=4. 962, 5. 416, 4. 734, 5. 627, all P<0. 05), compared with control group. Conclusions Apelin-13 inhibits the proliferation of 3T3-L1 prea-dipocytes in a concentration and time dependent manner. In addition, Apelin-13 could inhibit the formation of lipid droplets, and reduce lipid accumulation and triglyceride content during the differentiation of 3T3-L1 preadipocytes. The possible mechanism is related to inhibit the expression of PPARγ.