The Effect Of Glucagon Like Peptide-1on The3T3-L1Preadipocyte Differentiation And The Involved Mechanisms

Background:Nowadays, obesity and type2diabetes had been the serious public health problems in China. They are both metabolic diseases that have the similar basic reasons. Abnormal lipid deposition and insulin resistance are closely related to them [1]. Although a lot of genetic reasons contribute to the outcomes of obesity and insulin resistance, the most important factor in the environment is that the imbalance between energy intake and expenditure, thus causing too much fat accumulation in the body and causing obesity and lipid metabolism disorders[2.3]. The cellular mechanisms for obesity include the expansion of white adipose tissue via the hypertrophy of preexisting adipocytes and hyperplasia resulting from the adipogenesis of preadipocytes. Recent studies have shown that both the hypertrophy and hyperplasia have played the role in the pathology of obesity in human adults. However, there are significant differences in lipid and glucose metabolism between adipocyte hypertrophy and hyperplasia. Adipocyte hypertrophy is negatively correlated with dyslipidema and insulin resistance, independent of body composition. Interestingly, hyperplasia, which is characterized by an increased number of small subcutaneous adipocytes, may have a positive effect on lipid metabolism and insulin sensitivity through preadipocyte differentiation[4-6]. Glucagon-like peptide1(GLP-1) is the most effective incretin until now, which is secreted from intestinal L-cells and exerts multiple biological effects. Not only can it regulate the blood glucose, appetite, blood pressure and blood lipid; but also improve the cardiac and vascular endothelial function[7] Recent studies have revealed that GLP-1can significantly reduce fat mass and adipocyte hypertrophy, improve insulin sensitivity in adipocyte by up-regulating the expression of insulin receptor, insulin receptor substrate and Glut-4, reducing macrophage infiltration and inhibiting inflammatory adipocytes [8-10]. However, the effect of GLP-1on adipogenesis is less clear; and if GLP-1can improve the information of small adipocyte is also don’t have enough research. In this study, using3T3-L1preadipocyte, we examined the effect of GLP-1on preadipocyte differentiation and the mechanisms involved.Objective:1. To observe the effect of GLP-1on the adipocyte-specific proteins LPL, aP2and the transcription factors PPAR-y, CEBP/a during the process of3T3-L1preadipocyte differentiation.2. To observe the Effect of GLP-1on lipid accumulation and the cytomorphology of adipocyte.3. To investigate the possible mechanism involved in the GLP-1induced effect.Methods:1.3T3-L1preadipocytes culture and differentiation:3T3-L1preadipocytes were cultured in aseptic condition and the traditional "Cocktail method" was adopted to induce the3T3-L1preadipocyte differentiation.2. GLP-1was added to the medium of3T3-L1cells at different concentrations, MTT assay was used to test the cell viability.3. RT-PCR and Western-blot were used to exam the express levels of the adipocyte-specific markers.4. Oil Red O staining to examine the effect of GLP-1on lipid droplet accumulation at the8th day of differentiation, and the Image Pro plus5.02was used to analyse the size and number of lipid droplet.5. RT-PCR was used to exam the express level of Uncoupling protein-1(UCP-1) and Carnitine palmitoyltransferase1(CPT-1A) at the8th day of differentiation.6. Under the treatment of GLP-1, we investigated the levels of Akt, P38and ERK1/2 signaling pathway.Results:1. At varying times during differentiation, GLP-1enhanced the mRNA levels of the transcription factors PPAR-γ,CEBP/α and the adipocyte-specific markers LPL, aP2significantly in a dose-and time-dependent way, as compared to the control group(P<0.05).2. GLP-1enhanced the protein levels of the PPAR-γ and aP2significantly in a dose-and time-dependent way, as compared to the control group(P<0.05).3. During the first24h of differentiation, Akt, P38and ERK1/2signaling way were activated in different degrees. Compared to the control group, the protein level of p Akt in GLP-1group was increased markedly (P<0.05), while the protein levels of pP38and pERK1/2were not observed any significantly changes.4. Oil Red O staining results were shown that the lipid accumulation in experimental group were not increased as the enhanced concentrations of GLP-1. As compared to the control and other concentration groups, the cytomorphology of adipocyte in100nM GLP-1group was shown there were increased numbers of small adipocyte (P <0.05)5. Under the treatment of100nM GLP-1, the mRNA level of CPT-1A was increased significantly, while the mRNA level of UCP-1was not changed as compared to the control and other concentration groups (P<0.05).Conclusion:1. GLP-1enhanced the express of PPAR-γ, CEBP/α, LPL and aP2significantly in a dose-and time-dependent way during the period of3T3-L1preadipocyte differentiation,2. Akt signaling way may be involved in the above GLP-1-induced effects.3.100nM GLP-1can promote the formation of small adipocytes, which have more mRNA level of CPT-1A.