During The Differentiation, The Influence Of IGF1 Gene On The Proliferation Capacity Of Porcine Adipocytes

In this experiment, in order to reseach the influence of IGF1 gene on the proliferation capacity of porcine adipocytes during the differentiation, we chose 9 different time points during adipocytes inducing to differentiate to test the relative expression levels of IGF 1, IGFBP3, EN01, DCBLD2, APOE and BCL6 genes, using fluorescence quantitative, at frist. Then we analyzed genes’relative expression levels and their relevance, and we draught the growthcurve of adipocytes while this time. Among this genes, IGFBP3, EN01, DCBLD2, APOE and BCL6 were selected by our research team early, because they expressed high after adipocytes differentiation, using subtractive suppression hybridization. Second, we constructed a recombinanted eukaryotic expression vector pEGFP-N3-IGF1 and transfected it into porcine peradipocytes. Then we induced swine preadipocytes which was transfected successfully to differentiate, and draught the growthcurve on the 9 time points that was as same as fluorescence quantitative time points. Results were obtained as follows:1, The result of fluorescence quantitative show that the peak of expression in each of IGFBP3, EN01, APOE and BCL6 genes appeared after adipocytes differentiation. It proved that these four genes indeed expressed much more after inducing to differentiate than before. So it certified that those 4 genes were selected correctly by our research team in former time.2, Through a comprehensive analysised of genes relative expression levels, the correlation of expression levels, as well as cell growth curve, and contrasted the function of each gene in the existing research, we got that IGF1 in the process of adipocyte differentiation woked to promote cell proliferation and inhibit cell apoptosis. Meanwhile, during this time IGFBP3, EN01, DCBLD2 and BCL6 genes prefrom a role of inhibiting cell proliferation. And the role of APOE needed a futher study.3, We obtained the recombinant eukaryotic expression vector pEGFP-N3-IGF1 which was connected correctly and had no mutant site. After we transfected eukaryotic expression vector into porcine peradiocyte, we obtained transgene cells group A with empty vector pEGFP-N3, and group B with target vector pEGFP-N3-IGF1. The transfection efficiency of each was 75% and 73%.4, Analysising the growth curve of the adiocytes after overexpression 1GF1 we found that cells of group B which was transfected with pEGFP-N3-IGF1 plasmid proliferation more and apoptosis less compared with group A. It descripted that during adipocyte differentiation, IGF1 promoted cell proliferation and inhibited cell apoptosis indeed. So it certificated the conclusion that IGF1 in the process of adipocytes differentiation can promote cell proliferation and inhibit apoptosis...