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BACKGROUND:Adipose-derived stem cel s are totipotent stem cel s in the adipose tissue, and have the function of self-renewal and multi-directional differentiation. Human adipose-derived stem cel s are ideal seed cel s with stable genetic milieu and few rejections. OBJECTIVE:To extract human adipose-derived stem cel s from human omental adipose tissue and to identify the cel s by adipogenic and osteogenic induction. METHODS:Omental adipose tissues were col ected from surgical patients to isolate and culture adipose-derived stem cel s using type I col agenase digestion, filtration and centrifugation. Cel growth was observed and proliferative curve of human adipose-derived stem cel s were drawn by cel counting method to calculate the doubling time at logarithmic growth phase. After adipogenic and osteogenic induction, induced cel s were identified using oil red O and alizarin red staining, respectively. RESULTS AND CONCLUSION:Human adipose-derived stem cel s were successful y isolated from the omentum tissues of surgical patients. Adherent cel s were fusiform-shaped and like fibroblasts. The growth curve of passage 3 cel s was in S shape, and the doubling time was 45.90 hours. After adipogenic and osteogenic induction for 2 and 3 hours, respectively, oil red O staining showed unequal-sized orange fat droplets, and alizarin red staining showed typical calcified nodules that were in orange. These findings indicate that adipose-derived stem cel s have the adipogenic and osteogenic capacity.
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AIM: Some studies found that the glucose uptake ability of diabetes mellitus mice is stronger after feeding with taurine. Whether the taurine can affect adipocytes or not deserves further studies. This article investigates the effects of taurine on the differentiation of 3T3-L1 preadipocytes and detects the mechanics. METHODS: Experiments were performed in the Laboratory Center of Xiangya Third Hospital of Central South University from July to September 2006. ①3T3L1 preadipocytes were provided by Shanghai Cell Bank of China Academy of Sciences. ②3T3-L1 preadipocytes were cultured in high glucose DMEM medium containing fetal bovine serum of 0.10 volume fraction, 108 u/L benzylpenicillin and 80?1010 U/L streptaquaine. After confluence, cells at 5?107 L-1 were incubated in culture flask, and induced with 0.5 mmol/L IBMX, 0.5 mg/L insulin and 1 ?mol/L dexamethasone. The cells in an experimental group were intervened with taurine, whereas these in a control group were not treated with taurine. Oil O staining was used to observe the development of adipose cells. C3T3-L1 preadipocytes were treated with 10,20 mmol/L taurine respectively for 24 and 48 hours. RNA and protein were extracted in the control group and the experimental group. ③Genes related to adipose cells development was examined by RT-PCR and Western-blot. RESULTS: ①The number of cells stained by oil O was less in the experimental group than the control group in 3T3-L1 preadipocytes treated with 10 mmol/L taurine for 14 days. ②After C3T3-L1 preadipocytes were treated with 10,20 mmol/L taurine respectively for 24 and 48 hours, there were no changes in expression of Insig-2 protein, PPAR, Insing-2, adiponectin, adiponectin receptor, GLUT-4, AP-2 mRNA. CONCLUSION: Taurine inhibits the differentiation of 3T3-L1 preadipocytes, but the mechanics still need to be investigated.
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Objective To establish the cell line stably expressing INSIG2 and observe its effecet on fat metabolism after overexpression of INSIG2.Methods The eukaryotic expression plasmid pcDNA3.1(+)-INSIG2 was constructed,which was transfected into 3T3-L1 cells.The expression of INSIG2 and related genes were detected by RT-PCR and immunohistochemistry,the contents of FFA in cell culture medium and adipocyte differentiation were detected by ELISA and Oil Red "O"staining respectively.Results After pcDNA3.1(+)-INSIG2 was transfected into the 3T3-L1 cells,the expression of INSIG1 mRNA and FAS mRNA were down-regulated,the content of FFA in the cell culture medium was decreased and adipocyte differentiation was drepressed.Conclusion The cell line stably expressing INSIG2 was successfully established,the transfected INSIG2 may have a drepressant effect on fat metabolism.