Effects Of Berberine On Human Omental Preadipocytes Proliferation And Differentiation

Objective: To culture human omental preadipocytes primarily and investigate the proliferation and differentiation processes in vitro for better understanding the properties of hyperplasia and hypertrophy of human adipose tissues. To study the effects of berberine on human omental preadipocytes proliferation and differentiation . Methods: 1,Cell culture of human preadipocytes: Omental adipose tissue biopsies were obtained from patients undergoing elective open-abdominal surgery. None of the patients had diabetes or severe systemic illness or had any recent weight change. None was taking medications known to affect adipose tissue mass or metabolism. Age of patients was below 50, and body mass index was below 25. Adipocyte tissue was separated from omental adipose tissue and capillaries by dissection and then digested with collagenase type I .The digested tissue was subjected to progressive size filtration and centrifugation, followed by incubation . 2,Observation of cell proliferation process : Preadipocytes were seeded at a density of 10~4 cells/cm~2 and grown in base medium [DMEM:F12 (1:1),supplemented with 10% FCS, 100000IU/ml penicillin and streptomycin]. The proliferation of human preadipocytes was detected by MTT method and growth curve was drawn . 3,Observation of cell differrentiation process: To induce differentiation, 10~4 cells/cm~2 preadipocytes were placed in differentiated medium consisting of DMEM:F12 (1:1), supplemented with 5% FCS,100000IU/ml penicillin and streptomycin, 0.2nM thyroxine , 0.5uM insulin, 0.2uM dexamethasone(DEX), and 0.5 mM isobutylmethylxanthine (IBMX). The accumulation of lipid in the cytoplasm of differentiated adipocytes was observed by oil red O staining and photoed. The curve of time-OD value was drawn. 4,Effects of berberine on human omental preadipocytes proliferation and differentiation: Berberine was distinguished to add into the base medium or the differentiated medium to put up different dosages just as 0.1 μmol/ l,1 μmol/ l,10 μmol/ l . At proliferation and differentiation phases, we studied the effects of berberine on human omental preadipocytes at various times. The proliferation of human preadipocytes was detected by MTT method. The accumulation of lipid in the cytoplasm of differentiated adipocytes was observed by oil red O staining. 5,Statistical analysis: Values were given as means ±SE and analysized by SAS 8.2 and SPSS 11.5 softwares. Differences were considered significant at a value of P<0.05. Results: 1,Human preadipocytes were primary cultured successfully. The cells were highly homogeneous, proliferative and had a high differentiation rate. Their dynamic morphological changes ,growth curve ,extracting stained intracytoplasmic lipid with oil red O, all verified their preadipocyte identity. Under controlled conditions, the preadipocytes replayed their hyperplasia and hypertrophy process in vitro. 2 ,Berberine stimulated human preadipocytes proliferation and inhibited lipid accumulation in cell differentiation. Conclusion : 1,Human preadipocytes are separated from adipocyte tissue by using collagenase I and are investigated the proliferation and differentiation processes in vitro for better understanding the properties of hyperplasia and hypertrophy ofhuman adipose tissues . 2,Human preadipocytes respond to berberine characteristically. Our results suggest that berberine should have a potential to serve as a fat-reducing drug. It make a deeper understanding about the mechanisms which berberine control blood glucose and increase insulin sensitivity, and a clue of improving curative effect of berberine .