Regulation Of Mice Preadipocytes' Proliferation And Differentiation By ECM Components And CAMP

Normal and healthy male SD mice were employed to systematically explore the regulation of ECM Component ?collagen-IV , LN, cAMP, INS and adrenal hormone, on the proliferation and differentiation of mice preadipocytes, and influences were compared between the effects of Collagen-IV and LN on the expression of type I, III, IV of collagen genes at transcriptional level in mice adipocytes.Through repeated experiments, conditions required for the culture of mice preadipocytes were founded. In digestion culture, mice preadipocytes increase fast and vigorously, but in tissue culture, they increase slowly and evenly; when compared the influences of 4 substitutes on the culture of mice preadipocyte, it was found that growing conditions go down in the order of adding mice tale collagen, without substitutes, under glass slides and under plastic slides. By HE dyeing, Oil Red O dyeing and Immunohistochemical dyeing, it can all be found that mice preadiopocytes can be transformed to mature adipocytes with many lipid drops of various number and volume in cytoplasma, and the cellular nucleus by the side of the cells. Chemical indexes of protein, DNA, glucose, triglyceride were measured after mice preadipocytes were cultured for different times, it was found that the indexes increase gradually before the mice preadipocytes being cultured for 6 days, and they decline gradually after 6 days.By MTT method, Flow cytometer, GPDH activity detection, TE analysis and dot hybridization, regulation were investigated of collagen-IV and LN on the proliferation and differentiation of mice preadipocytes. The results showed that collagen-IV and LN can enhance the proliferation of mice preadipocytes, especially with a concentration of 100 ul/ml and 10 ul/ml respectively; and the two ECM components can reduce the cellular ratio during Gl period of the adipocytes cell cycle, increase it during PrI period, both are remarkable. The DNA content ratio of G2 to G1 period is very close to 2, it can be proven that the cell mitosis is in the way of bisplit; collagen-IV and LN and remarkably improve the content of protein, DNA, glucose, triglyceride in the mice adipocytes, especially at the concentration of 100 ul/ml and 10 ul/ml respectively. GPDH activities were much high than the control after the mice preadipocytes were treated with collagen-IV and LN, especially at the concentration of 100 ul/ml and 10 ul/ml respectively, it shows that they can both enhance the differentiation of preadipocytes. Various concentrations of collagen-IV and LN can differently influence the morphology and super microstructure of mice adipocytes, of which 100 ul/ml of collagen-IV and 10 ul/ml of LN and mostly improve the morphology and super microstructure. Collagen-IV and LN both can increase the positive hybridization signals of cDNA probe of I, III, IV types of collagen with the total RNA in adipocytes, and LN acts more obviously than collagen-IV do.By cellular numbering, MTT method, GPDH activity detection, TE analysis, the regulation of cAMP on the proliferation and differentiation of mice preadipocytes were investigated. The results are as follows: cAMPs of various concentrations can all remarkably increase the cell numbers ofpreadipocytes, reduce the cell double time, especially at the concentration of 107PM. Growth curve of preadipocytes treated with cAMP moves to the left of the control, the cell number increases before being cultured for 6 days, decreases after that. The cellular ratio is remarkably lower than the control in Gl period, higher in PrI period, indicating that cAMP can enhance the proliferation of mice preadipocytes. The contents of proteins, DNA, glucose, triglyceride are obviously higher in mice preadipocytes treated with cAMP than the control, especially when cAMP is at the concentration of of 107PM, showing that cAMP can improve the contents of the 4 substances in preadipocytes. The GPDH activities of adipocytes treated with cAMP are much more active than the control, and cAMP can have obvious influences on the morphology and super mic...