The Primary And Subculture Of Grass Carp Adipocyte From Adipose Tissue

Preadipocytes is a distinctive cell which, has the ability to proliferat and differentiat to adipocytes,It exists in animal's body and could be effective through their whole life. The study on the law of proliferation and differentiation in adipocytes could lay the foundation of adipocytes metabolism disorder.Grass Carp (Ctenopharyngodon idellus), one of the four family fish in China, was a fresh water dweller. Currently, Grass carp fat excessive accumulation is the most critical problem in aquaculture. Research on Grass carp adipocytes proliferation and differentiation is a meaningful and practical study for aquacultural and economic use.The tissue mass and the digested method was used in culture of Grass Carp adipocytes tissue in this experiments.The optimal primary Grass Carp preadipocytes culture was established,and the biology characteristics of the cells were determined. The tissue mass and the digested method were all able to outgrow Grass Carp preadipocytes,The mature adipocytes number was less from the tissue mass and the cells were highly homogeneous from the digested method,which could differentiate mature adipocytes by oil red O staining identification.During the culture by digested method,theâ…£colllagenase digest method with 200 item mesh,1000r centrifugal force for 10 min could get more homogeneous Grass carp preadipocytes.The attempt to aquire the in vitro primary culture of Grass Carp preadipocytes was successful, Grass Carp adipocytes went down to the fourth generation sucessfully. Grass Carp adipocytes biology characteristics was as follows: preadipocytes could proliferate rapidly between 2nd day to 5th day,and began to differentiate on the 3nd day.Cells became bigger and bigger especially in the later culture time.The morphology of fibroblast-like preadipocytes changed to mature adipocytes which was round or ovoid.The isolated cell we obtained was preadipocyte with typical characteristics, which was proved by grow curve determination, subculture and oil red O staining identification. It follows the 3 standards that addressed by Van RLR: 1.it comes from the adipocytes, with the morphological character of fibroblast-like appearance and the cytoplasm doesn't or rarely have lipid pellets. 2. It proliferated rapidly and the double up time was similar with fibroblast-like cell that from the same experimental individual. 3. It became adipocytes when many cells confluenced together,and cytoplasm contained lots of lipid pellets.Our study has primarily established the optimal conditions of Grass Carp preadipocytes in vitro culture which proved to be a better model for studying lipogenesis and its regulation.