草鱼前体脂肪细胞原代、传代培养及鉴定  

The Primary and Subculture and Identification of Grass Carp Preadipocyte from Adipose Tissue

作　　者：曹锴[1] 李影[1] 蔡志华[1] 

机构地区：[1]重庆师范大学生命科学学院重庆市动物生物学重点实验室,重庆400047

出　　处：《重庆师范大学学报（自然科学版）》2010年第6期20-22,F0002,共4页Journal of Chongqing Normal University:Natural Science

基　　金：国家自然科学基金(No.30800843);重庆师范大学重庆市动物学重点学科拓展研究项目(2009)

摘　　要：建立草鱼（Ctenopharyngodon idellus）前体脂肪细胞的培养体系,体外重现草鱼脂肪细胞的增殖分化过程。实验采用油红O染色法对脂肪细胞进行鉴定;采用倒置显微镜（NIKON）和CCD对细胞进行观察摄像;选用的健康草鱼体重为800~900 g;利用体外组织培养技术在温度为28℃,CO2浓度为5%,血清浓度为20%,pH值为7.0~7.2的条件下,以DMEM/F12培养基对来源于草鱼腹腔的脂肪组织进行原代培养;单层的原代培养细胞达到瓶底面积的70%~80%且汇合时,对细胞进行传代培养。研究发现在本实验条件下,培养48 h后组织块周围有梭形细胞迁出;3 d后细胞数量增多,多为梭形和多边形;培养8 d后大部分细胞融合;细胞内脂滴可被亲脂的油红O着色,证明为脂肪细胞。对融合后的细胞进行传代培养,可传至第4代;传代第3 d可形成致密单层;第8 d胞质内含有大量脂滴。研究初步确立了较为适宜的草鱼前体脂肪细胞离体培养体系。To establish the culture system of grass carp（Ctenopharyngodon idellus） preadipocyte,and redisplay the whole process of the proliferation and differentiation of grass carp preadipocytes in vitro,these experiments were carried out.The adipocytes were identified by oil red O staining,and the cells morphology was observed by inverted microscope and CCD.Grass carp selected was healthy and about 800～900 g.Primary culture of adipose tissue from grass carp abdominal cavity was done in the DMEM/F12 medium by using the tissue culture technology.All the experiments were done under the condition of 28℃,5% CO2,20% calf serum,pH 7.0～7.2.When monolayer cells occupied 70%～80%of the bottle＇s bottom and confluenced,the subculture was done.The results showed that the spindle cells emigrated from the tissues after 48 hours,the cell number increased and the cell morphology were mainly spindle and polygon after 3 days,cells confluenced most after 8 days and oil red O staining proved that stained cells were adipocytes.Confluenced cells could be passed at fourth passage.The cells could formed monolayer at the third day,and cytoplasm contained large quantities of lipid droplets at the eighth day.In summary,the sutiable culture system of grass carp preadipocyte was estabilished initially.

关 键 词：草鱼 脂肪细胞 原代培养 传代培养 

分 类 号：Q813.1[生物学—生物工程]