八眉猪肌内前体脂肪细胞的分离培养及诱导分化

Culture and induced differentiation of intramuscular preadipocytes from Bamei pigs in vitro

以Ⅱ型胶原酶消化5日龄八眉猪仔猪背最长肌组织,分离肌内前体脂肪细胞,经差速贴壁法纯化后接种培养,以RT-PCR和油红O染色法对分化前后的细胞进行鉴定,通过观察细胞形态变化,细胞贴壁率测定,及用MTT法检测增殖活性,油红O染色提取法检测成脂诱导分化过程中细胞脂肪含量的变化,分析前体脂肪细胞生长特性和分化能力。结果表明,分离的八眉猪肌内前体脂肪细胞呈梭状、形似成纤维细胞,贴壁性能好,贴壁率最高可达90%;细胞生长良好,增殖曲线呈"S"型,符合正常的细胞生长规律;肌内前体脂肪细胞体外自发分化能力弱,但经成脂诱导,可以分化为具有脂肪细胞典型特征的成熟脂肪细胞,分化水平以时间依赖方式显著提高。

Intramuscular （IM） preadipocytes were isolated from longissimus dorsi muscle obtained from 5-day old piglet of Bamei pigs by dtigestion of eollagenase type Ⅱ, and purified with differential adhesion method. The cells before and after differentiation were identified by RT-PCR and oil red O staining, responsively. The adherence rate and proliferation of the preadipocytes were detected, and variation in fat content in the cells induced differentiation process was determined by oil red O staining extraction, to study the growth characteristics and differentiation of the IM preadipoeytes. The results showed that the IM preadipocytes isolated from Bamei pigs appeared to spindle-shape and fibroblast-like and well attached with 90% of maximum adherent rate. The cells grew well, and had S-type of growth curve which was in accord with normal growth pattern of cells. Though having poorly potential of spontaneous differentiation in vitro, cultured IM preadipocytes could differentiate into adipocytes with typical characteristics after adipogenic induced, and differentiation level significantly increased in time-dependent manner.