牛前体脂肪细胞的分离培养及诱导分化

Primary culture and differentiation of bovine preadipocytes

【目的】建立牛前体脂肪细胞的体外分离培养方法,研究牛前体脂肪细胞的生物学特性和分化特征,为研究牛脂肪发育和脂肪沉积的机制提供一种简便有效的细胞模型。【方法】采用Ⅰ型胶原酶消化法自新生牛脂肪组织中获得前体脂肪细胞,对培养的牛前体脂肪细胞进行形态学观察、生长曲线测定、油红O染色及脂肪细胞标志基因LPL、PPAR-r和脂联素表达研究。【结果】80%的牛前体脂肪细胞接种后12h开始贴壁,4d后开始向脂肪细胞转化,10d后绝大部分细胞脂滴融合,细胞脂肪含量增加;分离的牛前体脂肪细胞接种后1~2d生长缓慢,3~8d进入对数生长期,9d后进入平台期,之后细胞数目开始下降。经胰岛素诱导分化过程中,LPL在牛前体脂肪细胞分化的早期开始表达,PPAR-γ在分化的中期开始高度表达,而脂联素在牛前体脂肪细胞分化的前期未检测到,在细胞分化后期高度表达。【结论】用Ⅰ型胶原酶消化法可获得大量的牛前体脂肪细胞。培养的牛前体脂肪细胞成分均一、生长旺盛,经胰岛素诱导后能稳定的向脂肪细胞分化。

[Objective] The study was preadipocytes in vitro and study the biologi [Method] Preadipocytes tured preadipocytes were expression of lipoprotein were isolated from to explore and establish a stable culturing system for bovine cal and differentiation characteristics of bovine preadipoeytes. adipose tissue of newborn bovine by type I collagenase. Cul- identified by morphological change, growth lipase gene （LPL）,peroxisome proliferators and adiponectin gene. [Result] 80 h,and the cells began to transfer to bovine prea fat cells after curve, oil red O staining, and mRNA activated receptor-）＇ gene （PPAR-r） pocytes began to paste the wall after inoculation 4 days. 10 days later,lipid droplets of most cells and fat contents increased. Bovine preadipocytes grew well in vitro. On days l--2, cells grew slow the cells entered logarithmic growth phase during days 3--8. The growth of cells became stable from since when the number of cells decreased. During the differentiation by insulin,LPL gene expressed for 12 fused, ly, and day 9, in the early stage and PPAR-γ gene expressed in the middle stage. The expression of adiponectin gene was not detected in the early stage, but it was highly expressed in mature adipocytes. [Conclusion] Preadipocytes isolated from adipose tissue of newborn bovine by type I collagenase grew well and could be differentiated into mature adipoeyte in vitro.