摘要
为了建立莱芜猪前体脂肪细胞的体外培养体系,探索猪脂肪组织增生的生物学特征。本研究采集1日龄莱芜猪仔猪颈、背部皮下脂肪组织,采用胶原酶与胰蛋白酶相结合的方法,分离原代前体脂肪细胞,进行前体脂肪细胞的原代和传代培养,以及前体脂肪细胞诱导分化的研究,并采用Real-time PCR方法检测前体脂肪细胞分化过程中关键基因PPARγ的表达。结果表明,分离的猪原代前体脂肪细胞约8h后开始贴壁,贴壁的细胞呈短梭形或不规则的三角形,第3天多数细胞进入指数生长期,呈成纤维细胞样形态,细胞生长曲线近似S形;诱导培养结果显示,少量细胞在第2天开始出现脂滴,大多数细胞在第3天出现脂滴,第7～8天小脂滴逐渐融合成大脂滴,在第12天左右融合成大脂滴,油红O染色呈橘红色。在前体脂肪细胞分化过程中PPARγ基因mRNA的表达量逐渐增加,在诱导后的48h表达量最高,随后其表达丰度逐渐下降。本试验成功建立了莱芜猪前体脂肪细胞培养体系和体外诱导分化模型,为进一步研究莱芜猪前体脂肪细胞增殖与分化机制和猪体脂肪的沉积奠定基础。
The study aimed to establish the primary culture system of Laiwu Black pig preadipo- cyte for better studying the properties of hyperplasia of Laiwu Black pig adipose tissue. Preadipo- cytes were isolated from subcutaneous adipose tissue of 1 day piglet by the combination of collage- nase and trypsin digestion and were successfully cultured. The induction and differentiation of preadipocytes were studied, the expression of the important transcription factor PPAR7 in the process of preadipocytes differentiation were detected. The result showed that the isolated cells started adherence at 8 h after initiation of primary culture and displayed short spindle or irregular triangle. On 3rd day, the cells entered logarithmic growth phase, displayed fibroblast-like, and the cells growth showed ＇S＇ shape; the induction result showed that fat droplets occurred in small amount of cells on 2nd day, in large amount on 3rd day, those small fat droplets gradually converged into big ones from 7th to 8th day, and on the 12th day become to one big fat droplets, the adipocytes showed jacinth by oil red O staining; the PPAR？, mRNA levels gradually increased during the process of the preadipocytes differentiation, and reached the top at 48 h after induceddifferentiation culture, then the PPAR？＂ mRNA levels gradually decreased. The Laiwu Black pig primary culture system and the model of the induced differentiation in vitro were established suc- cessfully. This study laid the basis for further researching Laiwu Black pig adipocyte prolifera- tion, differentiation and porcine fat deposition.
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2012年第9期1346-1352,共7页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
国家转基因重大专项(2011ZX08006-002)
国家高技术研究发展计划"863"重点项目(2008AA101008)
山东省现代农业(生猪)产业技术体系建设专项
山东省农业良种工程重大课题(2009LZ012-03)
关键词
莱芜猪
前体脂肪细胞
细胞培养
诱导分化
Laiwu Black pig
preadipocyte
cell culture
induced differentiation
