ADAMTS2和ADAMTS4基因对牛前脂肪细胞外基质重构及肌内脂肪沉积的调控研究

项目名称： ADAMTS2和ADAMTS4基因对牛前脂肪细胞外基质重构及肌内脂肪沉积的调控研究

项目编号： No.31301941

项目类型： 青年科学基金项目

立项/批准年度： 2014

项目学科： 农业科学

项目作者： 张小辉

作者单位： 河南科技大学

项目金额： 24万元

中文摘要： ADAMTSs是调控细胞外基质重构的新型金属蛋白酶，前期研究表明，ADAMTS2和ADAMTS4基因与南阳牛肌内脂肪沉积调控有关。本研究拟分离牛脂肪间充质干细胞并进行诱导培养，利用qRT-PCR分析细胞分化过程中ADAMTS2和ADAMTS4基因的表达变化。构建并筛选ADAMTS2和ADAMTS4基因RNAi的CPP-PNA非病毒投送系统，转染牛前脂肪细胞，利用激光共聚焦显微镜、扫描电镜和透射电镜分析RNAi对前脂肪细胞外基质重构的影响；通过油红O 染色分析RNAi对细胞内脂肪沉积的影响；利用qRT-PCR分析RNAi对细胞分化、细胞外基质重构和脂肪沉积关键基因表达的影响；通过肌内注射CPP-PNA进行活体试验，分析RNAi对肌肉组织结构和肌内脂肪的调控作用。本研究将细胞外基质重构与前脂肪细胞的分化、脂肪沉积和基因表达联系起来，为牛前脂肪细胞分化和脂肪沉积的调控研究提供新的思路和依据。

中文关键词： 牛；脂肪细胞；ADAMTS2；ADAMTS4；细胞外基质

英文摘要： The ADAMTSs are new members of metalloprotease,which regulate the remodeling of extracellular matrix(ECM). The ADAMTS2 and ADAMTS2 participate the regulation of intramuscular fat(IMF) in Nanyang cattle. In this study, the cattle adipose-derived stromal cells (ADSCs) will be isolated and induced to adipocyte in vitro. The expression patterns of ADAMTS2 and ADAMTS4 will be analyzed by qRT-PCR during the differentiation of preadipocyte. The nonviral delivery system (CPP-PNA) will be designed and synthesized. The effect of ADAMTS2 and ADAMTS4 RNAi by CPP-PNA during the differentiation of preadipocyte will be analyzed by the following methods: the remoding of extracellular matrix of preadipocyte will be analyzed by laser scanning confocal microscope(LSCM), scanning electron microscope(SEM) and transmission electron microscope(TEM); the lipid deposition will be analyzed by oil red O accumulation in cytoplasm; the expression of key genes, which regulate the cell differentiation, remodeling of extracellular matrix and lipid deposition, will be analyzed by qRT-PCR during the differentiation of preadipocyte. The IMF and muscular histological structure will be analyzed after intramuscular injection of CPP-PNA in vivo. The relationship of extracellular matrix remodeling and cell differentiation, lipid deposition, gene expre

英文关键词： Cattle；Adipocytes；ADAMTS2；ADAMTS4；Extracellular matrix