摘要
脂肪酸转位酶(CD36)具有广泛的生物学功能,能够促进长链脂肪酸转入脂肪细胞、心肌细胞和骨骼肌细胞,对细胞脂质代谢有重要调控作用。为探究CD36对鸭原代肝细胞和PC3细胞脂质效应,本试验采用Ⅳ型胶原酶消化法成功分离孵化了21日龄鸭胚原代肝细胞并进行培养,RT-PCR法获取鸭CD36基因序列,构建了携带HIS标签的pEGFP-N3-CD36真核表达载体pEGFP-N3-CD36-His,脂质体法转染至鸭胚原代肝细胞和PC3细胞,His标签检测试剂盒测定CD36在2种细胞中的过表达量,探讨鸭CD36基因过表达与原代肝细胞和PC3细胞中脂质性状的关联性。结果显示,pEGFP-N3-CD36-His重组质粒在鸭原代肝细胞和PC3细胞中均能发出绿色荧光,说明构建的载体转染成功;CD36过表达对2种细胞中脂质指标的影响相似,均与细胞中的甘油三酯(TG)呈极显著正相关(P<0.01),与总胆固醇(TCH)、高密度脂蛋白(HDL)和低密度脂蛋白(LDL)呈极显著负相关(P<0.01);揭示CD36基因过表达可能引起2种细胞中脂质代谢活动的不平衡而引起脂质代谢相关疾病的发生,为进一步研究CD36基因对细胞脂质代谢的调控奠定了基础。
Fatty acid transposase(CD36) plays a widely biological functions,which can transport long-chain fatty acids into adipocytes,cardiomyocytes and skeletal muscle cells,and has important regulatory effects on cells lipid metabolism.In order to explore effect of CD36 on duck primary hepatocytes and PC3 cells lipid traits.The primary hepatocytes were successfully isolated from embryo hatched 21 days and cultured using type Ⅳ collagenase digestion method.Duck CD36 gene sequence was obtained by RT-PCR approach.Eukaryotic expression vector pEGFP-N3-CD36-His was constructed and transfected into duck embryo primary hepatocytes and PC3 cells using liposome method.His-gag detection kit was used to determine the overexpression of CD36 in two kinds of cells,and to explore relationship between duck CD36 gene overexpression and lipid traits in primary hepatocytes and PC3 cells,respectively.The results showed that the recombinant plasmid pEGFP-N3-CD36-His could emit green fluorescence in primary hepatocytes and PC3 cells,respectively,indicated that the constructed vector was successfully transfected.The overexpression of CD36 had similar effects on lipid parameters in two kinds of cells.CD36 overexpression level was significantlypositively correlated with triglyceride(TG)(P <0.01),and significantly negatively correlated with total cholesterol(TCH),high density lipoprotein(HDL),low density lipoprotein(LDL)in two kinds of cells(P<0.01),respectively.This study suggests that the overexpression of CD36 gene may cause disorder of lipid metabolism of duck primary hepatocytes and PC3 cells,and lead to develop lipid metabolism-related diseases,and lays a foundation for further study on the regulation of CD36 gene in cellular lipid metabolism.
作者
覃媛钰
罗华伦
吴磊
张依裕
QIN Yuan-yu;LUO Hua-lun;WU Lei;ZHANG Yi-yu(Key Laboratory of Animal Genetics , Breeding and Reproduction in the Plateau Mountainous Region ,Ministry of Education ,College of Animal Science ,Guizhou University,Guiyang 550025,China)
出处
《中国兽医学报》
CAS
CSCD
北大核心
2019年第10期2075-2081,共7页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(31760663)
贵州省科技人才补助资助项目(黔科合平台人才[2017]5788)
