CD49f剪接异构体在不同细胞系中的表达

目的比较CD49f选择性剪接异构体在人不同来源干细胞以及结肠癌细胞系中的表达,并构建特定异构体过表达载体用于功能研究。方法 RT-PCR及real-time PCR检测CD49f选择性剪接异构体在不同细胞中的表达;分子克隆方法构建各异构体的慢病毒过表达载体;流式细胞计量术、Western blot及real-time PCR检测CD49f各异构体的过表达;Transwell方法检测细胞侵袭能力的改变。结果人胚胎干细胞系H9中表达CD49f异构体B,而上皮类细胞仅表达异构体A;间质细胞中二者均有表达,但异构体A表达多于异构体B;检测的4个结肠癌细胞系中CD49f异构体A和B均有表达,其中HT29和HCT116中以异构体A为主,而HCT8和LoVo中异构体B的表达更为明显。构建的慢病毒表达载体可使HT29中CD49f异构体A和B获得特异性过表达,其中异构体B的过表达使HT29的侵袭能力增加。结论不同类型细胞中CD49f选择性剪接异构体A和B的表达存在显著差异,二者的生物学功能具有差别。

Expressions of CD49f splicing isoforms in different cell lines

Objective To compare the expression of CD 49f splicing isoforms in different human stem cells and colon cancer cell lines , and construct lentiviral vectors overexpressing specific isoform .Methods The expression of CD49f isoforms in different cells was detected by RT-PCR and real-time PCR.The lentiviral vectors overexpressing CD49f isoforms were constructed by molecular cloning method .The overexpression of CD49f in colon cancer cell line HT29 was confirmed by FCM, Western blot and real-time PCR, the invasive ability was examined by transwell assay.Results CD49f splicing isoform B was highly expressed in H 9 human embryonic stem cell line , while iso-form A expressed in epithelial cells .Both isoform A and B were expressed in mesenchymal cells .CD49f isoforms A and B were also expressed in colorectal cancer cell lines , while HT29 and HCT116 showed higher expression of iso-form A than isoform B , HCT8 and LoVo showed higher expression of isoform B than isoform A .Overexpression of CD49f isoform B greatly increased the invasive ability of HT 29 cells, while isoform A showed no effect .Conclusions The expressions of CD49f splicing isoforms A and B in different types of cells are significantly different , which sug-gests that CD49f isoforms play different biological functions in cells .