重组人Ⅸ因子逆转录病毒载体在肌细胞中的表达

Expression of recombinant human factor Ⅸ gene in muscle cells in vitro

目的：观察肌细胞特异性表达ｈＦⅨ重组逆转录病毒载体（ＲＶ－ｈＦⅨ）体外转染成肌细胞后的表达效果。方法：采用产病毒细胞株ｐｄＬⅨｍ １ βＡ２００Ｍｅ１ ／ＰＡ３１７ 和ｐｄＬⅨｍ ２ βＡ２００Ｍｅ１ ／ＰＡ３１７ 培养上清，体外转染原代人胎、联合免疫缺陷（ＳＣＩＤ）小鼠及Ｃ５７ 鼠成肌细胞，ＥＬＩＳＡ法检测细胞培养上清中ｈＦⅨ分泌水平。结果：不同来源成肌细胞经两种载体转染后均有ｈＦⅨ蛋白分泌，其中人胎成肌细胞表达优于鼠源成肌细胞，１０６ 个细胞，２４ ｈ 表达最高值为（７９２±４９） ｎｇ。成肌细胞分化后ｈＦⅨ分泌水平高于分化前。结论：ＲＶ－ｈＦⅨ体外转染成肌细胞后可表达较高量的目的蛋白。

Objective: To observe the expression of muscle specific RV hFⅨ in myoblasts and myotubes. Methods: After primary myoblasts were obtained,they were transfected with RV hFⅨ pdLⅨm 1βA200Me 1 and pdLⅨm 2βA200Me 1. The hFⅨ protein secreted by the transfected cells in the medium were assayed by enzyme linked immunosordent assay with monocolonal anti human hFⅨ antibody. Results:Recombinant protein were secreted by myoblasts transfected with RV hFⅨ, the transfected human fetal ones were the best among myoblasts isolated from 3 sourses,which produced (792±49) ng hFⅨ/10 6 cells per 24 h. Conclusion: Myoblasts transfected with RV hFⅨ can effectively secrete hFⅨ protein.