Research of gene therapy for liver cirrhosis and fibrosis, using solble TGF-beta receptor.

我们利用RT-PCR技术从小鼠cDNA文库中获得了TGF-β Ⅱ型受体胞外区的cDNA。并将其导入COS细胞中，在培养上清中表达4 OkD蛋白。通过与TGF-β的结合及TGF-β功能抑制试验，证实了该40 kD蛋白为TGF-β Ⅱ型受体蛋白，但由于cDNA太长，不能转化大肠杆菌，未能获得大量TGF-β Ⅱ型受体蛋白。因此，我们利用杆状病毒，并获得40 kD的蛋白。我们又尝试通过添加干扰素的信号肽和免疫球蛋白Fc区的方法对cDNA进行改造。从COS细胞培养上清中获得了3倍有效的4 OkD蛋白。本实验拟采用柱层析法纯化蛋白质，并将其应用于肝硬化模型大鼠。

We obtained cDNA of extra cellular domain of TGF-beta type II receptor using RT-PCR from mouse cDNA library. And it was introduced to COS cells and 4OkD protein was expressed in culture supernatant. We confirmed that this 40kD protein was TGF-beta type II receptor protein by binding to TGF-beta , and by inhibition test of TGF-beta function.And we failed to earn much TGF-beta type II receptor protein by introducinig cDNA to E.coli, because that cDNA was too long for F.coil. So we used Baculovirus, and obtained 4OkD protein. But it was not available in function.Next we tried to improve cDNA by adding signal peptide of Interferon and adding Immunoglobulin Fc region. We obtained three times degree of available 4OkD protein from COS cells culture supernatant. Now we are going to purify protein by column, and should like to use it in model rat of liver cirrhosis.