Liang Z, He Z
Hepatobiliary Surgery Center, Southwestern Hospital, Third Military Medical University, Chongqing.
Zhonghua Yi Xue Za Zhi. 1998 Nov;78(11):850-2.
To investigate possible role of antisense TGF-beta 1 RNA in the regulation of TGF-beta 1 and ECM production in Ito cells.
A human transforming growth factor-beta 1(TGF-beta 1) cDNA (1467 bp) was inserted in reverse orientation into the retroviral vector, constructed retroviral vector pLATSN of antisense RNA for TGF-beta 1. A higher-titer, recombinant retroviral vector carried antisense RNA for TGF-beta 1 produced in PA317 packaging cells has been introduced into human Ito cells lines LI90. After selection with G418, resistant colonies were obtained.
Stable integration of retrovirus in infectants was shown the presence of antisense RNA was detected by RT-PCR. The expression of TGF-beta 1 protein and the production of extracellular matrix such as FN, Co1A1 were markedly decreased in the antisense TGF-beta 1 transfected cultured cells by ELISA, immunohistochemistry and in situ hybridization.
Antisense RNA of TGF-beta 1 can be successfully used to inhibit Ito cells activated, endogenous TGF-beta 1 mRNA and extracellular matrix produced, and may provide a basis for the development of anti-fibrosis gene therapy.
研究反义转化生长因子-β1（TGF-β1）RNA在肝星状细胞（Ito细胞）中对TGF-β1及细胞外基质（ECM）产生的调节作用。
将人转化生长因子-β1（TGF-β1）cDNA（1467bp）反向插入逆转录病毒载体，构建TGF-β1反义RNA的逆转录病毒载体pLATSN。将在PA317包装细胞中产生的携带TGF-β1反义RNA的高滴度重组逆转录病毒载体导入人Ito细胞系LI90。经G418筛选后，获得抗性克隆。
通过RT-PCR检测到反义RNA的存在，表明逆转录病毒稳定整合入感染细胞。采用ELISA、免疫组织化学和原位杂交法检测发现，反义TGF-β1转染的培养细胞中TGF-β1蛋白表达及细胞外基质如纤连蛋白（FN）、I型胶原（ColA1）的产生均明显减少。
TGF-β1反义RNA可成功用于抑制Ito细胞活化、内源性TGF-β1 mRNA及细胞外基质的产生，可能为抗纤维化基因治疗的发展提供依据。
