Arias Monica, Sauer-Lehnen Sibille, Treptau Jens, Janoschek Nora, Theuerkauf Ingo, Buettner Reinhard, Gressner Axel M, Weiskirchen Ralf
Institute of Clinical Chemistry and Pathobiochemistry, RWTH-University Hospital, Aachen, D-52074, Germany.
BMC Gastroenterol. 2003 Oct 18;3:29. doi: 10.1186/1471-230X-3-29.
Transforming growth factor-beta (TGF-beta) is a key mediator in establishing liver fibrosis. Therefore, TGF-beta as a causative agent may serve as a primary target for antifibrotic gene therapy approaches. We have previously shown that the adenoviral delivery of a transgene constitutively expressing a TGF-beta1 antisense mRNA blocks TGF-beta synthesis in culture-activated hepatic stellate cells and effectively abolishes ongoing fibrogenesis in vitro.
Ligature of the common bile duct was used to induce liver fibrosis in rats. The effect of the TGF-beta1 antisense on fibrogenesis was analyzed in this model of liver injury.
In the present study, we demonstrate that the adenoviral vector directs the synthesis of mRNA quantities that are approximately 8000-fold more abundant than endogenous TGF-beta1 mRNA. In experimentally injured rat livers induced by ligature of the common bile duct, a model for persistent fibrogenesis and cirrhosis, administration of the adenoviral vector abrogates TGF-beta-enhanced production of collagen and alpha-smooth muscle actin. Furthermore, the number of cells positive for alpha-smooth muscle actin resulting from active recruitment of activated hepatic stellate cells around the bile ductular structures was significantly reduced in animals after application of Ad5-CMV-AS-TGF-beta1. However, the observed elevated serum levels of aspartate aminotransferase, alanine aminotransferase, and bilirubin induced in this obstructive liver injury model were not significantly altered in the presence of the TGF-beta antagonist.
Taken together, our data provides in vivo evidence that the delivery of TGF-beta1 antisense mRNA specifically abolishes the diverse effects of direct TGF-beta function in ongoing liver fibrogenesis. Therefore, we conclude that the expressed transgene is therapeutically useful for inhibition of TGF-beta effects in diverse applications, ranging from clarification of TGF-beta function in the course of liver injury to the development of novel gene therapeutic approaches.
转化生长因子-β（TGF-β）是肝纤维化形成过程中的关键介质。因此，作为致病因子的TGF-β可能成为抗纤维化基因治疗方法的主要靶点。我们之前已经表明，组成型表达TGF-β1反义mRNA的转基因通过腺病毒载体递送，可阻断培养激活的肝星状细胞中TGF-β的合成，并有效消除体外持续的纤维化形成。
采用胆总管结扎法诱导大鼠肝纤维化。在该肝损伤模型中分析TGF-β1反义基因对纤维化形成的影响。
在本研究中，我们证明腺病毒载体指导合成的mRNA量比内源性TGF-β1 mRNA丰富约8000倍。在由胆总管结扎诱导的实验性损伤大鼠肝脏中，这是一个持续性纤维化形成和肝硬化的模型，给予腺病毒载体可消除TGF-β增强的胶原蛋白和α-平滑肌肌动蛋白的产生。此外，在应用Ad5-CMV-AS-TGF-β1后，动物胆管结构周围因激活的肝星状细胞的活跃募集而导致的α-平滑肌肌动蛋白阳性细胞数量显著减少。然而，在该梗阻性肝损伤模型中观察到的升高的血清天冬氨酸转氨酶、丙氨酸转氨酶和胆红素水平，在存在TGF-β拮抗剂的情况下并未显著改变。
综上所述，我们的数据提供了体内证据，即TGF-β1反义mRNA的递送特异性消除了直接TGF-β功能在持续肝纤维化形成中的多种作用。因此，我们得出结论，所表达的转基因在多种应用中对抑制TGF-β作用具有治疗价值，从阐明肝损伤过程中TGF-β的功能到开发新的基因治疗方法。
