Study on gene therapy for liver regeneration. -Effect of gene transduction of soluble HB-EGF and soluble TGF-β type II receptor on liver regeneration-

加速肝再生对肝功能衰竭的治疗具有重要意义。我们已经报道，HB-EGF是一种强大的肝细胞有丝分裂原，在肝再生中发挥重要作用。另一方面，生长抑制因子转化生长因子-β刺激肝细胞凋亡，抑制肝再生。在本研究中，我们证明了肝部分切除后，肝非实质细胞中的核因子-kB被激活，并刺激了肝细胞生长因子基因的表达。为了探索促进肝再生的新方法，我们研究了双重导入可溶性HB-EGF基因和阻断转化生长因子-β作用的转化生长因子-β的作用。我们构建了含有人免疫球蛋白Fc区或可溶性HB-β基因的可溶性转化生长因子-EGFII型受体基因的表达载体。利用体内电穿孔的方法将这两种基因导入小鼠骨骼肌。用新开发的酶联免疫吸附试验系统检测血浆中可溶性HB-β和可溶性转化生长因子-EGF-II型受体的浓度。经透明质酸酶预处理后，肝组织中可溶性HB-β和sTGFII型受体的表达水平提高了6倍，为进一步开展促进肝再生的基因治疗奠定了基础。

Acceleration of liver regeneration is important for the treatment of hepatic failure. We have reported that HB-EGF is a potent mitogen of hepatocyte and plays an important role in liver regeneratian. On the other hands, TGF-β, a growth inhibitory factor stimulates apoptosis of hepatocyte and inhibits liver regeneration.In this study, we demonstrated that NF-kB is activated in hepatic non-parenchymal cells after partial hepatectomy and stimulates the expression of HB-EGF gene. We also indicated that soluble form of HB-EGF is essential for liver regeneration using HB-EGF transgenic mice.To develop the new methods for acceleration the liver regeneration, dual introduction of soluble HB-EGF gene and soluble TGF-βtype II receptor that blocks the action of TGF-β is investigated. We developed the expression plasmid containing soluble TGF-βtype II receptor cDNA attached with human immunoglobulin Fc region or soluble HB-EGF cDNA. Both genes were introduced into mice skeletal muscle using in vivo electroporation methods. The plasma concentration of soluble HB-EGF and soluble TGF-β type II receptor were measured with newly developed ELISA systems. Pretreatment with hyaluronidase increased the expression levels and obtains 6-fold higher concentrations of soluble HB-EGF or soluble TGF-βtype II receptor.Our results give a basis for development of the gene therapy to accelerate the liver regeneration.