Establishment of new generation liver tissue engineering approach

使用肝细胞移植的肝组织工程已被提出作为全器官移植或肝脏导向基因治疗的替代方案，以纠正各种类型的肝功能不全。同种异体小鼠肾被膜下移植肝细胞不能维持。然而，当我们将从Engelbreth-Holm-Spot细胞中提取的细胞外基质成分移植到肝细胞中时，肝细胞至少存活了140天，并形成了小的肝组织。在血友病A小鼠中进行肝脏工程重建了5%至10%的正常凝血活性，足以减少出血时间，并具有治疗效益。相反，皮下间隙并不支持使用Engelbreth Holm-Sarm凝胶基质的肝细胞的持续存活。我们假设，在移植部位建立局部血管网络将减少移植物丢失。为了验证这一想法，我们在肝细胞移植到皮下间隙之前提供了一种有效的血管生成剂。通过这种方法，在整个实验过程中(120天)实现了持续存活。为了确定这些工程化肝组织在体内也保持了其天然的再生潜力，我们对幼稚肝诱导了两种不同的增殖刺激模式，并证实了肝外组织内的肝细胞再生的活性与幼稚肝相似。总之，我们的研究表明，肝组织可以在肝外部位被工程化和维护，在体内保持其再生能力，并成功地用于治疗遗传性疾病。

Liver tissue engineering using hepatocyte transplantation has been proposed as an alternative to whole-organ transplantation or liver-directed gene therapy to correct various types of hepatic insufficiency. Hepatocytes are not sustained when transplanted under the kidney capsule of syngeneic mice. However, when we transplanted hepatocytes with the extracellular matrix components extracted from Engelbreth-Holm-Swarm cells, hepatocytes survived for at least 140 days and formed small liver tissues. Liver engineering in hemophilia A mice reconstituted 5% to 10% of normal clotting activity, enough to reduce the bleeding time and have a therapeutic benefit. Conversely, the subcutaneous space did not support the persistent survival of hepatocytes with Engelbreth Holm-Swarm gel matrix. We hypothesized that establishing a local vascular network at the transplantation site would reduce graft loss. To test this idea, we provided a potent angiogenic agent before hepatocyte transplantation into the subcutaneous space. With this procedure, persistent survival was achieved for the length of the experiment (120 days). To establish that these engineered liver tissues also retained their native regeneration potential in vivo, we induced two different modes of proliferative stimulus to the naive liver and confirmed that hepatocytes within the extrahepatic tissues regenerated with activity similar to that of naive liver. In conclusion, our studies indicate that liver tissues can be engineered and maintained at extrahepatic sites, retain their capacity for regeneration in vivo, and used to successfully treat genetic disorders.