Takeda T, Murphy S, Uyama S, Organ G M, Schloo B L, Vacanti J P
Department of Surgery, Children's Hospital and Harvard Medical School, Boston, Massachusetts 02115., Department of Traumatology and Critical Care Medicine, Kyorin University, Tokyo, Japan.
Tissue Eng. 1995 Fall;1(3):253-62. doi: 10.1089/ten.1995.1.253.
Hepatocyte transplantation shows promise as therapy to support liver function. We have shown that hepatocytes can be transplanted into prevascularized synthetic polymers in rat models. While there are many studies in rats showing the benefits of hepatocyte transplantation, there are few in larger animal models more akin to humans. Therefore, we developed a swine model of hepatocyte transplantation using prevascularized synthetic polymers. Polyvinyl alcohol sponges measuring 2 x 3 x 0.5 cm were implanted in preperitoneal, mesenteric, and subcutaneous spaces for prevascularization as a transplantation bed. On postimplantation day 0, 4, 8, or 12 the sponges were removed and examined histologically. New tissue ingrowth was quite satisfactory on day 8 and 12 in preperitoneal and mesenteric sites, but the sponges in subcutaneous tissue were compressed, leaving little space for hepatocyte engraftment. Mesenteric sponges were irritating to intraabdominal organs and induced peritoneal adhesions. Thus, the preperitoneal sponges seemed to be best for hepatocyte transplantation. Hepatocytes isolated from donor livers using collagenase perfusion were transplanted into the preperitoneal sponges prevascularized for 0, 4, 8, or 12 days as allografts with cyclosporine immunosuppression, and the number of hepatocytes was evaluated on posttransplantation day 4. The optimal period for prevascularization for subsequent hepatocyte implantation was 8 days. The number of hepatocytes implanted in the preperitoneal sponges prevascularized for 8 days was counted on day 0, 1, 4, or 8 after transplantation. Hepatocytes were lost mainly in the first day after implantation, but still many hepatocytes maintained their shape histologically and there were mitotic figures confirming growth of the transplanted hepatocytes. Areas of hepatocytes within the sponge devices showed tissue remodeling with plates of hepatocytes lined with sinusoid-like capillaries and evidence of early tubular formation. Positive staining by immunohistochemical examination using antipig albumin indicated albumin production by implanted hepatocytes. The effect of a portacaval shunt as a hepatotrophic stimulation to maintenance of the implanted hepatocytes was evaluated on day 4, 8, or 12 after implantation. Total hepatocyte number in sponges was significantly increased by portacaval shunt compared to controls treated by a sham operation. These results suggest that significant numbers of hepatocytes can engraft and function using a prevascularized polymer bed as a site for transplantation and ongoing hepatotrophic stimulation with portacaval shunting.
肝细胞移植作为一种支持肝功能的治疗方法显示出前景。我们已经表明，在大鼠模型中，肝细胞可以被移植到预先血管化的合成聚合物中。虽然在大鼠中有许多研究显示了肝细胞移植的益处，但在更类似于人类的大型动物模型中的研究却很少。因此，我们利用预先血管化的合成聚合物开发了一种肝细胞移植的猪模型。将尺寸为2×3×0.5厘米的聚乙烯醇海绵植入腹膜前、肠系膜和皮下空间进行预先血管化，作为移植床。在植入后的第0、4、8或12天取出海绵并进行组织学检查。在腹膜前和肠系膜部位，第8天和第12天新组织长入情况相当令人满意，但皮下组织中的海绵被压缩，为肝细胞植入留下的空间很小。肠系膜海绵对腹腔内器官有刺激性并导致腹膜粘连。因此，腹膜前海绵似乎最适合肝细胞移植。使用胶原酶灌注从供体肝脏分离的肝细胞作为同种异体移植物，在环孢素免疫抑制下被移植到预先血管化0、4、8或12天的腹膜前海绵中，并在移植后第4天评估肝细胞数量。后续肝细胞植入的最佳预先血管化时间为8天。在移植后第0、1、4或8天对预先血管化8天的腹膜前海绵中植入的肝细胞数量进行计数。肝细胞主要在植入后的第一天丢失，但仍有许多肝细胞在组织学上保持其形态，并且有有丝分裂图像证实移植肝细胞的生长。海绵装置内肝细胞区域显示出组织重塑，肝细胞板排列有类似窦状的毛细血管，并存在早期管状形成的证据。使用抗猪白蛋白的免疫组织化学检查阳性染色表明植入的肝细胞产生白蛋白。在植入后第4、8或12天评估门腔分流作为对植入肝细胞维持的肝营养刺激的效果。与假手术治疗的对照组相比，门腔分流显著增加了海绵中肝细胞的总数。这些结果表明，使用预先血管化的聚合物床作为移植部位，并通过门腔分流进行持续的肝营养刺激，大量肝细胞可以植入并发挥功能。
