次世代型人工臓器設計のための腹膜機能の測定とモデル化

杂交型腹膜模型不仅可以用来研究溶质和水在腹膜中的转运，还可以用来研究内源性和/或外源性刺激引起的纤维化过程。然而，克隆腹膜成纤维细胞及其在细胞外基质中的3D培养等技术要求是需要克服的。在本研究中，我们对腹膜驻留细胞的三维培养用于体外重建腹膜进行了综述，除了一些利用培养的腹膜细胞进行宿主防御机制的研究外，很少有关于这方面的研究活动。通过改进他们的技术，我们建立了传代传代培养(t-DSC)，调节与培养塑料的接触时间，将成纤维细胞从原代培养的腹膜细胞中分离出来。在增加胶原浓度的胶原凝胶中加入PRFB(1×105/ml)与凝胶收缩呈负相关，而增加胶原凝胶中PRFB的细胞密度(2 mg/ml)与胶原凝胶收缩呈正相关。在3D-RPFB培养体系中加入转录抑制剂放线菌素D，可抑制凝胶收缩，且呈剂量依赖关系。然而，加入翻译抑制剂嘌呤霉素后，凝胶收缩并未受到抑制，同样，与2，2‘-联吡啶孵育后，凝胶收缩也没有明显影响。从这些结果可以推测，胶原凝胶收缩的机制可能与长期腹膜透析患者的溶质通透性改变有关。在本实验条件下，我们发现了转化生长因子-βmRNA的表达，可以反映整合素的表达。结论：在体外三维腹膜模型中，不仅可以理解溶质通过腹膜的渗透机制，而且可以理解纤维化过程的机制。

The hybrid-type peritoneal membrane model is a promising tool to understand not only solute and water transport via the peritoneum but also fibrotic process due to endogenous and/or exogenous stimuli. There exist, however, technological requirements to be overcome such as a cloning of peritoneal fibroblasts and their 3D cultures in the extra cellular matrices.In this study we have surveyed the 3 dimensional culture of the peritoneal resident cells to reconstruct the peritoneum in vitro and found few research activitie on it except for some studies of host defense mechanizms using cultured peritoneal cells. Modifying their techniques we established a time elapsed-differential subculture (t-DSC), regulating contact time to the culture plastics, to separate fibroblasts from sub-confluent primary cultured peritoneal cells. Establishing PRFB (1*10^5/ml) in collagen gels composed of increasing concentrations of collagen resulted in an inverse correlation with gel contraction while increasing the PRFB cell density within the collagen gels (2mg/ml) resulted in a direct correlation with collagen gel contraction. Addition of the transcriptional inhibitor actinomycin D to 3D-RPFB cultures resulted in a dose dependent inhibition of gel contraction. Gel contraction, however, was not inhibited following the addition of the translation inhibitor, puromycin and similarly no significant effect was seen following incubation with 2,2'-bipyridyl. From these results, it could be hypothesized that mechanisma of collagen gel contraction may relate to change in solute permeability of the peritoneal patients in a long-term peritoneal dialysis. We found an expression of TGF-beta mRNA expresssion in this experimental setting which could indicate an expression of integrin.It is concluded that in vittro 3D peritoneal model is promissing method to understand mechanism of not only solute permeability through the peritoneum but also fibrotic process.