IMPROVED BIOARTIFICIAL LIVER FUNCTION IN HEPATIC FAILURE

生物人工肝(BAL)已被提出作为一种支持装置 在可逆性肝损伤期间或作为肝脏中的桥梁技术 移植。调查人员已经开发出一种BAL，可以提供 在无肝兔模型中的支持。为了扩大对人类的影响 使用，改善肝细胞功能和增加活细胞密度是 必填项。我们的BAL是一个三隔室中空纤维生物反应器 两个腔内隔室，其中一个包含凝胶包裹的大鼠 肝细胞和一个含有肝细胞凝胶周围的培养液； 肝功能衰竭患者或动物的血液在 (三)毛细血管外腔。胶原蛋白的初步研究- 静态培养中包埋的肝细胞显示50%以上 8天后存活，细胞与胆小管聚集 形成、尿失禁、氨基酸摄取、白蛋白合成和 结合胆盐和代谢利多卡因的能力。当经过测试时 在急性无肝兔实验中，BAL与 凝血或溶血；它维持正常的血糖平衡和 血浆氨基酸正常化；尿素水平稳定，利多卡因 被生物改造了。没有兔抗体到达内腔凝胶- 包埋的大鼠细胞，反映了生物反应器膜的孔径大小。 目前的BAL应该支持将可存活细胞密度增加3倍。 我们假设肝细胞功能和活细胞密度 通过使用细胞外基质、生长因子、药物来改善 诱导和细胞-细胞共培养；此外，增强的BAL将 证明无肝兔的生化功能有所改善，并 延长D-氨基半乳糖肝损伤模型的存活时间。初步 静态培养中胶原蛋白包裹的肝细胞的数据显示 提高了生存能力，白蛋白合成，胆汁酸结合和 利多卡因代谢时特定的基质因子或生长因子 已添加到系统中。这些观察结果将在修改后的 确定最佳组合的线性优化协议 对下列肝细胞功能标志物的影响：细胞 活力、细胞聚集、白蛋白合成、胆汁酸和胆红素 结合，以及4-甲基伞形酮和利多卡因的代谢。 肝细胞活性和功能的特异性效应物包括牛 肠用硫酸肝素，猪肝糖胺多糖提取物， Matrigel，重组肝细胞生长因子，激素增强型培养基 脂肪细胞与苯巴比妥共培养及药物诱导 酸，或β-萘黄酮类。对以下对象影响最大的效应器 活性和功能将用D-氨基半乳糖进行评估 一项随机前瞻性研究中的肝损伤模型。这一过程 应实现大幅(50%-70%)的BAL纵向扩展，即 对于更大的动物以及最终的人体测试来说都是必要的。

The bioartificial liver (BAL) has been proposed as a support device during reversible hepatic injury or as a bridge technology in liver transplantation. The investigators have developed a BAL that provides support in an anhepatic rabbit model. In order to scale-up for human use, improved liver cell function and increased viable cell density are required. Our BAL is a three compartment hollow fiber bioreactor with two intraluminal compartments, one containing gel-entrapped rat hepatocytes and one containing media surrounding the hepatocyte gel; blood from a patient or animal in liver failure circulates in the (third) extracapillary compartment. Initial studies of the collagen- entrapped hepatocytes in static culture demonstrated more than 50% viability at eight days, cell aggregation with bile canaliculus formation, ureagenesis, amino acid uptake, albumin synthesis, and the ability to conjugate bile salts and metabolize lidocaine. When tested in acute anhepatic rabbit experiments, the BAL was not associated with clotting or hemolysis; it maintained normal glucose homeostasis and normalized plasma amino acids; urea levels were stabilized and lidocaine was biotransformed. No rabbit antibody reached the inner lumen gel- entrapped rat cells, a reflection of the bioreactor membrane pore size. The current BAL should support a 3-fold increase in viable cell density. We hypothesize that hepatocyte function and viable cell density can be improved through the use of extracellular matrix, growth factors, drug induction and cell-cell coculture; in addition, the enhanced BAL will demonstrate improved biochemical function in the anhepatic rabbit and prolonged survival in a D-galactosamine liver injury model. Preliminary data with the collagen-entrapped hepatocytes in static culture showed improved viability, albumin synthesis, bile acid conjugation and lidocaine metabolism when specific matrix factors or growth factors were added to the system. These observations will be extended in a modified linear optimization protocol to identify the best combination of effectors on the following markers of hepatocyte function: cell viability, cell aggregation, albumin synthesis, bile acid and bilirubin conjugation, and 4-methylumbelliferone and lidocaine metabolism. Specific effectors of hepatocyte viability and function include bovine intestinal heparin sulfate, pig liver glycosaminoglycan extract, Matrigel, recombinant hepatocyte growth factor, hormone enhanced media, lipocyte coculture, and drug induction with phenobarbital, clofibric acid, or beta-naphthoflavone. Effectors with the greatest influence on viability and function will be evaluated with the D-galactosamine hepatic injury model in a randomized, prospective study. This process should enable a substantial (50-70%) reduction in BAL scale-up that is necessary for larger animal and, eventually, human testing.