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 研究人员开发了一种BAL， 在无肝兔模型中支持。 为了扩大人类的 使用，改善肝细胞功能和增加活细胞密度， 必需的. 我们的BAL是一种三室中空纤维生物反应器， 两个腔内隔室，一个含有凝胶包埋大鼠 肝细胞和一个包含围绕肝细胞凝胶的培养基; 来自肝功能衰竭的患者或动物的血液在肝脏中循环。 （三）毛细血管外室。 对胶原蛋白的初步研究- 静态培养的肝细胞包埋率超过50%， 第8天的存活率，细胞与胆小管聚集 形成，尿素生成，氨基酸摄取，白蛋白合成， 结合胆汁盐和代谢利多卡因的能力。 测试时 在急性无肝兔实验中，BAL与 凝血或溶血;它维持正常的葡萄糖稳态， 血浆氨基酸正常化;尿素水平稳定，利多卡因 被生物转化了 没有兔抗体到达内腔凝胶- 截留的大鼠细胞，反映了生物反应器膜孔径。 当前BAL应支持活细胞密度增加3倍。 我们假设肝细胞功能和活细胞密度可以 通过使用细胞外基质、生长因子、药物 诱导和细胞-细胞共培养;此外，增强的BAL将 在无肝兔中表现出改善的生化功能， 延长D-半乳糖胺肝损伤模型中的存活。 初步 静态培养中胶原包埋肝细胞的数据显示， 改善的生存力、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， 添加到系统中。 这些观察将在修改后的 线性优化协议，以确定最佳组合， 对以下肝细胞功能标志物的效应物： 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代谢。 肝细胞活力和功能的特异性效应物包括牛 肠硫酸肝素，猪肝糖胺聚糖提取物， 基质胶，重组肝细胞生长因子，激素增强培养基， 脂肪细胞共培养，以及苯巴比妥、氯贝酸的药物诱导 酸或β-萘酮。 影响最大的效应器 将用D-半乳糖胺评价活力和功能 肝损伤模型的随机前瞻性研究。 这个过程 应该能够大幅减少（50 - 70%）BAL规模扩大， 更大的动物实验以及最终的人体实验。

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.