Use of primary human liver cells originating from discarded grafts in a bioreactor for liver support therapy and the prospects of culturing adult liver stem cells in bioreactors: a morphologic study.

INTRODUCTION: The development of a bioreactor providing a three-dimensional network of interwoven capillary membranes with integrated oxygenation and decentralized mass exchange enables the culture of primary human liver cells from discarded donor organs for extracorporeal liver support. METHODS: Primary liver cells were isolated from 54 discarded organs (donor age 56.7+/-13.2 years). Between 2.8x10(10) and 6.4x10(10) parenchymal cells (PC) were cocultured with nonparenchymal cells (NPC) of the same organ in bioreactors (n=36). The metabolic activity of the cells was regularly determined during culture. The cell morphology and ultrastructure were investigated after culture periods of 1 to 5 weeks. RESULTS: Cell metabolism was maintained over at least 3 weeks after a phase of adaptation lasting 2 to 3 days. Through the use of transmission electron microscopy and immunohistochemistry, it was demonstrated that PC and NPC spontaneously formed tissue-like structures. Vascular cavities (CD 31 immunoreactivity [IR]) and bile duct-like channels (CK 19 IR), both exhibiting proliferation activity (Ki-67 IR), were regularly distributed. Some of the bile duct-like channels showed similarities to the Canals of Hering found in the natural liver. Cells expressing morphologic and antigenic characteristics of adult liver stem cells (CD 34 IR and c-kit IR) and areas with cells that showed both hepatocyte and biliary characteristics were detected. CONCLUSION: The results show that primary human liver cells obtained from discarded donor organs recover and can be maintained in bioreactors for clinical liver support therapy. In addition, initial observations on adult liver stem-cell culture in bioreactors are presented.

Evaluation of primary human liver cells in bioreactor cultures for extracorporeal liver support on the basis of urea production.

Primary human liver cells from donor organs unsuitable for transplantation were cultivated in bioreactors developed for extracorporeal liver support. Because each system contains cells originating from an individual organ, each bioreactor culture must be individually characterized. The objective of this study was to identify suitable decisive parameters for the evaluation of cell culture performance. We analyzed the data from 47 bioreactor cultures containing 437 +/- 110 g of cells. Choosing urea production as the decisive parameter, the bioreactor cultures were divided into high-performance (daily urea production > or = 110 mg per bioreactor between culture days 3 and 14) and low-performance cultures. Comparing the mean courses of the groups revealed a significant distinction in most other investigated biochemical parameters. In conclusion, urea production seems to be an appropriate parameter for evaluating the performance of liver cell cultures in bioreactors because it corresponds to all other evaluated parameters of cell function.