Abstract
Bioartificial liver (BAL) support systems are currently expected to be novel therapeutic devices for the end stage of hepatic failure. Treatment with BAL systems will improve recipient conditions as a bridge to successful transplantation and will support the critical stage of the post-operational period. We developed a BAL system containing the human hepatoblastoma cell line HepG2 with the addition of an ammonia removal function by transfecting a glutamine synthetase (GS) gene. After transfection of a hamster GS gene into HepG2, the resulting GS-HepG2 cells showed 15% ammonia removal activity of porcine hepatocytes, while unmodified HepG2 cells had no such activity. The established GS-HepG2 cells were grown in a circulatory flow bioreactor and used when the cell numbers reached 3.5 – 4.1 × 109. We used pigs with ischemic liver failure to estimate the efficiency of the BAL system. The BAL treatment was started 3 hrs after the completion of total liver ischemia. The survival time of the animals treated with GS-HepG2 BAL was significantly longer than that of the cell-free control group (14.5 hrs vs. 8.5 hrs) and the group treated with BAL consisting of unmodified HepG2 (9.6 hrs). The BAL groups containing the GS-HepG2 cells had significantly fewer incidents of increased brain pressure and abnormalities of coagulation indices during the plasma exchange treatment.
Since the efficacy of BAL is regulated not only by cell capability but also by module specification, we estimated the xenobiotic clearance of our BAL system by using newly developed GS-3A4-HepG2, which was transduced with the CYP3A4 gene. The 3A4 activity reached 430 pmol/min/mg-protein in GS-3A4-HepG2, which was approximately 1.7 times that of human hepatocytes in primary culture. The lidocaine clearance of the entire BAL system was calculated to be 8.0 ml/min, while that of another BAL system with isolated porcine hepatocytes was reported to be 46 ml/min [1]. Further improvements will require control of the rate of blood circulation and the use of high-density cultures.
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References
Iwata, H., Sajiki, T., Maeda, H., Park, Y.G., Zhu, B., Satoh, S., Uesugi, T., Ikai, I., Yamaoka, Y., and Y. (1999). In Vitro evaluation of metabolic functions of a bioartificial liver. ASAIO Journal 45, 299–306.
Rozga, J. and Demetriou, A. A. (1995) Artificial liver. Evolution and future perspectives ASAIO Journal 41, 831–837.
Sussuman, N.L. and Kelly, J.H. (1993) Artificial liver: a forthcoming attraction Hepatology, 17, 1163–1164.
Enosawa, S., Suzuki, S., Kakefuda, T., and Amemiya, H (1996) Examination of 7-ethoxycoumarin deethylation and ammonia removal activities in 31 hepatocyte cell lines Cell transplant. 5, S3 9- S40.
Omasa, T., Higashiyama, K., Shioya, S., and Suga, K. (1992) Effect of lactate concentration on hybridoma culture in lactate-controlled fed-batch operation Biotechnoi Bioeng. 39,556–564.
Miyashita, T., Enosawa, S., Suzuki, S., Tamura, A., Tanaka, H., Amemiya, H., Matsumura, T„ Omasa, T., Suga, K., Aoki, T., and Koyanagi, Y. (2000) Development of a bioartificial liver with glutamine synthetase-transduced recombinant human hepatoblastoma cell line, HepG2 Transplant Proc., 32, 2355–2358.
Enosawa, S., Mukaiyama, T., Miyashita, T., Li, X.,-K., Suzuki, S., Amemiya, H., Matsumura, T., Omasa, T., and Suga, K. (2001) Application of circulatory flow bioreactor for long-term and large-scale culture of glutamine synthetase transduced CHO cells and its ammonia removal activity with an aim of development for a bioartificial liver assist system, Journal of Artificial Organ 4, 61–66.
Enosawa, S., Miyashita, T., Fujita, Y., Suzuki, S., Amemiya, H., Omasa, T., Hiramatsu, S., Suga, K., and Matsumura, T. (2001) In vitro estimation of bioartificial liver with recombinant HepG2 cells using pigs with ischemic liver failure, Cell transplant. 10, 429–433
Hoener, B.A. (1994) Predicting the hepatic clearance of xenobiotics in humans from in vitro data. Biopharmaceutics drug disposition 15, 295–304.
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© 2003 Springer Science+Business Media Dordrecht
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Omasa, T., Enosawa, S. (2003). Construction of Liver Model with Genetically Engineered Human HepG2 Cells. In: Yagasaki, K., Miura, Y., Hatori, M., Nomura, Y. (eds) Animal Cell Technology: Basic & Applied Aspects. Animal Cell Technology: Basic & Applied Aspects, vol 13. Springer, Dordrecht. https://doi.org/10.1007/978-94-017-0726-8_5
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DOI: https://doi.org/10.1007/978-94-017-0726-8_5
Publisher Name: Springer, Dordrecht
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