Porownanie aktywnosci metabolicznej bioreaktora watrobowego z perfundowana watroba

• Autorzy: Karlik W , Sztuk P. , Bakala A. , Glowala A. , Grono D. , Wiechetek M.

Warianty tytułu

EN

Comparative study of metabolic activity of liver bioreactor with perfused liver

• Języki publikacji: PL

Abstrakty

EN

The aim this study was to compare metabolic activity of a hollow fiber bioreactor with a perfused liver. A special construction of a hollow fiber bioreactor was used with freshly isolated rat hepatocytes. After isolation, hepatocytes were incubated in the bioreactor for the duration of 3 hrs in the following conditions: 100 ml medium Hanks Balances Salts Solution supplemented with 4% albumin, 2 mM ornithine, 10 mM ammonium chloride and 6 mM ethanol were used; the medium was perfused in a circulated system for 25 ml/min; samples of the medium were taken to estimate ammonia, urea, glucose, lactate, ethanol, AST and ALT activity before and after 5 min and every 15 min of perfusion time. The same experimental condition was used in the perfused rat liver system. Utilization of ammonia was different in both systems and amounted to: 8.89 and 5.23 µmol/h/g hepatocytes in the bioreactor and perfused liver, respectively. Urea production was: 2.35 and 8.22 µmol/h/g hepatocytes, respectively. The largest differences between the compared systems were observed in the glucose and lactate metabolism. The bioreactor did not release glucose and lactate during the entire time of perfusion. In contrast, perfused liver intensively released glucose (31.32 µmol/hr/g cells) and produced lactate (29.42 µmol/hr/g cells). On the other hand, there was no statistically significance difference in the rate of ethanol metabolism between both systems, which amounted to 2.55 and 2.04 umol/h/g hepatocytes in the bioreactor and perfused liver, respectively. The results indicate that a bioreactor with freshly isolated hepatocytes is not bioequivalent to a perfused liver if estimation is made on the basis of ureogenesis and/or glucose utilization. However, ethanol utilization gives evidence that the metabolic activity of the bioreactor is comparable with a perfused liver. On the basis of the obtained results it can be concluded that the difference in metabolic activity of the bioreactor and perfused liver is connected with catabolic state and disturbances of energy metabolism in freshly isolated hepatocytes. In such a condition HBSS is not the proper medium for the recovery of homeostasis. To estimate the metabolic activity of freshly isolated hepatocytes cultivated in various in vitro systems such as a marker of model usefulness it is suggested to use the activity of inducible enzymes, but not constituent enzymes.

Słowa kluczowe

PL

bioreaktory; badania modelowe; watroba perfundowana; bioreaktor watroby; modele in vitro; hepatocyty; aktywnosc metaboliczna

• Wydawca: -
• Czasopismo: Medycyna Weterynaryjna
• Rocznik: 2008
• Tom: 64
• Numer: 03
• Opis fizyczny: s.354-358,rys.,tab.,bibliogr.

Twórcy

autor

Karlik W

• Szkola Glowna Gospodarstwa Wiejskiego, ul.Ciszewskiego 8, 02-786 Warszawa

autor

Sztuk P.

autor

Bakala A.

autor

Glowala A.

autor

Grono D.

autor

Wiechetek M.

Bibliografia

• 1. Benford D. J., Hubbard S. A.: Preparation and culture of mammalian cells, [w:] Snell K., Mullock B. (red.): Biochemical Toxicology. A Practical Approach, IRL Press Ltd., Oxford 1987, 57-82.
• 2. Berry M. N., Edwards A. M., Barritt G. J.: Utilization of hepatocytes for drug studies, [w:] Burdon R. H., van Knippenberg P. H. (red.): Isolated Hepatocytes. Preparation, Properties and Applications. Elsevier, Amsterdam, New York, Oxford 1991, 179-200.
• 3. Dixit V. D.: Development of a bioartificial liver using isolated hepatocytes. Artificial Organs 1994, 18, 371-384.
• 4. Flendrich L. M. F., Soe J. S., Jorning G. G. A.: In vitro evaluation of a novel bioreactor based on an integral oxynegator and a spirally wound nonwoven polyester matrix for hepatocyte culture as small aggregates. J. Hepatol. 1997, 26, 1379-1392.
• 5. Garwacki S., Karlik W., Wiechetek W., Weryñski A.: Evaluation of metabolic activity of hepatocytes encapsulated in the hollow fiber, [w:] Capocaccia L., Merli M., Riggio O. (red.): Advances in Hepatic Encephalopathy and Metabolic Nitrogen Exchange. CRC Press, Inc., Boca Raton, Florida 1995, 495-499.
• 6. Gerlach J. G.: Development of a hybrid liver support system: a review. Int. J. Artif. Organs 1996, 19, 645-654.
• 7. Gerlach J. G., Encke J. E., Hole O. H., Muller C. M., Ryan C. R., Neuhaus P. N.: Bioreactor for a large scale hepatocyte in vitro perfusion. Transplantation 1994, 58, 984-988.
• 8. Griffin S. J., Houston J. B.: Comparison of fresh and cryopreserved rat hepatocyte suspensions for the prediction of in vitro intrinsic clearance. Drug Metabolism Disposition 2004, 32, 552-558.
• 9. Jóźwiak A., Karlik W., Weryński A., Wiechetek M.: Polisulfone membrane promotes hepatocytes adhesion in non-serum, non-hormones medium. XIth World Congress of the International Soc. Artif. Organs, Providence. Artif. Organs 1997, 21, 207.
• 10. Jóźwiak A., Karlik W., Wiechetek M., Weryński A.: Attachment and metabolic activity of hepatocytes cultivated on selected polymeric membranes. Int. J. Artif. Organs 1998, 21, 460-466.
• 11. Kamlot A. K., Rozga J. R., Watanabe F. W., Demetriou A. D.: Review: artificial liver support systems. Biotechnol. Bioeng. 1996, 50, 382-391.
• 12. Karlik W., Jóźwiak A., Weryński A., Wiechetek M.: Polisulfone membrane improves hepatocytes metabolism and reduce damaging effect of heparin on hepatocytes. XIth World Congress of the International Soc. Artif. Organs, Providence. Artif. Organs 1997, 21, 209.
• 13. Karlik W., Jóźwiak A., Wiechetek M., Weryński A.: A simple method for hepatocyte attachment in hollow fibre bioreactors. Int. J. Artif. Organs 1999, 22, 566-572.
• 14. Karlik W., Jóźwiak A., Wiechetek M., Weryński A.: Hollow fiber hepatic bioreactor as an in vitro model for metabolism. A comparative study with monolayer and suspension culture. The Wessex Conference Centre Sparhholt, Winchester, UK, 14-18 IX. Abstract Book of 10th International Workshop on In Vitro Toxicology 1998, 3, 11.
• 15. Kowalska-Pyłka H., Muzyczuk-Piekarska A.: Komórki wątroby - model w badaniach in vitro. Medycyna Wet. 2002, 58, 189-191.
• 16. Li A. P.: Primary hepatocyte culture as an in vitro toxicological system of the liver, [w:] Gad S. C. (red.): In Vitro Toxicology, Raven Press, New York 1994, 195-220.
• 17. Naritomi Y., Terashita S., Kagayama A., Sugiyama Y.: Utility of hepatocytes in predicting drug metabolism: comparision of hepatoc intrinsic clearance in rats and humans in vivo and in vitro. Drug Metabolism Disposition 2003, 31, 580-588.
• 18. Nyberg S. L. N., Shirabe K. S., Peshwa M. V. P., Sielaff T. D. S., Crotty P. L. C., Mann H. J. M., Remmel R. P. R., Payne W. D. P., Hu W. H., Cerra F. B. C.: Extracorporeal application of a gel-entrapment bioartificial liver: demonstration of drug metabolism and other biochemical functions. Cell Transplantation 1993, 2, 441-452.
• 19. Seglen P. O. S.: Preparation of isolated rat liver cells. Methods Cell Biolog. 1976, 13, 29-83.
• 20. Tyson C. A., Green C. E.: Application of liver and kidney cell system that reduce animal usage, [w:] Salem H. (red.): Animal Test Alternatives: Refinement, Reduction, Replacement. Marcel Dekker, Inc., New York 1995, 9-17.
• 21. Wiechetek M., Karlik W.: Izolowane hepatocyty: alternatywny model w badaniach toksyczności. Medycyna pracy 1996, XLVII (supl. 6), 105-114.