Technique for the kinetic characterization of the metabolic reactions of hepatocytes in adhesion culture.
In this paper, we report on the development of a technique for the kinetic characterization of the metabolic reactions of liver cells in adhesion culture. The technique is based on the use of a continuous-flow bioreactor which is designed and operated in such a way as to ensure a uniform distribution of metabolite at the cell site: hence, the metabolite concentration at the surface of cells cultured in adhesion at the bottom of the bioreactor equals that in the stream leaving the bioreactor. Under steady conditions, the rate of a given cell reaction is directly estimated from the metabolite concentration difference in the streams entering and leaving the bioreactor and can be correctly related to the actual concentration at the cell surface. Such a technique was used for a preliminary investigation of the kinetics of ammonia elimination, urea synthesis, and phenolsulfonphthalein (PSP) elimination by primary rat hepatocytes cultured in adhesion on collagen, with respect to ammonia and PSP concentration, respectively. The rate at which the hepatocytes eliminated ammonia increased with increasing ammonia concentrations according to a Michaelis-Menten kinetics. The hepatocytes synthesized urea also in the absence of ammonia in the medium: as ammonia concentration increased, the cells synthesized urea at a rate that increased according to a saturation kinetics. In the concentration range investigated, the hepatocytes eliminated PSP at a rate that increased linearly with the actual PSP concentration in the medium. Such kinetic information can be coupled to the mechanism of metabolite transport in a hybrid liver support device to yield an effective device design for the treatment of acute liver failure.
Catapano
G
G
Department of Chemical and Materials Engineering, University of Calabria, I-87036 Arcavacata di Rende (CS), Italy. catapano@pobox.unical.it
De Bartolo
L
L
D000641:Ammonia / Q000378:metabolism; D000818:Animals; D019149:Bioreactors; D002448:Cell Adhesion; D002478:Cells, Cultured; D007700:Kinetics; D008099:Liver / Q000166:cytology* / Q000378:metabolism*; D008297:Male; D010637:Phenolsulfonphthalein / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar
D000641:Ammonia; D010637:Phenolsulfonphthalein