Biotransformation of trichloroethylene in collagen gel sandwich cultures of rat hepatocytes

2000 ◽  
Vol 74 (10) ◽  
pp. 587-592 ◽  
Author(s):  
Karen De Smet ◽  
Thomas Brüning ◽  
Meinolf Blaszkewicz ◽  
Hermann M. Bolt ◽  
Antoine Vercruysse ◽  
...  
Keyword(s):  
Collagen Gel ◽  
Rat Hepatocytes ◽  
Collagen Gel Sandwich

Collagen-Gel Cultures of Rat Hepatocytes: Collagen-Gel Sandwich and Immobilization Cultures

2003 ◽  
pp. 303-310 ◽  
Author(s):  
Sonja Beken ◽  
Tamara Vanhaecke ◽  
Karen De Smet ◽  
Marleen Pauwels ◽  
Antoine Vercruysse ◽  
...  
Keyword(s):  
Collagen Gel ◽  
Rat Hepatocytes ◽  
Collagen Gel Sandwich

Collagen Type I Gel Cultures of Adult Rat Hepatocytes as a Screening Induction Model for Cytochrome P450-dependent Enzymes

2001 ◽  
Vol 29 (2) ◽  
pp. 179-192 ◽  
Author(s):  
Karen De Smet ◽  
Christophe Cavin ◽  
Antoine Vercruysse ◽  
Vera Rogiers
Keyword(s):  
Cytochrome P450 ◽  
Collagen Type ◽  
Collagen Gel ◽  
Rat Hepatocytes ◽  
Collagen Type I ◽  
Type I ◽  
Albumin Secretion ◽  
Activity Measurements ◽  
Collagen Gel Sandwich

Albumin secretion, expression of cytochrome P450-dependent mono-oxygenases (CYPs) and their inducibility by well-known inducers were evaluated during 1 week in collagen type I gel sandwich and immobilisation cultures of adult primary rat hepatocytes. Albumin secretion increased during culture and, following an initial decrease, CYP biotransformation activities remained stable for at least 7 days. Better preservation results were observed in the collagen gel sandwich culture than in the immobilisation model. The inducibility of CYPs by β-naphthoflavone (β-NF), 3-methylcholanthrene (3-MC), phenobarbital (PB) and dexamethasone (DEX) was studied in both collagen gel hepatocyte cultures. Exposure of the cells to either 5μM 3-MC or 25μM β-NF, added to the culture medium, resulted in strong increases of CYP1A1/2 activity in both culture models. Treatment with PB (3.2mM) resulted in an increase in the CYP2B activity and a higher hydroxylation of testosterone in the 16α-position (CYP2B1/2 and CYP2C11), the 7α-position (CYP2A1/2), and the 6β-position (CYP3A1). DEX (10μM) markedly increased testosterone 6β- and 7α-hydroxylation. Expression and induction experiments on CYP proteins exposed to these molecules confirmed the results of the CYP activity measurements. The patterns of CYP induction in collagen gel cultures of rat hepatocytes were similar to those observed in vivo. Consequently, collagen gel cultures and, more specifically, collagen gel sandwich cultures seem to be suitable as in vitro models for evaluating xenobiotics as potential inducers of CYP-enzymes.

Collagen gel sandwich and immobilization cultures of rat hepatocytes: Problems encountered in expressing glutathione S-transferase activities

1997 ◽  
Vol 11 (6) ◽  
pp. 741-752 ◽  
Author(s):  
S. Beken ◽  
M. Pauwels ◽  
S. Pahernik ◽  
H.-G. Koebe ◽  
A. Vercruysse ◽  
...  
Keyword(s):  
Collagen Gel ◽  
Rat Hepatocytes ◽  
Glutathione S Transferase ◽  
Collagen Gel Sandwich

scholarly journals Increase of l-serine dehydratase activity under gluconeogenic conditions in adult-rat hepatocytes cultured on collagen gel/nylon mesh

1981 ◽  
Vol 198 (3) ◽  
pp. 499-504 ◽  
Author(s):  
W W N Mak ◽  
H C Pitot
Keyword(s):  
Enzyme Activity ◽  
Hormonal Regulation ◽  
Collagen Gel ◽  
Glucose Production ◽  
Rat Hepatocytes ◽  
Cultured Hepatocytes ◽  
Adult Rat ◽  
Nylon Mesh ◽  
Serine Dehydratase ◽  
Dehydratase Activity

Hormonal regulation of L-serine dehydratase [L-serine hydro-lyase (deaminating), EC 4.2.1.13] was studied in primary cultures of adult-rat hepatocytes. The hepatocytes were isolated by collagenase perfusion and maintained in culture on collagen-gel/nylon-mesh substrata. L-Serine dehydratase activity was measured with [14C]threonine as substrate. The enzyme activity in hepatocytes of normal adult rats was low and declined rapidly in culture in L-15 medium containing 0.1 micro M-insulin and even more in the presence of glucose. L-Serine dehydratase activity in hepatocytes of rats with streptozotocin-induced diabetes was initially 20-fold higher than that of normal rats, but fell rapidly to a low value by 4 days in culture. Hormonal regulation of the enzyme activity was manifested by treatment of the cultured hepatocytes with insulin (0.1 micro M), glucagon (0.3 micro M), dexamethasone (10 micro M) and combinations of these hormones. Either glucagon or dexamethasone in the absence of insulin enhanced the activity of L-serine dehydratase, but failed to do so in the presence of insulin. Treatment with both hormones resulted in a 2-3-fold increase in enzyme activity in culture on days 3 and 4. Under conditions in which the enzyme activity was enhanced, glucose production by the cultured hepatocytes was concomitantly increased. Glucose production resulted in part from gluconeogenesis from pyruvate and not entirely from glycogenolysis. The gluconeogenic conditions of culture resulted in a decrease in cellular lipids in the cultured hepatocytes, as evidenced by ultrastructural studies.

scholarly journals Hepatocytes in collagen sandwich: evidence for transcriptional and translational regulation.

1992 ◽  
Vol 116 (4) ◽  
pp. 1043-1053 ◽  
Author(s):  
J C Dunn ◽  
R G Tompkins ◽  
M L Yarmush
Keyword(s):  
Extracellular Matrix ◽  
Transcriptional Control ◽  
Collagen Gel ◽  
Single Layer ◽  
Mrna Translation ◽  
Normal Liver ◽  
Rat Hepatocytes ◽  
Albumin Production ◽  
Albumin Mrna ◽  
Gel System

The influence of extracellular matrix configuration on the tissue-specific function of cultured hepatocytes was investigated. Adult rat hepatocytes sandwiched between two layers of collagen gel were compared to cells cultured on a single layer of collagen gel for differences in the total RNA content, the level of albumin-specific mRNA, the rate of albumin gene transcription, and the rate of albumin mRNA translation. Adult hepatocytes in the sandwich system maintained the level of albumin mRNA similar to that found in the normal liver for at least six weeks, whereas the level of albumin mRNA declined rapidly in the single gel system. After one week of culture, hepatocytes in the single gel system could be induced to recover the high level of albumin mRNA and albumin production when a second layer of collagen gel was overlaid at that time. Furthermore, sandwiched hepatocytes maintained significantly higher transcriptional activity compared to cells in the single gel system. In addition to transcriptional control, the ultimate rate of albumin production was shown to depend on the rate of translation, which increased with culture time and reached a plateau in one to two weeks. This increase in translational activity over time in culture was observed in both the sandwich and the single gel systems and, thus, appeared to be independent of the configuration of extracellular matrix.

scholarly journals Hemodynamic flow improves rat hepatocyte morphology, function, and metabolic activity in vitro

2013 ◽  
Vol 304 (11) ◽  
pp. C1053-C1063 ◽  
Author(s):  
A. Dash ◽  
M. B. Simmers ◽  
T. G. Deering ◽  
D. J. Berry ◽  
R. E. Feaver ◽  
...  
Keyword(s):  
Collagen Gel ◽  
Rat Hepatocytes ◽  
Fold Increase ◽  
Multidrug Resistant ◽  
Metabolic Phenotype ◽  
Metabolic Function ◽  
Transport Parameters ◽  
Differentiation Markers

In vitro primary hepatocyte systems typically elicit drug induction and toxicity responses at concentrations much higher than corresponding in vivo or clinical plasma Cmax levels, contributing to poor in vitro-in vivo correlations. This may be partly due to the absence of physiological parameters that maintain metabolic phenotype in vivo. We hypothesized that restoring hemodynamics and media transport would improve hepatocyte architecture and metabolic function in vitro compared with nonflow cultures. Rat hepatocytes were cultured for 2 wk either in nonflow collagen gel sandwiches with 48-h media changes or under controlled hemodynamics mimicking sinusoidal circulation within a perfused Transwell device. Phenotypic, functional, and metabolic parameters were assessed at multiple times. Hepatocytes in the devices exhibited polarized morphology, retention of differentiation markers [E-cadherin and hepatocyte nuclear factor-4α (HNF-4α)], the canalicular transporter [multidrug-resistant protein-2 (Mrp-2)], and significantly higher levels of liver function compared with nonflow cultures over 2 wk (albumin ∼4-fold and urea ∼5-fold). Gene expression of cytochrome P450 (CYP) enzymes was significantly higher (fold increase over nonflow: CYP1A1: 53.5 ± 10.3; CYP1A2: 64.0 ± 15.1; CYP2B1: 15.2 ± 2.9; CYP2B2: 2.7 ± 0.8; CYP3A2: 4.0 ± 1.4) and translated to significantly higher basal enzyme activity (device vs. nonflow: CYP1A: 6.26 ± 2.41 vs. 0.42 ± 0.015; CYP1B: 3.47 ± 1.66 vs. 0.4 ± 0.09; CYP3A: 11.65 ± 4.70 vs. 2.43 ± 0.56) while retaining inducibility by 3-methylcholanthrene and dexamethasone (fold increase over DMSO: CYP1A = 27.33 and CYP3A = 4.94). These responses were observed at concentrations closer to plasma levels documented in vivo in rats. The retention of in vivo-like hepatocyte phenotype and metabolic function coupled with drug response at more physiological concentrations emphasizes the importance of restoring in vivo physiological transport parameters in vitro.

Effect of Epidermal Growth Factor in Collagen Gel Cultures of Rat Hepatocytes

1999 ◽  
Vol 13 (4-5) ◽  
pp. 579-585 ◽  
Author(s):  
K. De Smet ◽  
S. Beken ◽  
M. Depreter ◽  
F. Roels ◽  
A. Vercruysse ◽  
...  
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Collagen Gel ◽  
Rat Hepatocytes ◽  
Epidermal Growth
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