Primary culture of rat hepatocytes entrapped in cylindrical collagen gels: An in vitro system with application to the bioartificial liver

1992 ◽  
Vol 10 (3) ◽  
pp. 205-215 ◽  
Author(s):  
Scott L. Nyberg ◽  
Russell A. Shatford ◽  
William D. Payne ◽  
Wei-Shou Hu ◽  
Frank B. Cerra
Keyword(s):  
Primary Culture ◽  
Rat Hepatocytes ◽  
Bioartificial Liver ◽  
Collagen Gels ◽  
In Vitro System

Manipulation of the phenotype of immortalised rat hepatocytes by different culture configurations and by dimethyl sulphoxide

2000 ◽  
Vol 19 (5) ◽  
pp. 309-317 ◽  
Author(s):  
M I Grant ◽  
K Anderson ◽  
G McKay ◽  
M Wills ◽  
C Henderson ◽  
...  
Keyword(s):  
Primary Cultures ◽  
Toxicity Testing ◽  
Normal Liver ◽  
Rat Hepatocytes ◽  
In Vitro Toxicity ◽  
Collagen Gels ◽  
Testosterone Metabolism ◽  
Glutathione S Transferases ◽  
Cells Cultured

The liver-specific phenotype of immortalised rat hepato-cytes is not irretrievably lost as they age in culture but can be manipulated by modifying the culture environment. Testosterone metabolism was used to investigate the profile of cytochrome P450 isoenzymes present in two immortalised cell lines, P9 and LQC, and in primary cultures of rat hepatocytes, cultured on collagen films, gels and double gel cultures (sandwich configuration). The extent of testosterone metabolism, and the range of metabolites produced, was increased in immortalised cells by the presence of collagen as a substratum film or gel but survival was poorer and the range of metabolites was reduced in sandwich culture. In contrast, testosterone metabolism was retained in primary hepatocytes in sandwich cultures at a higher level than in collagen film or gel cultures. Expression of alpha class glutathione-S-transferases (GSTs) increased and that of GSTP1 decreased (changes which indicate a recovery of normal liver GST phenotype) when the medium of immortalised cell cultures was supplemented with dimethyl sulph-oxide (DMSO). DMSO also improved ethoxyresorufin 0-deethylation (EROD) and testosterone metabolism in immortalised cells. It also markedly inhibited prolifera-tion, DNA, RNA and protein synthesis. Maximal testosterone metabolism was observed in immortalised cells cultured on collagen gels in the presence of l1o (v/v) DMSO. Development of a protocol for treating immortalised liver cells cultured on collagen gels with DMSO to switch between proliferation and differentiation may provide a convenient system expres-sing the xenobiotic metabolising enzymes required for in vitro toxicity testing.

Effect of thyroid hormones on angiotensinogen production in the rat in vivo and in vitro

1987 ◽  
Vol 115 (2) ◽  
pp. 311-315 ◽  
Author(s):  
M. Ruiz ◽  
M. Montiel ◽  
E. Jimenez ◽  
M. Morell
Keyword(s):  
Thyroid Hormones ◽  
Time Course ◽  
Plasma Renin ◽  
Rat Hepatocytes ◽  
In Vitro System ◽  
Adult Rat ◽  
Monolayer Cultures ◽  
Thyroid Dysfunctions

ABSTRACT The influence of thyroid hormones on angiotensinogen production was studied in vitro and in vivo. In the in-vitro system, angiotensinogen production rate (APR) of monolayer cultures of rat hepatocytes in response to tri-iodothyronine (T3) and thyroxine (T4) was assayed. In the in-vivo system, plasma angiotensinogen concentration (PAC) and liver angiotensinogen content (LAC) were measured in hyper- and hypothyroid rats. In both thyroid dysfunctions, a significant decrease of PAC was found compared with that in control animals; however, LAC showed a significant increase in hyperthyroidism and a marked decrease in hypothyroidism. As PAC is dependent upon both angiotensinogen production by the liver and angiotensinogen degradation by renin, the decrease in PAC observed in hyperthyroidism could be due to an increase in plasma renin concentration, which would overcome the increased synthesis of liver angiotensinogen observed in these animals. In fact, addition of various concentrations of T4 or T3 to monolayer cultures of adult rat hepatocytes significantly enhanced APR. This increase was greater and started earlier with T3 (1196·1 ± 143·7 (s.d.) pg/mg protein per 6-h incubation; significant differences at the third hour of incubation) than with T4 (858·3 ± 88·2 pg/mg protein per 6-h incubation; significant differences at the sixth hour of incubation). In addition, a close dose–response relationship was found in the cultures supplemented with T3. The different time-course in the response elicited by T3 and T4 on APR could be a consequence of the necessary transformation of T4 into T3 to acquire biological activity. J. Endocr. (1987) 115, 311–315

Extracorporeal Application of a Gel-Entrapment, Bioartificial Liver: Demonstration of Drug Metabolism and Other Biochemical Functions

1993 ◽  
Vol 2 (6) ◽  
pp. 441-452 ◽  
Author(s):  
Scott L. Nyberg ◽  
Ken Shirabe ◽  
Madhusudan V. Peshwa ◽  
Timothy D. Sielaff ◽  
Paul L. Crotty ◽  
...  
Keyword(s):  
Plasma Concentrations ◽  
Scale Up ◽  
Rabbit Model ◽  
Small Animal ◽  
Glucose Production ◽  
Rat Hepatocytes ◽  
Bioartificial Liver ◽  
Large Animal ◽  
Gel Entrapment

Metabolic activity of a gel-entrapment, hollow fiber, bioartificial liver was evaluated in vitro and during extracorporeal hemoperfusion in an anhepatic rabbit model. The bioartificial liver contained either 100 million rat hepatocytes (n = 12), fibroblasts (n = 3), or no cells (n = 7) during hemoperfusion of anhepatic rabbits. Eight other anhepatic rabbits were studied without hemoperfusion as anhepatic controls, and three sham rabbits served as normal controls. Albumin production rates (mean ± SEM) were similar during in vitro (17.0 ± 2.8 μg/h) and extracorporeal (18.0 ± 4.0 μg/h) application of the hepatocyte bioartificial liver. Exogenous glucose requirements were reduced (p < 0.01) and euglycemia was prolonged (p < 0.001) in anhepatic rabbits treated with the hepatocyte bioartificial liver. The maximum rate of glucose production by the hepatocyte bioartificial liver ranged from 50-80 μg/h. Plasma concentrations of aromatic amino acids, proline, alanine, and ammonia were normalized in anhepatic rabbits during hepatocyte hemoperfusion. Gel-entrapped hepatocytes in the bioartificial liver performed sulfation and glucuronidation of 4-methylumbelliferone. P450 activity was demonstrated during both in vitro and extracorporeal application of the BAL device by the formation of 3-hydroxy-lidocaine, the major metabolite of lidocaine biotransformation by gel-entrapped rat hepatocytes. In summary, a gel-entrapment, bioartificial liver performed multiple hepatocyte-specific functions without adverse side effects during extracorporeal application in an anhepatic, small animal model. With its potential for short term support of acute liver failure, scale-up of the current bioartificial liver device is indicated for further investigations in large animal, preclinical trials.

scholarly journals A role of asialoglycoproteins for plasma-membrane-induced inhibition of the switching from α1 to β subtypes in adrenergic response during primary culture of rat hepatocytes

1996 ◽  
Vol 316 (3) ◽  
pp. 743-749 ◽  
Author(s):  
Yasuo KAJIYAMA ◽  
Yutaka SANAI ◽  
Michio UI
Keyword(s):  
Primary Culture ◽  
Rat Liver ◽  
Plasma Membranes ◽  
Rat Hepatocytes ◽  
Prior Treatment ◽  
Adult Rats ◽  
Adult Rat ◽  
Liver Membranes ◽  
Young Rat

Adrenergic responses of rat hepatocytes were studied by measuring Ins(1,4,5)P3 (for the response via α1-subtype receptors) and cAMP (for β-subtype response) generation during brief incubation of cells with respective agonists. Hepatocytes from young rats with an age of 1 week displayed a very high β response without a significant α1 response. The β response decreased and the α1 response increased progressively as the age increased; the response was almost exclusively via α1 receptors in hepatocytes of adult rats 9 weeks or more old. The β response developed, again at the expense of the α1 response, in hepatocytes from adult rats during the primary culture at low cell densities [(1–2.5)×104 cells/cm2]. Such ‘α1 to β subtype switching’ of adrenergic responses in vitro was totally inhibited by adding plasma membranes prepared from adult rat liver into the low-cell-density culture, but not inhibited at all by membranes from young rat liver. The inhibitory effect of adult rat liver membranes was lost when the membranes had been exposed to endoglycosidase F or β-galactosidase but was not affected by prior treatment with sialidase. On the contrary, young rat liver membranes became inhibitory to ‘α1 to β subtype switching’ after prior treatment with sialidase. Thus glycoproteins with unsialylated galactosyl termini on the surface of adult rat hepatocytes are likely to function as a determinant of the relative development of α1/β subtypes of adrenergic responses; the β response is predominant in hepatocytes in the juvenile, presumably as a result of sialylation of the galactosyl termini of the functional glycoproteins.

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