Propranolol metabolism by isolated hepatocytes from normal and cirrhotic rat livers: the effect of albumin

1990 ◽  
Vol 68 (6) ◽  
pp. 657-662 ◽  
Author(s):  
Louise Gariepy ◽  
Daphna Fenyves ◽  
Jean-Luc Petit ◽  
Ginette Raymond ◽  
Jean-Pierre Villeneuve
Keyword(s):  
Free Fraction ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Hepatic Uptake ◽  
Albumin Concentration ◽  
Isolated Rat Hepatocytes ◽  
Subsequent Elimination ◽  
Rat Livers ◽  
Mediated Catalysis ◽  
Isolated Rat

Several recent reports have shown that the hepatic uptake and subsequent elimination of some substrates is faster in the presence of albumin than in its absence, as if some of the substrate bound to albumin was also available for uptake. In the present study, we examined the effect of albumin on the clearance of propranolol by isolated rat hepatocyte suspensions. The clearance of total drug decreased progressively as albumin concentration increased. There was also a progressive decrease in the free fraction of propranolol and the net result was an increase in the clearance of unbound drug (+50% at 40 g/L albumin). This increase was not due to an oncotic pressure effect of albumin, nor to the presence of fatty acids bound to albumin. The clearance of propranolol by isolated hepatocytes from cirrhotic rats was decreased compared with controls (−50%), and albumin also increased propranolol free clearance, albeit to a lesser extent than in control animals. Our results indicate that albumin facilitates the elimination of propranolol by hepatocytes, possibly because of surface-mediated catalysis of the albumin–propranolol complexes.Key words: propranolol clearance, albumin, isolated rat hepatocytes, cirrhosis.

Uptake of unconjugated bilirubin by isolated rat hepatocytes

1979 ◽  
Vol 236 (1) ◽  
pp. C9-C14 ◽  
Author(s):  
T. Iga ◽  
D. L. Eaton ◽  
C. D. Klaassen
Keyword(s):  
Lipid Membrane ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Hepatic Uptake ◽  
Unconjugated Bilirubin ◽  
Metabolic Inhibitors ◽  
Isolated Rat Hepatocytes ◽  
Intracellular Binding ◽  
High Degree ◽  
Isolated Rat

The mechanism responsible for the hepatic uptake of unconjugated bilirubin was examined in isolated rat hepatocytes from control and phenobartital-pretreated rats. The uptake was extremely rapid and the equilibrium between cell and medium was attained within 60 s with a 100-fold higher concentration in the cell than the medium. The initial velocity of uptake (Vo) exhibited a linear relationship to the bilirubin concentration in the medium. Pretreatment of cells with various metabolic inhibitors had no effect on the uptake of unconjugated bilirubin. Ouabain did significantly decrease Vo, but replacement of sodium ion with choline or lithium had no effect on bilirubin uptake. The organic acids sulfobromophthalein (112 muM) and taurocholic acid (50 (muM) and two steroidal compounds, diethylstilbestrol (50 muM) and spironolactone (50 muM), had no effect on the uptake of bilirubin. It is suggested that bilirubin gains access to the hepatocyte interior by passive diffusion into and through the lipid membrane and that intracellular binding may explain the high degree of bilirubin accumulation associated with the isolated hepatocytes.

scholarly journals Sugar uptake by fluid-phase pinocytosis and diffusion in isolated rat hepatocytes

1987 ◽  
Vol 243 (3) ◽  
pp. 655-660 ◽  
Author(s):  
P B Gordon ◽  
H Høyvik ◽  
P O Seglen
Keyword(s):  
Plasma Membrane ◽  
Fluid Phase ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Free Sugar ◽  
Sugar Uptake ◽  
Isolated Rat Hepatocytes ◽  
Fluid Phase Endocytosis ◽  
And Diffusion ◽  
Isolated Rat

Measurements of sugar pinocytosis (fluid-phase endocytosis of radiolabelled sucrose, lactose and raffinose) in freshly isolated rat hepatocytes are disturbed by sugar diffusing into the cells through plasma-membrane blebs. Non-pinocytic entry may be even more pronounced at 0 degrees C, and is a major contributor to ‘background’ radioactivity. By electrodisruption of the plasma membrane, a distinction can be made between pinocytotically sequestered sugar and free sugar that has entered the cytosol by diffusion. Pinocytosis proceeds at a rate of 2%/h (relative to the intracellular fluid volume), whereas the rate of sucrose entry by diffusion is more than twice as high. Three pinocytotic compartments are distinguishable in isolated hepatocytes: (1) a rapidly recycling compartment, which is completely destroyed by electrodisruption, and which may represent pinocytic channels continuous with the plasma membrane; (2) a non-recycling (or very slowly recycling) electrodisruption-resistant compartment, which allows accumulation of the lysosomally hydrolysable sugar lactose, and which therefore must represent non-lysosomal vacuoles (endosomes?); (3) a lysosomal compartment (non-recycling, electrodisruption-resistant), which accumulates raffinose and sucrose, but which hydrolyses lactose. The last two compartments can be partially resolved in metrizamide/sucrose density gradients by the use of different sugar probes.

scholarly journals Homocysteine enhances the inhibitory effect of extracellular adenosine on the synthesis of proteins in isolated rat hepatocytes

1995 ◽  
Vol 310 (3) ◽  
pp. 893-896 ◽  
Author(s):  
S Tinton ◽  
P Buc-Calderon
Keyword(s):  
Protein Synthesis ◽  
Kinase Inhibitor ◽  
Adenine Nucleotides ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Adenosine Kinase ◽  
Isolated Rat Hepatocytes ◽  
Extracellular Adenosine ◽  
Inhibition Of Protein Synthesis ◽  
Isolated Rat

Previous work has shown that extracellular adenosine inhibits the incorporation of radiolabelled leucine into proteins in isolated rat hepatocytes [Tinton, Lefebvre, Cousin and Buc Calderon (1993) Biochim. Biophys. Acta 1176, 1-6]. In this study, we investigated whether its metabolism into adenine nucleotides, inosine or S-adenosylhomocysteine (AdoHcy) is required to induce such an impairment. Incubation of isolated hepatocytes in the presence of adenosine at 0.5 or 1 mM reduces the synthesis of proteins by about 45% after 120 min of incubation. Such an inhibition occurred without cell lysis and was not modified by adding the adenosine kinase inhibitor 5-iodotubercidin (15 microM) or the adenosine deaminase inhibitor coformycin (0.1 microM). It is therefore unlikely that the anabolic and catabolic pathways of adenosine are involved in the inhibition of protein synthesis. Adenosine (1 mM) increased the level of AdoHcy and S-adenosylmethionine by 20- and 5-fold respectively after 60 min of incubation and reduced the methylation index. These events as well as the inhibition of protein synthesis were strongly enhanced in the presence of L-homocysteine (2 mM). It is therefore concluded that the metabolism of adenosine into AdoHcy, which is known to be a potent inhibitor of cellular methylation reactions, may play an important role in the control of translation.

Role of extracellular Ca2+ in hepatic bile formation and taurocholate transport

1985 ◽  
Vol 249 (6) ◽  
pp. G711-G718 ◽  
Author(s):  
M. S. Anwer ◽  
L. M. Clayton
Keyword(s):  
Control Period ◽  
Rat Hepatocytes ◽  
Experimental Period ◽  
Isolated Hepatocytes ◽  
Hepatic Uptake ◽  
Hepatic Bile ◽  
Bile Formation ◽  
Isolated Rat Hepatocytes ◽  
Diffusional Permeability

The role of extracellular Ca2+ in hepatic bile formation, biliary membrane permeability, and taurocholate (TC) transport was studied in isolated perfused rat livers and in isolated rat hepatocytes to determine the functional importance of paracellular permeability in biliary bile acid excretion. Each liver was perfused for 1 h with perfusate containing 1.3 mM Ca2+ (control period) followed by another hour with 1.3, 0.5, 0.1, 0.05, 0.03, or 0.01 mM Ca2+ (experimental period). Basal bile flow and biliary excretion of added TC declined significantly only at and below 0.05 mM perfusate Ca2+ and was associated with an increase in bile-to-perfusate concentration ratio of [3H]inulin (B/P inulin ratio). A twofold increase in the diffusional permeability coefficient at 0.05 mM and a sixfold increase at 0.03 and 0.01 mM perfusate Ca2+ could explain the increased in B/P inulin ratios. Time-dependent increases in cell-to-medium concentration ratios of inulin were less in the absence than in the presence of Ca2+. Hepatic uptake rates of TC determined in isolated hepatocytes and from perfusate disappearance of added TC and efflux rates of TC from preloaded hepatocytes were not significantly affected by Ca2+ removal. It is possible that the observed decline in biliary TC excretion at low perfusate Ca2+ is due to regurgitation of secreted TC back into the perfusate followed by reuptake. This was supported by an accumulation of perfusate radioactivity when TC uptake inhibitors (furosemide and bumetanide) were added to the perfusate (0.03 mM Ca2+) 60 min after the addition of [14C]TC.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals N-alkylation of exogenous haem analogues caused by drugs in isolated hepatocytes. Structural isomerism and chirality of the resulting porphyrins

1986 ◽  
Vol 238 (1) ◽  
pp. 263-268 ◽  
Author(s):  
F De Matteis ◽  
C Harvey ◽  
S R Martin
Keyword(s):  
Direct Evidence ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Isomeric Composition ◽  
Isolated Rat Hepatocytes ◽  
Structural Isomerism ◽  
Cytochrome P 450 ◽  
Isolated Rat ◽  
Suicide Substrates

Isolated rat hepatocytes incubated with two suicide substrates of cytochrome P-450, 2-allyl-2-isopropylacetamide and 3,5-diethoxycarbonyl-4-ethyl-1,4-dihydro-2,6-dimethylpyridine(4-ethyl-DD C), convert exogenous mesohaem and deuterohaem into N-alkylated mesoporphyrins and deuteroporphyrins respectively. The N-alkylated mesoporphyrins can be separated by h.p.l.c. from the corresponding N-alkylated protoporphyrins originating from endogenous haem; in this way the contribution of both endogenous and exogenous pools of haem can be studied in the same experiment. N-Alkylated mesoporphyrin exhibits chiral properties, and its isomeric composition and/or amount are dependent on the particular cytochrome P-450 enzyme predominating in the cell. These findings provide additional and more direct evidence that exchangeable haem is taken up by cytochrome P-450 before being N-alkylated.

Effect of Bile Acids on Calcium Efflux from Isolated Rat Hepatocytes and Perfused Rat Livers

1989 ◽  
Vol 191 (2) ◽  
pp. 147-152 ◽  
Author(s):  
M. S. Anwer ◽  
J. M. Little ◽  
D. G. Oelberg ◽  
P. Zimniak ◽  
R. Lester
Keyword(s):  
Bile Acids ◽  
Rat Hepatocytes ◽  
Calcium Efflux ◽  
Isolated Rat Hepatocytes ◽  
Rat Livers ◽  
Isolated Rat

scholarly journals Insulin stimulates synthesis of soluble proteins in isolated rat hepatocytes

1980 ◽  
Vol 190 (3) ◽  
pp. 615-619 ◽  
Author(s):  
R L Clark ◽  
R J Hansen
Keyword(s):  
Protein Synthesis ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Cellular Protein ◽  
Soluble Proteins ◽  
Leucine Incorporation ◽  
Isolated Rat Hepatocytes ◽  
Hepatic Protein Synthesis ◽  
Isolated Rat ◽  
Stimulation Of

The incorporation of [3H]leucine into soluble cellular protein was measured in isolated hepatocytes at extracellular leucine concentrations ranging from 0.15 to 20.0 mM. Insulin caused a 12—15% stimulation of [3H]leucine incorporation in the presence of high extracellular leucine concentrations. It is concluded that insulin causes a small but significant increase in the rate of hepatic protein synthesis.

Role of bicarbonate in biliary excretion of diisothiocyanostilbene disulfonate

1988 ◽  
Vol 255 (6) ◽  
pp. G713-G722
Author(s):  
M. S. Anwer ◽  
K. Nolan ◽  
W. G. Hardison
Keyword(s):  
Biliary Excretion ◽  
Rat Hepatocytes ◽  
Hepatic Uptake ◽  
Disulfonic Acid ◽  
Base Line ◽  
Hepatic Transport ◽  
Isolated Rat Hepatocytes ◽  
Canalicular Membrane ◽  
Major Mechanism ◽  
Rat Livers

Hepatic transport of 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) was studied in isolated perfused rat livers and in isolated rat hepatocytes to determine if DIDS-induced decrease in biliary HCO3- excretion is due to a DIDS-HCO3- exchange and/or due to inhibition of Cl(-)-HCO3- exchange. In isolated perfused rat livers, DIDS reversibly decreased biliary HCO3- concentration and excretion. The changes in biliary HCO3- concentration were inversely related to biliary DIDS concentration. DIDS was concentrated in bile, indicating active hepatic transport. Replacement of perfusate HCO3- with equimolar dimethyloxazolidinedione (DMO) or tricine decreased biliary excretion, but not hepatic uptake, of DIDS. Biliary excretion of DIDS was also associated with a decrease in bile pH, and this decrease in pH was greater in the presence of HCO3-. HCO3-, but not DMO or tricine, stimulated DIDS efflux from preloaded hepatocytes. DIDS efflux was also temperature dependent and increased with increasing extracellular pH. Collectively, these results are consistent with the presence of a DIDS-HCO3- (OH-) exchange mechanism at the canalicular membrane. HCO3(-)-dependent Cl- uptake in hepatocytes was competitively inhibited by DIDS (Ki = 0.24 mM), confirming the presence of DIDS-inhibitable Cl(-)-HCO3- exchange. However, the ability of DIDS to decrease biliary HCO3- excretion persisted when perfusate Cl- was replaced by isethionate. Moreover, biliary HCO3- concentration returned to base line despite the presence of 2-6 mM DIDS in bile. Thus it seems unlikely that the inhibition of Cl(-)-HCO3- exchange by DIDS is a major mechanism of inhibition of HCO3- excretion. We, therefore, conclude that a DIDS-HCO3- (OH-) exchange at the canalicular membrane is the most likely explanation for the observed decrease in biliary HCO3- excretion.

scholarly journals Glutathione analogues as novel inhibitors of rat and human glutathione S-transferase isoenzymes, as well as of glutathione conjugation in isolated rat hepatocytes and in the rat in vivo

1995 ◽  
Vol 308 (1) ◽  
pp. 283-290 ◽  
Author(s):  
S Ouwerkerk-Mahadevan ◽  
J H van Boom ◽  
M C Dreef-Tromp ◽  
J H T M Ploemen ◽  
D J Meyer ◽  
...  
Keyword(s):  
Ethyl Ester ◽  
Rat Liver ◽  
Rat Hepatocytes ◽  
Isolated Hepatocytes ◽  
Glutathione S Transferase ◽  
Isolated Rat Hepatocytes ◽  
Glutathione S Transferases ◽  
Gamma S ◽  
Isolated Rat

Inhibitors of rat and human Alpha- and Mu-class glutathione S-transferases that effectively inhibit the glutathione (GSH) conjugation of bromosulphophthalein in the rat liver cytosolic fraction, isolated rat hepatocytes and in the rat liver in vivo have been developed. The GSH analogue (R)-5-carboxy-2-gamma-(S)-glutamylamino-N-hexylpentamide [Adang, Brussee, van der Gen and Mulder (1991) J. Biol. Chem. 266, 830-836] was used as the lead compound. To obtain more potent inhibitors, it was modified by replacement of the N-hexyl moiety by N-2-heptyl and by esterification of the 5-carboxy group with ethyl and dodecyl groups. In isolated hepatocytes, the branched N-2-heptyl derivatives were stronger inhibitors of GSH conjugation of bromosulphophthalein than the N-hexyl derivatives. The ethyl ester compounds were more efficient than the corresponding unesterified derivatives. The dodecyl ester of the N-2-heptyl analogue was the most effective inhibitor in isolated hepatocytes, but was relatively toxic in vivo. However, the corresponding ethyl ester was a potent in vivo inhibitor: GSH conjugation of bromosulphophthalein (as assessed by biliary excretion of the conjugate) was decreased by 70% after administration of a dose of 200 mumol/kg. The isoenzyme specificity of the inhibitors towards purified rat and human glutathione S-transferases was also examined. The unesterified compounds were more potent than the esterified analogues, and inhibited Alpha- and Mu-class isoenzymes of both rat and human glutathione S-transferase (Ki range 1-40 microM). Other GSH-dependent enzymes, i.e. GSH peroxidase, GSH reductase and gamma-glutamyltranspeptide, were not inhibited. Thus (R)-5-ethyloxycarbonyl-2-gamma-(S)-glutamylamino-N-2-hept ylpentamide, the in vivo inhibitor of GSH conjugation, may be useful in helping to assess the role of the Alpha and Mu classes of glutathione S-transferases in cellular biochemistry, physiology and pathology.

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