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Isolated Perfused Rat Liver (isolated + perfused_rat_liver)
Selected AbstractsUptake and Dispersion of Metformin in the Isolated Perfused Rat LiverJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 8 2000CHEN-HSI CHOU Although metformin is a widely used oral antihyperglycaemic, the exact mechanisms of its cellular uptake and action remain obscure. In this study the hepatic extraction and disposition kinetics of metformin were investigated by use of an isolated in-situ rat liver preparation. The liver was perfused in single-pass mode with protein-free Krebs bicarbonate medium at a flow rate of 20mLmin,1. During constant infusion with 1 mgL,1 metformin hydrochloride the hepatic uptake of metformin approached equilibrium within 10 min. The steady-state availability, F, determined from the ratio of outflow concentration to input concentration, was 0.99±0.02 (mean±s.d., n=4). The outflow profile of metformin resulting from a bolus injection of 25 ,g into the portal vein, had a sharp peak then a slower declining terminal phase. The mean transit time (MTT; 49.5±14.5, n = 6) and normalized variance (CV2; 4.13±0.05) of the hepatic transit times of metformin were estimated by numerical integration from the statistical moments of the outflow data. The volume of distribution of metformin in the liver (1.58±0.28 mL (g liver),1) was estimated from its MTT. The volume of distribution is greater than the water space of liver, indicating that metformin enters the hepatic aqueous space and becomes distributed among cellular components. The magnitude of CV2 for metformin is greater than for the vascular marker sucrose, suggesting that distribution of metformin into hepatic tissue is not instantaneous. In conclusion, hepatic uptake of metformin is rate-limited by a permeability barrier. Although metformin is accumulated in the liver, the organ does not extract it. [source] Hepatic Kupffer cell phagocytotic function in rats with erythrocytic-stage malariaJOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY, Issue 5 2002MICHAEL S NOBES AbstractBackground: In the erythrocytic phase of malaria, Kupffer cells show marked hypertrophy and hyperplasia and are filled with malarial pigment. However, phagocytic function in this state has not been well characterized. The aim of the present study was to use mouse Plasmodium berghei to infect rats with malaria and study the phagocytic function and morphology of Kupffer cells. Methods: We used a recirculating isolated perfused rat liver (IPRL) to quantitate Kupffer cell phagocytic clearance of radiolabeled albumin,latex over 120 min in high parasitemia (53 ± 6%; n = 7) and low parasitemia (,1%; n = 4) malaria-infected rats and littermate controls (n = 7 and n = 4, respectively). In a further group of high-parasitemic rats, perfusion was ceased after 7 min and liver radioactivity also measured. Electron microscopy was performed after perfusions. Results: In high-parasitemia malaria rats, clearance of radiolabeled latex from IPRL perfusate over 120 min was significantly (P < 0.01) faster than in controls, with a lower area under the curve (0.19 ± 0.02 vs 0.43 ± 0.07 /mL per min, respectively) and shorter half-life (t1/2k; 2.4 ± 0.6 vs 10.0 ± 2.3 min, respectively). Low-parasitemia rats were identical to controls. After 7 min perfusion in high-parasitemic rats (n = 4), total radioactivity in liver homogenates was higher than in controls (n = 4; 33.1 ± 6.2 vs 18.4 ± 1.9% of injected radiolabel; P < 0.05). Electron microscopy showed latex in Kupffer cells, more abundantly seen in high-parasitemic animals. Conclusions: Total Kupffer cell phagocytic activity of the liver is markedly increased in rats with a high parasitemic load of malarial P. berghei infection. This is presumed to reflect an upregulation of scavenger activity phagocytosing erythrocytes and their breakdown products. © 2002 Blackwell Publishing Asia Pty Ltd [source] Extrapolating in vitro metabolic interactions to isolated perfused liver: Predictions of metabolic interactions between R -bufuralol, bunitrolol, and debrisoquineJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 10 2010Sami Haddad Abstract Drug,drug interactions (DDIs) are a great concern to the selection of new drug candidates. While in vitro screening assays for DDI are a routine procedure in preclinical research, their interpretation and relevance for the in vivo situation still represent a major challenge. The objective of the present study was to develop a novel mechanistic modeling approach to quantitatively predict DDI solely based upon in vitro data. The overall strategy consisted of developing a model of the liver with physiological details on three subcompartments: the sinusoidal space, the space of Disse, and the cellular matrix. The substrate and inhibitor concentrations available to the metabolizing enzyme were modeled with respect to time and were used to relate the in vitro inhibition constant (Ki) to the in vivo situation. The development of the liver model was supported by experimental studies in a stepwise fashion: (i) characterizing the interactions between the three selected drugs (R -bufuralol (BUF), bunitrolol (BUN), and debrisoquine (DBQ)) in microsomal incubations, (ii) modeling DDI based on binary mixtures model for all the possible pairs of interactions (BUF,BUN, BUF,DBQ, BUN,DBQ) describing a mutual competitive inhibition between the compounds, (iii) incorporating in the binary mixtures model the related constants determined in vitro for the inhibition, metabolism, transport, and partition coefficients of each compound, and (iv) validating the overall liver model for the prediction of the perfusate kinetics of each drug determined in isolated perfused rat liver (IPRL) for the single and paired compounds. Results from microsomal coincubations showed that competitive inhibition was the mechanism of interactions between all three compounds, as expected since those compounds are all substrates of rat CYP2D2. For each drug, the Ki values estimated were similar to their Km values for CYP2D2 indicative of a competition for the same substrate-binding site. Comparison of the performance between the novel liver physiologically based pharmacokinetic (PBPK) model and published empirical models in simulating the perfusate concentration,time profile was based on the area under the curve (AUC) and the shape of the curve of the perfusate time course. The present liver PBPK model was able to quantitatively predict the metabolic interactions determined during the perfusions of mixtures of BUF,DBQ and BUN,DBQ. However, a lower degree of accuracy was obtained for the mixtures of BUF,BUN, potentially due to some interindividual variability in the relative proportion of CYP2D1 and CYP2D2 isoenzymes, both involved in BUF metabolism. Overall, in this metabolic interaction prediction exercise, the PBPK model clearly showed to be the best predictor of perfusate kinetics compared to more empirical models. The present study demonstrated the potential of the mechanistic liver model to enable predictions of metabolic DDI under in vivo condition solely from in vitro information. © 2010 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 99:4406,4426, 2010 [source] Metabolism and disposition of resveratrol in the isolated perfused rat liver: Role of Mrp2 in the biliary excretion of glucuronidesJOURNAL OF PHARMACEUTICAL SCIENCES, Issue 4 2008Alexandra Maier-Salamon Abstract In this study, the hepatic metabolism and transport system for resveratrol was examined in isolated perfused livers from Wistar and Mrp2-deficient TR, rats. Based on extensive metabolism to six glucuronides and sulfates (M1,M6), the hepatic extraction ratio and clearance of resveratrol was very high in Wistar and TR, rats (E: 0.998 vs. 0.999; Cl: 34.9 mL/min vs. 36.0 mL/min). However, biliary excretion and efflux of conjugates differs greatly in TR, rats. While cumulative biliary excretion of the glucuronides M1, M2, M3, and M5 dropped dramatically to 0,6%, their efflux into perfusate increased by 3.6-, 1.8-, 2.5-, and 1.5-fold. In contrast, biliary secretion of the sulfates M4 and M6 was partially maintained in the Mrp2-deficient rats (61% and 39%) with a concomitant decline of their efflux into perfusate by 33.2% and 78.1%. This indicates that Mrp2 exclusively mediates the biliary excretion of resveratrol glucuronides but only partly that of sulfates. Cumulative secretion of unconjugated resveratrol into bile of TR, rats was only reduced by 40%, and into perfusate by 19%, suggesting only a minor role of Mrp2 in resveratrol elimination. In summary, resveratrol was dose-dependently metabolized to several conjugates whereby the canalicular transporter Mrp2 selectively mediated the biliary excretion of glucuronides. © 2007 Wiley-Liss, Inc. and the American Pharmacists Association J Pharm Sci 97:1615,1628, 2008 [source] The effects of the phytoestrogenic isoflavone genistein on the hepatic disposition of preformed and hepatically generated gemfibrozil 1- O -acyl glucuronide in the isolated perfused rat liverJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 10 2003Anthony N. Lucas ABSTRACT Foods and complementary medicines contain phytoestrogenic isoflavones such as genistein, which undergo hepatic glucuronidation and excretion into bile and can potentially interfere with the hepatic elimination of other compounds. To investigate this potential, livers from Sprague-Dawley rats were perfused in single-pass mode with preformed gemfibrozil 1- O -acyl glucuronide (GG) (1 ,M, n = 12) for 60 min followed by a 30-min washout phase, or with gemfibrozil (1 ,M n = 10) for 120 min. Half of each group of livers were co-perfused with genistein (10 ,M) throughout the experiment. Perfusate and bile were analyzed for GG and gemfibrozil by HPLC. Co-perfusion with genistein significantly (P < 0.05) decreased the biliary extraction ratio of preformed GG from a mean of 0.82 to 0.65 and the first-order rate constant for transport of GG into bile from 0.054 + 0.010 to 0.032 + 0.008 min,1, but increased the first-order rate constant for sinusoidal efflux of GG from 0.128 + 0.023 to 0.227 + 0.078 min,1. Co-perfusion with genistein also significantly decreased the biliary extraction ratio of hepatically generated GG from 0.95 + 0.01 to 0.83 + 0.05. The findings confirm that genistein increases the potential for hepatic and systemic exposure to hepatically generated glucuronides, which may be important for patients on conventional drugs who consume isoflavones. [source] Determination of antiviral nucleoside analogues AM365 and AM188 in perfusate and bile of the isolated perfused rat liver using HPLCBIOMEDICAL CHROMATOGRAPHY, Issue 3 2006Jiping Wang Abstract Development, validation and application of an HPLC assay for new antiviral nucleoside analogues AM365 and AM188 in isolated perfused rat liver perfusate and bile were performed. An analytical column (Phenosphere-NEXT, 250 × 4.6 mm, C18, 4 µm, Phenomenex) was used in tandem with a guard column (4 × 3 mm, C18, Phenomenex) and operated at 25°C. The mobile phase [methanol:10 mmol/L sodium orthophosphate buffer (pH 7.0), 15:85, v/v] was pumped at 1 mL/min. The signal from a diode array detector was collected from 190 to 300 nm. The chromatogram was processed at 220 and 252 nm for AM365 and AM188, respectively. The HPLC method was validated by six intraday and seven interday runs. Standard curves were linear in the range 0.125,8.00 µg/mL for AM365 and AM188, and the lower limit of quantification for AM365 and AM188 was 0.125 µg/mL. Mean interday precision and accuracy of IPL perfusate quality control samples were within 8.8%, and mean intraday precision and accuracy were within 13.1%. The assay has been successfully used in the study of metabolism and disposition of AM365 in the isolated perfused rat liver. Copyright © 2005 John Wiley & Sons, Ltd. [source] Induction of cellular resistance against Kupffer cell,derived oxidant stress: A novel concept of hepatoprotection by ischemic preconditioningHEPATOLOGY, Issue 2 2003Rolf J. Schauer Ischemic preconditioning (IP) triggers protection of the liver from prolonged subsequent ischemia. However, the underlying protective mechanisms are largely unknown. We investigated whether and how IP protects the liver against reperfusion injury caused by Kupffer cell (KC)-derived oxidants. IP before 90 minutes of warm ischemia of rat livers in vivo significantly reduced serum alanine aminotransferase (AST) levels and leukocyte adherence to sinusoids and postsinusoidal venules during reperfusion. This protective effect was mimicked by postischemic intravenous infusion of glutathione (GSH), an antioxidative strategy against KC-derived H2O2. Interestingly, no additional protection was achieved by infusion of GSH to preconditioned animals. These findings and several additional experiments strongly suggest IP mediated antioxidative effects: IP prevented oxidant cell injury in isolated perfused rat livers after selective KC activation by zymosan. Moreover, IP prevented cell injury and pertubations of the intracellular GSH/GSSG redox system caused by direct infusion of H2O2 (0.5 mmol/L). IP-mediated resistance against H2O2 could neither be blocked by the adenosine A2a antagonist DMPX nor mimicked by A2a agonist CGS21680. In contrast, H2O2 resistance was abolished by the p38 mitogen-activated protein kinase (p38 MAPK) inhibitor SB203580, but induced when p38 MAPK was directly activated by anisomycin. In conclusion, we propose a novel concept of hepatoprotection by IP: protection of liver cells by enhancing their resistance against KC-derived H2O2. Activation of p38 MAPK and preservation of the intracellular GSH/oxidized glutathione (GSSG) redox system, but not adenosine A2a receptor stimulation, seems to be pivotal for the development of H2O2 resistance in preconditioned livers. [source] Role of mitogen-activated protein kinases in tauroursodeoxycholic acid-induced bile formation in cholestatic rat liverHEPATOLOGY RESEARCH, Issue 7 2008Gerald Ulrich Denk Aim:, Ursodeoxycholic acid exerts anticholestatic effects in various cholestatic disorders and experimental models of cholestasis. Its taurine conjugate (TUDCA) stimulates bile salt secretion in isolated perfused rat livers (IPRL) under physiological, non-cholestatic conditions, in part by mitogen-activated protein kinase (MAPK)-dependent mechanisms. The role of MAPK in the anticholestatic effect of TUDCA, however, is unclear. Therefore, we studied the role of MAPK in the anticholestatic effect of TUDCA in IPRL and isolated rat hepatocytes (IRH) in taurolithocholic acid (TLCA)-induced cholestasis. Methods:, Bile flow, biliary levels of 2,4-dinitrophenyl-S-glutathione (GS-DNP) as a marker of hepatobiliary organic anion secretion and activity of lactate dehydrogenase (LDH) in hepatovenous effluate as a marker of hepatocellular damage in IPRL perfused with TUDCA and/or TLCA were determined in the presence or absence of MAPK inhibitors. In addition, phosphorylation of Erk 1/2 and p38MAPK induced by TUDCA and/or TLCA was studied by Western immunoblot in IPRL and IRH. Results:, TUDCA-induced bile flow was impaired by the Erk 1/2 inhibitor PD98059 in normal livers (,28%), but not in livers made cholestatic by TLCA. GS-DNP secretion was unaffected by PD98059 under both conditions. TUDCA-induced bile formation and organic anion secretion both in the presence and absence of TLCA were unaffected by the p38MAPK inhibitor SB202190. Erk 1/2 phosphorylation in liver tissue was unchanged after bile salt exposure for 70 min, but was transiently enhanced by TUDCA in IRH. Conclusion:, MAPK do not mediate the anticholestatic effects of TUDCA in TLCA-induced cholestasis. [source] | |||||||||||||