Gram Of Liver Research Articles

Transporter Expression in Liver Tissue from Subjects with Alcoholic or Hepatitis C Cirrhosis Quantified by Targeted Quantitative Proteomics.

Although data are available on the change of expression/activity of drug-metabolizing enzymes in liver cirrhosis patients, corresponding data on transporter protein expression are not available. Therefore, using quantitative targeted proteomics, we compared our previous data on noncirrhotic control livers (n = 36) with the protein expression of major hepatobiliary transporters, breast cancer resistance protein (BCRP), bile salt export pump (BSEP), multidrug and toxin extrusion protein 1 (MATE1), multidrug resistance-associated protein (MRP)2, MRP3, MRP4, sodium taurocholate-cotransporting polypeptide (NTCP), organic anion-transporting polypeptides (OATP)1B1, 1B3, 2B1, organic cation transporter 1 (OCT1), and P-glycoprotein (P-gp) in alcoholic (n = 27) and hepatitis C cirrhosis (n = 30) livers. Compared with control livers, the yield of membrane protein from alcoholic and hepatitis C cirrhosis livers was significantly reduced by 56 and 67%, respectively. The impact of liver cirrhosis on transporter protein expression was transporter-dependent. Generally, reduced protein expression (per gram of liver) was found in alcoholic cirrhosis livers versus control livers, with the exception that the expression of MRP3 was increased, whereas no change was observed for MATE1, MRP2, OATP2B1, and P-gp. In contrast, the impact of hepatitis C cirrhosis on protein expression of transporters (per gram of liver) was diverse, showing an increase (MATE1), decrease (BSEP, MRP2, NTCP, OATP1B3, OCT1, and P-gp), or no change (BCRP, MRP3, OATP1B1, and 2B1). The expression of hepatobiliary transporter protein differed in different diseases (alcoholic versus hepatitis C cirrhosis). Finally, incorporation of protein expression of OATP1B1 in alcoholic cirrhosis into the Simcyp physiologically based pharmacokinetics cirrhosis module improved prediction of the disposition of repaglinide in liver cirrhosis patients. These transporter expression data will be useful in the future to predict transporter-mediated drug disposition in liver cirrhosis patients.

Changes in cytochrome P450s-mediated drug clearance in patients with hepatocellular carcinoma in vitro and in vivo: a bottom-up approach.

Hepatocellular carcinoma (HCC) accompanied by severe liver dysfunction is a serious disease, which results in altered hepatic clearance. Generally, maintenance doses depend upon drug clearance, so individual dosage regimens should be customized for HCC patients based on the condition of patients. Based on clearance of CYP isoform-specific substrates at the microsomal level (CLM), microsomal protein per gram of liver (MPPGL), liver weight, hepatic blood flow, hepatic clearance values (CLH) for 10 CYPs in HCC patients (n=102) were extrapolated using a predictive bottom-up pharmacokinetic model. Compared with controls, the CLM values for CYP2C9, 2D6, 2E1 were significantly increased in HCC patients. Additionally, CYP1A2, 2C8, 2C19 CLM values decreased while the values for CYP2A6, 2B6, 3A4/5 were unchanged. The MPPGL values in HCC tissues were significantly reduced. CLH values of HCC patients for CYP1A2, 2A6, 2B6, 2C8, 2C19, and 3A4/5 were significantly reduced, while this for CYP2E1 were markedly increased and those for CYP2C9 and 2D6 did not change. Moreover, disease (fibrosis and cirrhosis) and polymorphisms of the CYP genes have influenced the CLH for some CYPs. Prediction of the effects of HCC on drug clearance may be helpful for the design of clinical studies and the clinical management of drugs in HCC patients.

Content and activity of human liver microsomal protein and prediction of individual hepatic clearance in vivo.

The lack of information concerning individual variation in content and activity of human liver microsomal protein is one of the most important obstacles for designing personalized medicines. We demonstrated that the mean value of microsomal protein per gram of liver (MPPGL) was 39.46 mg/g in 128 human livers and up to 19-fold individual variations existed. Meanwhile, the metabolic activities of 10 cytochrome P450 (CYPs) were detected in microsomes and liver tissues, respectively, which showed huge individual variations (200-fold). Compared with microsomes, the activities of liver tissues were much suitable to express the individual variations of CYP activities. Furthermore, individual variations in the in vivo clearance of tolbutamide were successfully predicted with the individual parameter values. In conclusion, we offer the values for MPPGL contents in normal liver tissues and build a new method to assess the in vitro CYP activities. In addition, large individual variations exist in predicted hepatic clearance of tolbutamide. These findings provide important physiological parameters for physiologically-based pharmacokinetics models and thus, establish a solid foundation for future development of personalized medicines.

Effects of Strain and Production-Cycle on Indices of Lysine Catabolism in Turkeys

In typical turkey production diets, lysine is frequently the second limiting amino acid. Understanding its catabolism could provide opportunities to increase the efficiency of lysine use for protein synthesis. We hypothesize that indices of lysine catabolism in turkey liver vary throughout the production cycle. Two commercial strains of turkey, Strain A and B, were analyzed 8 times over a period of 17 weeks (n = 8 birds/sampling time/strain) for lysine alpha-ketoglutarate (LKR) and saccharopine dehydrogenase (SDH) activities and mRNA abundance, in vitro lysine oxidation (LOX), L-amino acid oxidase (LAAO) activity, lysyl oxidase (LO) activity and mRNA expression of cationic amino acid transporters (CAT) 1, 2 and 3. We found differences in hepatic LKR activity (p = 0.007), LKR mRNA (p = 0.0004), SDH activity (p = 0.008), LOX (p<0.0001), lysyl oxidase activity (p<0.01), LAAO (p<0.0001) and CAT1 (p = 0.005) and 2 mRNA abundance (p = 0.0022) throughout the production cycle. We also found an effect of strain on SDH (p = 0.046) and CAT-2 mRNA abundance (p = 0.02); while no age or strain effects were detected for CAT3 mRNA abundance. Interestingly, the average LKR and SDH activities across strains and weeks was 240 and 420 nmol per minute per gram of liver, respectively, as opposed to the average LAAO activity, lysyl oxidase activity and L OX which were 0.70, 0.10 and 13.5 nmol per minute per gram liver, respectively. These data indicate that the saccharopine-dependent pathway is the predominant pathway of lysine degradation in turkey liver and that indices of hepatic lysine catabolism vary throughout the production cycle.

Microsomal protein per gram of liver (MPPGL) in paediatric biliary atresia patients

The microsomal protein per gram of liver (MPPGL) is an important scaling factor in the in vitro-in vivo extrapolation of metabolic data obtained in liver microsomes. This study aimed to determine the MPPGL in four biliary atresia patients (0.6-1.6 years old) undergoing liver transplantation, as it is known that the MPPGL is affected by age and possibly by liver disease. Due to the presence of bilirubin in the homogenates and microsomes, the NADPH-cytochrome reductase activity was used to determine the recovery factor, rather than methods using the dithionite difference spectrum. A mean value of 18.73 (± 2.82) mg/g (geometric mean ± SD, n = 4) was observed, which is lower than the expected MPPGL based on the age of the patients (26.60 ± 0.40 mg/g). This suggests a decreased amount of microsomal protein in the livers of biliary atresia patients. Moreover, no differences in MPPGL between different zones of the liver could be detected.

Extrapulmonary transport of MWCNT following inhalation exposure

BackgroundInhalation exposure studies of mice were conducted to determine if multi-walled carbon nanotubes (MWCNT) distribute to the tracheobronchial lymphatics, parietal pleura, respiratory musculature and/or extrapulmonary organs. Male C57BL/6 J mice were exposed in a whole-body inhalation system to a 5 mg/m3 MWCNT aerosol for 5 hours/day for 12 days (4 times/week for 3 weeks, lung burden 28.1 ug/lung). At 1 day and 336 days after the 12 day exposure period, mice were anesthetized and lungs, lymph nodes and extrapulmonary tissues were preserved by whole body vascular perfusion of paraformaldehyde while the lungs were inflated with air. Separate, clean-air control groups were studied at 1 day and 336 days post-exposure. Sirius Red stained sections from lung, tracheobronchial lymph nodes, diaphragm, chest wall, heart, brain, kidney and liver were analyzed. Enhanced darkfield microscopy and morphometric methods were used to detect and count MWCNT in tissue sections. Counts in tissue sections were expressed as number of MWCNT per g of tissue and as a percentage of total lung burden (Mean ± S.E., N = 8 mice per group). MWCNT burden in tracheobronchial lymph nodes was determined separately based on the volume density in the lymph nodes relative to the volume density in the lungs. Field emission scanning electron microscopy (FESEM) was used to examine MWCNT structure in the various tissues.ResultsTracheobronchial lymph nodes were found to contain 1.08 and 7.34 percent of the lung burden at 1 day and 336 days post-exposure, respectively. Although agglomerates account for approximately 54% of lung burden, only singlet MWCNT were observed in the diaphragm, chest wall, liver, kidney, heart and brain. At one day post exposure, the average length of singlet MWCNT in liver and kidney, was comparable to that of singlet MWCNT in the lungs 8.2 ± 0.3 versus 7.5 ± 0.4 um, respectively. On average, there were 15,371 and 109,885 fibers per gram in liver, kidney, heart and brain at 1 day and 336 days post-exposure, respectively. The burden of singlet MWCNT in the lymph nodes, diaphragm, chest wall and extrapulmonary organs at 336 days post-exposure was significantly higher than at 1 day post-exposure.ConclusionsInhaled MWCNT, which deposit in the lungs, are transported to the parietal pleura, the respiratory musculature, liver, kidney, heart and brain in a singlet form and accumulate with time following exposure. The tracheobronchial lymph nodes contain high levels of MWCNT following exposure and further accumulate over nearly a year to levels that are a significant fraction of the lung burden 1 day post-exposure.

Quasi-Experimental Designs in Practice-based Research Settings: Design and Implementation Considerations

Although randomized controlled trials are often a gold standard for determining intervention effects, in the area of practice-based research (PBR), there are many situations in which individual randomization is not possible. Alternative approaches to evaluating interventions have received increased attention, particularly those that can retain elements of randomization such that they can be considered "controlled" trials. Methodological design elements and practical implementation considerations for two quasi-experimental design approaches that have considerable promise in PBR settings--the stepped-wedge design, and a variant of this design, a wait-list cross-over design, are presented along with a case study from a recent PBR intervention for patients with diabetes. PBR-relevant design features include: creation of a cohort over time that collects control data but allows all participants (clusters or patients) to receive the intervention; staggered introduction of clusters; multiple data collection points; and one-way cross-over into the intervention arm. Practical considerations include: randomization versus stratification, training run in phases; and extended time period for overall study completion. Several design features of practice based research studies can be adapted to local circumstances yet retain elements to improve methodological rigor. Studies that utilize these methods, such as the stepped-wedge design and the wait-list cross-over design, can increase the evidence base for controlled studies conducted within the complex environment of PBR.

Production‐cycle‐related changes in indices of lysine catabolism in turkey liver

In typical turkey production diets, lysine is frequently the second limiting amino acid. Understanding its catabolism could provide opportunities to increase the efficiency of lysine use for protein synthesis. We hypothesize that indices of lysine catabolism in turkey liver vary throughout the production cycle. Two commercial strains of turkey, Hybrid and Nicolas, were analyzed 8 times over a period of 17 weeks (n=8 birds/sampling time/strain) for lysine α‐ketoglutarate (LKR) and saccharopine dehydrogenase (SDH) activity and mRNA abundance, in vitro lysine oxidation (LOX) and amino acid oxidase (AAOX) activity. We found differences in LKR mRNA (P&lt;0.01), SDH activity (P&lt;0.05) and LOX (P&lt;0.0001) and a trend for differences in LKR activity (P=0.09), SDH mRNA (P=0.07) and AAOX activity (P=0.06) throughout the production cycle. No consistent strain differences were detected for any variable. Interestingly, the average LKR and SDH activities across strains and weeks was 230 and 400 nmol per minute per gram of liver, respectively, as opposed to the average AAOX activity and LOX was 1.25 and 13.6 nmol per minute per gram liver, respectively. These data indicate that the saccharopine‐dependent pathway is the predominant pathway of lysine degradation in turkey liver and that indices of hepatic lysine catabolism vary throughout the production cycle. (Support; HATCH WVA 470, Virginia Poultry Growers Cooperative)

Mitochondrion is a novel site of biotransformation of dichloroacetate by glutathione transferase zeta

Dichloroacetate (DCA) is an environmental contaminant as well as a potential therapeutic agent through its ability to stimulate the pyruvate dehydrogenase complex. Repeated doses of DCA result in reduced drug clearance, probably through inhibition of glutathione transferase zeta 1 (GSTz1), a cytosolic enzyme that converts DCA to glyoxylate. Because DCA is known to be taken up into mitochondria, its principal site of action, it is of interest to explore the role of mitochondria in DCA biotransformation. Incubation of human and rat liver mitochondria with 14C‐DCA resulted in biotransformation only when glutathione was added, and glyoxylate was identified as the major metabolite. Protein expression of GSTz was detected in human and rat liver mitochondria by Western Blotting against human GSTz1 antibody. In rats, the expression of GSTz per gram of liver is 14 times as much in cytosol as in mitochondria. Similar to cytosolic GSTz, the expression and activity of mitochondrial GSTz was markedly reduced in rats treated with 500 mg DCA/kg for 8 weeks. Studies of sub‐mitochondrial fractions of rat liver showed that GSTz is localized in the mitochondrial matrix. We conclude that the mitochondrion is a site of DCA biotransformation catalyzed by GSTz, an enzyme shown for the first time to be co‐localized in cytosol and mitochondrial matrix. Supported in part by the US Public Health Service, ES 07355, ES 014617, and 1UL1 RR029890.

Gold Nanoparticle Contrast in a Phantom and Juvenile Swine: Models for Molecular Imaging of Human Organs using X-ray Computed Tomography

The purpose of this study was to demonstrate the application of gold nanoparticles (AuNP) as a contrast agent for a clinical x-ray computed tomography (CT) system using a phantom and juvenile swine. A tissue-mimicking phantom with spherical inclusions containing known concentrations of Au was scanned. Swine were injected with gum Arabic stabilized Au nanoparticles (GA-AuNP), up to 85 mg kg(-1) body weight. CT scans were performed before and after the injections. Changes in Hounsfield unit (HU) values between pre- and post- injection scans were evaluated and compared to postmortem determinations of Au uptake. Average uptake of GA-AuNP in the liver of the swine was 380 microg per gram of liver and 680 microg per gram of spleen. Concentrations of Au in tissues increased the CT numbers in liver by approximately 22 HU per mg Au concentration at 80 kVp and 27 HU per mg Au concentration at 140 kVp. These data were consistent with HU changes observed for similar concentrations in the phantom. AuNP-based contrast agents may be useful in x-ray based CT. This study provides data for determining concentrations of AuNP in comparison to other contrast materials.

Adipocyte Apoptosis, a Link between Obesity, Insulin Resistance, and Hepatic Steatosis

Adipocyte death has been reported in both obese humans and rodents. However, its role in metabolic disorders, including insulin resistance, hepatic steatosis, and inflammation associated with obesity has not been studied. We now show using real-time reverse transcription-PCR arrays that adipose tissue of obese mice display a pro-apoptotic phenotype. Moreover, caspase activation and adipocyte apoptosis were markedly increased in adipose tissue from both mice with diet-induced obesity and obese humans. These changes were associated with activation of both the extrinsic, death receptor-mediated, and intrinsic, mitochondrial-mediated pathways of apoptosis. Genetic inactivation of Bid, a key pro-apoptotic molecule that serves as a link between these two cell death pathways, significantly reduced caspase activation, adipocyte apoptosis, prevented adipose tissue macrophage infiltration, and protected against the development of systemic insulin resistance and hepatic steatosis independent of body weight. These data strongly suggest that adipocyte apoptosis is a key initial event that contributes to macrophage infiltration into adipose tissue, insulin resistance, and hepatic steatosis associated with obesity in both mice and humans. Inhibition of adipocyte apoptosis may be a new therapeutic strategy for the treatment of obesity-associated metabolic complications.

Phenylalanine-pyruvate aminotransferase activity in chicks subjected to phenylalanine imbalance or phenylalanine toxicity

Two experiments were done to determine the influence of Phe imbalance and excess on Phe-pyruvate aminotransferase (PAT) activity in the chick. Five replicates of 3 chicks (experiment 1) or 2 chicks (experiment 2) of a commercial brown egg layer strain were fed a semipurified diet for 1 wk and then received experimental diets for 10 d. Three diets were used in experiment 1: the basal diet contained 0.46% Phe; the imbalance diet was similar to the basal diet except that it contained a 10% mixture of indispensable amino acids lacking Phe (IAA - Phe) to create a Phe imbalance; the imbalance corrected diet was similar to the imbalance diet except that it was supplemented with 1.12% Phe to correct the imbalance. A 3 x 2 factorial arrangement of treatments in experiment 2 provided 3 dietary levels (0.46, 1.58, and 2.46%) of Phe and either no supplement or 10% supplement of IAA - Phe. Nonfasted chicks were killed and livers were sampled in experiment 1, and livers, kidneys, brains, and pectoralis major muscles were sampled in experiment 2. In experiment 1, liver PAT activity per gram of liver was 80 and 55% higher (P < 0.01) in chicks fed the imbalance and imbalance corrected diets than in chicks fed the basal diet. In experiment 2, the livers and kidneys, but not brains and muscles, of chicks that received the 10% supplement of IAA - Phe had higher activities of PAT per gram of tissue per minute and per milligram of tissue protein extract per minute than chicks that did not receive IAA - Phe (P < 0.001). No effect of dietary Phe on PAT activity was detected (P > 0.05). Phenylalanine-pyruvate aminotransferase activity appears to be regulated in response to dietary content of indispensable amino acids but not by the dietary level of Phe.

Occurrence of Ascocotyle ( Phagicola) longa Ransom, 1920 (Digenea: Heterophyidae) in Mugil incilis from Cartagena Bay, Colombia

Fishborne metacercariae have not been reported in Colombia. During an 8-month period, 250 mullet/lisa ( Mugil incilis) were examined for trematode metacercariae in the liver. Average prevalence and intensity were 93.4 ± 2.5% and 8476 ± 1238 cysts per gram of liver, respectively. Monthly prevalence of the trematode, identified as Ascocotyle ( Phagicola) longa Ransom (1920), was always high (>79%) and there was no difference between sampling periods ( P > 0.05). Trematode intensity in lisa had a significant negative effect on host size ( R = −0.325, P < 0.001), condition factor ( R = −0.352, P < 0.001), and liver-somatic index ( R = −0.372, P < 0.001); in contrast, a positive relationship was found with gill-somatic index ( R = 0.446, P < 0.001). Histopathological analysis indicated the presence of abundant cysts together with inflammatory reactions involving macrophage aggregates and necrosis. These results suggest that trematodes infecting the liver of M. incilis affect the growth and well-being of these fish, and their consumption could represent a major risk for human infection.

STUDIES IN IRON METABOLISM

An enzyme system incorporating inorganic iron into protoporphyrin has been demonstrated in human and rat liver, kidney, spleen and in human bone marrow and measured, using radioiron as a tracer. Studies on the enzyme content of the intracellular fractions of human and rat liver showed that while there was some activity in all fractions, the mitochondrial fraction was the most active. Dialysis of tissue homogenates led to a loss of enzymic activity which could be restored by the addition of ascorbic acid, GSH, cysteine or DPNH. The effect of these substances was not entirely due to an effect of reduction of the ferric iron since potentiation of activity resulted when these substances were added to a system containing iron in the ferrous form. Observations have also been made on the in vitro biosynthesis of haem from protoporphyrin and iron bound to siderophilin or to rat liver homogenate. Rat liver homogenate or a preparation from it of mitochondria served as the source of iron-incorporating enzyme. The reducing substances namely ascorbic acid, GSH, cysteine and DPNH, which were effective in increasing the incorporation into protoporphyrin of inorganic ferric iron also substantially increased the transfer of protein bound iron from haem biosynthesis. ATP enhanced the transfer of iron bound to siderophilin for incorporation into protophyrin but had no effect on the transfer of iron bound to rat liver protein. From this it is clear that physiological reducing substances play an important role in the transfer of iron from its protein-bound form for incorporation into protoporphyrin in the process of haem biosynthesis. A technique has been established for the measurement of iron incorporating enzyme activity in haemolysates of rabbit bone marrow containing a known number of red cell precursors. The method has been applied to marrows of normal and bled rabbits. In the bled animal, the enzymic activity of a unit number of cells was greatly decreased as compared to normal but was increased by the addition of reducing substances. This would indicate that the marrow responds to blood loss by an increase in the production of cells rather than an increased capacity of each cell to form haemoglobin. The enzymic activity of liver homogenates was also measured in normal and bled rabbits. In this tissue there was no difference in the enzymic activity per gram of liver between the normal and the bled animals. There was, however, a marked decrease in iron incorporation by liver cells from animals which had been injected with 300 mgs. Imferon. In two rabbits made iron deficient and anaemic there was an increase in iron incorporation as compared to normal. From this it is clear that gross changes in the size of the iron pool will vitiate the correct interpretation of results on the measurement of haem synthesis using radio-iron as a tracer. Nakao et al (1960) have suggested that plasma samples from a variety of haematological disorders possess a factor (or factors) stimulating haem synthesis as compared to normal plasma. The 'haem-stimulating activity' was measured by an in vitro technique in which radio-iron incorporation was used as a measure of haem synthesis. This work has been repeated using the same technique and results show that any marked increase in haem formation, measured in this way, could be explained by the diminution of the residual iron-binding capacity of the plasma sample. It is concluded that no definite evidence has yet been obtained for the presence of a specific haem-stimulating factor in human plasma. A statistically significant difference has been shown between the absorption of radio-iron given along with a standard meal, between patients with acid present in their gastric juice and patients with a histamine-fast achlorhydria. Both of these groups of patients were anaemic and there was an absence of stainable iron in the bone marrow of both groups. The mean absorption from the meal of the patients with acid in their gastric juice was 57.5 per cent while the mean absorption of those patients with histamine-fast achlorhyrdria was 18.5 per cent.

Covariation of Human Microsomal Protein Per Gram of Liver with Age: Absence of Influence of Operator and Sample Storage May Justify Interlaboratory Data Pooling

Scaling of metabolic clearance values from liver microsomal data or recombinantly expressed cytochrome P450 enzymes to predict human hepatic clearance requires knowledge of the amount of microsomal protein per gram of liver (MPPGL). Identification of physiological covariates of MPPGL requires analysis of values from large diverse populations, which necessitates pooling of data from numerous sources. To ensure compatibility between results obtained within and between studies, the impact of interoperator differences and sample storage on values of MPPGL was investigated. With use of triplicate samples from one liver (HL86), no statistically significant difference was detected between values of MPPGL prepared from samples stored at -80 degrees C (23.5 +/- 1.2 mg g(-1)) and those determined using fresh tissue (21.9 +/- 0.3 mg g(-1)). Although there was a significant difference in the yield of microsomal protein obtained from another liver sample (HL43) by three different operators (17 +/- 1, 19 +/- 2, and 24 +/- 1 mg g(-1); p = 0.004, analysis of variance), no difference was observed in the estimated MPPGL after application of appropriate correction factors for each operator (28 +/- 1, 30 +/- 5, and 31 +/- 4 mg g(-1)). The result provided justification for pooling reported values of MPPGL for use in covariate analysis. Investigation of the relationship between age and MPPGL provided preliminary evidence that MPPGL values increase from birth to a maximum of 40 mg g(-1) [95% confidence interval for the geometric mean (95% CI mean(geo)): 37-43 mg g(-1) at approximately 28 years followed by a gradual decrease in older age (mean of 29 mg g(-1) at 65 years; 95% CI mean(geo): 27-32 mg g(-1)). Accordingly, appropriate age-adjusted scaling factors should be used in extrapolating in vitro clearance values to clinical studies.

Recognizing, treating and understanding pernicious anaemia.

Recognizing, treating and understanding pernicious anaemia.

Obesity-Induced Lymphocyte Hyperresponsiveness to Chemokines: A New Mechanism of Fatty Liver Inflammation in Obese Mice

Hepatic lipid retention (steatosis) predisposes hepatitis. We investigated the mechanisms of lymphocyte homing to fatty liver and the role of lipopolysaccharide (LPS) in the onset of inflammation in ob/ob mice. We decreased intestinal bacterial compounds by oral antibiotic treatment to test the role of endogenous LPS in liver inflammation. Adoptive transfer of lymphocytes was used to study the respective contributions of steatosis and lymphocytes to liver inflammation. We tested lymphocyte response to chemokines by in vitro chemotaxis assays in ob/ob, their lean controls, and "non-obese ob/ob" mice, generated by controlling caloric intake to distinguish between the effects of obesity and leptin deficiency. Antibiotic treatment decreased liver infiltration with CD4(+) T, CD8(+) T, natural killer (NK)T, B, and NK cells. Adoptive transfer of lymphocytes from ob/ob or control mice showed that (1) steatosis increased lymphocyte recruitment to the liver; (2) CD4(+) T, CD8(+) T, and B cells from ob/ob mice had a greater propensity to migrate specifically to the liver. This migration was enhanced by LPS. These results were also observed in a model of high-fat diet-induced obesity. CD4(+) T and B cells were hyperresponsive to CXCL12 and CXCL13, respectively. Weight normalization in "non-obese ob/ob" mice decreased liver inflammation, lymphocyte response to chemokines, and homing to the liver. Our study provides the first evidence that liver inflammation in mice with genetic or diet-induced obesity results from both steatosis and lymphocyte hyperresponsiveness to chemokines expressed in the liver. These abnormalities are reversible with weight normalization.

Polyunsaturated fat in the methionine-choline-deficient diet influences hepatic inflammation but not hepatocellular injury

Methionine-choline-deficient (MCD) diets that cause steatohepatitis in rodents are typically enriched in polyunsaturated fat. To determine whether the fat composition of the MCD formula influences the development of liver disease, we manufactured custom MCD formulas with fats ranging in PUFA content from 2% to 59% and tested them for their ability to induce steatohepatitis. All modified-fat MCD formulas caused identical degrees of hepatic steatosis and resulted in a similar distribution of fat within individual hepatic lipid compartments. The fatty acid composition of hepatic lipids, however, reflected the fat composition of the diet. Mice fed a PUFA-rich MCD formula showed extensive hepatic lipid peroxidation, induction of proinflammatory genes, and histologic inflammation. When PUFAs were substituted with more saturated fats, lipid peroxidation, proinflammatory gene induction, and hepatic inflammation all declined significantly. Despite the close relationship between PUFAs and hepatic inflammation in mice fed MCD formulas, dietary fat had no impact on MCD-mediated damage to hepatocytes. Indeed, histologic apoptosis and serum alanine aminotransferase levels were comparable in all MCD-fed mice regardless of dietary fat content. Together, these results indicate that dietary PUFAs promote hepatic inflammation but not hepatotoxicity in the MCD model of liver disease. These findings emphasize that individual dietary nutrients can make specific contributions to steatohepatitis.

Scaling Factors for the Extrapolation of In Vivo Metabolic Drug Clearance From In Vitro Data: Reaching a Consensus on Values of Human Micro-somal Protein and Hepatocellularity Per Gram of Liver

Reported predictions of human in vivo hepatic clearance from in vitro data have used a variety of values for the scaling factors human microsomal protein (MPPGL) and hepatocellularity (HPGL) per gram of liver, generally with no consideration of the extent of their inter-individual variability. We have collated and analysed data from a number of sources, to provide weighted meangeo values of human MPPGL and HPGL of 32 mg g-1 (95% Confidence Interval (CI); 29-34 mg.g-1) and 99x10(6) cells.g-1 (95% CI; 74-131 mg.g-1), respectively. Although inter-individual variability in values of MPPGL and HPGL was statistically significant, gender, smoking or alcohol consumption could not be detected as significant covariates by multiple linear regression. However, there was a weak but statistically significant inverse relationship between age and both MPPGL and HPGL. These findings indicate the importance of considering differences between study populations when forecasting in vivo pharmacokinetic behaviour. Typical clinical pharmacology studies, particularly in early drug development, use young, fit, healthy male subjects of around 30 years of age. In contrast, the average age of patients for many diseases is about 60 years of age. The relationship between age and MPPGL observed in this study estimates values of 40 mg.g-1 for a 30 year old individual and 31 mg.g-1 for a 60 year old individual. Investigators may wish to consider the reported covariates in the selection of scaling factors appropriate for the population in which estimates of clearance are being predicted. Further studies are required to clarify the influence of age (especially in paediatric subjects), donor source and ethnicity on values of MPPGL and HPGL. In the meantime, we recommend that the estimates (and their variances) from the current meta-analysis be used when predicting in vivo kinetic parameters from in vitro data.

Effect of dietary protein/carbohydrate ratio on activities of hepatic enzymes involved in the amino acid metabolism of Nile tilapia, Oreochromis niloticus (L.)

This study investigated the influence of different dietary protein/carbohydrate ratios on activities of enzymes involved in the amino acid metabolism as well as on growth performance and body composition of Nile tilapia (Oreo- chromis niloticus). Fish were fed semi-synthetic diets differing in their protein/carbohydrate ratio. The diets were formulated to be isolipidic and isoenergetic (on the basis of metabolizable energy). Fish were reared individually. After a prefeeding phase, tilapia were randomly assigned to three groups and were fed the experimental diets at a relatively low level. The study lasted 9 weeks. Proximate composition, gross energy content, and activities of aspartate aminotransferase (ASAT) and alanine aminotransferase (ALAT) in the liver of fish were determined. While the mean growth rate and feed conversion efficiency did not differ significantly among the feeding groups, the body composition showed significant differences. The dietary protein/carbohydrate ratio had no significant effect on the protein and energy gain, but the lipid gain was significantly higher in tilapia fed the low-protein diet. The activities of ASAT and ALAT per gram of liver and per gram of soluble protein were significantly increased in fish fed the high-protein diet. There was a significant positive correlation between the activities of ASAT and ALAT. The activities of both enzymes in the liver were significantly higher in fish with a higher protein gain. Measuring the activities of enzymes involved in the amino acid metabolism can provide more information about the metabolic utilized dietary protein.