Gram Of Liver Research Articles

Graft Inflow Modulation by Splenic Artery Ligation for Portal Hyper Perfusion does not Decrease Rates of Early Allograft Dysfunction in Adult Live Donor Liver Transplantation: A Randomized Control Trial.

The primary objective was to compare the rates of early allograft dysfunction (EAD) in patients undergoing elective adult live donor liver transplantation (ALDLT) with and without graft portal inflow modulation (GIM) for portal hyper-perfusion. The secondary objectives were to compare time to normalization of bilirubin and International Normalized Ratio (INR), day 14 ascitic output more than 1liter, small-for-size syndrome (SFSS), intensive care unit / high dependency unit and total hospital stay, and 90 day morbidity and mortality. GIM can prevent EAD in ALDLT patients with portal hyper-perfusion. A single-center randomized trial with and without GIM for portal hyper-perfusion by splenic artery ligation (SAL) in ALDLT was performed. After reperfusion, patients with portal venous pressure (PVP)>15mm Hg with a gradient (PVP - central venous pressure) of ≥ 7mm Hg and/or portal venous flow (PVF)>250mL/min/100 grams of liver were randomized into two groups: GIM and No GIM. 75 of 209 patients satisfied the inclusion criteria, and 38 underwent GIM. Baseline PVF and PVP were comparable between the GIM and no GIM groups. SAL significantly reduced the PVF and PVP (P<0.001). There were no differences in the primary and secondary outcomes between the two groups. In the subgroup analysis, with a Graft to Recipient Weight Ratio (GRWR)≤0.8, there were no significant differences in the primary and secondary outcomes. SAL significantly decreased PVP and PVF, but did not decrease rates of EAD in adult LDLT.

Ontogeny of Scaling Factors for Pediatric Physiologically Based Pharmacokinetic Modeling and Simulation: Cytosolic Protein Per Gram of Liver.

Scaling factors are necessary for translating in vitro drug biotransformation data to in vivo clearance values in physiologically-based pharmacokinetic modeling and simulation. Values for microsomal protein per gram of liver are available from several sources for use as a scaling factor to estimate hepatic clearance from microsomal drug biotransformation data. However, data regarding the distribution of cytosolic protein per gram of liver (CPPGL) values across the lifespan are limited, and sparse pediatric data have been published to date. Thus, CPPGL was determined in 160 liver samples from pediatric (n = 129) and adult (n = 31) donors obtained from multiple sources: the University of Maryland Brain and Tissue Bank, tissue retrieval services at the University of Minnesota and University of Pittsburgh, and Sekisui-XenoTech. Tissues were homogenized and subjected to differential centrifugation to isolate cytosolic fractions. Cytosolic protein content was determined by BCA assay. CPPGL varied from two- to sixfold within each age group/developmental stage. Tissue source and sex did not contribute substantially to variability in protein content. Regression analyses revealed minimal change in CPPGL over the first two decades of life (logCPPGL increases 0.1 mg/g per decade). A mean ± S.D. CPPGL value of 44.4 ± 17.4 mg/g or median 41.0 mg/g is representative of values observed between birth and early adulthood (0-18 years, n = 129). SIGNIFICANCE STATEMENT: Cytosolic protein per gram of liver (CPPGL) is a scaling factor required for physiologically based pharmacokinetic modeling and simulation of drug biotransformation by cytosolic enzymes, but pediatric data are limited. Although CPPGL varies from two- to sixfold within developmental stages, a value of 44.4 ± 17.4 mg/g (mean ± S.D.) is representative of the pediatric period (0-18 years, n = 129).

Sex- and lifestyle-related factors are associated with altered hepatic CYP protein levels in people diagnosed with mental disorders.

In this study, we used human postmortem tissue to investigate hepatic protein expression levels of CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4 by LC‒MS/MS in a population of people suffering from mental disorders (n=171). We report hepatic protein levels of these six CYP isoforms in 171 individuals in total, and define a focused population dataset of 116 individuals after excluding 55 samples due to low Microsomal Protein Per Gram of Liver (MPPGL) yield. Postmortem decay was most likely the reason for the low MPPGL yield in the 55 samples. In the focused population, we found women to have significantly higher protein levels of CYP3A4 than men in addition to decreased CYP3A4 protein levels among obese individuals. Furthermore, MPPGL was negatively correlated with body mass index (BMI). An increase in CYP1A2 protein levels was observed among smokers and increased CYP2E1 protein levels were observed among individuals with a history of alcohol abuse. Finally, individuals who received phenobarbital (CYP3A4 inducer) had significantly higher CYP3A4 levels. In conclusion, lifestyle-related factors prevalent among people suffering from mental disorders are associated with altered CYP protein levels which may alter drug metabolism and affect the efficacy of commonly prescribed drugs. Furthermore, this investigation demonstrates that postmortem hepatic tissue can be used to study how lifestyle and effectors affect hepatic CYP-levels in a large cohort of patients. Significance Statement Using a large number of postmortem hepatic tissue specimens (n=116) originating from the autopsy of individuals diagnosed with mental disorders, we were able to show that hepatic CYP-levels were affected by alcohol, smoking, BMI, and sex and that MPPGL was affected by BMI. Theses lifestyle-related changes may alter drug metabolism and affect the efficacy of commonly prescribed drugs. It is a novel approach to use a large postmortem cohort to investigate how lifestyle and effectors affect hepatic CYP-levels.

Establishing allometric relationships between microsomal protein and cytochrome P450 content with body weight in vertebrate species

Data from in vitro studies are routinely used to estimate in vivo hepatic clearance of chemicals and this information is needed to parameterise physiologically based kinetic models. Such clearance data can be obtained from laboratory experiments using liver microsomes, hepatocytes, precision-cut liver slices or recombinant enzymes. Irrespective of the selected test system, scaling factors are required to convert the in vitro measured intrinsic clearance to a whole liver intrinsic clearance. Scaling factors such as the hepatic microsomal protein per gram of liver and/or the amount of cytochrome P450 per hepatocyte provide a means to calculate the whole liver intrinsic clearance. Here, a database from the peer-reviewed literature has been developed and provides quantitative metrics on microsomal protein (MP) and cytochrome P450 contents in vertebrate orders namely amphibians, mammals, birds, fish and reptiles. This database allows to address allometric relationships between body weight and MP content, and body weight and cytochrome P450 content. A total of 85 and 74 vertebrate species were included to assess the relationships between log10 body weight versus log10 MP, and between log10 body weight and log10 cytochrome P450 content, respectively. The resulting slopes range from 0.76 to 1.45 in a range of vertebrate species. Such data-driven allometric relationships can be used to estimate the MP content necessary for in vitro to in vivo extrapolation of in vitro clearance data. Future work includes applications of these relationships for different vertebrate taxa using quantitative in vitro to in vivo extrapolation models coupled to physiologically based kinetic models using chemicals of relevance as case studies including pesticides, contaminants and feed additives.

Schistosoma japonicum translationally controlled tumour protein, which is associated with the development of female worms, as a target for control of schistosomiasis

Schistosomiasis is a globally important helminthic disease of both humans and animals, and is the second most common parasitic disease after malaria. Although praziquantel is extensively used for treatment of parasitic diseases, drug resistance has been reported. Therefore, new drugs and effective vaccines are needed for continuous control of schistosomiasis. Eggs produced by schistosomes are responsible for the occurrence and spread of schistosomiasis. Revealing the reproductive mechanism of schistosomes will help to control this disease. In this study, the proteomic profiles of single-sex infected female worms and bisexual infected mature female worms of Schistosoma japonicum at 18, 21, 23 and 25 days p.i. were identified with isobaric tags for relative quantitation-coupled liquid chromatography–tandem mass spectrometry. Differentially expressed proteins were subsequently used for bioinformatic analysis. Six highly expressed differentially expressed proteins in mature female worms were selected and long-term interference with small interfering RNA (siRNA) was conducted to determine biological functions. SiRNA against S. japonicum translationally controlled tumour protein (SjTCTP) resulted in the most significant effect on the growth and development of MF worms. Sjtctp mRNA expression gradually increased over time with a high level of expression maintained at 25–42 days p.i., while levels were significantly higher in mature female worms than male and SF worms. The subsequent animal immune protection experiments showed that recombinant SjTCTP (rSjTCTP) reduced the number of adults by 44.7% (P < 0.01), average egg burden per gram of liver by 57.94% (P < 0.01), egg hatching rate by 47.57% (P < 0.01), and oviposition of individual females by 43.16%. rSjTCTP induced higher levels of serum IgG, IL-2, and IL-10 in mice. Collectively, these results show that SjTCTP is vital to reproduction of female worms and, thus, is a candidate antigen for immune protection.

Ontogeny of Scaling Factors for Pediatric Physiology-Based Pharmacokinetic Modeling and Simulation: Microsomal Protein Per Gram of Liver.

Microsomal protein per gram of liver (MPPGL) is an important scaling factor for bottom-up physiology-based pharmacokinetic modeling and simulation, but data in pediatrics are limited. Therefore, MPPGL was determined in 160 liver samples from pediatric (n = 129) and adult (n = 31) donors obtained from four sources: the University of Maryland Brain and Tissue Bank (UMBTB), tissue retrieval services at the University of Minnesota and University of Pittsburgh, and Sekisui-Xenotech. Tissues were homogenized and subjected to differential centrifugation to prepare microsomes, and cytochrome c reductase activities in tissue homogenates and microsomes were used to estimate cytochrome P450 reductase (POR) activity as a marker of microsomal recovery; microsomal POR content was also assessed by quantitative proteomics. MPPGL values varied 5- to 10-fold within various age groups/developmental stages, and tissue source was identified as a contributing factor. Using a "trimmed" dataset comprised of samples ranging from 3 to 18 years of age common to the four sources, POR protein abundance and activity in microsomes and POR activity in homogenates was lower in UMBTB samples (autopsy) compared with other sources (perfused/flash-frozen). Regression analyses revealed that the UMBTB samples were driving an apparent age effect as no effect of age on log-transformed MPPGL values was observed when the UMBTB samples were excluded. We conclude that a mean±SD MPPGL value of 30.4±1.7 mg/g is representative between one month postnatal age and early adulthood. Potential source effects should be considered for studies involving tissue samples from multiple sources with different procurement and processing procedures. SIGNIFICANCE STATEMENT: Microsomal protein per gram of liver (MPPGL) is an important scaling factor for bottom up PBPK modeling and simulation, but data in pediatrics are limited. Although MPPGL varies 5- to 10-fold at a given developmental stage, a value of 30.4 ± 1.7 mg/g (mean ± SD) is representative between one month postnatal age and early adulthood. However, when tissue samples are obtained from multiple sources, different procurement and processing procedures may influence the results and should be taken into consideration.

In Vivo Assessment of the Antischistosomal Activity of OPUNTIA FICUS-INDICA Flowers and Their Chemical Constituents

Hepatic schistosomiasis is the most well-known form of chronic disease, with a wide variety of clinical symptoms. The aim of this study was to assess the schistosomicidal effect of the aqueous methanolic extract of Opuntia ficus-indica flowers in-vivo regarding disease progression in a comparative experimental study to praziquantel (PZQ). Phytochemical investigation of flowers extract resulted in thirteen phenolic and flavonoid compounds. Structures of these compounds were elucidated by UV and 1D⁄2D 1H⁄ 13C NMR spectroscopy and compared with the literature data. From the 49th day after infection, mice infected with Schistosoma mansoni were given the extract (200 mg/kg) orally every day for 5 days, whereby the Praziquantel (500 mg/kg) was used as reference drug. The parasitological parameters (total worm burden, tissue egg load and oogram pattern) per gram of liver or intestinal tissue of the infected and the treated mice were counted. A histopathological examination of the liver granuloma took place as well. Opuntia ficus-indica extract caused a substantial decrease in the number of worms and eggs (%); the extract also decreased worm and egg burdens moderately. The results demonstrated the flower extract as a promising antischistosomal activity due to the parasitological and histopathological changes induced, besides it highlights the importance of its polyphenolic constituents.

Hepatic Scaling Factors for In Vitro-In Vivo Extrapolation of Metabolic Drug Clearance in Patients with Colorectal Cancer with Liver Metastasis.

In vitro-in vivo extrapolation (IVIVE) linked with physiologically based pharmacokinetics (PBPK) modeling is used to predict the fates of drugs in patients. Ideally, the IVIVE-PBPK models should incorporate systems information accounting for characteristics of the specific target population. There is a paucity of such scaling factors in cancer, particularly microsomal protein per gram of liver (MPPGL) and cytosolic protein per gram of liver (CPPGL). In this study, cancerous and histologically normal liver tissue from 16 patients with colorectal liver metastasis were fractionated to microsomes and cytosol. Protein content was measured in homogenates, microsomes, and cytosol. The loss of microsomal protein during fractionation was accounted for using corrections based on NADPH cytochrome P450 reductase activity in different matrices. MPPGL was significantly lower in cancerous tissue (24.8 ± 9.8 mg/g) than histologically normal tissue (39.0 ± 13.8 mg/g). CPPGL in cancerous tissue was 42.1 ± 12.9 mg/g compared with 56.2 ± 16.9 mg/g in normal tissue. No correlations between demographics (sex, age, and body mass index) and MPPGL or CPPGL were apparent in the data. The generated scaling factors together with assumptions regarding the relative volumes of cancerous versus noncancerous tissue were used to simulate plasma exposure of drugs with different extraction ratios. The PBPK simulations revealed a substantial difference in drug exposure (area under the curve), up to 3.3-fold, when using typical scaling factors (healthy population) instead of disease-related parameters in cancer population. These indicate the importance of using population-specific scalars in IVIVE-PBPK for different disease states. SIGNIFICANCE STATEMENT: Accuracy in predicting the fate of drugs from in vitro data using IVIVE-PBPK depends on using correct scaling factors. The values for two of such scalars, namely microsomal and cytosolic protein per gram of liver, is not known in patients with cancer. This study presents, for the first time, scaling factors from cancerous and matched histologically normal livers. PBPK simulations of various metabolically cleared drugs demonstrate the necessity of population-specific scaling for model-informed precision dosing in oncology.

Scaling Drug Clearance from Adults to the Young Children for Drugs Undergoing Hepatic Metabolism: A Simulation Study to Search for the Simplest Scaling Method

Previous research showed that scaling drug clearance from adults to children based on body weight alone is not accurate for all hepatically cleared drugs in very young children. This study systematically assesses the accuracy of scaling methods that, in addition to body weight, also take age-based variables into account for drugs undergoing hepatic metabolism in children younger than five years, namely scaling with (1) a body weight-based function using an age-dependent exponent (ADE) and (2) a body weight-based function with fixed exponent of 0.75 (AS0.75) combined with isoenzyme maturation functions (MFPBPK) similar to those implemented in physiologically based pharmacokinetic (PBPK) models (AS0.75 + MFPBPK). A PBPK-based simulation workflow was used, including hypothetical drugs with a wide range of properties and metabolized by different isoenzymes. Adult clearance values were scaled to seven typical children between one day and four years. Prediction errors of ± 50% were considered reasonably accurate. Isoenzyme maturation was found to be an important driver of changes in hepatic metabolic clearance in children younger than five years, which prevents the systematic accuracy of ADE scaling. AS0.75 + MFPBPK, when accounting for maturation of isoenzymes and microsomal protein per gram of liver (MPPGL), can reasonably accurately scale hepatic metabolic clearance for all low and intermediate extraction ratio drugs except for drugs binding to alpha-1-acid glycoprotein in neonates. As differences in the impact of changes in system-specific parameters on drugs with different properties yield differences in clearance ontogeny, it is unlikely that for the remaining drugs, scaling methods that do not take drug properties into account will be systematically accurate.

Impact of Ethnicity-Specific Hepatic Microsomal Scaling Factor, Liver Weight, and Cytochrome P450 (CYP) 1A2 Content on Physiologically Based Prediction of CYP1A2-Mediated Pharmacokinetics in Young and Elderly Chinese Adults.

The vast majority of physiological and biological data required for physiologically based predictions are primarily available in Caucasians rather than other ethnic populations, which leads to a lack of confidence in the application of physiologically based pharmacokinetic (PBPK) modeling for ethnicity-specific prediction of pharmacokinetics in the Chinese population. In this study we recalibrate the system parameters of Chinese-specific PBPK modeling and explore for the first time the relative importance of ethnicity-specific microsomal protein per gram of liver (MPPGL), liver weight, and cytochrome P450 (CYP) 1A2 abundance to the projection of drug disposition mediated by CYP1A2 in young and elderly Chinese adults. Chinese MPPGL levels and associated variability were parameterized and incorporated for the first time into ethnicity-specific PBPK models for the Chinese adults. Parameterization of Chinese liver weights was also recalibrated on the basis of autopsy data from Chinese individuals (n = 4081) across the entire adult age range. Uncertainty surrounding the Chinese-specific CYP1A2 content has also been explored and clarified by conducting ethnicity-related PBPK simulations under different scenarios. Various ethnicity-related or 'what-if' scenarios for PBPK modeling were implemented to assess the predictive performance and explore the relative importance of ethnicity-specific MPPGL and liver weight to the projection of drug disposition mediated by CYP1A2 in terms of two typical CYP1A2 substrates, caffeine and theophylline, in young and elderly Chinese adults by comparing the predicted concentration-time data and associated pharmacokinetic parameter estimates with observations. Compared with 0.85, the liver scalar of 0.9 generally produced more accurate liver weight levels in virtual Chinese peers. Additionally, simulated MPPGL levels on the basis of Caucasian data were not able to reflect the age-independent pattern observed in Chinese adults, dissimilar to that on the basis of Chinese-specific adult MPPGL data. The modeling Scenarios A and B provided similar predictions for theophylline pharmacokinetics in young Chinese adults across different age groups, while Scenario B provided the most accurate prediction for theophylline pharmacokinetics in elderly Chinese adults. However, the use of a stratified value of CYP1A2 content derived from a Han Chinese cohort with a small sample size instead of the pooled value of all Chinese cohorts involved regardless of Chinese sub-ethnicity resulted in inadequate prediction of CYP1A2-mediated pharmacokinetics in terms of caffeine and theophylline in either young or elderly Chinese subjects. Additionally, the impact of ethnic-specific MPPGL on predictive accuracy of theophylline pharmacokinetics in elderly Chinese subjects is more evident than that of liver weight. We provided quantitative information pertaining to Chinese-specific levels of liver weight and MPPGL, and recalibrated these system parameters for PBPK modeling for young and elderly Chinese subjects. Uncertainty surrounding the Chinese-specific CYP1A2 content has also been clarified. PBPK modeling based on the recalibrated system parameters can accurately simulate CYP1A2-mediated pharmacokinetics in both young and elderly Chinese adults, particularly in elderly individuals.

Liver Antioxidants in Relation to Beak Morphology, Gizzard Size and Diet in the Common Eider Somateria mollissima.

Antioxidants in the liver are particularly abundant in capital breeders that rely on stored resources for egg production. Capital breeders like eider (hereafter common eider) Somateria mollissima have disproportionately large livers with low levels of coenzyme Q10 when compared to other bird species. Concentrations of total carotenoids and vitamin E in the livers of eiders were smaller than predicted for similarly sized bird species. Eiders with high body condition estimated as body mass relative to skeletal body size had high levels of total carotenoids and low levels of coenzyme Q10. The concentration of total carotenoids per gram of liver increased with age, and vitamin E and total carotenoids accumulated during the winter onwards from February to peak at the start of incubation in April. Total vitamin E, total carotenoids, and coenzyme Q10 per gram of liver decreased with increasing beak volume. The size of the empty gizzard increased with increasing liver mass but decreased with total carotenoids and coenzyme Q10. The main components of the diet were blue mussels Mytilus edulis (40%), draft whelk Nassarius reticulatus (27%), and periwinkle Littorina littorea (10%). The concentration of vitamin E increased with the number of razor clams Ensis sp. and draft whelks in the gizzard and the concentration of total carotenoids increased with the number of beach crabs Carcinus maenas. These observations are consistent with the hypothesis that eiders are limited in their levels of antioxidants through food limitation. Furthermore, they imply that diet and morphological characters involved in food acquisition and processing are important determinants of the level of antioxidants in the liver.

Effect of regulatory T cells on the efficacy of the fatty acid-binding protein vaccine against Schistosoma japonicum.

Schistosomiasis is one of the most devastating parasitic diseases, making it imperative to develop efficient vaccines to control the causative flatworms called schistosomes. Regulatory T cells (Tregs) and the Th1 immune response have been implicated in the effectiveness of vaccines to control schistosomiasis, but the mechanisms underlying their effects are unclear. In this study, we evaluated the role of Tregs on the efficacy of the 14kDa FABP (fatty acid-binding protein) vaccine against Schistosoma japonicum. BALB/c female mice were randomly divided into five groups: an uninfected group, infected control group, anti-CD25 monoclonal antibody (anti-CD25 mAb) group, FABP group, and combined anti-CD25 mAb and FABP group. Compared with FABP alone, a combined treatment with FABP and anti-CD25 mAb increased the rate of S. japonicum inhibition in mice from 30.3 to 56.08% and decreased the number of eggs per gram of liver. Compared with that of the infected control group, the percentage of Tregs in the spleen decreased significantly after single or combined treatment with FABP and anti-CD25 mAb, while it increased gradually in the anti-CD25 mAb group. Further, the secretion of Th1 cytokines, IFN-γ, and IL-2 increased in splenocytes in the anti-CD25 mAb group. Our results indicate that anti-CD25 mAb partially blocks Tregs and concomitantly enhances the Th1 type immune response, thereby enhancing the protective effect of the FABP vaccine.

Allometric scaling of hepatic biotransformation in rainbow trout

Biotransformation may substantially impact the toxicity and accumulation of xenobiotic chemicals in fish. However, this activity can vary substantially within and among species. In this study, liver S9 fractions from rainbow trout (4–400 g) were used to evaluate relationships between fish body mass and the activities of phase I and phase II metabolic enzymes. An analysis of log-transformed data, expressed per gram of liver (g liver−1), showed that total cytochrome P450 (CYP) concentration, UDP-glucuronosyltransferase (UGT) activity, and glutathione S-transferase (GST) activity exhibited small but significant inverse relationships with fish body weight. In contrast, in vitro intrinsic clearance rates (CLIN VITRO,INT) for three polycyclic aromatic hydrocarbons (PAHs) increased with increasing body weight. Weight normalized liver mass also decreased inversely with body weight, suggesting a need to express hepatic metabolism data per gram of body weight (g BW−1) in order to reflect the metabolic capabilities of the whole animal. When the data were recalculated in this manner, negative allometric relationships for CYP concentration, UGT activity, and GST activity became more pronounced, while CLIN VITRO,INT rates for the three PAHs showed no significant differences across fish sizes. Ethoxyresorufin O-deethylase (EROD) activity normalized to tissue weight (g liver−1) or body weight (g BW−1) exhibited a non-monotonic pattern with respect to body weight. The results of this study may have important implications for chemical modeling efforts with fish.

Differential growth of Mycobacterium leprae strains (SNP genotypes) in armadillos

Leprosy (Hansen's Disease) has occurred throughout human history, and persists today at a low prevalence in most populations. Caused by Mycobacterium leprae, the infection primarily involves the skin, mucosa and peripheral nerves. The susceptible host range for Mycobacterium leprae is quite narrow. Besides humans, nine banded armadillos (Dasypus novemcinctus) and red squirrels (Sciurus vulgaris) are the only other natural hosts for M. leprae, but only armadillos recapitulate the disease as seen in humans. Armadillos across the Southern United States harbor a single predominant genotypic strain (SNP Type-3I) of M. leprae, which is also implicated in the zoonotic transmission of leprosy. We investigated, whether the zoonotic strain (3I) has any notable growth advantages in armadillos over another genetically distant strain-type (SNP Type-4P) of M. leprae, and if M. leprae strains manifest any notably different pathology among armadillos. We co-infected armadillos (n = 6) with 2 × 109 highly viable M. leprae of both strains and assessed the relative growth and dissemination of each strain in the animals. We also analyzed 12 additional armadillos, 6 each individually infected with the same quantity of either strain. The infections were allowed to fulminate and the clinical manifestations of the disease were noted. Animals were humanely sacrificed at the terminal stage of infection and the number of bacilli per gram of liver, spleen and lymph node tissue were enumerated by Q-PCR assay. The growth of M. leprae strain 4P was significantly higher (P < 0.05) than 3I when each strain was propagated individually in armadillos. Significantly (P < 0.0001) higher growth of the 4P strain also was confirmed among animals co-infected with both 3I and 4P strain types using whole genome sequencing. Interestingly, the zoonotic strain does not exhibit any growth advantage in these non-human hosts, but the varied proliferation of the two M. leprae strains within armadillos suggest there are notable pathological variations between M. leprae strain-types.

The use of 99mTc-phytate for assessment the protective effect of vitamin E against hepatotoxicity induced by methotrexat in rat.

In this study, we investigated the protective effect of vitamin E against methotrexate (MTX)-induced hepatotoxicity by quantitative liver 99mTc-phytate uptake and liver imaging and to compare its effect with histopathology in rat. Rats were divided into five groups as control, solvent, Vit E (100 mg/kg), MTX (20 mg/kg), Vit E + MTX and. Vit E was intraperitoneally administrated for 17 days before MTX injection and continued for 4 days. 99mTc-phytate was injected through the tail of rats after the drug administration. The percentage of the injected dose per gram of liver and spleen tissues (%ID/g) was calculated. Liver imaging was obtained with gamma camera. In other experiment, liver of treated rats were assessed for histopathology. 99mTc-phytate uptake per gram tissue of the livers as %ID/g in control, solvent, MTX, Vit E, Vit E + MTX and MTX groups were 8.99% ± 1.37, 8.53% ± 2.91, 8.65% ± 3.84, 3.22% ± 1.09 and 8.38% ± 2.68. Vit E administration with MTX resulted in a significant increasing in the level of %ID/g. Vit E treatment improved the shape of live in planner image. Histophatological examinations showed a protective effect of Vit E against MTX-induced hepatoxicity in rats. The results showed that Vit E significantly attenuates the MTX-induced hepatotoxicity in rats, and 99mTc-phytate uptake in liver as well as liver image to be acceptable techniques for assessment of liver and spleen damages and/or their tissues protective effects in animal model.

Quantification of Hepatic Lipid Using 7.0T Proton Magnetic Resonance Spectroscopy and Computed Tomography in Mild Alcoholic Steatotic Mice.

Background:In vivo proton magnetic resonance spectroscopy (1H MRS) has been used to semi-quantify hepatic lipids in preclinical and clinical studies of fatty liver disease. Quantifying absolute amount of liver lipids utilizing 1H MRS and computerized tomography (CT) is essential to accurately interpret hepatic steatosis.Purpose:To establish reliable parameters to convert relative hepatic lipid levels obtained by 1H-MRS and liver volumes by CT to the absolute amount of liver lipids in a mild hepatic steatosis, and to determinate the correlation between these absolute liver lipids with liver triglyceride (TG) and cholesterol (Chol) measured by biochemistry assays.Methods:Mild steatosis was induced in mice by a 3 week ethanol diet containing standard lipids. Evaporated liver water was measured after baking liver tissues and volume of liver was measured using water displacement. 1H MRS semiquantitation of hepatic lipids and CT measurement of liver volume were performed and then used to calculate amount of liver lipids. These data were compared with liver TG and Chol.Results:Percentage of liver water and liver density were persistent in two groups and were used to convert the percentage of liver lipids to liver water by 1H-MRS to the absolute amount of liver lipids per gram of liver or per milliliter of CT volume. Using 1H-MRS and biochemical assays, an increase of liver lipids was confirmed in mild steatosis mice compared to controls (P<0.01). The amounts of imaging detected liver lipids were strongly correlated to liver TG and Chol measured by biochemical assays in mild steatosis mice.Conclusion:1H MRS and CT liver imaging techniques are able to quantify absolute hepatic lipid levels utilizing relative persistent parameters percentage of liver water and liver density in a preclinical mild steatosis setting.

The effects of aging on hepatic microsomal scaling factor and hepatocellularity number in the horse

1. Scaling factor values for the in vitro-in vivo extrapolation of hepatic metabolic clearance for xenobiotics have not yet been determined in horses. Scaling factors were determined by comparing the total protein and or cytochrome (CYP) P450 content in microsomes and cryopreserved hepatocytes against the content in the liver.2. Microsomal protein per gram of liver (MPPGL) and hepatocellularity number per gram of liver (HPGL) using CYP P450 content method ranged 41–73 mg/gram of liver (mean= 57 mg/gram of liver, n = 39) and 146–320 × 106 cells/g of liver (mean = 227× 106 cells/g of liver, n = 18), respectively and 156–352 × 106 cells/g of liver (mean = 232× 106 cells/g of liver) using total protein method.3. A non-monotonic and inverse relationship between age and MPPGL and HPGL, respectively, was observed. Between one and 20 y of age, the liver cell size decreases as age increases. Subsequently, the cell size increases until the hepatocytes of the oldest horses approached the size found in the youngest horses. Hepatocyte density was inversely related to the size of the hepatocytes.4. This study provides the first extensive and comprehensive data demonstrating the relationship between the size of hepatocytes and HPGL in any species.

From Genotype to Phenotype: Cytochrome P450 2D6-Mediated Drug Clearance in Humans.

How genotypic variation results in phenotypic differences is still a challenge for biology. In the field of drug metabolism, the means by which specific cytochrome P4502D6 (CYP2D6) genotypes yield different phenotypes at various levels (molecular, cellular, and organismal) is an important question, as differences in CYP2D6 activity can contribute to adverse drug reactions. Herein, the genotype of CYP2D6 was determined along with the absolute content of CYP2D6 and microsomal protein per gram of liver in human liver microsomes, the molecular, cellular (microsomal, tissue, organ), and organismal phenotype of CYP2D6 determined; the effect of genotype on each phenotype of CYP2D6-mediated dextromethorphan clearance (CL) was delineated, and the overall genotype-phenotype relationship for CYP2D6 was charted. We demonstrate that changes in the cellular and organismal CL phenotypes are markedly greater than changes seen at the molecular level. With individuals carrying the 1661CC polymorphism, for example, the most noticeable change took place in organ CL phenotype (4.17-fold), followed by tissue (3.75-fold), organism (3.69-fold), microsomal (3.09-fold), and molecular (1.66-fold) phenotypes. In addition, the biggest intragenotype individual coefficient of variation in organismal phenotype was observed in the 1661GG individuals, which reached 104.5%, followed by that of 100TT, 100CT, 1661GC, 100CC, and 1661CC polymorphisms (102.7%, 62.4%, 53.5%, 49.7%, and 44.8%, respectively). Our study has allowed us to chart the genotype-phenotype relationship for CYP2D6 from the molecular to the organismal level as well as allowed us to determine intragenotype individual variation in phenotype with each genotype.

Discarded Livers Find a New Life: Engineered Liver Grafts Using Hepatocytes Recovered From Marginal Livers.

Treatment for end-stage liver failure is restricted by the critical shortage of donor organs; about 4000 people die in the USA while waiting for a transplantable organ. This situation has been a major driving force behind the rise of tissue engineering to build artificial tissues/organs. Recent advancements in creating transplantable liver grafts using decellularized liver scaffolds bring the field closer to clinical translation. However, a source of readily available and highly functional adult hepatocytes in adequate numbers for regenerative liver therapies still remains unclear. Here, we describe a new method to utilize discarded livers to make transplantable new liver grafts. We show that marginal donor livers damaged due to warm ischemia could be treated with machine perfusion to yield 39 million viable hepatocytes per gram of liver, similar to fresh livers, and these cells could be used to repopulate decellularized liver matrix (DLM) scaffolds to make transplantable liver grafts. The hepatocytes from recovered livers sustained their characteristic epithelial morphology while they exhibited slightly lower protein synthesis functions both in plate cultures and in recellularized liver grafts. The dampened protein synthesis was attributed to residual endoplasmic reticulum stress found in recovered cells. The results here represent a unique approach to reengineer transplantable liver grafts solely from discarded organs.

The impact of variation in scaling factors on the estimation of internal dose metrics: a case study using bromodichloromethane (BDCM)

A rate for hepatic metabolism (Vmax) determined in vitro must be scaled for in vivo use in a physiologically based pharmacokinetic (PBPK) model. This requires the use of scaling factors such as mg of microsomal protein per gram of liver (MPPGL) and liver mass (FVL). Variation in MPPGL and FVL impacts variation in Vmax, and hence PBPK model-derived estimates of internal dose used in dose response analysis. The impacts of adult human variation in MPPGL and FVL on estimates of internal dose were assessed using a human PBPK model for bromodichloromethane (BDCM), a water disinfection byproduct, for multiple internal dose metrics for two exposure scenarios (single 0.25 liter drink of water or 10 min shower) under plausible (5 μg/L) and high level (20 μg/L) water concentrations. For both concentrations, all internal dose metrics were changed less than 5% for the showering scenario (combined inhalation and dermal exposure). In contrast, a 27-fold variation in area under the curve (AUC) for BDCM in venous blood was observed at both oral exposure concentrations, whereas total amount of BDCM metabolized in liver was relatively unchanged. This analysis demonstrates that variability in the scaling factors used for in vitro to in vivo extrapolation (IVIVE) for metabolic rate parameters can have a significant route-dependent impact on estimates of internal dose under environmentally relevant exposure scenarios. This indicates the need to evaluate both uncertainty and variability for scaling factors used for IVIVE.