Evaluation of the xenobiotic biotransformation capability of six rodent hepatoma cell lines in comparison with rat hepatocytes

Summary Phase I and II activities were examined in six rodent hepatoma cell lines and compared with those of cultured rat hepatocytes both in basal conditions and after exposure to 5μM methylcholanthrene, 2 mM phenobarbital, and 15μMβ-naphtoflavone. The metabolic profile of testosterone was also studied. The highest aryl hydrocarbon hydroxylase and 7-ethoxycoumarinO-deethylase activities were found in MH1C1 cells. Comparable values for 7-ethoxyresorufinO-deethylase activity, ranging from 21.6 to 42.9 pmol/mg × min, were observed in the hepatocytes and hepatoma cells, except the HTC cells. In contrast, only Fao cells showed 7-pentoxyresorufinO-depentylase activity at levels similar to those of hepatocytes (6.2±1.0 and 7.4±1.2 pmol/mg × min, respectively). Rat hepatocytes actively hydroxylatedp-nitrophenol, but this activity was not measurable in hepatoma cells. Glutathione transferase activity was maintained in all the hepatoma cell lines at similar levels to those found in hepatocytes (684 ± 56 nmol/mg × min). The seven hydroxylated metabolites of testosterone produced by cultured hepatocytes were negligible in hepatoma cells. Exposure of cells to inducers revealed that aryl hydrocarbon hydroxylase activity was mainly increased after treatment with 3-methylcholanthrene andβ-naphtoflavone, and the highest values were found in rat hepatocytes followed by MH1C1 and Fao cells. 3-Methylcholanthrene and naphtoflavone treatment also resulted in a marked increase in 7-ethoxyresorufinO-deethylase activity in hepatocytes as well as in H4IIC3, McA-Rh7777, MH1C1, and Fao cells. An enhancement of 7-ethoxycoumarinO-deethylase activity due to the three inducers was observed in both rat hepatocytes and hepatoma cells, with MH1C1 cells treated with methylcholanthrene showing the highest activity (727±74 pmol/mg × min). Increases in 7-pentoxyresorufinO-depentylase activity were detected after phenobarbital treatment of hepatocytes, MH1C1, and Fao cells, whereas a low response was observed in the other hepatoma cells. Of the six hepatoma cell lines examined, MH1C1 and Fao cells are the ones that are most similar to cultured rat hepatocytes in their expression of biotransformation activities.     

Influence of branched-chain amino acid composition of culture media on the synthesis of plasma proteins by serum-free cultured rat hepatocytes

Summary Supplementation of Ham's F12 culture medium with essential amino acids (EAA) up to the rat plasma levels increased the rates of synthesis of albumin and transferrin by cultured rat hepatocytes by 1.3 and 1.7, respectively. Fifty percent of this increase could be attributed to three of the EAA: the branched-chain amino acids (BCAA: Leu Ile and Val). Non-branched-chain essential amino acids (non-BC-EAA) stimulated only 25% of the increase produced by the whole EAA mixture. When each EAA was tested individually, none of them caused an appreciable increase in albumin and transferrin in culture medium. When the concentrations of all EAA were raised to rat postprandial portal levels, albumin and transferrin synthesis rates reached a maximum, increasing by 3.2 and 3.5, respectively. Supplementation with BCAA at postprandial portal concentrations increased albumin and transferrin synthesis rates by 2.2 and 2.0, respectively, and had no noteworthy effect on the synthesis of cellular proteins. Non-BC-EAA at their postprandial portal concentrations increased albumin and transferrin synthesis rates by 1.7 and 1.9, respectively. Supplementation with alanine to reach a nitrogen content equal to that of the modified EAA-enriched medium had no stimulatory effect. Our results show that EAA have a specific effect on the synthesis of plasma proteins by cultured hepatocytes, and that BCAA at physiologic concentrations account for the major part of this stimulatory effect. Consequently, EAA and particularly BCAA concentration should be elevated in serum-free nutrient media to sustain maximum plasma protein synthesis.     

Worldwide genetic relationships of pigs as inferred from X chromosome SNPs

The phylogeography of the porcine X chromosome has not been studied despite the unique characteristics of this chromosome. Here, we genotyped 59 single nucleotide polymorphisms (SNPs) in 312 pigs from around the world, representing 39 domestic breeds and wild boars in 30 countries. Overall, widespread commercial breeds showed the highest heterozygosity values, followed by African and American populations. Structuring, as inferred from F_ST and analysis of molecular variance, was consistently larger in the non‐pseudoautosomal (NPAR) than in the pseudoautosomal regions (PAR). Our results show that genetic relationships between populations can vary widely between the NPAR and the PAR, underscoring the fact that their genetic trajectories can be quite different. NPAR showed an increased commercial‐like genetic component relative to the PAR, probably because human selection processes to obtain individuals with high productive parameters were mediated by introgressing boars rather than sows.     

Effect of glucocorticoids on the expression of gamma-glutamyltransferase and tyrosine aminotransferase in serum-free-cultured hepatocytes

Glucocorticoids exert a known beneficial effect on cultured hepatocytes when present in culture medium, maintaining their polygonal morphology and ultrastructural organization throughout the days of culture. Parallel to this excellent morphology, hepatocytes cultured in serum-free conditions, but with continuous presence of Dexamethasone, retained after a week the ability to express tyrosine aminotransferase when stimulated by glucagon and glucocorticoids. The rise of gamma-glutamyltransferase was blocked in cultures supplemented by Dexamethasone.     

Location of phosphoglycerate mutase in rat skeletal muscle. An immunocytochemical and biochemical study

The subcellular distribution of phosphoglycerate mutase was studied by immunogold techniques. With the aid of highly affinity-purified anti-phosphoglycerate mutase antibodies, the enzyme was found in both cytosol and nucleus of rat skeletal muscle. No evidence of interaction with contractile proteins was observed in cytosol. Nuclear location was also confirmed biochemically using purified nuclear preparations from rat skeletal muscle. Only one immunoreactive nuclear band was observed by Western blot experiments and corresponded to that of phosphoglycerate mutase mobility. Activity measurements from nuclear extracts showed that 25% of total specific activity is found in the nuclei.     

Fluorometric assays in the study of nucleic acid-protein interactions : I. The use of diaminobenzoic acid as a reagent of DNA

The microfluorometric determination of DNA with diaminobenzoic acid in combination with a filter binding assay offers an easy and accurate procedure to study the interaction of proteins with any source of DNA. Using a highly polymerized commercial preparation of calf thymus DNA, the binding curve of histones or protamines changes from hyperbolic to increasingly sigmoidal depending on the length and temperature of incubation. The presence in the DNA preparation of small amounts of contaminating proteases, undetectable by conventional methods, is responsible for this change in the binding curve, since the presence of phenylmethylsulfonylfluoride in the reaction mixture or the removal of the proteases from the DNA produces only hyperbolic curves.     

Migration and Dependency: Mexican Countryside Proletarianization and the Seasonal Agricultural Worker Program

Dialectical Anthropology - This article addresses the Seasonal Agricultural Worker Program (SAWP) between Mexico and Canada by examining the forms of disposability and job insecurity of Mexicans...     

Temperature dependence for development of the whitefly predator <Emphasis Type="Italic">Clitostethus arcuatus</Emphasis> (Rossi)

This work aimed to study the biology of Clitostethus arcuatus (Rossi) (Coleoptera: Coccinellidae) under different temperatures and evaluate the optimum temperature for its mass rearing. Studies were carried out in the laboratory at four constant temperatures (15°C, 20°C, 25°C and 30°C), 75 ± 5% relative humidity and a photoperiod of 16 h light:8 h dark, in which C. arcuatus was fed ad libitum with nymphs of all instar of Aleyrodes proletella L. (Homoptera: Aleyrodidae) on Brassicae oleracea L. (var. Costata). The following biological parameters were evaluated: development time and survival rates of pre-imaginal stages, adult longevity (female and male), length of the pre-oviposition and oviposition periods, fecundity, fertility and percentage of egg hatching. Population growth parameters, the lower development threshold and the sum of effective temperatures were estimated. Temperatures ranging from 20°C to 30°C were suitable for the development of C. arcuatus, suggesting that this species is well adapted to the temperatures usually found inside greenhouses or in open fields in temperate regions. Although the intrinsic rate of natural increase and doubling time were similar at 25°C and 30°C, the temperature of 25°C was shown to be the most suitable for mass rearing and development of populations under field conditions, since the percentage of egg hatching and the accumulated survival rates of the pre-imaginal stages were the highest. Considering the estimated lower threshold for pre-imaginal development (7.9°C) and the sum of effective temperatures [293.6 degree-days (°D)], it is predicted for Ponta Delgada (Azores, Portugal) that the first adults of C. arcuatus should emerge by the first fortnight of February and that up to 12 generations per year can occur.     

Isolation and sequencing of a cDNA encoding the B isozyme of rat phosphoglycerate mutase.

Phosphoglycerate mutase consists of two kinds of different subunits, M and B. We previously sequenced a rat cDNA encoding the type-M subunit. Here, we report the sequence of the type-B subunit-encoding cDNA. This cDNA has 1754 bp and contains a long 3'-untranslated region of 897 bp.