Molecular cloning and sequencing of the cDNA encoding the catalytic subunit of mouse glutamate-cysteine ligase.

Reverse transcription-polymerase chain reaction (RT-PCR) was used for the enzymatic synthesis of cDNA sequences encompassing the open reading frame for the catalytic subunit of mouse kidney glutamate-cysteine ligase (Glclc). Comparison of the mouse Glclc cDNA sequence and predicted protein sequence with that of rat Glclc and human GLCLC revealed between 94.8% and 88.4% cDNA homology and 98.4% to 95% amino acid identity, respectively.

Molecular cloning and sequencing of the cDNA encoding mouse glutamate-cysteine ligase regulatory subunit.

Reverse transcription-polymerase chain reaction (RT-PCR) was used to amplify and clone the regulatory subunit of mouse glutamate-cysteine ligase (Glclr) using primers adapted from the published rat Glclr cDNA sequence, and from mouse genomic DNA. Amplified cDNA was cloned into a plasmid vector, and additional RT-PCR reactions coupled with 3' RACE were used to amplify and sequence 3' regions covered by the rat primer. Comparison of the mouse Glclr cDNA sequence and predicted protein sequence with that of rat Glclr and human GLCLR revealed extensive homology in cDNA and amino acid sequences among these species.

The effect of the sex composition of jobs on starting wages in an organization: findings from the NLSY.

We show that individuals in a job with a higher percentage of females earn lower starting wages with an employing organization. This holds true with controls for individuals' human capital, job demands for skill or difficult working conditions, and detailed industry. We use a measure of sex composition that applies to detailed jobs: cells in a three-digit census occupation by three-digit census industry matrix. We use pooled panel data from the 1979-1987 waves of the National Longitudinal Survey of Youth. The unit of analysis is the spell-the time in which a person worked for one organization. The dependent variable is the first wage in the spell. We use models with fixed-effects to control for unmeasured, unchanging individual characteristics; we also show results from OLS and weighted models for comparison. The negative effect on wages of the percentage female in one's job is robust across procedures for black women, white women, and white men. For black men the sign is always negative but the coefficient is often nonsignificant.

Further examination of the selective toxicity of CCl4 in rat liver slices.

Lipid peroxidation and loss of enzymes located predominantly in either periportal or centrilobular hepatocytes were investigated in precision-cut liver slices from male Sprague-Dawley rats. Pretreatment of animals with 80 mg/kg phenobarbital for the site-specific enzyme studies enhanced and accelerated CCl4 toxicity in slices resulting from increased radical formation. Liver slices were exposed to 0.57 mM CCl4 by vaporization using a roller incubation system at 37 degrees C for a total of 9 hr. Conjugated diene formation, an index of lipid peroxidation, was detected 15 min following CCl4 administration and increased over time. Loss of cytochrome P450 occurred in a time-dependent manner relative to controls where levels in treated slices were 42% of controls at 9 hr. A 48-hr fast prior to termination increased intracellular K+ leakage relative to that present in slices from fed animals. Significant leakage of glucose-6-phosphate dehydrogenase and beta-glucuronidase from centrilobular hepatocytes occurred 9 hr following CCl4 administration. The content of the periportal enzymes (lactate dehydrogenase and sorbitol dehydrogenase) was unchanged in the same slices over the duration of the experiment. Reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide, a mitochondrial selective dye and indicator of viability, was significantly lower in treated slices from phenobarbital-treated animals at 9 hr relative to controls. These studies demonstrate that precision-cut slices are an ideal in vitro system for mechanistic studies and the investigation of site-specific toxicants since the integral architecture of the liver and cellular identity are maintained.

Use of staphylococcal enterotoxin A-induced lymphoproliferation and interleukin 2 production as indicators of immunotoxicity.

Suppression of mitogen-induced splenocyte lymphoproliferation and interleukin 2 (IL-2) production can be used as indicators of immunotoxicity. Staphylococcal enterotoxin A (SEA) is both a potent mitogen and the most potent in vitro inducer of IL-2 production that has been described. An in vitro system was used to measure impairment of SEA-induced lymphoproliferation and IL-2 production using splenocytes from female C57BL/6 mice dosed with either cyclosporin A (30 mg/kg/day, 14 days), benzene (220, 440, or 880 mg/kg/day, 14 days), or vehicle. Splenocytes were stimulated with either concanavalin A (con A) or SEA. Benzene- and cyclosporin A-treated mice demonstrated significant decreases in splenocyte proliferation. IL-2 production was determined by incubating splenocyte culture supernatants with IL-2 dependent cytotoxic T-cells (CTLL-2), pulsing with 3H-thymidine, and determining amount of incorporated label. Cell proliferation and IL-2 production were inhibited by both benzene and cyclosporin A, effects more clearly demonstrated using SEA than con A. SEA was a superior mitogen compared to con A in the assays evaluated here.