H-ras gene is expressed at the G1 phase in primary cultures of hepatocytes

The expression of c-H-ras and proliferating cell nuclear antigen (PCNA) in primary cultures of rat hepatocytes was determined in order to elucidate the relationship between the c-H-ras gene and the S phase of the cell cycle. In cells treated with EGF, elevation of c-H-ras expression was detected at the 22nd, 34th, 44th, and 54th h after plating, PCNA expression and DNA synthesis were detected at the 44th and 54th h. In cells without EGF treatment, only c-H-ras expression was detected at the 44th and 54th h. In our previous report, we showed that c-myc expression increased within several hours after plating, suggesting that isolated hepatocytes traverse from G0 to G1 under culture conditions, regardless of EGF treatment. These results clearly showed that the c-H-ras gene of adult rat hepatocytes was expressed in the mid-to-late G1 phase of the cell cycle as well as in the early S phase in primary culture.     

Isolation and Characterization of an Enterobacter cloacae Strain That Reduces Hexavalent Chromium under Anaerobic Conditions

An Enterobacter cloacae strain (HO1) capable of reducing hexavalent chromium (chromate) was isolated from activated sludge. This bacterium was resistant to chromate under both aerobic and anaerobic conditions. Only the anaerobic culture of the E. cloacae isolate showed chromate reduction. In the anaerobic culture, yellow turned white with chromate and the turbidity increased as the reduction proceeded, suggesting that insoluble chromium hydroxide was formed. E. cloacae is likely to utilize toxic chromate as an electron acceptor anaerobically because (i) the anaerobic growth of E. cloacae HO1 accompanied the decrease of toxic chromate in culture medium, (ii) the chromate-reducing activity was rapidly inhibited by oxygen, and (iii) the reduction occurred more rapidly in glycerol- or acetate-grown cells than in glucose-grown cells. The chromate reduction in E. cloacae HO1 was observed at pH 6.0 to 8.5 (optimum pH, 7.0) and at 10 to 40°C (optimum, 30°C).     

Molecular cloning and nucleotide sequence of human pancreatic prechymotrypsinogen cDNA

The cDNA clone encoding human prechymotrypsinogen was isolated from a human pancreas cDNA library and its nucleotide sequence was determined. The sequence consists of a 16 bp 5' non-coding region, a 789 bp amino acid coding region and a 60 bp 3' non-coding region. The predicted product consists of 263 amino acids, including 18 amino acids for a signal peptide and 15 amino acids possible for an activation peptide. Southern blot analyses using the cloned cDNA as a probe revealed that human genomic DNA carries at least two genes that are related to chymotrypsinogen.     

Hypoxia-induced fibre type transformation in rat hindlimb muscles Histochemical and electro-mechanical changes

Twelve male Sprague-Dawley rats (21 days old) were randomly assigned into two experimental groups: sea level control (CONT) and hypobaric hypoxia (HYPO). The HYPO rats were kept in an hypobaric chamber maintaining a simulated altitude of 4000 m (61.1 kPa). After 10 weeks of treatment, the rat hindlimb muscles [soleus (SOL) and extensor digitorum longus (EDL)] were subjected to histochemical and electro-mechanical analyses. Results indicated that compared to CONT the HYPO SOL muscle had a significantly greater relative distribution of fast-twitch-oxidative-glycolytic (FOG) fibres (28.9% SEM 2.0 vs 18.3% SEM 1.8, P less than 0.01) with a significant decrease in slow twitch oxidative fibre distribution (69.5% SEM 2.4 vs 82.9% SEM 3.1, P less than 0.01). Compared to CONT the HYPO EDL muscle also manifested a significant increase in FOG fibre distribution (51.6% SEM 0.8 vs 46.6% SEM 1.1, P less than 0.01), but this was accompanied by a significant decrease in fast twitch glucolytic fibres (44.3% SEM 0.9 vs 49.2% SEM 1.7, P less than 0.05). These histochemical fibre type transformations accompanied significant and expected changes in the electro-mechanical parameters tested in situ, e.g. maximal twitch force, maximal rate of force development, contraction time, half relaxation time, force: frequency curve, and fatigability. It was concluded that chronic hypobaric hypoxia could have a potent influence upon the phenotype expression of muscle fibres.     

Multiple expression of keratins, vimentin, and S-100 protein in pleomorphic salivary adenomas.

Immunohistochemical staining for S-100 protein and the intermediate filaments keratin and vimentin, was made in 41 salivary adenomas. In pleomorphic adenomas, great heterogeneity in the staining, as well as multiple and co-expressions of these proteins were found in the outer tumor cells of tubulo-ductal structures and modified myoepithelial cells, but not in the luminal tumor cells. All the outer tumor cells stained for S-100 protein, 97% for K8.12 keratin and 85% for vimentin. Of these cells, 29% showed multiple expression of K8.12 keratin, vimentin, and S-100 protein, and 17% showed co-expression of K8.12 and S-100 protein. Modified and neoplastic myoepithelial cells showed similar expressions of these proteins to those of outer tumor cells; myoepithelioma cells displayed the most complicated pattern, being positive for KL1, PKK1, and K8.12 keratins, vimentin and S-100 protein. In luminal tumor cells there was a heterogeneous expression of KL1 and PKK1 in 82%, and of KL1, PKK1, and K8.12 in only 14.7%. Based on the immunohistochemical findings obtained with different monoclonal antibodies in pleomorphic salivary adenomas, outer tumor cells may be derived from ductal basal cells and luminal tumor cells from intercalated duct cells.     

Regional excitatory and inhibitory amino acid levels in epileptic El mouse brain

Inbred mutant El mice are highly susceptible to convulsive seizures upon tossing stimulation. The levels of excitatory (e.g. glutamate and aspartate) and inhibitory amino acids [e.g. -aminobutyrate (GABA)] were examined in discrete regions of stimulated El mice [El(+)] non-stimulated El mice [El(-)] and ddY mice, which do not have convulsive disposition. In comparison with ddY, a general increased levels of aspartate, glutamate, glutamine, and taurine were detected in brain regions of El(-). The levels of GABA and glycine were almost the same in ddY and El(-). Compared to El(+), the levels of aspartate, glutamate, glutamine, and GABA in El(-) were either the same or higher. In the case of taurine and glycine, the levels in El(-) were either the same or lower than El(+). Alanine is special in that El(-) have a higher level than El(+) in hippocampus but lower in cerebellum. Furthermore, while marked changes were registered in several brain regions, none of the amino acids investigated showed any significant differences in the hypothalamus of three different groups of mice.     

Effects of muscimol and baclofen on levels of monoamines and their metabolites in the El mouse brain

We compared the changes in monoamines and their metabolites in the El mouse brain induced by GABA-A and GABA-B receptor agonists. Muscimol was used as a GABA-A receptor agonist, and baclofen as a GABA-B receptor agonist. Muscimol (3 mg/kg) significantly increased the DOPAC level in all parts of the mouse brain and the HVA level in the cortex, striatum, and midbrain. No significant change was observed in the dopamine (DA) level. These findings suggest that muscimol may accelerate both the synthesis and catabolism of DA. Baclofen (20 mg/kg) increased the DA level in the hippocampus and midbrain, and the DOPAC level in the hippocampus. Muscimol increased 5-HIAA levels and decreased 5-HT levels. This result suggests that 5-HT metabolism is accelerated by muscimol. No change in 5-HT or 5-HIAA levels was induced by baclofen. The GABA-A receptor system seems to have a potent effect not only on DA neurons, but on 5-HT neurons. However, the GABA-B receptor system appears to have almost no effect on 5-HT neurons, though it appears to have some effect on DA neurons.     

Inhibition of ovulation, steroidogenesis and collagenolytic activity in rabbits by sulpiride-induced hyperprolactinaemia

Ovulation induced by hCG in rabbits was reduced significantly (P less than 0.005) by sulpiride-induced hyperprolactinaemia. The pre- and post-ovulatory increases in peripheral and ovarian venous progesterone (but not oestradiol or testosterone) were suppressed in the treated animals. The condition of hyperprolactinaemia also prevented the usual changes in 2,4-dinitrophenyl-Pro-Gln-Gly-Ile-Ala-Gly-Gln-D-Arg-OH peptidase (DNP-peptidase) and alpha-N-benzoyl-DL-Arg-beta-naphthylamide hydrolase (BANA-hydrolase) activities in follicular tissue that had been stimulated by an ovulatory dose of hCG. These results suggest that inhibition of progesterone production and collagenolytic enzyme activity by sulpiride-induced hyperprolactinaemia may be responsible for the ovulatory dysfunction that occurs when a mammal has a high level of circulating prolactin.     

The genotoxicity of a variety of aniline derivatives in a DNA repair test with primary cultured rat hepatocytes

The genotoxicity of a variety of aniline derivatives was examined by a DNA repair test with rat hepatocytes. Out of 37 aniline derivatives, 6 chemicals, i.e., 2,4,6-trimethylaniline (mesidine), 2,4-xylidine, 3,5-diaminobenzoic acid, 3,4-diaminochlorobenzene, 2-chloro-4-methylaniline and 4-chloro-N-methylaniline, elicited positive DNA repair responses. The results are in agreement with the bacterial mutagenicities with or without norharman of these compounds. Positive compounds of unknown carcinogenicity in the present assay, i.e., 3,5-diaminobenzoic acid, 2-chloro-4-methylaniline and 4-chloro-N-methylaniline are suspected of being potentially carcinogenic.     

Immunocytochemical localization of ornithine transcarbamylase in rat intestinal mucosa. Light and electron microscopic study

We investigated light and electron microscopic localization of ornithine transcarbamylase (OTC) in rat intestinal mucosa. In the immunoblotting assay of OTC-related protein, a single protein band with a molecular weight of about 36,500 is observed in extracts of liver and small intestinal mucosa but is not observed in those of stomach and large intestine. For light microscopy, tissue slices of the digestive system were embedded in Epon and stained by using anti-bovine OTC rabbit IgG and the immunoenzyme technique. For electron microscopy, slices of these and the liver tissues were embedded in Lowicryl K4M and stained by the protein A-gold technique. By light microscopy, the absorptive epithelial cells of duodenum, jejunum, and ileum stained positively for OTC, but stomach, large intestine, rectum, and propria mucosa of small intestine were not stained. Electron microscopy showed that gold particles representing the antigenic sites for OTC were confined to the mitochondrial matrix of hepatocytes and small intestinal epithelial cells. However, the enzyme was detected in mitochondria of neither liver endothelial cells, submucosal cells of small intestine, nor large intestinal epithelial cells. Labeling density of mitochondria in the absorptive epithelial cells of duodenum, jejunum, and ileum was about half of that in liver cells.