Mucin secretion and PKC isoforms in SPOC1 goblet cells: differential activation by purinergic agonist and PMA

SPOC1 cells, which are a mucin-secreting model of rat airway goblet cells, possess a luminal P2Y2 purinoceptor through which UTP, ATP, and ATPgammaS stimulate secretion with EC50 values of approximately 3 microM. PMA elicits mucin secretion with high EC50 (75 nM) and saturation (300 nM) values. For the first time in airway mucin-secreting cells, the PKC isoforms expressed and activated by a secretagogue were determined using RT-PCR/restriction-enzyme mapping and Western blotting. Five isoforms were expressed: cPKCalpha, nPKCdelta and -eta, and aPKCzeta and -iota/lambda. PMA caused cPKCalpha and nPKCdelta to translocate to the membrane fraction of SPOC1 cells; only nPKCdelta so responded to ATPgammaS. Membrane-associated nPKCdelta and mucin secretion increased in parallel with ATPgammaS concentration and yielded EC50 values of 2-3 microM and maximum values of 100 microM. Effects of PMA to increase membrane-associated cPKCalpha and nPKCdelta saturated at 30 nM, whereas mucin secretion saturated at 300 nM, which suggests a significant PKC-independent effect of PMA on mucin secretion. A prime alternate phorbol ester-receptor candidate is the C1-domain protein MUNC13. RT-PCR revealed the expression of ubiquitous (ub)MUNC13-2 and its binding partner, DOC2-gamma. Hence, P2Y2 agonists activate nPKCdelta in SPOC1 cells. PMA activates cPKCalpha and nPKCdelta at high affinity and stimulates a lower affinity PKC-independent pathway that leads to mucin secretion.     

Identification of tissue-specific microRNAs from mouse

MicroRNAs (miRNAs) are a new class of noncoding RNAs, which are encoded as short inverted repeats in the genomes of invertebrates and vertebrates. It is believed that miRNAs are modulators of target mRNA translation and stability, although most target mRNAs remain to be identified. Here we describe the identification of 34 novel miRNAs by tissue-specific cloning of approximately 21-nucleotide RNAs from mouse. Almost all identified miRNAs are conserved in the human genome and are also frequently found in nonmammalian vertebrate genomes, such as pufferfish. In heart, liver, or brain, it is found that a single, tissue-specifically expressed miRNA dominates the population of expressed miRNAs and suggests a role for these miRNAs in tissue specification or cell lineage decisions. Finally, a miRNA was identified that appears to be the fruitfly and mammalian ortholog of C. elegans lin-4 stRNA.     

Culture conditions influencing phytase production of Mitsuokella jalaludinii,a new bacterial species from the rumen of cattle

AIMS: To determine the effectiveness of combined treatments with chemicals, heat and ultrasound in killing or removing Salmonella and Escherichia coli O157:H7 on alfalfa seeds intended for sprout production. METHODS AND RESULTS: Alfalfa seeds inoculated with Salmonella or E. coli O157:H7 were treated with ultrasound (38.5-40.5 kHz) in solutions containing 1% Ca(OH)(2), 1% Tween 80, 1% Ca(OH)(2) plus 1% Tween 80, 160 microg ml(-1) Tsunami 200 and 0.5% Fit at 23 and 55 degrees C for 2 and 5 min. Highest reductions were in chemical solutions at 55 degrees C, but seed viability was also reduced compared with treatment at 23 degrees C. Inactivation of Salmonella and E. coli O157:H7 was generally enhanced by simultaneous treatments with ultrasound, chemicals and heat. CONCLUSIONS: Ultrasound treatment, in combination with chemicals and heat, had a modest enhancing effect on the effectiveness of chemicals in killing or removing pathogens on alfalfa seeds. Overall, treatment with 1% Ca(OH)(2) was most effective in killing Salmonella and E. coli O157:H7. SIGNIFICANCE AND IMPACT OF THE STUDY: The use of 1% Ca(OH)(2) instead of 20,000 microg ml(-1) chlorine, which is currently recommended as a sanitizer for seeds intended for sprout production in the US, should be considered. Ultrasound treatment of alfalfa seeds containing Salmonella or E. coli O157:H7, in combination with chemical treatment, contributes to achieving greater reductions in populations of these pathogens, thereby reducing the risk of contamination and the presence of pathogens in sprouts produced from these seeds.     

Nomega-nitro-L-arginine methylester ameliorates myocardial toxicity induced by doxorubicin

The effects of Nomega-nitro-L-arginine methylester (L-NAME) and L-arginine on cardiotoxicity that is induced by doxorubicin (Dox) were investigated. A single dose of Dox 15 mg/kg i.p. induced cardiotoxicity, manifested biochemically by a significant elevation of serum creatine phosphokinase (CPK) activity [EC 2.7.3.2]. Moreover, cardiotoxicity was further confirmed by a significant increase in lipid peroxides, measured as malon-di-aldehyde (MDA) in cardiac tissue homogenates. The administration of L-NAME 4 mg/kg/d p.o. in drinking water 5 days before and 3 days after the Dox injection significantly ameliorated the cardiotoxic effects of Dox, judged by the improvement in both serum CPK activity and lipid peroxides in the cardiac tissue homogenates. On the other hand, the administration of L-arginine 70 mg/kg/d p.o. did not protect the cardiac tissues against the toxicity that was induced by the Dox treatment. The findings of this study suggest that L-NAME can attenuate the cardiac dysfunction that is produced by the Dox treatment via the mechanism(s), which may involve the inhibition of the nitric oxide (NO) formation. L-NAME may, therefore, be a beneficial remedy for cardiotoxicity that is induced by Dox and can then be used to improve the therapeutic index of Dox.     

Tellurium-induced dose-dependent impairment of antioxidant status: differential effects in cerebrum,cerebellum, and brainstem of mice

The effect of various doses of sodium tellurite (0.4, 0.8, and 2.0 mg/kg body weight, orally) on the activity of antioxidant enzymes (glutathione peroxidase, glutathione reductase, glutathione-S-transferase, and catalase) and content of glutathione and thiobarbituric acid reactive substances (TBARSs) in the cerebrum, cerebellum, and brainstem of male albino mice was studied after 15 d of treatment. All of the doses of tellurium (0.4, 0.8, and 2.0 mg/kg body weight, orally) have depleted the activity of antioxidant enzymes and the content of glutathione dose dependently in the cerebrum, cerebellum, and brainstem and it was significant with the dose of 2.0 mg/kg. On the other hand, the 2.0-mg/kg dose of tellurium has significantly elevated the content of TBARSs in the cerebrum and cerebellum. The 0.8-mg/kg dose of tellurium has significantly depleted the activities of glutathione peroxidase in the cerebrum and brainstem, glutathione-S-transferase in the cerebrum and cerebellum, catalase in the brainstem, and the content of glutathione in the cerebrum and cerebellum. In contrast, this dose has significantly elevated the content of TBARSs in the cerebrum and cerebellum. However, the depletion in the activity of glutathione reductase with various doses of sodium tellurite was not significant in any brain part of mice. The result suggests that sodium tellurite differentially affects the antioxidant status within various parts of the mice brain.     

Effects of zinc deficiency and supplementation on malondialdehyde and glutathione levels in blood and tissues of rats performing swimming exercise

The aim of the study was to investigate the effects of zinc deficiency and supplementation on lipid peroxidation and glutathione levels in blood and in some tissues of rats performing swimming exercise. Forty adult male Sprague-Dawley rats were divided into four groups: group 1, zinc-deficient consisted of swimming rats; group 2 consisted of zinc-supplemented swimming rats; groups 3 and 4 were the swimming and nonswimming controls, respectively. The levels of malondialdehyde and glutathione were measured after 4 wk of zinc-deficient or zinc-supplemented diet and 30 min of swimming exercise daily. The erythrocyte glutathione levels of groups 2 and 4 were significantly higher than those of groups 1 and 3 (p<0.01). The plasma malondialdehyde level of group 1 was significantly higher than all other groups. The glutathione levels in liver, kidney, striated muscle, and testes of group 2 were higher than in the other groups (p<0.01) and higher in kidney and striated muscle of group 3 than in groups 1 and 4 (p<0.01). The tissue malondialdehyde levels of striated muscle, liver, kidney, and testes of group 1 were significantly higher than for all other groups (p<0.01). Our findings suggest that both swimming exercise and zinc deficiency result in an increase of lipid peroxidation in tissues and that zinc supplementation prevents these alterations by the activation of the antioxidant system.     

Heavy metal concentrations in the breast milk of Saudi women

Lead, cadmium, and mercury concentrations were determined in breast milk of Saudi lactating mothers from Riyadh and Al-Ehssa regions in Saudi Arabia who were not occupationally exposed. The mean levels for cadmium, lead, and mercury were 1.732 μg/L, 31.671 μg/L, and 3.100 μg/L, respectively. In contrast to mercury, mothers living in the Al-Ehssa region had significantly higher cadmium and lead concentrations in their breast milk than those in the Riyadh region. The estimated weekly intakes of cadmium, lead, and mercury of breast-fed infants in this study were in some cases higher than the Provisional Tolerance Weekly Intake (PTWI) recommended by FAO/WHO, which pose a threat to their health. This necessitates the urgent need to undertake a comprehensive study to determine the sources of exposure to these heavy metals. Breast-feeding is of great benefical value for the infant’s development; therefore, efforts should be made to prevent its contamination with environmental pollutants.     

Melatonin reduces glial reactivity in the hippocampus, cortex, and cerebellum of streptozotocin-induced diabetic rats

Hyperglycemia plays a critical role in the development and progression of diabetic neuropathy. One of the mechanisms by which hyperglycemia causes neural degeneration is via the increased oxidative stress that accompanies diabetes. Metabolic and oxidative insults often cause rapid changes in glial cells. Key indicators of this response are increased synthesis of glial fibrillary acidic protein (GFAP) and S100B, both astrocytic markers. In the present study, we examined glial reactivity in hippocampus, cortex, and cerebellum of streptozotocin (STZ)-induced diabetic rats by determining the expression of GFAP and S-100B and we evaluated the effect of melatonin on the glial response. Western blot measurement of contents in brain regions after 6 weeks of STZ-induced diabetes indicated significant increases in these constituents compared with those in nondiabetic controls. Administration of melatonin prevented the upregulation of GFAP in all brain regions of diabetic rats. Using GFAP immunohistochemistry, we observed an increase in GFAP immunostaining in the hippocampus of STZ-diabetic rats relative to levels in the control brains. Treatment with melatonin resulted in an obvious reduction of GFAP-immunoreactive astrocytes in hippocampus. Like GFAP, S100B levels also were increased in all three brain areas of diabetic rats, an effect also reduced by melatonin treatment. Finally, the levels of lipid peroxidation products were elevated as a consequence of diabetes, with this change also being prevented by melatonin. These results suggest that diabetes causes increased glial reactivity possibly due to elevated oxidative stress, and administration of melatonin represents an achievable adjunct therapy for preventing gliosis.     

Estimation of 16S rRNA gene copy number in several probiotic Lactobacillus strains isolated from the gastrointestinal tract of chicken

The copy numbers of 16S rRNA genes in 12 probiotic Lactobacillus strains of poultry origin were analyzed. Genomic DNA of the strains was digested with restriction endonucleases that do not cut within the 16S rRNA gene of the strains. This was followed by Southern hybridization with a biotinylated probe complementary to the 16S rRNA gene. The copy number of the 16S rRNA gene within a Lactobacillus species was found to be conserved. From the hybridization results, Lactobacillus salivarius I 24 was estimated to have seven copies of the 16S rRNA gene, Lactobacillus panis C 17 to have five copies and Lactobacillus gallinarum strains I 16 and I 26 four copies. The 16S rRNA gene copy numbers of L. gallinarum and L. panis reported in the present study are the first record. Lactobacillus brevis strains I 12, I 23, I 25, I 211, I 218 and Lactobacillus reuteri strains C 1, C 10, C 16 were estimated to have at least four copies of the 16S rRNA gene. In addition, distinct rRNA restriction patterns which could discriminate the strains of L. reuteri and L. gallinarum were also detected. Information on 16S rRNA gene copy number is important for physiological, evolutionary and population studies of the bacteria.