Nitric oxide and thiol reagent modulation of Ca2+-activated K+ (BKCa) channels in myocytes of the guinea-pig taenia caeci.
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The modulation of large conductance Ca2+-activated K+ (BKCa) channels by the nitric oxide (NO) donors S-nitroso-L-cysteine (NOCys) and sodium nitroprusside (SNP) and agents which oxidize or reduce reactive thiol groups were compared in excised inside-out membrane patches of the guinea-pig taenia caeci. When the cytosolic side of excised patches was bathed in a physiological salt solution (PSS) containing 130 mM K+ and 15 nM Ca2+, few BKCa channel openings were recorded at potentials negative to 0 mV. However, the current amplitude and open probability (NPo) of these BKCa channels increased with patch depolarization. A plot of ln(NPo) against the membrane potential (V) fitted with a straight line revealed a voltage at half-maximal activation (V0.5) of 9.4 mV and a slope (K) indicating an e-fold increase in NPo with 12.9 mV depolarization. As the cytosolic Ca2+ was raised to 150 nM, V0.5 shifted 11.5 mV in the negative direction, with little change in K (13.1 mV). NOCys (10 microM) and SNP (100 microM) transiently increased NPo 16- and 3. 7-fold, respectively, after a delay of 2-5 min. This increase in NPo was associated with an increase in the number of BKCa channel openings evoked at positive potentials by ramped depolarizations (between -60 and +60 mV). Moreover, this NOCys-induced increase in NPo was still evident in the presence of 1H-[1,2,4]oxadiazolo[4, 3-a]quinoxalin-1-one (ODQ; 10 microM), the specific blocker of soluble guanylyl cyclase. The sulfhydryl reducing agents dithiothreitol (DTT; 10 and 100 microM) and reduced glutathione (GSH; 1 mM) also significantly increased NPo (at 0 mV) 7- to 9-fold, as well as increasing the number of BKCa channel openings evoked during ramped depolarizations. Sulfhydryl oxidizing agents thimerosal (10 microM) and 4,4'-dithiodipyridine (4,4DTDP; 10 microM) and the thiol-specific alkylating agent N-ethylmaleimide (NEM; 1 mM) significantly decreased NPo (at 0 mV) to 40-50% of control values after 5-10 min. Ramped depolarizations to +100 mV evoked relatively few BKCa channel openings. The effects of thimerosal on NPo were readily reversed by DTT, while the effects of NOCys were prevented by NEM. It was concluded that both redox modulation and nitrothiosylation of cysteine groups on the cytosolic surface of the alpha subunit of the BKCa channel protein can alter channel gating. (+info)
The dissociation between upregulated endothelins and hemodynamic responses during polymicrobial sepsis.
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Polymicrobial sepsis is characterized by an early, hyperdynamic phase followed by a late, hypodynamic phase. Although studies have suggested that endothelins (ETs) contribute to the development of shock after a bolus injection of endotoxin, little is known about the role of ETs in the transition from the hyperdynamic phase to the hypodynamic phase of sepsis. To study this, male adult rats were subjected to sepsis by cecal ligation and puncture (CLP) followed by fluid resuscitation. Plasma levels of ET-1 and ET-2 were measured by radioimmunoassay at 2, 5, 10 h (i.e. the early stage of sepsis), and 20 h (late stage) following CLP or sham operation. Tissue levels of ET-1 and ET-2 were determined in the heart, lungs, small intestine, and spleen at 5 h after CLP or sham operation. In addition, preproendothelin-1 (precursor of ET-1) gene expression was analyzed by reverse transcription-polymerase chain reaction (RT-PCR) at 5 h in the heart, lungs, small intestine, spleen, and liver. The results indicate that plasma levels of ET-1 and ET-2 were not different from values of sham groups at 2 and 20 h, but were significantly higher than the sham values at 5 and 10 h after CLP. While there were no significant increases in tissue levels of ET-1 and ET-2 at 5 h post-CLP, RT-PCR analysis indicates a significant upregulation of preproendothelin-1 gene expression in the heart, spleen, and liver (but not in the lungs or small intestine) at 5 h after the onset of sepsis. These results indicate that the heart, spleen, and liver appear to be important ET-producing organs during the early stage of sepsis. The lack of significant increases in tissue ET levels could be due to the possibility that the newly converted peptide is quickly transferred to the bloodstream. Since the hyperdynamic phase of sepsis occurs at 2-10 h and the hypodynamic phase occurs at 20 h after CLP, the increased plasma levels of ET at 5 and 10 h suggest that mediators other than ETs (such as adrenomedullin) are responsible for producing the biphasic hemodynamic responses during the progression of polymicrobial sepsis. (+info)
Coarse brown rice increases fecal and large bowel short-chain fatty acids and starch but lowers calcium in the large bowel of pigs.
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Young male pigs were fed a diet formulated from human foods including either boiled white rice plus rice bran or heat-stabilized brown rice at equivalent levels of fiber for 3 wk. Stool and starch excretion were low in pigs fed white rice during the first 2 wk of the experiment. In pigs fed brown rice, their excretion was high during wk 1 but declined in wk 2 while short-chain fatty acid (SCFA) excretion was higher at both times. Large bowel digesta mass, measured during wk 3, was higher in pigs fed brown rice but only in the proximal colon. Large bowel and fecal starch concentrations were higher in pigs fed brown rice but the difference was insufficient to explain the increase in large bowel digesta mass. In pigs with a cecal cannula, digesta starch concentrations were equally higher when white or brown rice was fed compared with the corresponding rice which had been finely milled, indicating that particle size was a determinant of ileal digestibility. Concentrations and pools of total and individual SCFA were higher in all regions of the colon but not the cecum of pigs fed brown rice. Large bowel Ca(2+) concentrations were lower in pigs fed brown rice, suggesting greater absorption. The data confirm earlier findings that brown rice raises large bowel digesta mass and SCFA through greater fermentation of starch but show that starch itself makes a relatively small contribution to digesta and stool mass. Apparently, the rate of passage of digesta is a determinant of the concentrations and pools of SCFA in the distal colon and in feces. (+info)
Immune responses of specific pathogen-free and gnotobiotic mice to antigens of indigenous and nonindigenous microorganisms.
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Strains of indigenous Escherichia coli, Bacteroides, and Lactobacillus were isolated from the gastrointestinal tracts of specific pathogen-free (SPF) mice. Nonvaccinated SPF mice exhibited in their spleens low numbers of plaque-forming cells (PFC) and rosette-forming cells reacting with antigens of these andigenous bacteria. PFC reacting with these bacterial antigens were not detected in infant SPF mice until 7 days after birth. Compared with nonvaccinated controls, SPF mice vaccinated parenterally with indigenous E. coli or Bacteroides produced a moderate increase in the numbers of specific PFC. Thus, the SPF mouse is capable of responding immunologically after vaccination with microbes indigenous to its intestinal tract. However, more PFC reacting with homologous vaccine antigens were detected after parenteral vaccination of SPF mice with nonindigenous E. coli O127:B8, E.coli O14, or B. fragilis than after parenteral vaccination with indigenous E. coli or Bacteroides. Gnotobiotic mice orally monoassociated with these nonindigenous bacteria exhibited greater immune responses to antigens of the bacteria used to monoassociation than did gnotobiotes monoassociated with the indigenous microbes. The results are consistent with the hypothesis that mice are more responsive immunologically to antigens of nonindigenous bacteria than they are to antigens of certain microbes indigenous to their gastrointestinal tracts. (+info)
Chronological recognition by chicken of antigenic polypeptides in Salmonella enteritidis with different plasmid profiles: relationship to infection rate.
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The antigenic polypeptides in Salmonella Enteritidis (SE) were chronologically recognized by the chicken immune system, using Western immunoblotting. Broiler chicks challenged at three days of age with SE strain carrying the most prevalent plasmid profile of 14.1 and about (approximately) 50 kb were bled at 17, 24, 31, 38 and 45 days of age. Pooled sera of blood collected at each age was reacted by Western immunoblotting with banded polypeptides of three predominant SE strains that acquired the following respective plasmid profiles: 14.1 kb; 14.1 and approximately 50.0 kb; and 1.8, 14.1 and approximately 50.0 kb. The immunoblots of each pooled sera collected at a specific age against the three SE strains were similar. More specifically, the early immune response at 17 days of age had antibodies recognizing only one polypeptide in the three SE strains namely, the 35.8 kDa. At 24 or 31 days of age, the acquired immunity to infection had antibodies recognizing five similar polypeptides in the three SE strains namely, the 14.4 (fimbriae protein), 21.5 (fimbriae protein), 30.5, 35.8, and 66.2 kDa. At 38 and 45 days of age, the antibodies recognized additional polypeptides namely, the 41.5 and 55.6 kDa, respectively. The recognition of the 41.5 and 55.6 kDa polypeptides at 45 days of age was associated with higher invasiveness of SE to spleens and livers (15.6%) and in higher cecal colonization (59.4%) in comparison to absence of recognition to the two polypeptides at 31 days of age associated with low infectivity to spleens (0%), livers (3.1%), and ceca (9.4%). (+info)
Effects of skin pressure by clothing on digestion and orocecal transit time of food.
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In order to reveal the influence of clothing skin pressure on digestion of food through the gastrointestinal tract, we examined the absorption of dietary carbohydrate and orocecal transit time of a test meal by means of a breath hydrogen test on 7 healthy young women. In this experiment, we collected breath samples from the participants wearing loose-fitting experimental garment on the second day of the experiment and from the same participants but wearing an additional tight-fitting girdle on the following day for 16 hours and 9 hours, respectively. Skin pressure applied by a girdle on participant's waist, abdomen and hip region was 15.5 +/- 0.4 mmHg (mean +/- SE), 11.0 +/- 0.2 mmHg, and 13.6 +/- 0.6 mmHg, respectively, and the values were 2-3 times larger than those of the experimental garment. The hydrogen concentration vs. time curve showed that breath hydrogen levels at its peaks (15:00, 15:30, 16:00, 16:30, and 17:00 hr) on the third day of the experiment were significantly higher than those of the corresponding time on the second day (p < 0.05 at 17:00 and 15:00, p < 0.01 at 15:00, 16:00 and 16:30). Consequently, significantly pronounced breath hydrogen excretion was observed under the "pressure" clothing condition (p < 0.01). On the other hand, the transit time of the test meal for the subjects wearing a girdle did not differ significantly from that for the subjects wearing the garment of less pressure (270 +/- 18 minutes and 263 +/- 21 minutes, respectively). These results indicate that the clothing skin pressure has an inhibitory effect on the absorption of dietary carbohydrate in the small intestine, but no effect on the orocecal transit time of a meal. (+info)
Clinical outcome of patients with familial hypercholesterolemia and coronary artery disease undergoing partial ileal bypass surgery.
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Familial hypercholesterolemia is characterized by high serum levels of total cholesterol and LDL-cholesterol. It may be homozygous or heterozygous. In homozygous patients, LDL-cholesterol levels range from 500 to 1000 mg/dL and coronary artery disease is precocious, usually manifesting itself between the 2nd and 3rd decades of life. The diagnosis is often made by the presence of xanthoma tuberosum and tendinous xanthomas that appear between the 1st and 2nd decades of life. The use of high doses of statins or even unusual procedures (apheresis, partial ileal bypass surgery, liver transplantation, gene therapy), or both, is necessary for increasing survival and improving quality of life, because a reduction in cholesterol levels is essential for stabilizing the coronary artery disease and reducing xanthomas. We report our experience with 3 patients with xanthomatous familial hypercholesterolemia and coronary artery disease, who underwent partial ileal bypass surgery. Their follow-up over the years (approximately 8 years) showed a mean 30% reduction in total cholesterol, with a significant reduction in the xanthomas and stabilization of the coronary artery disease. (+info)
Heritabilities of and genetic relationships between salmonella resistance traits in broilers.
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Using experimental infections, three traits for salmonella resistance were studied: mortality, survival time (in animals that died by infection), and quantitative cecal salmonella carriage at the end of the rearing period (in animals that did not die). In total, 548 animals were used; mortality was 29.2%, mean survival time was 5.97 d (n = 160), and the mean 10log of colony forming units per gram of cecal contents was 1.62 (n = 387). Genetic parameters were evaluated in bivariate threshold-linear models to account for the selective measurement of survival time and cecal carriage. Heritabilities were .06 for survival time, .09 for cecal carriage, and .12 for mortality. The genetic correlation between mortality and cecal carriage was weak (.26), which suggests that these traits are largely controlled by different genes. The genetic correlation between mortality and survival time was relatively strong (-.68). Simultaneous study of multiple traits seems to be of particular importance in judging epidemiological consequences of a possible selection for resistance. Results here indicate that selection on decreased mortality could be unfavorable for the spread of salmonella because the resulting correlated increase in survival time, implying longer shedding by infected animals, is relatively stronger than the correlated decrease in level of cecal carriage. Selection to reduce the level of cecal salmonella carriage could be done while keeping survival time constant, if so desired, because the correlation between these traits is weak (-.15). (+info)