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PubMed1998

Processing of Mycobacterium tuberculosis bacilli by human monocytes for CD4+ alphabeta and gammadelta T cells: role of particulate antigen.

Mycobacterium tuberculosis readily activates both CD4+ and Vdelta2+ gammadelta T cells. Despite similarity in function, these T-cell subsets differ in the antigens they recognize and the manners in which these antigens are presented by M. tuberculosis-infected monocytes. We investigated mechanisms of antigen processing of M. tuberculosis antigens to human CD4 and gammadelta T cells by monocytes. Initial uptake of M. tuberculosis bacilli and subsequent processing were required for efficient presentation not only to CD4 T cells but also to Vdelta2+ gammadelta T cells. For gammadelta T cells, recognition of M. tuberculosis-infected monocytes was dependent on Vdelta2+ T-cell-receptor expression. Recognition of M. tuberculosis antigens by CD4+ T cells was restricted by the class II major histocompatibility complex molecule HLA-DR. Processing of M. tuberculosis bacilli for Vdelta2+ gammadelta T cells was inhibitable by Brefeldin A, whereas processing of soluble mycobacterial antigens for gammadelta T cells was not sensitive to Brefeldin A. Processing of M. tuberculosis bacilli for CD4+ T cells was unaffected by Brefeldin A. Lysosomotropic agents such as chloroquine and ammonium chloride did not affect the processing of M. tuberculosis bacilli for CD4+ and gammadelta T cells. In contrast, both inhibitors blocked processing of soluble mycobacterial antigens for CD4+ T cells. Chloroquine and ammonium chloride insensitivity of processing of M. tuberculosis bacilli was not dependent on the viability of the bacteria, since processing of both formaldehyde-fixed dead bacteria and mycobacterial antigens covalently coupled to latex beads was chloroquine insensitive. Thus, the manner in which mycobacterial antigens were taken up by monocytes (particulate versus soluble) influenced the antigen processing pathway for CD4+ and gammadelta T cells.

Ammonium Chloride↗

PubMed2001

Hypokalemic metabolic acidosis attributed to cough mixture abuse.

This report describes a patient with mixed normal anion gap hyperchloremic metabolic and respiratory acidosis associated with hypokalemia attributed to cough mixture abuse. Metabolic acidosis was likely related to an overdose of ammonium chloride, whereas respiratory acidosis was probably related to the effect of hypokalemia on respiratory muscles, causing hypoventilation. Hypokalemia was caused by a transcellular shift of potassium induced by ephedrine and pseudoephedrine. Both ammonium chloride and ephedrine were probably present in the cough mixture obtained by our patient as an over-the-counter medication. Physicians should be aware of the potential for cough mixture abuse to cause major electrolyte disturbances that may carry the risk for major cardiac arrhythmias, particularly in youth.

Acidosis↗

PubMed1988

Biosynthesis of vitamin B-12 in anaerobic bacteria. Experiments with Eubacterium limosum on the origin of the amide groups of the corrin ring and of N-3 of the 5,6-dimethylbenzimidazole part.

The pathway of vitamin B-12 biosynthesis in anaerobic bacteria differs in several respects from the pathway found in aerobic or aerotolerant microorganisms. The aim of this investigation was to elucidate the formation of the 5,6-dimethylbenzimidazole part and the amide groups of vitamin B-12 in anaerobic bacteria. [15N]Ammonium chloride or L-[amido-15N]glutamine or a mixture of [15N]ammonium sulfate and [15N]glycine was added to fermentations with Eubacterium limosum. The vitamin B-12 isolated from these fermentations was methylated and degraded to cobinamide and 1,5,6-trimethylbenzimidazole. The amide groups of cobinamide were hydrolyzed and the amide nitrogen of the side chains a, b, c, d, e and g trapped as benzamide. The 15N incorporation was determined by mass spectroscopy. Thus in the experiment with [15N]ammonium chloride the benzamide and the 1,5,6-trimethylbenzimidazole contained 9.6% 15N, whereas in the experiment with L-[amido-15N]glutamine 37.5% of the molecules were 15N labeled. The 1H-NMR spectrum of 1,5,6-trimethylbenzimidazole revealed that the 15N from the ammonium salts and from glutamine was incorporated into N-3 of the 5,6-dimethylbenzimidazole moiety of vitamin B-12. With a mixture of [15N]ammonium sulfate and [15N]glycine both nitrogens of 5,6-dimethylbenzimidazole became 15N-labeled. These experiments demonstrate that in E. limosum the amide nitrogen of glutamine is not only the precursor of the six amide groups of the corrin ring, but also of N-3 of the 5,6-dimethylbenzimidazole moiety of vitamin B-12.

Amides↗

PubMed1982

The kidney of chicken adapts to chronic metabolic acidosis: in vivo and in vitro studies.

Renal adaptation to chronic metabolic acidosis was studies in Arbor Acre hens receiving ammonium chloride by stomach tube 0.75 g/kg/day during 6 days. During a 14-day study, it was shown that the animals could excrete as much as 60% of the acid load during ammonium chloride administration. At the same time urate excretion fell markedly but the renal contribution to urate excretion (14%) did not change. During acidosis, blood glutamine increased twofold and the tissue concentration of glutamine rose in both liver and kidney. Infusion of L-glutamine led to increased ammonia excretion and more so in acidotic animals. Glutaminase I, glutamate dehydrogenase, alanine aminotransferase (GPT), and malic enzyme activities increased in the kidney during acidosis but phosphoenolpyruvate carboxykinase (PEPCK) activity did not change. Glutaminase I was not found in the liver, but hepatic glutamine synthetase rose markedly during acidosis. Glutamine synthetase was not found in the kidney. Renal tubules incubated with glutamine and alanine were ammoniagenic and gluconeogenic to the same degree as rat tubules with the same increments in acidosis. Lactate was gluconeogenic without increment during acidosis. The present study indicates that the avian kidney adapts to chronic metabolic acidosis with similarities and differences when compared to dog and rat. Glutamine originating from the liver appears to be the major ammoniagenic substrate. Our data also support the hypothesis that hepatic urate synthesis is decreased during acidosis.

Acidosis, Renal Tubular↗

PubMed2002

Formation of vinyl halides via a ruthenium-catalyzed three-component coupling.

The ruthenium-catalyzed three-component coupling of an alkyne, an enone, and halide ion to form E- or Z-vinyl halides has been investigated. Through systematic optimization experiments, the conditions effecting the olefin selectivity were examined. In general, more polar solvents such as DMF favored the formation of the E-isomer, and less polar solvents such as acetone favored formation of the Z-isomer. The optimized conditions for the formation of E-vinyl chlorides were found to be the use of cyclopentadienyl ruthenium (II) cyclooctadiene chloride, stannic chloride pentahydrate as a cocatalyst, and for a chloride source, either ammonium chloride in DMF/water mixtures or tetramethylammonium chloride in DMF. A range of several other ruthenium (II) catalysts was also shown to be effective. A wide variety of vinyl chlorides could be formed under these conditions. Substrates with tethered alcohols or ketones either five or six carbons from the alkyne portion gave instead diketone or cyclohexenone products. For formation of vinyl bromides, a catalyst system involving the use of cyclopentadienylruthenium (II) tris(acetonitrile) hexafluorophosphate with stannic bromide as a cocatalyst was found to be most effective. The use of ammonium bromide in DMF/acetone mixtures was optimal for the synthesis of E-vinyl bromides, and the use of lithium bromide in acetone was optimal for formation of the corresponding Z-isomer. Under either set of conditions, a wide range of vinyl bromides could be formed. When alkynes with propargylic substituents are used, enhanced selectivity for formation of the Z-isomer is observed. When aryl acetylenes are used as the coupling partners, complete selectivity for the Z-isomer is obtained. A mechanism involving a cis or trans halometalation is invoked to explain formation of the observed products. The vinyl halides have been shown to be precursors to alpha-hydroxy ketones and cyclopentenones, and as coupling partners in Suzuki-type reactions.

Catalysis↗

PubMed2005

Evidence that membrane transduction of oligoarginine does not require vesicle formation.

The involvement of vesicular formation processes in the membrane transduction and nuclear transport of oligoarginine is currently a subject of controversy. In this report, a novel quantitative method which allows for the selective measurement of membrane transduction excluding concurrent endocytosis was used to determine the effects of temperature, endosomal acidification, endosomolysis, and several known inhibitors of endocytic pathways on the internalization of oligoarginine. The results show that, unlike endocytosis, transduction of oligoarginine was not affected by incubation at 16 degrees C as compared to the 37 degrees C control, and was only partially inhibited at 4 degrees C incubation. Additionally, membrane transduction was not inhibited to the same extent as endocytosis following treatment with ammonium chloride, hypertonic medium, amiloride, or filipin. The endosomolytic activity of oligoarginine was investigated by examining the leakage of FITC-dextran into the cytosolic compartment, which was not higher in the presence of oligoarginine. Furthermore, ammonium chloride showed no effect on the nuclear transport of oligoarginine. The data presented in this report indicate that membrane transduction is likely to occur at the plasma membrane without the formation of membrane vesicles, and the nuclear localization involves membrane transduction, rather than endocytosis of oligoarginine.

Amiloride↗

PubMed1987

Transport of a fluorescent macromolecule via endosomes to the vacuole in Saccharomyces cerevisiae.

Fluorescein isothiocyanate-conjugated dextran (FITC-dextran) is internalized by endocytosis into the lysosome-like vacuoles of Saccharomyces cerevisiae (Makarow, M., 1985, EMBO (Eur. Mol. Biol. Organ.) J. 4:1861-1866). Here we show that under energy depletion conditions FITC-dextran accumulated in a cytoplasmic compartment, from which it could be chased to the vacuole when the energy block was removed. The internal pH of the intermediate compartment under energy depletion was determined by fluorometry to be 5.8. The pH could be raised by the lysosomotropic agent ammonium chloride, the protonophore carbonyl cyanide p-trifluoromethoxyphenyl-hydrazone (CCCP) and the ATPase inhibitors dicyclohexylcarbodiimide (DCCD) and sodium vanadate. The pH of the vacuole was found to be 6.5. It was raised by ammonium chloride, CCCP, and DCCD, but not with sodium vanadate. Efrapeptin had no effect on the internal pH of either compartment. By dissecting the endocytic pathway, two portions of the route leading to the vacuole could be studied separately. The internalization of FITC-dextran from the extracellular fluid to the intermediate compartment followed linear kinetics, was independent of energy, and occurred at temperatures of between 15 degrees and 37 degrees C. Transfer of the marker from the intermediate compartment to the vacuole required energy, took place at temperatures between 19 degrees and 37 degrees C, and had a half-time of 7 min at 37 degrees C. Transport of the marker from the exterior of the cell to the vacuole did not require acidic pH values in the intermediate compartment or the vacuole. We suggest that the cytoplasmic compartment revealed by FITC-dextran, under energy depletion, represents the equivalent of the endosomes of mammalian cells.

Adenosine Triphosphatases↗

PubMed2002

The internal pH of synaptic-like microvesicles in rat pinealocytes in culture.

Synaptic-like microvesicles (SLMVs) are morphological and functional equivalents of neuronal synaptic vesicles, and are responsible for the storage and secretion of classical neurotransmitters in various endocrine cells. Vacuolar H+-ATPase acidifies the internal space of these organelles and provides a driving force for the uptake of neurotransmitters. Thus, the luminal pH is an important determinant of the function of SLMVs, although its value in living cells is unknown. Here, we determined the luminal pH of SLMVs in living rat pinealocytes by means of an immunoelectronmicroscopic procedure basedon the distribution of an amphipathic amine, 3-(2,4-dinitroanilino)-3'-amino-N-methyldipropylamine (DAMP). Use of double-labeling techniques with antibodies against 2,4-dinitrophenol for DAMP and synaptophysin for SLMVs, and of frozen ultrathin sections enabled us to determine the number of immunogold particles for DAMP per microm2 of SLMVs. Using the density of gold particles, the luminal pH of SLMVs was calculated to be 5.11 +/- 0.01. Treatment with either 1 microm bafilomycin A1, a specific inhibitor of vacuolar H+-ATPase, or 50 mm ammonium chloride, a dissipater of the transmembrane pH gradient, increased the luminal pH to 6.04 +/- 0.07 and 6.05 +/- 0.11, respectively. Simultaneously, the lysosomal pH was found to be 5.14 +/- 0.07, which increased to 5.77 +/- 0.09 and 5.93 +/- 0.13 with bafilomycin A1 and ammonium chloride, respectively. It is concluded that the luminal pH of SLMVs is comparable to that of lysosomes in vivo.

Ammonium Chloride↗

PubMed1988

In vivo metabolism of nitrogen precursors for urea synthesis in the postprandial rat.

(1) Adult postprandial rats were given a continuous, intravenous infusion of 15N-labelled glutamate, alanine, ammonium chloride and glutamine amide for 6 h. The enrichment in the free hepatic pool was measured for ammonia, glutamine amide, urea, aspartate, glutamate and alanine. (2) Glutamine and glutamate supplied significantly more nitrogen to urea than ammonium chloride or alanine. (3) Glutamate was not a significant source of hepatic ammonia, hence in this situation it is not necessary to impute a major role to glutamate dehydrogenase in hepatic ammoniagenesis for urea synthesis. (4) Glutamine and ammonia, mostly of intestinal origin in the postprandial state, were major precursors of hepatic ammonia. (5) The nitrogen of glutamate and alanine moved to urea primarily through aspartic acid.

Alanine↗

PubMed1976

Mutant strains (nit) of Salmonella typhimurium with a pleiotropic defect in nitrogen metabolism.

We have isolated mutant strains (nit) of Salmonella typhimurium that are defective in nitrogen metabolism. They have a reduced ability to use a variety of compounds including glutamate, proline, arginine, N-acetyl-glucosamine, alanine, and adenosine as sole nitrogen source. In addition, although they grow normally on high concentrations of ammonium chloride (greater than 1 mM) as nitrogen source, they grow substantially more slowly than wild type at low concentrations (less than 1 mM). We postulated that the inability of these strains to utilize low concentrations of ammonium chloride accounts for their poor growth on other nitrogen sources. The specific biochemical lesion in strains with a nit mutation is not known; however, mutant strains have no detectable alteration in the activities of glutamine synthetase, glutamate synthetase, or glutamate dehydrogenase, the enzymes known to be involved in assimilation of ammonia. A nit mutation is suppressed by second-site mutations in the structural gene for glutamine synthetase (glnA) that decrease glutamine synthetase activity.

Acetylglucosamine↗

PubMed1981

Mechanism by which enhanced ammonia production reduces urinary potassium excretion.

To determine the mechanism whereby an increase in ammonia production decreases urinary potassium excretion, we perfused isolated rat kidneys at a pH of either 7.0 or 7.4. After 45 min of perfusion at either pH, glutamine or ammonium chloride was added to the perfusate to result in concentration of 5 and 0.8 mM, respectively and observations were continued for 50 min. Control kidneys were perfused at both pH's without further additions to the perfusate. At pH 7.0 glutamine increased ammonia production and increased urinary ammonium excretion strikingly; whereas the addition of ammonium chloride did not change ammonia production but increased urinary ammonium excretion to a comparably degree. Both maneuvers resulted in a reciprocal fall in urinary potassium excretion in comparison with control perfusions. The decreases in potassium excretion could not be accounted for by differences in perfusate or urinary acid-base parameters, or by changes in urinary sodium, water, or chloride excretion. At pH 7.4, glutamine also significantly increased ammonia production and perfusate ammonia concentration. In contrast to the studied at pH 7.0 in which the urine pH was acid (5.9), the urine remained alkaline (pH 7.2), and both urinary ammonium excretion and urinary potassium excretion were unaltered. Thus, potassium sparing is not a nonspecific effect of glutamine, its metabolism to ammonia, or perfusate ammonia concentration but is directly related to an increase in urinary ammonium excretion.

Ammonia↗

PubMed1971

Ammonium and chloride extrusion: hyperpolarizing synaptic inhibition in spinal motoneurons.

The reversal potential of postsynaptic inhibition shifts toward resting membrane potentials in cat spinal motoneurons after intravenous infusion of ammonium salts(1 to 3 millimoles per kilogram of body weight). Simultaneously, the depolarizing action of intracellularly injected chloride ions on the inhibitory membrane is enhanced and recovery therefrom is prolonged. Passive membrane properties remain unaltered. The results indicate a blocking of active extrusion of chloride which normally maintains a high ionic gradient for hyperpolarizing inhibition.

Action Potentials↗

PubMed1989

Repression of sporulation: isolation and characterization of repression-resistant mutants of Bacillus subtilis.

The sporulation of Bacillus subtilis B34 was repressed by 24 h if glutamine or ammonium chloride but not glutamate was added at late log phase (70 h) when glucose and glutamate were nearly exhausted. Glutamine-resistant mutants were isolated by selective heat treatment during the delay period induced by glutamine. Glutamine-resistant mutants showed cross resistance not only against ammonium chloride but also against glucose just as glucose-resistant mutants showed resistance against nitrogenous catabolites.

Ammonium Chloride↗

PubMed2006

[Effect of Rongshi granule on renal stone formation and osteopontin expression in rat urolithiasis model].

OBJECTIVE: To investigate the effect of Rongshi granule on osteopontin(OPN) expression. METHOD: The urlisthiasis rats were induced by ethylene glycol (EG) and ammonium chloride, the control group rats were non-treated, and the Rongshi granule groups (low-dose group, middle-dose group and high-dose group) were administered Rongshi granule in addition to EG and ammonium chloride in 21 days. Pooled 24 h urine samples from each group were collected weekly with the use of metabolic cages, the concentration of uric calcium and oxalic acid were respectively measured by EDTA and photoelectric colorimetric method. Eight animals from each group were killed at 0, 7, 14, and 21 days, kidneys were histologic examinaed and immunohistochemical staining. RESULT: The expression of kidney osteopontin in model group was obviously higher than that of control group (P <0.01), and was up to the highest at 21 days with 1.4 times (0.281 3/0.201 8) of the control group. The expression of kidney osteopontin in all of the Rongshi granule groups were lower than those of model group (P < 0.05), with an obvious dose-dependent manner. The degree of the kidney calcium oxalate crystal of the rats in all the Rongshi granule groups was much lower than that of model group, and the uric calcium and oxalic acid were much lower than those of model group (P < 0.01). CONCLUSION: The Rongshi granule could inhibit the expression of osteopontin in rat urolithiasis model.

Ammonium Chloride↗

PubMed1991

CD4-Pseudomonas exotoxin hybrid proteins: modulation of potency and therapeutic window through structural design and characterization of cell internalization.

Replacing the Pseudomonas exotoxin A (PE) cell binding domain with the human immunodeficiency virus (HIV) gp120 binding domain from CD4 yields a hybrid toxin (CD4-PE) with potential therapeutic use in treating acquired immunodeficiency syndrome (AIDS). To find the most therapeutically potent combination of CD4 and PE four different hybrid toxins composed of one [CD4(122)] or two [CD4(181)] Ig-like CD4 domains and sequences of PE where the binding domain was partially [PE(392)] or completely [PE(364)] removed were constructed and expressed in Escherichia coli. The number of CD4 domains determined the binding affinity to gp120 and in cell viability assays the window between specific and nonspecific cytotoxicity. The length of PE determined the potency of the drug. The optimal hybrid toxin was composed of two Ig-like domains of CD4 and PE(392). Investigation of the internalization mechanism of CD4-PE revealed that the hybrid toxin binds to target cells and is endocytosed within one hour. However, more than 6 hours are required for maximum translation inhibition. In contrast to PE which is inhibited by ammonium chloride treatment, cell toxicity of CD4-PE is not affected by ammonium chloride. Further investigations showed that the acid-induced hydrophobicity change which is required for membrane translocation is also observed with CD4-PE but at significantly higher pH than with PE.

ADP Ribose Transferases↗

PubMedDate not supplied

Resolution of a left ureteral stone using electrohydraulic lithotripsy in a thoroughbred colt.

A 3-year-old Thoroughbred colt was presented for evaluation of azotemia and anorexia. Physical examination revealed a ureterolith in the left ureter, approximately 10 cm from the bladder, which was thought to obstruct urine flow by approximately 90% when viewed cystoscopically. Ultrasonographic examination of both kidneys revealed indistinct corticomedullary junctions, and the right kidney was more hyperechoic. A percutaneous biopsy of the right kidney revealed chronic interstitial nephritis with marked interstitial medullary fibrosis. Medical therapy consisting of IV fluids, sodium chloride PO, and ammonium chloride PO was initiated. Ureteroscopic electrohydraulic lithotripsy via a perineal urethrostomy was used to successfully remove the stone. Klebsiella oxytoca, which responded to oral enrofloxacin therapy, was cultured from the urine after surgery. Azotemia resolved and the horse resumed training.

Animals↗

PubMed1973

The effect of some molecules and ions on gastric function in the milk-fed calf.

1. The abomasum of the milk-fed calf has been examined using an adaptation of the Serial Test Meal method devised by Hunt & Spurrell (1951). The emptying process, acid secretion and pepsin secretion were studied.2. Using serial test meals of simple solutions instilled into the abomasum via a cannula, our investigation leaves no doubt that the osmolarity of the abomasal contents significantly modifies the rate of abomasal emptying.3. Hypotonic and isotonic solutions of sodium chloride and sodium bicarbonate increase abomasal emptying but bicarbonate is most effective.4. Increasing the concentration of solutes in the abomasal contents slows abomasal emptying. Sodium chloride, sodium bicarbonate, ammonium chloride and urea do not delay abomasal emptying until hypertonic concentrations are attained. Hypotonic solutions of potassium chloride, calcium chloride, glucose, lactose, hydrochloric acid and acetic acid delay abomasal emptying.5. The results obtained in the calf show that the abomasum is under restraint probably from duodenal receptors as is the simple stomach (Hunt & Knox, 1968) and that an osmoreceptor as postulated by Hunt (1956) is an important factor in this mechanism.6. Acid secretion is inhibited when hypertonic solutions are instilled into the abomasum.7. Pepsin secretion is not affected by simple solutions in the abomasum.8. Gastric function in the milk-fed calf appears to be controlled by mechanisms essentially similar to those already demonstrated in the simple stomach.

Abomasum↗

PubMed1985

Sodium bioavailability from phosphate sources.

Two experiments were conducted with day-old broiler chicks from 1 to 21 days of age to measure the relative bioavailability of sodium from three commercially available defluorinated phosphates and two reagent sources (monosodium and dibasic sodium phosphate) compared to sodium chloride as the standard. Ten chicks were assigned to each pen in the first experiment and eight chicks in the second. Each diet was fed to three pens of males and three pens of females within an experiment. Feed-grade sodium chloride was added to a basal feed containing dicalcium phosphate in increments of .05% (0 to .25%) to provide total sodium levels from 180 to 967 ppm. Each of the defluorinated materials was substituted into the dicalcium basal at a level sufficient to provide one-half of the supplemental phosphorus. Reagent phosphate sources were added to the basal in amounts sufficient to supply .059% sodium, the same amount as provided by .15% sodium chloride. Reagent grade ammonium chloride was added to treatment diets to ensure a level of .156% total chloride. Mortality was related to sodium level and was excessive only for the treatment receiving no supplementation. Significant manure moisture increases were associated with up to .10% supplemental sodium chloride. Slope-ratio calculations of sodium availability, using body weight and daily sodium intake, revealed average availability values over the experiment ranging from 83.9 to 89.8% for the three defluorinated sources. Availability for monobasic sodium phosphate was calculated to be 91.8%, while that of the dibasic source was somewhat higher at 97.2%.

Animal Feed↗