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J L Bailey

Publications and source records attributed to J L Bailey.

At least 19 recordsLinked to original sources
PubMed1999

Evaluation of signals activating ubiquitin-proteasome proteolysis in a model of muscle wasting.

The ubiquitin-proteasome proteolytic system is stimulated in conditions causing muscle atrophy. Signals initiating this response in these conditions are unknown, although glucocorticoids are required but insufficient to stimulate muscle proteolysis in starvation, acidosis, and sepsis. To identify signals that activate this system, we studied acutely diabetic rats that had metabolic acidosis and increased corticosterone production. Protein degradation was increased 52% (P < 0.05), and mRNA levels encoding ubiquitin-proteasome system components, including the ubiquitin-conjugating enzyme E214k, were higher (transcription of the ubiquitin and proteasome subunit C3 genes in muscle was increased by nuclear run-off assay). In diabetic rats, prevention of acidemia by oral NaHCO3 did not eliminate muscle proteolysis. Adrenalectomy blocked accelerated proteolysis and the rise in pathway mRNAs; both responses were restored by administration of a physiological dose of glucocorticoids to adrenalectomized, diabetic rats. Finally, treating diabetic rats with insulin for >/=24 h reversed muscle proteolysis and returned pathway mRNAs to control levels. Thus acidification is not necessary for these responses, but glucocorticoids and a low insulin level in tandem activate the ubiquitin-proteasome proteolytic system.

Acidosis↗
PubMed1998

Metabolic acidosis and protein catabolism: mechanisms and clinical implications.

Metabolic acidosis increases protein degradation resulting in muscle wasting and a negative nitrogen balance. The branched-chain amino acids serve as useful markers of these changes and their catabolism is increased in acidosis, particularly for the spontaneous acidosis associated with renal failure. As a result, the neutral nitrogen balance is compromised and malnutrition results. Glucocorticoids mediate these changes through the recently discovered ATP-dependent ubiquitin-proteasome pathway. Therapy necessitates correction of the underlying acidosis either through adjustment of the alkalinity of the dialysate for the patient on dialysis or through dietary protein restriction and sodium bicarbonate supplements for the predialysis patient.

Acidosis↗
PubMed1998

Tissue-specific responses of branched-chain alpha-ketoacid dehydrogenase activity in metabolic acidosis.

In adrenalectomized rats, acidosis does not increase whole-body leucine oxidation unless a physiologic amount of glucocorticoids (dexamethasone) is also provided; an equivalent dose of dexamethasone without acidosis does not change leucine catabolism. Because the influences of acidification and glucocorticoids on branched-chain amino acid metabolism in specific organs are unknown, the function of branched-chain alpha-ketoacid dehydrogenase (BCKAD), the rate-limiting enzyme in branched-chain amino acid catabolism, in adrenalectomized rat skeletal muscle and liver, the two major tissues that degrade branched-chain amino acid was measured. In muscle of acidotic adrenalectomized rats receiving dexamethasone, basal and total BCKAD activities were increased 2.6- (P < 0.05) and 2.8-fold (P < 0.05), respectively. Neither acidosis nor dexamethasone alone increased these activities. BCKAD E1alpha subunit mRNA in muscle of acidotic rats given dexamethasone was increased 1.89-fold (P < 0.05) in parallel with the change in BCKAD activity; BCKAD E2 subunit mRNA was increased by acidosis, dexamethasone, or a combination of both stimuli. In contrast, basal BCKAD activity in liver of rats with acidosis or dexamethasone was nearly threefold lower (P < 0.05) and changes in enzyme activity reflected reduced subunit mRNA. Thus, there are reciprocal, tissue-specific changes in BCKAD function in response to acidosis.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMed1998

Mechanisms contributing to muscle-wasting in acute uremia: activation of amino acid catabolism.

Acute uremia (ARF) causes metabolic defects in glucose and protein metabolism that contribute to muscle wasting. To examine whether there are also defects in the metabolism of essential amino acids in ARF, we measured the activity of the rate-limiting enzyme for branched-chain amino acid catabolism, branched-chain ketoacid dehydrogenase (BCKAD), in rat muscles. Because chronic acidosis activates muscle BCKAD, we also evaluated the influence of acidosis by studying ARF rats given either NaCl (ARF-NaCl) or NaHCO3 (ARF-HCO3) to prevent acidosis, and sham-operated, control rats given NaHCO3. ARF-NaCl rats became progressively acidemic (serum [HCO3] = 21.3 +/- 0.7 mM within 18 h and 14.7 +/- 0.8 mM after 44 h; mean +/- SEM), but this was corrected with NaHCO3. Plasma valine was low in ARF-NaCl and ARF-HCO3 rats. Plasma isoleucine, but not leucine, was low in ARF-NaCl rats, and isoleucine tended to be lower in ARF-HCO3 rats. Basal BCKAD activity (a measure of active BCKAD in muscle) was increased more than 17-fold (P < 0.01) in ARF-NaCl rat muscles, and this response was partially suppressed by NaHCO3. Maximal BCKAD activity (an estimate of BCKAD content), subunit mRNA levels, and BCKAD protein content were not different in ARF and control rat muscles. Thus, ARF increases branched-chain amino acid catabolism by activating BCKAD by a mechanism that includes acidosis. Moreover, in a muscle-wasting condition such as ARF, there is a coordinated increase in protein and essential amino acid catabolism.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗
PubMed1998

Effect of serum-clot contact time on clinical chemistry laboratory results.

The effect of serum-clot contact time on laboratory results was studied by dividing each blood specimen into four blood collection tubes. The control sera were separated from the clot within 30 min of the collection. The other tubes were incubated at 32 degrees C, and the sera were separated at 3, 6, and 24 h. The sera were stored at 4 degrees C and analyzed at the same time. The stability of the tests was determined by comparing the results of the 3-, 6-, and 24-h samples with the values from the 30-min samples. The acceptable limits around the 30-min values were derived from the analytical and intraindividual biological variation of the tests. A total of 63 analytes were studied. Potassium, phosphorous, and glucose were the least stable, and the serum should be separated from the clot within 3 h for these analytes. Albumin, bicarbonate, chloride, C-peptide, HDL-cholesterol, iron, LDL-cholesterol, and total protein should be separated within 6 h. The other analytes were stable for 24 h of serum-clot contact.

Analysis of Variance↗
PubMed1997

The search for the uremic toxin: the case for metabolic acidosis.

Much effort has been expended on determining which compound, hormone or metabolic condition causes the uremic syndrome. Byproducts of protein metabolism that can cause uremic symptoms, including loss of lean body mass, have been a focus of research but specific toxins have been difficult to identify. Evidence is provided that implicates metabolic acidosis as the prime signal initiating muscle wasting in uremia since it activates branched-chain ketoacid dehydrogenase and the ubiquitin proteasome pathway. These responses degrade the essential branched chain amino acids and protein in muscle, leading to loss of muscle mass. Correction of the metabolic acidosis with sodium bicarbonate supplements has significant therapeutic implications for uremic patients with even mild degrees of metabolic acidosis.

Acidosis↗
PubMed1997

The flk gene of Salmonella typhimurium couples flagellar P- and L-ring assembly to flagellar morphogenesis.

The flagellum of Salmonella typhimurium is assembled in stages, and the negative regulatory protein, FlgM, is able to sense the completion of an intermediate stage of assembly, the basal body-hook (BBH) structure. Mutations in steps leading to the formation of the BBH structure do not express the flagellar filament structural genes, fliC and fljB, due to negative regulation by FlgM (K. L. Gillen and K. T. Hughes, J. Bacteriol. 173:6453-6459, 1991). We have discovered another novel regulatory gene, flk, which appears to sense the completion of another assembly stage in the flagellar morphogenic pathway just prior to BBH formation: the completion of the P- and L-rings. Cells that are unable to assemble the L- or P-rings do not express the flagellin structural genes. Mutations by insertional inactivation in either the flk or flgM locus allow expression of the fljB flagellin structural gene in strains defective in flagellar P- and L-ring assembly. Mutations in the flgM gene, but not mutations in the flk gene, allow expression of the fljB gene in strains defective in all of the steps leading to BBH formation. The flk gene was mapped to min 52 of the S. typhimurium linkage map between the pdxB and fabB loci. A null allele of flk was complemented in trans by a flk+ allele present in a multicopy pBR-based plasmid. DNA sequence analysis of the flk gene has revealed it to be identical to a gene of Escherichia coli of unknown function which has an overlapping, divergent promoter with the pdxB gene promoter (P. A. Schoenlein, B. B. Roa, and M. E. Winkler, J. Bacteriol. 174:6256-6263, 1992). An open reading frame of 333 amino acids corresponding to the flk gene product of S. typhimurium and 331 amino acids from the E. coli sequence was identified. The transcriptional start site of the S. typhimurium flk gene was determined and transcription of the flk gene was independent of the FlhDC and sigma28 flagellar transcription factors. The Flk protein observed in a T7 RNA polymerase-mediated expression system showed an apparent molecular mass of 35 kDa, slightly smaller than the predicted size of 37 kDa. The predicted structure of Flk is a mostly hydrophilic protein with a very C-terminal membrane-spanning segment preceded by positively charged amino acids. This finding predicts Flk to be inserted into the cytoplasmic membrane facing inside the cytoplasm.

Amino Acid Sequence↗
PubMed1997

Glucocorticoids mediate a decrease in AVP-regulated urea transporter in diabetic rat inner medulla.

Providing glucocorticoids to adrenalectomized (Adx) rats results in downregulation of the vasopressin (AVP)-regulated urea transporter (VRUT) in the renal inner medullary (IM) tip. To examine the physiological relevance of this response, we studied rats with uncontrolled diabetes mellitus induced by streptozotocin (STZ), since these rats have increased corticosterone production and urea excretion. We measured VRUT protein in extracts from the IM tip or base of pair-fed control and diabetic rats by Western analysis using an antibody to rat VRUT. In the IM tip, VRUT was significantly reduced by 39% in diabetic compared with control rats. In the IM base, there was no significant difference between diabetic and control rats. To determine whether the decrease in VRUT in the IM tip was mediated by glucocorticoids, the experiment was repeated using the following three groups of rats: 1) Adx alone, 2) Adx + STZ, and 3) Adx + STZ + replacement with a physiological dose of glucocorticoid. There was no significant difference in VRUT between Adx and Adx + STZ rats. However, VRUT was significantly reduced by 32% in the IM tip of glucocorticoid-treated Adx + STZ rats compared with control Adx + STZ rats. We conclude that glucocorticoids regulate the abundance of VRUT protein independently of insulin in diabetic rats.

Adrenalectomy↗
PubMed1997

Safety and efficacy of low-dose subcutaneous erythropoietin in hemodialysis patients.

Anemia in hemodialysis patients is effectively treated by intravenous (IV) injections of recombinant human erythropoietin (rHuEPO) at each dialysis session. Because the hormone is effective by subcutaneous (SC) administration, it was decided that this study would evaluate low-dose weekly SC rHuEPO therapy. To determine the safety and efficacy of weekly SC rHuEPO administration to hemodialysis patients, only one third the weekly IV dose was given and the effects were compared with those from an age-, gender-, and nephrologic disease-matched control group treated in the standard fashion. Forty-four patients entered the trial and 27 completed the protocol along with 27 control subjects. During Phase 1, experimental and control subjects received standard IV rHuEPO at dialysis for 6 months. During Phase 2, experimental patients received weekly SC rHuEPO at one third the weekly IV dose for 10 months; control subjects continued to receive IV therapy. In Phase 3, both groups were treated for 6 more months with IV rHuEPO. In Phase 2, there was no significant reduction in hematocrit value, reticulocyte count, transferrin saturation, or ferritin level in the experimental group, even with only one third the weekly rHuEPO IV dose over the 10-month period. There were no significant differences between IV and SC rHuEPO administration or between experimental and control subjects in blood pressure, serum chemistries, or parameters of "dialysis adequacy." It was concluded that low-dose weekly SC rHuEPO administration is a safe and effective method for maintaining the hematocrit level of stable hemodialysis patients. This therapy could enhance the efficacy of rHuEPO and substantially reduce costs while preserving patient care outcomes.

Anemia↗
PubMed1997

Cellular mechanisms controlling protein degradation in catabolic states.

The daily turnover of protein amounts to 280 g in an adult weighing 70 kg but the metabolic processes responsible for protein turnover are only just beginning to be understood. In cells, the major pathway of protein degradation is the ubiquitin-proteasome pathway and protein flux through this pathway is precisely regulated. In catabolic conditions such as uremia, activity of the ubiquitin-proteasome pathway increases, resulting in degradation of muscle protein. In addition to increased protein degradation, gene transcription is activated, resulting in higher levels of the mRNAs encoding ubiquitin and proteasome subunits. The signals activating this pathway include metabolic acidosis and glucocorticoids but must be more diverse since the pathway is also activated in response to starvation, sepsis, cancer, muscle denervation, thermal injury, and acute diabetes. Understanding how the pathway is controlled could lead to the prevention of muscle loss in uremia and other conditions.

Animals↗
PubMed1997

Signals regulating accelerated muscle protein catabolism in uremia.

In chronic renal failure (CRF), the ATP-dependent, ubiquitin-proteasome proteolytic pathway is activated with concurrent increases in the transcription of genes encoding proteins of this pathway in muscle. We have shown that the stimuli for these responses include acidosis and glucocorticoids, but other endocrine abnormalities in CRF (e.g., insulin resistance) could contribute to these responses. In fact, a major effect of insulin in muscle is to suppress protein degradation. To examine whether insulin influences the ubiquitin-proteasome pathway, we measured protein degradation in incubated epitrochlearis muscles of diabetic and pair-fed control rats. Muscle proteolysis was increased in pathways that do not involve lysosomes or Ca(2+)-dependent proteases; but MG132, a protease inhibitor that blocks ATP synthesis, eliminated the accelerated rate of protein degradation in diabetic rat muscles. Diabetes mellitus also increased levels of mRNAs encoding ubiquitin (334%), E2 ubiquitin-conjugating enzyme (247%), and the C3 (320%), C5 (349%), and C9 (216%) proteasome subunits in muscle. Finally, transcription of the ubiquitin gene in diabetic rat muscles was increased. Diabetic rats were acidotic, but eliminating acidemia by giving NaHCO3 did not block the increase in muscle proteolysis. Giving diabetic rats insulin prevented the excessive muscle proteolysis, suggesting that insulin acts as a suppressor of the ubiquitin-proteasome pathway. Thus, the insulin resistance of uremia could contribute to muscle protein wasting in CRF.

Animals↗
PubMed1996

Muscle wasting in insulinopenic rats results from activation of the ATP-dependent, ubiquitin-proteasome proteolytic pathway by a mechanism including gene transcription.

In normal subjects and diabetic patients, insulin suppresses whole body proteolysis suggesting that the loss of lean body mass and muscle wasting in insulinopenia is related to increased muscle protein degradation. To document how insulinopenia affects organ weights and to identify the pathway for accelerated proteolysis in muscle, streptozotocin-treated and vehicle-injected, pair-fed control rats were studied. The weights of liver, adipose tissue, and muscle were decreased while muscle protein degradation was increased 75% by insulinopenia. This proteolytic response was not eliminated by blocking lysosomal function and calcium-dependent proteases at 7 or 3 d after streptozotocin. When ATP synthesis in muscle was inhibited, the rates of proteolysis were reduced to the same level in insulinopenic and control rats suggesting that the ATP-dependent, ubiquitin-proteasome pathway is activated. Additional evidence for activation of this pathway in muscle includes: (a) an inhibitor of proteasome activity eliminated the increased protein degradation; (b) mRNAs encoding ubiquitin and proteasome subunits were increased two- to threefold; and (c) there was increased transcription of the ubiquitin gene. We conclude that the mechanism for muscle protein wasting in insulinopenia includes activation of the ubiquitin-proteasome pathway with increased expression of the ubiquitin gene.

Adenosine Triphosphate↗
PubMed1996

The acidosis of chronic renal failure activates muscle proteolysis in rats by augmenting transcription of genes encoding proteins of the ATP-dependent ubiquitin-proteasome pathway.

Chronic renal failure (CRF) is associated with negative nitrogen balance and loss of lean body mass. To identify specific proteolytic pathways activated by CRF, protein degradation was measured in incubated epitrochlearis muscles from CRF and sham-operated, pair-fed rats. CRF stimulated muscle proteolysis, and inhibition of lysosomal and calcium-activated proteases did not eliminate this increase. When ATP production was blocked, proteolysis in CRF muscles fell to the same level as that in control muscles. Increased proteolysis was also prevented by feeding CRF rats sodium bicarbonate, suggesting that activation depends on acidification. Evidence that the ATP-dependent ubiquitin-proteasome pathway is stimulated by the acidemia of CRF includes the following findings: (a) An inhibitor of the proteasome eliminated the increase in muscle proteolysis; and (b) there was an increase in mRNAs encoding ubiquitin (324%) and proteasome subunits C3 (137%) and C9 (251%) in muscle. This response involved gene activation since transcription of mRNAs for ubiquitin and the C3 subunit were selectively increased in muscle of CRF rats. We conclude that CRF stimulates muscle proteolysis by activating the ATP-ubiquitin-proteasome-dependent pathway. The mechanism depends on acidification and increased expression of genes encoding components of the system. These responses could contribute to the loss of muscle mass associated with CRF.

Acidosis↗
PubMed1996

Glucocorticoids and acidosis stimulate protein and amino acid catabolism in vivo.

We have shown that chronic metabolic acidosis in awake rats accelerates whole body protein turnover using stochastic modeling and a continuous infusion of L-[1-13C] leucine. To delineate the role that glucocorticoids play in mediating these catabolic responses, we measured protein turnover in awake, chronically catheterized, adrenalectomized rats in the presence or absence of glucocorticoids and/or a NH4Cl feeding regimen which induced chronic metabolic acidosis. In adrenalectomized rats receiving no glucocorticoids there was no statistical difference in amino acid oxidation, protein degradation or synthesis whether or not the rats had acidosis. In contrast, chronically acidotic, adrenalectomized rats receiving glucocorticoids demonstrated accelerated whole body protein turnover with a 84% increase in amino acid oxidation and a 26% increase in protein degradation, compared to rats not receiving glucocorticoids or those given the same dose of glucocorticoids but without acidosis. We conclude that metabolic acidosis accelerates amino acid oxidation and protein degradation in vivo, and that glucocorticoids are necessary but not sufficient to mediate the catabolic effects of metabolic acidosis.

Acidosis↗
PubMed1996

A comparison of three methods for detecting the acrosome reaction in human spermatozoa.

This study was designed to compare three different fluorescent probes to assay the acrosome reaction in human spermatozoa: chlortetracycline (CTC), mannosylated bovine serum albumin (BSA) labelled with fluorescein (MAF), and quinacrine (QN). Normal human sperm ejaculates were washed and allowed to swim up for 30-60 min. Samples were examined under epifluorescence for the percentage of the acrosome reacted spermatozoa, as detected by the three probes. There was no significant differences between samples of fresh, uncapacitated spermatozoa evaluated with CTC, MAF or QN; all gave < 10% reacted. Following capacitation for 3 h, the percentage of spontaneously reacted spermatozoa was higher than in fresh spermatozoa; CTC and MAF gave the same percentage (12%), while QN indicated a higher percentage (18%) of reacted spermatozoa (P < 0.001). Following exposure to ionophore A23187 at 1 h, the percentage of acrosome reactions increased to a mean of 31% as detected with CTC or MAF; the mean percentage (45%) was significantly higher with QN (P < 0.0001). Further incubation up to 2 h with A23187 did not change these percentages. These results suggest that the QN probe detects the onset stage of the acrosome reaction, whereas the CTC and MAF probes detect the later stages in which the acrosomal cap is lost. Use of the two types of probe provides a means for finer resolution of the time course of the acrosome reaction in the human spermatozoa.

Acrosome↗
PubMed1996

Influence of acid loading, extracellular pH and uremia on intracellular pH in muscle.

UNLABELLED: Our previous work has shown that chronic metabolic acidosis can induce changes in protein and amino acid metabolism in muscle. The relationship of these metabolic responses to changes in muscle pH is unknown. To examine the role that acute acidosis might have on intracellular pH in rats, we evaluated the influence an acute infusion of HCl had on intracellular pH using 31P-NMR. Male rats fed 22% protein and weighing between 150 and 170 g were infused with 150 mM HCl (4 mmol/kg) over a 2- or 4-hour period. Baseline blood pH was 7.36 and dropped to 7.04 at the end of the infusion. Despite this, no changes in intracellular pH were seen. In contrast, we found that chronic acid loading (4 mmol NH4Cl/100 g/day for 5 days) produces a small change in muscle pH (0.05 pH units, p < 0.05). Chronic uremia did not change resting muscle pH despite a decrease in extracellular pH to 7.23 even though metabolic changes are well documented. CONCLUSIONS: (1) An acute acid load does not alter intracellular pH while chronic metabolic acidosis does reduce intracellular pH. (2) The lack of an acute change in intracellular pH suggests that intracellular buffering capacity changes over time.

Acidosis↗
PubMed1996

Acidosis and glucocorticoids induce branched-chain amino acid catabolism.

Chronic renal failure (CRF) is frequently complicated by malnutrition and wasting. The loss of lean body mass in CRF is the result of accelerated protein and amino acid degradation. Both appear to occur via acidosis-induced, glucocorticoid-dependent processes. In skeletal muscle, acidosis stimulates the activity of the rate-limiting enzyme in branched-chain amino acid metabolism, branched-brain ketoacid dehydrogenase (BCKAD). The activation of BCKAD in acidosis is likely to be glucocorticoid-dependent.

3-Methyl-2-Oxobutanoate Dehydrogenase (Lipoamide)↗