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Biomedical subjects

R J Hughes

Publications and source records attributed to R J Hughes.

18 recordsLinked to original sources
PubMed1992

The effect of exercise training on salivary immunoglobulin A and cortisol responses to maximal exercise.

The salivary immunoglobulin A (s-IgA) and cortisol responses to maximal exercise were examined in 24 adult males (X +/- SD; 22.1 +/- 3.0 yrs) before and after 10 weeks of run training. The subjects performed an incremental treadmill test to exhaustion and were randomly assigned to one of three groups: control (CON; n = 5), low intensity training (LO; n = 8), or high intensity training (HI; n = 11). Following the ten weeks of training, the subjects performed a second maximal treadmill test. Saliva samples were collected before, as well as immediately and 1 hr following each of the maximal treadmill tests and were analyzed for s-IgA and salivary cortisol. Maximal oxygen consumption (VO2max) increased significantly (p < 0.05) in the LO and HI groups but remained unchanged in the CON group. The s-IgA levels decreased significantly (p < 0.05) immediately post-exercise but returned to pre-exercise levels by one hour recovery. In addition, s-IgA and cortisol levels were not significantly (p > 0.05) correlated at any of the sampling times. These findings indicated that the s-IgA response to maximal exercise was unaffected by moderate (70% of VO2 max) to heavy (86% of VO2max) training (designed to develop cardiorespiratory fitness in healthy non-athletic adults) and independent of salivary cortisol.

Adult↗
PubMed1992

The effect of exhaustive exercise on salivary immunoglobulin A.

The purpose of this study was to examine salivary immunoglobulin A (s-IgA) responses to exhaustive exercise. Twenty-nine, college age (means +/- SD = 21.45 +/- 3.1 yrs; range 18-29 yrs) moderately active males (running < 10 miles per week) performed an incremental treadmill test to exhaustion. Unstimulated salivary samples, collected before as well as immediately and one hour following the test, were analyzed for s-IgA using an enzyme-linked immunosorbent assay (ELISA). Mean s-IgA levels decreased significantly (24.4%, p < 0.05) immediately following the maximal test, and remained depressed (16.9%) one hour after the test. Of the 29 subjects, five exhibited an increase (range = 1.3 to 53.3%) in s-IgA following the exercise bout. The results of this study suggest that exhaustive exercise may temporarily reduce salivary IgA levels.

Adolescent↗
PubMed1991

Decreased beta 2-adrenergic receptor mRNA expression in receptor-deficient S49 lymphoma cells.

Many variants of the S49 mouse lymphoma cell have been isolated along the pathway of cyclic AMP generation and response. Two such variants, beta p and beta d, were isolated by Johnson and colleagues and described in 1979 [Mol. Pharmacol. 15:16-27 (1979)]. The beta p and beta d variants express one half and one quarter, respectively, of the wild-type number of beta 2-adrenergic receptors. This observation has now been extended through the use of DNA-excess solution hybridization. Using this exquisitely sensitive technique for quantitation of gene and mRNA, we have been able to demonstrate that the beta 2-adrenergic receptor-deficient variant cells contain the same quantity of the beta 2-adrenergic receptor gene as the wild-type cells. In contrast, the beta 2-adrenergic receptor-deficient variant cells express reduced quantities of beta 2-adrenergic receptor-specific mRNA. The amount of beta 2-adrenergic receptor-specific mRNA correlates very well with the reduction in receptor expression in these cells. Both gene and mRNA in the wild-type and variant cells appear to be the same size, as judged by Southern and Northern analysis. Thus, the diminution of beta 2-adrenergic receptors in the beta p and beta d variants appears to reflect primarily the relative paucity of gene transcripts in the variant cells. These data imply that variations in cellular content of beta 2-adrenergic receptor mRNA, which may occur among closely related cells, is one explanation for differences in receptor number.

Animals↗
PubMed1990

The effect of glycogen depletion and supercompensation on the physical working capacity at the fatigue threshold.

The purpose of this investigation was to determine the effect of glycogen depletion and supercompensation on the physical working capacity at the fatigue threshold (PWCFT). Ten adult males (mean age 23 years, SD 3) volunteered as subjects for this study. During the first laboratory visit the subjects performed a maximal bicycle ergometer test for the determination of maximum oxygen consumption (VO2max). Between 48 and 72 h later, the subjects pedaled to exhaustion at a power output which corresponded to a mean of 76% of VO2max (range, 72-80%) for the purpose of glycogen depletion. For the next 3 days, the subjects were fed a 10.5 MJ.day-1 low carbohydrate diet which consisted of 7.5% carbohydrates, 22.0% protein and 70.5% fat. The subjects then performed an incremental cycle ergometer test to the onset of fatigue or PWCFT, which was estimated from integrated electromyographic voltages of the vastus lateralis muscle. For the next 3 days the subjects were fed a 10.5 MJ high carbohydrate diet which consisted of 72.2% carbohydrates, 12.4% protein and 15.4% fats for the purpose of glycogen supercompensation. The subjects then performed a second PWCFT test. A paired t-test indicated that there was no significant (p greater than 0.05) difference between the means of the PWCFT values (depletion 246 W, SD 30; supercompensation 265 W, SD 28) and they were highly correlated at r = 0.884. The results of this investigation suggested that the methods commonly used to affect glycogen depletion or supercompensation had no effect on PWCFT.

Adaptation, Physiological↗
PubMed1989

Isokinetic strength and body composition of high school wrestlers across age.

The purpose of this study was to determine the changes in body composition as well as absolute and relative isokinetic forearm and leg strength of high school wrestlers across age. One hundred ninety-five wrestlers (means age +/- SD = 16.36 +/- 1.12 yr) volunteered to be measured for strength using a Cybex II dynamometer at 30, 180, and 300 degrees.s-1. In addition, underwater weighing was used to determine body composition characteristics. The subjects were divided into four age groups: group 1 (G1) = 14.17-15.00 yr (N = 20); group 2 (G2) = 15.01-16.00 yr (N = 60); group 3 (G3) = 16.01-17.00 yr (N = 52); and group 4 (G4) = 17.01-18.50 yr (N = 63). One-way ANOVA or ANCOVA with Tukey post hoc comparisons indicated significant (P less than 0.05) changes across age for height, body weight, and fat-free weight, as well as absolute and relative forearm and leg strength. The results of this study indicated that, while a large portion of the improvements in strength across age were associated with increases in lean tissue, there was an additional "age effect" which could not be accounted for by changes in fat-free weight. Although the mechanism responsible for the "age effect" is unclear, it is possible that neural development contributed to the strength increases across age.

Adolescent↗
PubMed1989

A mobile traction device for MR and CT imaging.

The development of a spinal traction device for use in Magnetic Resonance Imaging (MRI) and Computed Tomography (CT) imaging is described. Such a device has arisen from the need to apply continuous traction to patients having suffered acute cervical spine trauma. Previous traction techniques have proven unsuitable for use in the MRI scanner, particularly as they lack stability, are awkward to implement and prove quite cumbersome to manage. Traction may be applied to a patient for periods of time in excess of 1 hour with a minimum of discomfort. Materials are used which have a negligible proton density and a low atomic number, yielding no detectable magnetic resonance and very small x-ray cross sections respectively. Calibrated traction may be applied to patients via tongs or a head harness, the latter rendering the device suitable for general transport of injured patients with traction applied. The device is compact, light and strong and contains no ferrometallic parts. Thus far in excess of 40 patients have been scanned, most in a FONAR beta-3000M .3 Tesla electromagnetic MR imager and some also in a GE 9800 Quickscan CT scanner. No artefact has been demonstrated in any of the images to date for any of the MR pulse sequences or CT kVp. Modifications likely include refinement of the tensioning mechanism, inclusion of x-ray cassette facilities and improvement of aesthetic design.

Equipment Design↗
PubMed1988

Certain beta-blockers can decrease beta-adrenergic receptor number: II. Down-regulation of receptor number by alprenolol and propranolol in cultured lymphoma and muscle cells.

We have used two different cultured cell lines--S49 lymphoma cells and BC3H-1 muscle cells--to examine the regulation of beta-adrenergic receptors by receptor antagonists. Rather than an increase ("up-regulation") of receptor number that such antagonists often produce, we found that certain beta-blockers elicit a decrease ("down-regulation") of beta-adrenergic receptors. Alprenolol and propranolol, but not sotalol or ICI 118,551, at concentrations of 10-100 nM down-regulated beta-adrenergic receptors 20-70% following 16-20 hours of treatment of S49 or BC3H-1 cells. Several observations suggest that this phenomenon depends upon beta-receptor interaction, including stereoselectivity [(-)-enantiomers more potent than (+)-enantiomers], blockade of the effect by ICI 118,551, absence of down-regulation of alpha-adrenergic receptors in BC3H-1 cells, and lack of a decrease in beta-adrenergic receptor-independent (forskolin-stimulated) cyclic AMP accumulation in S49 cells. The possibility of retained antagonist interfering with receptor measurement was precluded by the fact that the antagonist-induced decrease in receptor number required several hours incubation and occurred without a prominent change in receptor affinity. The ability of the beta-blockers to elicit down-regulation did not correlate with hydrophobicity of the drugs. Antagonist-induced down regulation of beta-adrenergic receptors did not occur in S49 lymphoma cells that lack the alpha-subunit of Gs, the guanine nucleotide-binding regulatory protein, thus implying a requirement for receptor-alpha s interaction in eliciting beta-receptor down-regulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenergic beta-Antagonists↗
PubMed1988

Receptor-induced degradation of atrial natriuretic peptide by a rabbit carotid artery smooth muscle cell.

We have used a smooth muscle cell line isolated from rabbit carotid artery (RCA) as a model system with which to study the expression of atrial natriuretic peptide (ANP) receptors and, in addition, the receptor-mediated degradation of ANP. RCA cells bind rat alpha ANP-(1-28) reversibly at 37 C, apparently to a single class of high affinity (Kd approximately equal to 50 pM) binding sites (approximately equal to 20,000 sites per cell). Binding of rat alpha ANP-(1-28) elicits rapid accumulation of intracellular cGMP. However, the concentrations of rat alpha ANP-(1-28) and related peptides, abbreviated at the N- and C-terminals, required to stimulate cGMP synthesis are substantially greater than those required for binding. Analysis by HPLC of 125I-labeled rat alpha ANP-(1-28) bound to RCA cells at 37 C illustrates the rapid and continuous degradation of the radioiodinated rat alpha ANP-(1-28) to two radiolabeled products, one of which, 125I-labeled tyrosine is the major radiolabeled component that dissociates from the cells. Measurement of rat alpha ANP-(1-28) interaction with RCA cells by radioligand binding techniques therefore subsumes several processes. One of these processes is the rapid and continuous degradation of specifically bound ANP by these cells and perhaps also other target cells that respond to ANP.

Animals↗
PubMed1987

Regulation of the atrial natriuretic peptide receptor on a smooth muscle cell.

We have used a smooth muscle cell line isolated from rabbit carotid artery (RCA) as a model system with which to study the ligand-mediated regulation of the atrial natriuretic peptide (ANP) receptor. RCA cells bind rat alpha-ANP-(1-28) semi-irreversibly at 4 degrees C, apparently to a single class of high-affinity binding sites (Kd approximately equal to 5 pM). In contrast, ANP initially bound to RCA cells at 37 degrees C is fully reversible after the cells have been cooled to 4 degrees C. Although the measured dissociation constant for binding rat alpha-ANP-(1-28) is 10-fold lower at 4 degrees C than at 37 degrees C, the maximum amount of specifically bound radioligand is equivalent at both temperatures. We infer this to be suggestive of ANP receptor recycling occurring at 37 degrees C. This hypothesis is substantiated by experiments demonstrating the rapid disappearance of ANP receptors (measured at 4 degrees C) after exposure of RCA cells to ANP at 37 degrees C. We believe this reflects a loss of cell-surface receptors that is reversible on removal of ANP from the incubation medium at 37 degrees C. The concentration dependence and rank order of potency of ANP in eliciting receptor redistribution resembles that for binding to the ANP receptor and not for eliciting cyclic GMP formation. In contrast to this striking loss of presumed cell-surface receptors, incubation of RCA cells with ANP elicits only limited desensitization of the cyclic GMP response.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Alpha 1-adrenergic receptor-linked guanine nucleotide-binding protein in muscle and kidney epithelial cells.

We have studied the interaction of guanine nucleotides with alpha 1-adrenergic receptors of two cloned cell lines, the Madin Darby canine kidney (MDCK-D1) cells and BC3H-1 muscle cells. Although guanylylimidodiphosphate, Gpp(NH)p, had no effect on the affinity or the total number of [3H]prazosin-binding sites in membranes prepared from these cells, the nucleotide decreased the apparent affinity of the agonists (-)-epinephrine and (-)-norepinephrine in competing for [3H]prazosin-binding sites in both cell types. A maximal effect of Gpp(NH)p occurred at 10 microM. Guanine nucleotides were significantly more effective in shifting agonist affinity for the alpha 1 receptor than adenine nucleotides, and Mg2+ was required to observe a maximal effect. Binding of agonist to alpha 1-adrenergic receptors activated phosphatidylinositol (PI) hydrolysis in both cell types but had no effect on membrane adenylate cyclase activity. Incubation of MDCK cells for 19 hr with 100 ng/ml pertussis toxin, which eliminated the ability of pertussis toxin added to membranes to ADP-ribosylate 39-41-KDa substrate(s), failed to alter binding of agonists to alpha 1-adrenergic receptors, the ability of Gpp(NH)p to regulate agonist binding to these receptors, or epinephrine-stimulated PI hydrolysis and prostaglandin E2 production. Incubation of BC3H1 cells with pertussis toxin had no effect on the ability of epinephrine to stimulate PI turnover. These results show that binding of agonists to alpha 1-adrenergic receptors in mammalian kidney and muscle cells is regulated by guanine nucleotides, presumably by interaction with a guanine nucleotide-binding (G) protein. The failure of the G-protein to regulate adenylate cyclase activity and the lack of effect of pertussis toxin to alter receptor-mediated binding or functional activity suggests that a G-protein other than Gs, Gi, or Go interacts with alpha 1-adrenergic receptors in kidney and smooth muscle.

Adenylate Cyclase Toxin↗
PubMed1987

Interactions of amiloride with alpha- and beta-adrenergic receptors: amiloride reveals an allosteric site on alpha 2-adrenergic receptors.

Interaction of amiloride with adrenergic receptors was studied using radioligand binding techniques. Amiloride competed for [3H]prazosin binding to alpha 1-adrenergic receptors on rat renal cortical membranes and BC3H-1 muscle cell membranes. Non-linear regression analysis of radioligand binding isotherms showed that amiloride increased Kd without a change in Bmax, suggesting the drug binds competitively in a mutually exclusive manner with the radioligand at the receptor-binding site. Similarly, amiloride competitively blocked [125I]iodocyanopindolol binding to beta-adrenergic receptors on both tissues. The addition of guanylyl 5'-imidodiphosphate or sodium chloride did not alter the interaction of amiloride with alpha 1- or beta-adrenergic receptors. The interaction of amiloride with alpha 2-adrenergic receptors was more complex and revealed an allosteric site. In both rat renal cortical membranes and intact human platelets, amiloride increased the Kd for [3H]rauwolscine binding, as well as decreasing the apparent Bmax. In binding experiments where amiloride competed for [3H]rauwolscine-binding sites, pseudo-Hill slopes of less than 1.0 were obtained for both platelet and renal alpha 2 receptors. In addition, amiloride increased the rate of [3H]rauwolscine dissociation from renal alpha 2 receptors. In the presence of 100-120 mM sodium chloride, the Ki for amiloride competition was decreased an average of 54% in renal membranes; in contrast, sodium increased the Kd of the agonist epinephrine. Taken together, these data support the hypothesis that alpha 2-adrenergic receptors, but not alpha 1- or beta-adrenergic receptors, have an allosteric site to which amiloride binds and which we propose to be a cation-binding site.

Allosteric Regulation↗
PubMed1986

Agonist-mediated regulation of alpha 1- and beta 2-adrenergic receptor metabolism in a muscle cell line, BC3H-1.

We have compared the metabolism of alpha 1- and beta 2-adrenergic receptors which are both expressed in BC3H-1 muscle cells. During growth of the cells to confluence, the number of alpha 1-receptors per mg of membrane protein increases, whereas that of the beta 2-receptors remains constant. Experiments using cycloheximide and irreversible alpha 1- and beta 2-receptor antagonists, phenoxybenzamine and N-[2-hydroxy-3-(1-naphthoxy)-propyl]-N'-bromoacetylethylenediamine , respectively, yield disparate turnover rates (t1/2) for the two receptors: alpha 1 congruent to 25 hr, beta 2 congruent to 200 hr. These experiments suggest that synthesis of beta 2-receptors virtually ceases in confluent cells. Maximally effective doses of agonists down-regulated both receptor types 80-90% and enhanced the rates of loss of both receptors (t1/2 = 1-5 hr). The rates of down-regulation were not affected by cycloheximide, implying that agonists enhance receptor clearance rather than decrease receptor appearance. The rank orders of potencies of agonists for promoting receptor down-regulation were those characteristic of alpha 1- and beta 2-receptors. However, concentrations of agonists that resulted in down-regulation of each receptor subtype were 10- to 100-fold lower than those required for occupancy of receptors as assessed in radioligand binding studies. Receptor recovery following removal of agonists was blocked by cycloheximide and was much faster than the recovery that followed treatment of cells with irreversible antagonists. Therefore, protein synthesis (but perhaps not receptor synthesis per se) appears necessary for recovery from down-regulation. In addition, the rates of recovery of alpha 1- and beta 2-receptor-mediated functions (phosphatidylinositol turnover and cyclic AMP synthesis, respectively) following receptor down-regulation or irreversible blockade parallel the rates of receptor recovery. These data indicate that basal metabolism of alpha 1- and beta 2-receptors in BC3H-1 cells is substantially different, but that agonist-mediated changes in metabolism of the two receptor subtypes are similar. Thus, common mechanisms appear to mediate the regulation by agonists of alpha 1- and beta 2-receptors in these cells.

Animals↗
PubMed1979

Departure from ideality in saturation binding assays.

In this paper we show that the measurement of cyclic AMP by a protein saturation assay deviates from ideality due to the presence of two classes of binding sites. An equation was derived which completely describes saturation assays and the effects on these of varying each of the component parameters was measured. The relevance of these findings to other protein saturation assays is discussed.

Binding Sites↗
PubMedDate not supplied

Comparison of methods for the analysis of tricyclic antidepressants in small whole blood samples.

A variety of liquid-liquid and solid-phase extraction techniques have been compared for the extraction of tricyclic antidepressant drugs from small whole blood samples. Each of the procedures was evaluated with respect to extract cleanliness, extraction efficiency, and reproducibility. The three most efficient methods were compared on a variety of authentic whole blood case samples containing drugs at therapeutic and toxic levels. The results of the study indicated that a liquid-liquid extraction procedure involving the extraction of drugs by ether was the most suitable to use on poor quality forensic samples.

Antidepressive Agents, Tricyclic↗