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Biomedical subjects

T M Murray

Publications and source records attributed to T M Murray.

At least 73 records · Page 4Linked to original sources
PubMed1985

Binding of intact parathyroid hormone to chicken renal plasma membranes: evidence for a second binding site with carboxyl-terminal specificity.

We studied the binding of bovine PTH (bPTH) to chicken renal plasma membranes using an intact hormone radioligand, [125I]bPTH-(1-84). In contrast to previous studies using amino-terminal radioligands, our experiments revealed the presence of two distinct binding sites for intact bPTH. Scatchard analysis of competition curves consistently gave a biphasic curve, and computerized nonlinear regression analysis indicated a high affinity [dissociation constant (Kd) = 1.21 nM] and a low affinity (Kd = 333 nM) site (P less than 0.001). Analysis of binding of [125I] bPTH-(1-34) using identical techniques revealed only one site, similar to the high affinity sites seen with intact hormone tracer. The low affinity site for [125I]bPTH-(1-84) had carboxyl-terminal specificity since analysis of competition curves with unlabeled human PTH-(53-84) gave virtually identical binding parameters to the low affinity site obtained by competition with unlabeled native hormone. Binding to the low affinity site was inhibited in the presence of 10 mM Mg2+ and was reduced after storage of membranes for over 1 month at -70 C. Association of [125I]bPTH-(1-84) to the low affinity site took distinctly longer (approximately 4 h) than to the high affinity site (essentially complete by 2.25 h). Our data suggest that the high affinity site is coupled to adenylate cyclase in these membranes, while the low affinity site is not. The physiological significance of the low affinity carboxyl-terminal PTH binding is not known, and further studies are indicated.

Adenylyl Cyclases↗
PubMed1984

Effect of a 42.2-km footrace and subsequent rest or exercise on muscular strength and work capacity.

This study 1) quantitates the effect of a 42.2-km footrace (marathon) on leg extensor strength (maximal peak torque, MPT) and work capacity (WC, measured during a leg extensor fatigue test), and 2) describes the effect of either a rest or exercise regimen for 1 wk after the marathon on the recovery of MPT and WC. Ten trained male runners performed personal records in a marathon and were then randomly assigned to either a rest or exercise-recovery group. The rest group did not train, whereas the exercise group ran 20-45 min/day at their selected intensity of exercise [50-60% maximal O2 consumption (Vo2max)] during the recovery week. MPT was measured at 1.1, 3.2, and 5.3 rad X s-1. The total work generated during a 50-contraction active extension-passive flexion fatigue test conducted at 3.2 rad X s-1 was defined as WC. Reports of perceived soreness of the quadriceps were obtained before each strength-testing session. These measurements were obtained before the marathon and 15-20 min and 1, 3, 5, and 7 days postmarathon. A significant reduction in MPT and WC resulted and continued 1 day postmarathon. MPT of both groups improved through day 5 postmarathon at 1.1 and 3.2 rad X s-1. MPT of the rest group improved through day 7 postmarathon but remained less than premarathon MPT. Recovery of MPT was impaired in the exercise group through days 5-7 postmarathon after 40-45 min exercise at 60% Vo2max. WC was recovered 3 days postmarathon in the rest group but was still impaired 7 days postmarathon in the exercise group.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1983

Preparation and properties of biologically active radioiodinated parathyroid hormone.

A constant-current microelectrolytic radioiodination method was used to label bovine parathyroid hormone (BPTH) with 125I to an overall iodination ratio of 1:1 iodide atoms per PTH molecule. Such iodinated preparations were shown to be fully active in several bioassay systems: in vitro adenylate cyclase activation in rat renal and skeletal membranes, in vitro calcium release from rat calvaria, and the in vivo hypercalcemic response in chickens. Analysis by Sephadex G-15 chromatography after enzymatic digestion showed the radioiodine to be incorporated predominantly as monoiodotyrosine. Bioassay of iodinated preparations from which uniodinated hormone had been removed by isoelectric focusing showed essentially full hormonal activity. Such methods can be used to consistently produce radioiodinated biologically active preparations of BPTH 1-84 with high specific activity (2000 Ci/mmol).

Adenylyl Cyclases↗
PubMed1983

Elimination of cross-reactivity to some component of erythrocytes in an antiserum to parathyroid hormone by absorption with fixed erythrocytes.

Cross-reactivity to some component of rat erythrocytes with an antiserum to parathyroid hormone (anti-PTH) was detected in fixed demineralized sections of bone using the peroxidase-antiperoxidase method. The cross-reactivity was eliminated by the preabsorption of anti-PTH with fixed, washed rat erythrocytes. This technique provides an easy and rapid method with which to eliminate cross-reactivity to erythrocytes whenever such a situation is encountered in immunohistochemical procedures.

Absorption↗
PubMed1983

Binding of radioiodinated bovine parathyroid hormone-(1-84) to canine renal cortical membranes.

Binding of PTH to purified canine renal cortical membranes was investigated using biologically active radioiodinated bovine PTH-(1-84) [bPTH-(1-84)] as radioligand. PTH-(1-84) is thought to be the major circulating form of bioactive PTH, but oxidative inactivation upon iodination has prevented its use as a radioligand probe of PTH receptors. We have labeled PTH-(1-84) by a microelectrolytic constant current method to a high specific activity (180-220 muCi/micrograms), corresponding to an average ratio of 1 mol 125I/mol peptide. Affinity purification on chick renal membranes consistently improved the radioligand-binding properties, with a 6-fold increase in fraction specifically bound. Analysis of equilibrium (180 min at 15 C) competition curves showed two classes of binding sites for bPTH-(1-84). A high affinity binding site appeared to be coupled to activation of adenylate cyclase and exhibited affinity varying between 1.9 and 4.3 X 10(8) M-1. The affinity of this site for bPTH was decreased more than 50% by the nonhydrolyzable analog of GTP, guanyl-5'-yl-imidodiphosphate. A low affinity binding site also was detected (Ka = 0.6-6.2 X 10(6) M-1), and its affinity for bPTH was modulated by the concentration of magnesium. The high affinity sites exhibited hormonal specificity and guanine nucleotide dependency characteristic of peptide hormone receptors, while the low affinity sites did not. Analysis by polyacrylamide gel electrophoresis or high pressure liquid chromatography of the radioligand incubated with or released from purified canine renal cortical membranes revealed a single peak of radioactivity that comigrated with [125I]iodo-bPTH-(1-84). It appears from these studies that cleavage of amino-terminal to residue 43 ([125I]tyrosine) was not required for binding or release of hormone. These experiments indicate that radioiodinated bPTH-(1-84) is a useful probe for further characterization of PTH receptors in kidney and other organs.

Adenylyl Cyclases↗
PubMed1983

Immunohistochemical demonstration of parathyroid hormone binding to specific cell types in fixed rat bone tissue.

Deparaffinized sections of fixed decalcified neonatal rat radii were incubated in bovine PTH (bPTH; 1-10 MRC units/ml) or in PTH-solvent. They were then stained for PTH by the peroxidase-antiperoxidase method using guinea pig antiserum to bPTH and the substrate 3,3'-diaminobenzidine-H2O2. Staining caused by nonspecific binding of PTH to the bone matrix and the glass slides supporting the sections was eliminated completely by preincubation of the sections in 100% normal goat serum. Cross-reactivity of the antiserum to erythrocytes was eliminated by preabsorption of the antiserum with fixed rat erythrocytes. After the cross-reactivity of the anti-PTH antiserum to erythrocyte components and the nonspecific binding of PTH to bone matrix were eliminated, we were able to demonstrate intense staining over the cytoplasm of the osteoclasts in rat radii sections incubated with PTH. Less intense staining was observed over the osteocytes, periosteal osteoblasts, and, possibly, the endosteal osteoblasts. An explanation for this differential staining could be that osteoclasts have a greater receptor number and/or a greater receptor affinity for the bPTH than do osteocytes and osteoblasts. This study demonstrates that binding of PTH to bone tissue can be localized in all identifiable osteoclasts, osteocytes, and osteoblasts, and thus suggests that all three cell types can interact directly with PTH.

Animals↗
PubMed1983

Binding of radioiodinated parathyroid hormone to cloned bone cells.

We have investigated the binding of biologically active radioiodinated bovine PTH-(1-84) to cloned osteoblast-like rat osteosarcoma cells (ROS 17/2.8) and correlated binding with biological response assessed as cAMP production. The hormone was labeled with 125I on tyrosine-43 using a constant current microelectrolytic method which allows full retention of biological activity. At confluency, cells were removed from culture, and assays were carried out on intact cells in suspension. At 22 C, saturable binding reached equilibrium by 90 min. At that time, the apparent dissociation rate constant was 8 X 10(-8) min-1. At an earlier time, 10 min of incubation, the dissociation rate was twice that observed at equilibrium. Nonsaturable binding was 30-35% of the total binding. The dissociation constant derived from kinetic analysis was 41 nM and correlated with the half-maximal cAMP production at 36 nM. The results obtained suggest that cleavage amino-terminal to residue 43 is not required for binding to these bone-derived cells. Binding to receptors on the osteosarcoma cells was specific and reversible, and appeared biologically relevant, since it correlated closely with the biological response, cAMP production. The competitive inhibitor [Nle8,Nle18,Tyr34]bPTH-(3-34)amide showed the same apparent affinity in inhibiting receptor binding as in inhibiting cAMP production.

Animals↗
PubMed1982

Increased bone apposition in primary hyperparathyroidism: measurements based on short interval tetracycline labeling of bone.

Bone biopsies of 23 subjects suffering from primary hyperparathyroidism, which was proven by the demonstration of parathyroid pathology during neck surgery, were studied by morphometric analysis and short interval sequential tetracycline labelling and compared with those of 13 patients without metabolic bone disease. In 19 of these patients, abnormalities were found in the trabecular volume, resorption surface, formation surface, or in various combinations. However, no constant pattern of bone changes was observed. In 4 patients, the bone morphometric parameters were normal. The bone apposition rate, on the other hand, was elevated in all 23 subjects. The rate was re-evaluated in 4 patients 6 wk to 3 mo following successful parathyroid surgery. It fell back within the control range. These findings confirm the observation in experimental animals that parathyroid hormone stimulates bone apposition in vivo and give support to the rationale of using this hormone in the treatment of osteoporosis.

Adolescent↗
PubMed1982

Parathyroid hormone stimulates the bone apposition rate independently of its resorptive action: differential effects of intermittent and continuous administration.

The deposition of mineralized bone matrix by differentiated osteoblasts was studied in rats in vivo by labeling the bone with three doses of tetracycline given at 48-h intervals. Only bone formation loci bearing all three tetracycline doses were measured, thus eliminating sites where bone formation was not continuous during the labeling period. Using this technique, the effects of intact bovine parathyroid hormone [bPTH-(1-84)] and of a synthetic amino-terminal fragment of human PTH [hPTH-(1-34)] were measured in thyroparathyroidectomized animals. bPTH-(1-84), administered sc, and hPTH-(1-34), administered iv, caused a dose-dependent increase in the bone apposition. Subcutaneous administration of hPTH-(1-34) in doses varying from 2.7-173.0 pmol/rat.day had no effect, probably due to the degradation of the hormone when administered this way. We also compared the effects of bPTH-(1-84) when administered by either daily sc injections or continuous infusion. Continuous infusion of bPTH-(1-84) resulted in an increased apposition rate. Using a morphometric technique, we also found an increase in both formation and resorption surfaces and a net decrease in the trabecular bone volume in this group. Daily injection of the hormone caused an increase in the bone apposition rate, accompanied by an increase in the formation surface without an increase in the resorption surface. This resulted in a net increase in trabecular bone volume. The results thus suggest that the resorptive effects of bPTH-(1-84) can be separated from the effects of the hormone on the apposition rate.

Animals↗
PubMed1981

Hypercalcemia with ovarian carcinoma: evidence of a pathogenetic role for prostaglandins.

A 70-year-old white woman had a lower abdominal mass and hypercalcemia. Physical and radiologic evidence was found for the presence of nonmetastatic pelvic tumor. Biochemical tests confirmed the presence of hypercalcemia with evidence of active bone resorption. Plasma parathyroid hormone (PTH) and the nephrogenic urinary cyclic AMP excretion were low; levels of plasma prostaglandins were elevated. Bone biopsy revealed histologic evidence of extensive osteoclastic bone resorption. At operation, a papillary serous cystadenocarcinoma of the ovary was removed. Postoperatively, the serum calcium fell to normal, and plasma prostaglandins became undetectable. Short-term incubation of ovarian tumor fragments demonstrated the production by tumor tissue of a substance causing bone resorption in an in vitro bioassay. The production of this substance was blocked by indomethacin. Radioimmunoassay of the incubation medium revealed significant amounts of prostaglandins of the E + F series. Parathyroid hormone was not detected in the medium. These data implicate tumor-produced prostaglandins as mediators of the hypercalcemia in this patient.

Aged↗
PubMed1981

Treatment of renal osteodystrophy with 1,25-dihydroxycholecalciferol.

Nine patients with renal osteodystrophy were tested for 6.5 to 35 months with 1,25-dihydroxycholecalciferol (1,25-DHCC). A close biochemical follow-up was performed during the first 6 months of treatment, including biweekly determinations of serum calcium, phosphorus, magnesium, alkaline phosphatase and creatinine levels. A bone biopsy, radiologic investigations and determinations of plasma levels of immunoreactive parathyroid hormone (IPTH) and intestinal absorption of calcium 47 were performed before and after the 6 months. Although the five patients with osteitis fibrosa showed a significant improvement, the four with predominantly osteomalacic lesions showed no response to treatment. These four had a normal initial plasma iPTH level, higher serum calcium levels than the other five patients, extreme sensitivity to 1,25-DHCC, with frequent episodes of hypercalcemia, and only a slightly increased serum alkaline phosphatase level, which remained unchanged during treatment. All but one of the patients, irrespective of the histologic abnormality, showed a decrease in the uptake of radionuclide by bone after treatment. The renal function of one patient, a man with long-standing stable renal failure who had not undergone dialysis, deteriorated during treatment.

Calcitriol↗
PubMed1981

Observations on the mechanism of bone resorption induced by multiple myeloma marrow culture fluids and partially purified osteoclast-activating factor.

Supernatant fluids from the cultures of bone marrow cells from 10 of 12 patients with multiple myeloma (MM) caused bone resorption in organ cultures of fetal rat calvaria. In four patients, the marrow cells were cultured with and without indomethacin (1 muM). The supernatant fluids from indomethacintreated marrow cultures caused significantly less bone resorption than supernatant fluids of cell cultures without indomethacin. This inhibition of release of bone resorbing factor(s) by myeloma cultures is similar to the previously observed indomethacin-induced inhibition of osteoclast-activating factor (OAF) production by activated human leukocytes. None of the MM supernatants had any effect on cyclic (c)AMP accumulation in resorbing bone in vitro. Four separate preparations of partially purified OAF obtained from phytohemagglutinin-stimulated peripheral human leukocytes were tested for their ability (a) to cause bone resorption in organ cultures of fetal rat and neonatal mouse calvaria and (b) to cause accumulation of cAMP in rat and mouse skeletal tissue in vitro. Those dilutions of OAF that caused bone resorption had no effect on accumulation of cAMP in rat or mouse calvaria incubated in vitro. In addition, no stimulation of adenylate cyclase activity in membranes prepared from fetal rat calvaria could be found. Bone cell populations isolated by sequential collagenase digestion of fetal rat calvaria also showed no cAMP response to these dilutions of OAF. Parathyroid hormone caused a clear response in all three systems. Furthermore, no cAMP response to OAF was observed in calvaria in the presence of cholera toxin (1 mug/ml) and isobutyl-methylxanthine (0.3 mM). These observations demonstrate that (a) supernatant fluids from MM marrow cultures stimulate bone resorption but do not increase cAMP accumulation in vitro; (b) indomethacin interferes with the release of bone resorbing factors by MM bone marrow cultures suggesting that this process requires prostaglandins; and (c) Sephadex G100 or G75 purified OAF does not stimulate adenylate cyclase or increase cAMP accumulation at equivalent bone resorbing concentrations in rat and mouse skeletal tissue. The resorptive action of MM culture fluids is similar to that of partially purified OAF from activated cultured leukocytes, but different from those of other bone resorbing factors, parathyroid hormone and prostaglandin E(2), which stimulate cAMP production in skeletal tissue.

Aged↗
PubMed1981

A possible role of vitamin D in the genesis of parenteral-nutrition-induced metabolic bone disease.

Patients receiving long term parenteral nutrition may develop metabolic bone disease. In all 11 patients studied, histologic studies of bone showed excessive unmineralized bone tissue despite normal plasma 25-hydroxyvitamin D levels. Three patients also had bone pain and fractures and severe urinary loss of calcium and phosphate. Withdrawal of vitamin D from parenteral nutrition solutions was associated with improved histologic findings of bone in all patients, shown by a decrease in osteoid tissue and an increase in tetracycline uptake. In the three patients with symptoms, bone pain subsided, fractures healed, and urinary loss of calcium and phosphate decreased. Thus, vitamin D may be a factor in the genesis of parenteral nutrition-induced metabolic bone disease.

25-Hydroxyvitamin D 2↗
PubMed1980

Metabolic bone disease in patients receiving long-term total parenteral nutrition.

We have prospectively investigated calcium and bone metabolism in 16 patients receiving total parenteral nutrition for periods ranging from 7 to 89 months. In 12 patients, bone biopsies at 6 to 73 months after the start of parenteral nutrition showed osteomalacia. Plasma 25-hydroxyvitamin D levels were normal in all patients. Seven persons developed hypercalcemia, and 10 had hypercalciuria with a negative calcium balance. Serum phosphorus was normal and plasma parathyroid hormone level, normal or decreased. Three patients with the severest form of the disease had vitamin D withdrawn from their solutions. Subsequently, urinary calcium decreased, and serum calcium became normal; two persons reverted to a positive calcium balance. Thus, patients receiving total parenteral nutrition may develop metabolic bone disease characterized by osteomalacia, hypercalcemia, hypercalciuria, and a negative calcium balance. This may be caused by both defective mineralization and increased bone resorption induced by vitamin D, its metabolites, or another unrecognized factor.

Adult↗