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Biomedical subjects

J D Wark

Publications and source records attributed to J D Wark.

At least 109 records · Page 6Linked to original sources
PubMed1989

Effects of tri-iodothyronine, cortisol and transcriptional inhibitors on vitamin D3-enhanced thyrotrophin secretion by rat pituitary cells in vitro.

The hormone 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) has been shown to selectively enhance agonist-induced TSH release in the rat thyrotroph in vitro. The interaction of 1,25-(OH)2D3 with tri-iodo-thyronine (T3) and cortisol was studied in primary cultures of dispersed anterior pituitary cells. TRH (1 nmol/l)-induced TSH release over 1 h was enhanced by 70% (P less than 0.01) following exposure to 10 nmol 1,25-(OH)2D3/l for 24 h. Pretreatment with T3 (1 pmol/l-1 mumol/l) for 24 h caused a dose-dependent inhibition of TRH-induced TSH release. Net TRH-induced TSH release was inhibited by 85% at T3 concentrations of 3 nmol/l or greater. Co-incubation with 1,25-(OH)2D3 resulted in enhanced TRH-induced TSH release at all T3 concentrations tested (P less than 0.001). The increment of TRH-induced TSH release resulting from 1,25-(OH)2D3 pretreatment was equivalent in the presence or absence of maximal inhibitory T3 concentrations. At 1 nmol T3/l, there was a two- to threefold relative increase in 1,25-(OH)2D3-enhanced TRH-induced TSH release. Incubation with cortisol (100 pmol/l-100 nmol/l) had no effect on basal or TRH-induced TSH release, nor did it alter 1,25-(OH)2D3-enhanced TRH-induced TSH release when added 24 h before, or at the time of addition of 1,25-(OH)2D3. Actinomycin D and alpha-amanitin abolished 1,25-(OH)2D3-enhanced TSH secretion. These data demonstrate that the action of 1,25-(OH)2D3 in the thyrotroph required new RNA transcription, and was not affected by cortisol. In the presence of T3, the response of the thyrotroph to TRH induced by 1,25-(OH)2D3 was increased. We have shown that 1,25-(OH)2D3 has significant effects on the action of TRH and T3 in vitro. These findings support the proposal that 1,25-(OH)2D3 may modulate TSH secretion in vivo.

Amanitins↗
PubMed1988

Chromatography of serum on Sep-pak C18 corrects falsely elevated vitamin D metabolite levels measured by protein binding assay.

Using a commercial kit method we obtained a vitamin D metabolite level within the normal range in a patient with biopsy-proven osteomalacia. This suggested that the ethanol extraction method employed had not removed serum factors known to falsely elevate the measurement of vitamin D metabolites. We therefore compared the levels measured after ethanol extraction and after purification by chromatography on Sep-pak C18 or Sephadex LH-20. Vitamin D metabolite levels after ethanol extraction of sera correlated with, but were higher than, those after chromatography on Sep-pak C18 cartridges (r = 0.84; 134 +/- 76 vs 76 +/- 46 nmol/l: mean +/- SD; p less than 0.01). Results were similar after chromatography on Sep-pak C18 and Sephadex LH-20 (r = 0.95; 79 +/- 46 vs 68 +/- 41 nmol/l). Sera from 5 patients (4 with osteomalacia, 1 with chronic pancreatitis/malabsorption) had vitamin D metabolite levels in the normal range after ethanol extraction but had low levels after Sep-pak C18 chromatography; four of these sera also had low levels after chromatography on Sephadex LH-20. These findings indicate that chromatography of serum on Sep-pak C18 cartridges corrects falsely elevated vitamin D metabolite levels measured by protein binding assay.

Adult↗
PubMed1988

The metabolic clearance rate of osteocalcin in sheep.

Plasma osteocalcin has been proposed as a useful and convenient biochemical marker of bone formation. However, the effect on plasma osteocalcin due to variations in the rate of its removal from the circulation has been little investigated. We have measured the metabolic clearance rate of plasma osteocalcin in adult oophorectomized sheep. Two methods were used: intravenous bolus injection (six animals) and 6 hour constant intravenous infusion (four animals) of 125I-ovine osteocalcin. Using the bolus injection method, the plasma clearance of osteocalcin was found to be 3.3 liters/h. With the constant infusion method, the calculated value was 2.8 liters/h; based on this value and the mean ovine plasma osteocalcin concentration of 26.9 ng/ml (N = 29), a plasma production rate of 1.8 mg/day was derived. Osteocalcin clearance was relatively constant among animals in a basal state. Hence, the present approach should permit us to evaluate the relative contributions to changes in circulating osteocalcin levels from altered osteocalcin plasma clearance and production under various physiological and pathological conditions.

Animals↗
PubMed1988

Vitamin D-induction of secretory responses in rat pituitary tumour (GH4C1) cells.

1,25-Dihydroxyvitamin D3(1,25-(OH)2D3) selectively enhances prolactin gene expression in GH4C1 clonal rat pituitary tumour cells. Because this effect requires extracellular Ca2+, we studied the effect of 1,25-(OH)2D3 on another Ca2+-dependent process, agonist-induced hormone secretion. Pretreatment with 1,25-(OH)2D3 (1 nmol/l) caused at least 25-fold sensitization of GH4C1 cells to the voltage-sensitive Ca2+ channel agonist BAY K 8644 (methyl-1,4-dihydro-2,6-dimethyl-3-nitro-4-(2-trifluoromethylphenyl)-pyr idine -5-carboxylate) as a prolactin secretagogue. This inductive effect of 1,25-(OH)2D3 followed a similar time-course to the enhancement of prolactin production. 1,25-(OH)2D3 had no effect on basal or BAY K 8644-induced 45Ca2+ uptake. The Ca2+-selective divalent cation ionophore 11,19,21-trihydroxy-4,6,8,12,14,18,20- heptamethyl-9-oxo-22-(tetrahydro-5 methyl-5-tetra hydro-5-(1-hydroxyethyl)-5-methyl-2-furanyl)-10,16-docosadienoic acid (ionomycin; 12 nmol/l-1.2 mumol/l) caused no significant increase in prolactin secretion in the absence of 1,25-(OH)2D3, but in cells treated with 1,25-(OH)2D3-(1 nmol/l), it increased prolactin secretion by 73% at 12 nmol/l and by a maximum of 98% at 0.12 mumol/l. These data demonstrate that vitamin D markedly enhances the responsiveness of GH4C1 functional pituitary tumour cells to two secretagogues which acts primarily through Ca2+-dependent mechanisms. They support the proposal that 1,25-(OH)2D3 acts in this cultured cell model either by effecting a redistribution of intracellular Ca2+ or by increasing the response of a Ca2+-sensitive effector system, but not by enhancing agonist-induced Ca2+ uptake.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1987

Steroid inhibition of calcitriol-induced prolactin production in GH4C1 cells. Specificity and sensitivity.

The induction of prolactin (PRL)-gene expression by calcitriol (1,25-dihydroxyvitamin D3, 1,25-dihydroxycholecalciferol) in clonal rat pituitary tumour (GH4C1) cells was selectively inhibited by cortisol [IC50 (concentration causing 50% inhibition) = 3.2-4.1 nM]. The steroid specificity of this effect was investigated and various steroids were found to inhibit calcitriol-stimulated PRL production with the following relative potencies: cortisol, 1; dexamethasone, 8; 11-deoxycortisol, 0.5; corticosterone, 0.4; aldosterone, 0.07; testosterone and oestradiol, less than 0.003. The steroid antagonist RU 38486 did not affect basal or calcitriol-stimulated PRL production, but antagonized the effect of 10 nM-cortisol in a concentration-dependent manner. Neither progesterone nor 11-deoxycortisol antagonized the effect of 10 nM-cortisol. Calcitriol-induced PRL production was 14 times more sensitive to dexamethasone inhibition than was non-stimulated PRL production. Growth-hormone production was stimulated by dexamethasone, in the presence or absence of calcitriol, with a concentration-dependence similar to that of dexamethasone inhibition of basal PRL production. These data indicate that steroid inhibition of calcitriol-stimulated PRL production is a specific glucocorticoid effect. The sensitivity of calcitriol-stimulated PRL production to dexamethasone was 14-26-fold greater than that of other measured responses in the same cells. Two of the possible explanations for this selectively increased sensitivity to glucocorticoids are: amplification of the glucocorticoid effect via an induced mediator; and the presence of very-high-affinity glucocorticoid-receptor-binding sites on DNA.

Aldosterone↗
PubMed1987

Hypokalemic periodic paralysis of thyrotoxic origin.

A 50-year-old Vietnamese man suffered recurrent episodes of hypokalemic periodic paralysis during treatment for thyrotoxicosis. Suspected precipitants of the paralysis were oral prednisolone, strenuous exertion and poor compliance with medications. Propranolol prevented the periodic paralysis. Control of the thyrotoxicosis was complicated by a florid skin reaction attributed to carbimazole, and was ultimately achieved using radioiodine.

Australia↗
PubMed1987

1,25-Dihydroxyvitamin D3 enhances thyrotropin releasing hormone induced thyrotropin secretion in normal pituitary cells.

The findings of specific binding of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] in normal rat pituitary tissue and selective effects of 1,25-(OH)2D3 on gene expression in clonal pituitary tumour cells have suggested that vitamin D may regulate pituitary function. Therefore, the in vitro effect of 1,25-(OH)2D3 on normal pituitary cells was investigated. Primary anterior pituitary cell cultures prepared from female rats were maintained in experimental medium +/- 10(-8) M 1,25-(OH)2D3 for up to 24 h and then incubated with fresh experimental medium containing TRH (10(-10)-10(-8) M) or vehicle for 1 h. Pretreatment with 1,25-(OH)2D3 for 24 h led to increased TSH release at all TRH concentrations tested (P less than 0.0001), a decrease in the half-maximal stimulatory dose of TRH for TSH release from 2 X 10(-9) M to 0.4 X 10(-9) M, a 22% increase in maximal TSH release (P less than 0.01), and an 81% increase in TSH release at 10(-9) M TRH (P less than 0.001). 1 X 10(-9) M 1,25-(OH)2D3 increased TRH (10(-9) M)-induced TSH release by 20% (P less than 0.05) but 10(-7) M and 10(-6) M 25-hydroxyvitamin D3 (25-OH D3) had no effect. The effect of 1,25-(OH)2D3 on TRH (10(-9) M)-induced TSH release was evident within 8 h and was maximal by 16 h. There was no effect on basal TSH release, TSH accumulation in the medium in the preceding 24 h nor on cell-associated TSH. 1,25-(OH)2D3 pretreatment had no effect on TRH-induced PRL secretion, PRL accumulation in the medium nor on cell-associated PRL. We have shown that 1,25-(OH)2D3 acts selectively on the thyrotroph to enhance in vitro responsiveness to physiologically relevant concentrations of TRH. These findings are consistent with the reported autoradiographic localization of [3H]-1,25-(OH)2D3 in the thyrotroph and support a permissive or regulatory role of vitamin D in the normal pituitary gland.

Animals↗
PubMed1986

Glucocorticoids antagonize induction of prolactin-gene expression by calcitriol in rat pituitary tumour cells.

Clonal strains of rat pituitary tumour (GH4C1) cells are known to possess specific intracellular binding sites for calcitriol (1,25-dihydroxycholecalciferol, 1,25-dihydroxyvitamin D3). GH4C1 cells respond to calcitriol by a selective increase in prolactin(PRL)-gene expression. The interaction between calcitriol and glucocorticoids was studied by using this cultured-cell model. It was found that cortisol potently antagonized the induction of PRL mRNA and PRL production by calcitriol. The effects were concentration-dependent and were evident at glucocorticoid concentrations that did not alter basal PRL production. Inhibition was half-maximal at 3.2 nM-cortisol and 0.4 nM-dexamethasone. Calcitriol-induced PRL mRNA fell by more than 50% at 25 h and reached the control level 50 h after treatment with cortisol. The inhibition by cortisol of calcitriol induction of PRL production was selective when compared with effects on other inducers of PRL-gene expression [thyroliberin, epidermal growth factor and phorbol myristate acetate ('12-omicron-tetradecanoylphorbol 13-acetate')]. Potent antagonism by glucocorticoids of vitamin D action on specific gene expression has been demonstrated. Further studies with this cultured-cell model may help to explain the mechanism of this hormonal interaction, which assumes particular importance at major sites of vitamin D action such as the intestine.

Animals↗
PubMed1984

Isolation, purification and cell-free synthesis of calmodulin from the pig anterior pituitary gland.

Calmodulin was extracted and purified from pig anterior pituitary gland. The protein was characterized by its migration on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis in the presence of Ca2+ or EGTA, its U.V. spectrum between 240 and 290 nm and the activation of calmodulin-deficient cyclic AMP phosphodiesterase. The yield was 370 mg/kg wet wt. mRNA was also extracted from the same tissue and translated in a wheat-germ cell-free translation system. Translated calmodulin was identified by its heat-stability, its co-migration with authentic anterior-pituitary calmodulin on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, its acidic isoelectric point (4.15) on flat-bed isoelectric focusing, its Ca2+-dependent binding to fluphenazine-Sepharose 6B, and its co-elution from this gel with authentic unlabelled calmodulin with EGTA buffer. Calmodulin was not translated as a precursor form. In this tissue it was calculated that calmodulin accounted for 0.5-1% of the total translated protein.

Animals↗
PubMed1984

Biphasic action of prostaglandin E2 on conversion of 25-hydroxyvitamin D3 to 1,25-dihydroxyvitamin D3 in chick renal tubules.

The effect of PGE2 on the conversion of 25-hydroxyvitamin D3 (25 OH D3) to 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) by isolated renal tubules from vitamin D deficient chicks was studied under a variety of experimental conditions. In the absence of added vitamin D metabolites, PGE2 (2 x 10(-6)M) caused an immediate inhibition of formation of 1,25-(OH)2D3, followed by a delayed stimulation, apparent after 15 h exposure to PGE2. Pretreatment of the tubules with 1,25-(OH)-2D3 prevented the immediate inhibitory action of PGE2, and allowed the stimulation to be apparent after 4 h exposure to PGE2. The cyclic nucleotide phosphodiesterase inhibitor 3-isobutyl-1-methyl xanthine (IBMX) significantly stimulated the formation of 1,25-(OH)-2D3. PGE2 significantly inhibited 1,25-(OH)2D3 formation in tubules which had been stimulated by IBMX. PGE2 stimulated the adenylate cyclase activity in a crude particulate fraction from the chick kidney, and raised cyclic adenosine 3', 5'-monophosphate (cyclic AMP) levels in the renal tubules. It is concluded that PGE2 can either stimulate or inhibit 1,25-(OH)2D3 formation in chick renal tubules. The stimulatory effect may be partly due to elevation of cyclic AMP. The mechanism of the inhibitory effect requires further investigation.

Adenylyl Cyclases↗
PubMed1983

Regulation of prolactin mRNA by 1,25-dihydroxyvitamin D3 in GH4C1 cells.

GH4C1 cells, which possess specific cytosolic binding for 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3), have previously been found to respond to this hormone with a highly selective increase in prolactin synthesis. Here we extend our previous observations by measuring the effect of 1,25-(OH)2D3 on the accumulation of specific mRNA in GH4C1 cells, using cytoplasmic dot hybridization. In a hormone-free, chemically defined medium containing 0.4 mM Ca2+, stimulation of prolactin mRNA accumulation was 200-215% of control after 96 h of treatment with 1 X 10(-9) M 1,25-(OH)2D3. Thyrotropin-releasing hormone (1 X 10(-7) M) caused a similar increase in prolactin mRNA (206% of control). Neither hormone increased growth hormone mRNA. Stimulation of prolactin mRNA by 1,25-(OH)2D3 was 163% of control after 48 h and 200% after 96 h. No consistent effect was observed in the first 24 h of treatment. The same response to 48 h treatment with 1,25-(OH)2D3 was observed whether the cells were withdrawn from serum-containing medium for 24 h (as above) or 72 h. In cells withdrawn for 96 h, 1,25-(OH)2D3 raised prolactin mRNA within 24 h. The effect of 1,25-(OH)2D3 was significant at a concentration of 10(-11) M, and half-maximal at approximately 10(-10) M. In low-Ca2+ medium, the response to greater than 10(-10) M 1,25-(OH)2D3 was eliminated. Extracellular Ca2+ (0.4 mM) increased prolactin mRNA in the presence of, but not in the absence of, 1,25-(OH)2D3. These findings demonstrate that 1,25-(OH)2D3 enhanced prolactin gene expression with a distinctive time course and interactively with calcium.

Animals↗
PubMed1983

Opposing effects of cyclic adenosine-3',5'-monophosphate and cyclic guanosine-3',5'-monophosphate on the metabolism of 24-hydroxyvitamin D3 in isolated chick renal tubules.

The conversion of 25-hydroxyvitamin D3 (25 OH D3) to 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3), 24,25-dihydroxyvitamin D3 (24,25-(OH)2D3) and 1,24,25-trihydroxyvitamin D3 (1.24,25-(OH)3D3) was studied in renal tubules prepared from chicks raised on a vitamin D deficient diet with or without vitamin D supplementation. As described previously, in tubules from vitamin D deficient chicks, cyclic AMP caused an increase in the net accumulation of 1,25-(OH)2D3, the major metabolite formed under these circumstances. This stimulation was shown to be due to an increased maximum velocity of the hydroxylation reaction. There was also a significant inhibition of the net accumulation of 24,25-(OH)2D3. Cyclic GMP caused a significant inhibition of 1,25-(OH)2D3 formation and stimulation of the net accumulation of 24,25-(OH)2D3. In chicks supplemented with high doses of vitamin D, 24,25-(OH)2D3 was the major metabolite of 25 OH D3 detected and 1-hydroxylase activity was negligible. Under these circumstances, neither cyclic AMP nor cyclic GMP affected net accumulation of 24,25(OH)2D3. This suggested that the apparent effect of the nucleotides on formation of 24,25-(OH)2D3 may have been due to further metabolism of 24,25-(OH)2D3 when 1-hydroxylase activity was high. It is concluded that cyclic AMp and cyclic GMP have reciprocal effects on renal 25 OH D3-1-hydroxylase activity, and both should be considered potential intracellular regulators of 25 OH D3 metabolism.

24,25-Dihydroxyvitamin D 3↗
PubMed1983

A test of 1,25-dihydroxyvitamin D3 secretory capacity in normal subjects for application in metabolic bone diseases.

The circulating concentration of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] varies in man in response to a variety of physiological stimuli. In pathological states, random plasma 1,25-(OH)2D3 levels reflect the sum of the underlying pathological process and the homeostatic corrections to those perturbations. Hence random 1,25-(OH)2D3 levels do not allow differentiation as to whether changes in 1,25-(OH)2D3 production or metabolism are primary or secondary to other changes. Similarly random levels do not provide much insight into the reserve of 1,25-(OH)2D3 secretory capacity. We have developed a single infusion of parathyroid extract (PTE) as a test of 1,25-(OH)2D3 secretory capacity. Normal responses in twelve males and six females consisted of an increase in plasma 1,25-(OH)2D3 levels of 78% over basal in the male and 77% over basal in the female subjects. The peak response occurred 18-22 h after the PTE was administered. This test of 1,25-(OH)2D3 secretory capacity should prove useful in the evaluation of patients with metabolic bone disease. As part of this study, the diurnal plasma 1,25-(OH)2D3 level was examined in six males and seven females. There was no significant diurnal rhythm between 8.00 and 22.00 h.

Adult↗
PubMed1982

Vitamin D stimulates prolactin synthesis by GH4C1 cells incubated in chemically defined medium.

GH4C1 cells were evaluated as a model system for the investigation of the cellular and molecular mode of action of vitamin D. When cells were incubated in hormone-free, chemically defined medium, the metabolite 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) potently stimulated PRL synthesis (ED50 = 2 x 10(-10) M) with minimal effects on GH synthesis. The stimulation of PRL synthesis was vitamin D metabolite specific. The effect of 1,25-(OH)2D3 was dependent on the addition of extracellular calcium in the concentration range up to 0.1 mM. These results indicate the potential usefulness of this experimental system for the study of vitamin D action.

Animals↗
PubMed1981

Regulation of 25-hydroxy-vitamin D-1 alpha-hydroxylase in chick isolated renal tubules: effects of prostaglandin E2, frusemide and acetylsalicylic acid.

1. Isolated renal tubules were prepared from vitamin D-deficient chicks. The effects of added prostaglandin E2 (PGE2) and agents which modify prostaglandin metabolism on the metabolism of 25-hydroxy-vitamin D3 were studied. 2. Frusemide (0.1 mmol/l) raised the prostaglandin E (PGE) content of the tubule incubation medium; it significantly increased 1,25-dihydroxy-vitamin D3 [1,25-(OH)2D3] production from 25-hydroxy-vitamin D3 [25-(OH)D3] and significantly inhibited the net production of 24,25-dihydroxy-vitamin D3 [24,25-(OH)2D3]. 3. Acetylsalicylic acid (aspirin; 0.1 mmol/l) decreased the PGE content of the tubule incubation medium and significantly inhibited 1,25-(OH)2D3 production. 4. In the presence of 1,25-(OH)2D3, 4 h exposure of tubules from vitamin D-deficient chicks to concentrations of PGE2 between 2.8 X 10(-6) and 2.8 X 10(-8) mol/l significantly enhanced 1,25-(OH)2D3 production. 5. It is concluded that, in view of the effects of modulation of endogenous prostaglandin levels by frusemide in aspirin, and the stimulatory effect of exogenous PGE2, prostaglandins should be considered potential regulators of the renal 25-hydroxy-vitamin D-1 alpha-hydroxylase [25-(OH)D-1 alpha-hydroxylase] enzyme.

25-Hydroxyvitamin D3 1-alpha-Hydroxylase↗
PubMed1979

Modulation of plasma 1,25-dihydroxyvitamin D in man by stimulation and suppression tests.

In normal volunteers infusion of parathyroid extract raised plasma 1,25-dihydroxyvitamin D (1,25-[OH]2D) levels by 80% (peak 8--40 h after infusion) without producing any significant change in plasma calcium, phosphate, or magnesium. An oral calcium load suppressed plasma 1,25-(OH)2D to 63% of control values within 48--72 h. These responses will be useful in the diagnosis and management of disturbances of calcium and mineral metabolism in which vitamin D metabolism is abnormal.

Administration, Oral↗
PubMed1979

Chronic diphenylhydantoin therapy does not reduce plasma 25-hydroxy-vitamin D.

A randomized controlled study was performed to investigate the effect of 2 years' monitored diphenylhydantoin (DPH) therapy on plasma 25-hydroxyvitamin D (25-OHD) in non-epileptic, non-institutionalized subjects. Mean +/- SEM plasma 25-OHD of 18 DPH-treated subjects at the end of 2 years' drug treatment was 59 +/- 8 nmol/l (23.6 +/- 3.2 ng/ml), which was not decreased compared to that of eighteen control subjects (54 +/- 8 nmol/l, 21.6 +/- 3.2 ng/ml). In addition, mean plasma 25-OHD had not changed 1 month after ceasing DPH. The treated group had a higher mean serum alkaline phosphatase (SAP) during DPH treatment, attributable to hepatic enzyme induction. It is concluded that therapeutic doses of DPH without other anticonvulsants do not have a clinically significant effect on plasma 25-OHD.

Adult↗