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Biomedical subjects

D Feldman

Publications and source records attributed to D Feldman.

At least 217 records · Page 12Linked to original sources
PubMed1983

Ketoconazole blocks adrenal steroidogenesis by inhibiting cytochrome P450-dependent enzymes.

Ketoconazole has recently been shown to interfere with steroidogenesis in patients and rat in vitro systems. In this study we attempted to elucidate the site of inhibition in the adrenal gland. Although ketoconazole impaired adrenocorticotropic hormone stimulated cyclic (c)AMP production, dibutyrl cAMP addition did not bypass the steroidogenic blockade indicating that the critical ketoconazole-inhibited step was distal to cAMP. Addition of radiolabeled substrates to isolated adrenal cells and analysis of products by high performance liquid chromatography demonstrated a ketoconazole block between deoxycorticosterone (DOC) and corticosterone. This 11-hydroxylase step is carried out by a P450-dependent mitochondrial enzyme. No restriction of progesterone or pregnenolone conversion to DOC was detected, steps carried out by non-P450-dependent microsomal enzymes. Inhibition of cholesterol conversion to pregnenolone by mitochondrial fractions indicated a second block at the side chain cleavage step, another mitochondrial P450-dependent enzyme. Adrenal malate dehydrogenase, a non-P450-dependent mitochondrial enzyme was not inhibited while renal 24-hydroxylase, a P450-dependent mitochondrial enzyme in another organ, was blocked by ketoconazole. We conclude that ketoconazole may be a general inhibitor of mitochondrial P450 enzymes. This finding suggests that patients receiving ketoconazole be monitored for side effects relevant to P450 enzyme inhibition. Further, we raise the possibility that this drug action may be beneficially exploited in situations where inhibition of steroidogenesis is a therapeutic goal.

Adrenal Cortex Hormones↗
PubMed1983

Ketoconazole binds to glucocorticoid receptors and exhibits glucocorticoid antagonist activity in cultured cells.

We have recently found that ketoconazole inhibits adrenal steroidogenesis; in this paper we investigated whether imidazole antimycotic drugs additionally interact with glucocorticoid receptor sites in target tissues. Our approach was to assess the ability of three drugs: ketoconazole, clotrimazole, and RS 49910, to inhibit [3H]dexamethasone binding to hepatoma tissue culture (HTC) cell cytosol. The results indicated dose-dependent, competitive displacement of [3H]dexamethasone binding that was in the potency sequence: clotrimazole greater than ketoconazole greater than RS 49910. We then examined the functional response of this binding by measuring tyrosine aminotransferase (TAT) activity in HTC cells. The antimycotics did not exhibit TAT agonist activity and inhibition of basal enzyme levels was not detected. However, the drugs were potent antagonists of dexamethasone-induced TAT activity and the effect was temporally reversible. This antagonist activity was in the same sequence and closely correlated with the binding potency of the three drugs. We conclude that ketoconazole and other imidazole antimycotic drugs possess glucocorticoid antagonist activity by virtue of occupancy of glucocorticoid receptor sites in target tissues.

Animals↗
PubMed1983

Glucocorticoid modulation of cell proliferation in cultured osteoblast-like bone cells: differences between rat and mouse.

Glucocorticoids are important regulators of cell proliferation, but divergent effects have been noted in various systems. In this study, the actions of glucocorticoids on cell proliferation were examined in primary cultures of osteoblast-like cells from perinatal rodents. The aim of the study was to determine whether rat and mouse bone cells exhibit species differences in their responses to glucocorticoids and whether the response of either species was influenced by the cell density and time of treatment. Cultured osteoblast-like cells were treated with dexamethasone or ethanol vehicle (less than 0.05%). Changes in the proliferation rate were assessed by measurement of cell number, DNA content, and thymidine incorporation rate. We found that the response to dexamethasone was dependent on the cell density at the time of treatment in rat cultures: inhibitory in sparse and stimulatory in dense cultures. In contrast, dexamethasone inhibited mouse cell proliferation at all cell densities. The stimulatory effects in rat cells were seen after 4 days of treatment but not after 2 days. Both the stimulatory and inhibitory effects in rat and mouse cells occurred between dexamethasone concentrations of 1.3-13 nM, with half-maximal effects seen at 6 nM. The divergent responses in sparse and dense rat cultures were found not to result from the selective attachment of cell subpopulations at different plating densities. Epidermal growth factor was mitogenic in both species. The effects of dexamethasone were not modified by the presence of epidermal growth factor in either low (1%) or high (10%) serum concentrations. In conclusion, these findings reemphasize the importance of species differences and the difficulty in comparing one system to another, even with closely related species such as mouse and rat. Furthermore, the data show that the culture conditions at the time of treatment dramatically influence the responses of these cells to glucocorticoids. These factors must be considered in studies of the hormonal regulation of bone cell proliferation.

Animals↗
PubMed1983

Cutaneous vasculitis in adult polymyositis/dermatomyositis.

Seven (9.2%) of 76 patients with adult-onset polymyositis/dermatomyositis (PM/DM) seen over an 11-year period had cutaneous vasculitis. This was manifest by dermal and/or subcutaneous nodules in 4, periungual infarcts in 3 and digital ulceration in 2. When these 7 patients were compared to the remaining 69, a significant association was noted between cutaneous vasculitis and DM (p = .025); only 1 of 31 patients with primary PM and none of 18 with overlap syndromes had vasculitis. Furthermore, 2 (28.6%) of those with vasculitis had an associated malignancy compared to only 4 (5.8%) of those without vasculitis. These data document the occurrence of cutaneous vasculitis in adult-onset PM/DM and suggest that its presence may be a marker of an underlying malignancy.

Adult↗
PubMed1982

An estrogen-binding protein and endogenous ligand in Saccharomyces cerevisiae: possible hormone receptor system.

A protein macromolecule in the cytosol of the unicellular eukaryotic yeast Saccharomyces cerevisiae selectively binds the vertebrate estrogen hormone 17 beta-estradiol with high affinity. Lipid extracts of the yeast cells or the conditioned growth medium yield a substance that can bind competitively to the tritiated estradiol-binding sites in the yeast and to mammalian estrogen receptors. These findings suggest that the binding protein may be a primitive hormone receptor and that the lipid-extractable substance represents the endogenous ligand.

Animals↗
PubMed1982

Characterization of a unique corticosterone-binding protein in Candida albicans.

This paper further characterizes a protein we have demonstrated in Candida albicans which has the ability to bind corticosterone and related steroid hormones. Fungal cells are disrupted and cytosol is incubated with [3H]corticosterone for 3 h at which time peak steady state binding is achieved. Bound hormone is separated from free using Sephadex G-50 minicolumns or dextran-coated charcoal. Binding was found to be a linear function of protein concentration. The bound hormone co-migrates with authentic corticosterone in thin layer chromatographic systems indicating no metabolism of the radioprobe. Scatchard analysis of the binding in the pseudohyphal form of C. albicans yielded values of 6.3 nM for the Kd and a binding capacity of about 650 fmol/mg of cytosol protein; both determinations are comparable to our findings in the yeast form of this organism. A series of sterols were tested for their ability to displace [3H]corticosterone from the yeast binder, and the results show that the binder is remarkably selective and stereo specific. Physical-chemical studies show the binder to be degraded at high temperatures and that binding is destroyed by trypsin and sulfhydryl blockers. The protein sediments at 4 S on sucrose gradients and does not exhibit ionic dependent aggregation. The molecular weight is estimated to be approximately 43,000 daltons by gel chromatography. We hypothesize that this intracellular protein may represent a primitive form of either the mammalian glucocorticoid receptor or the plasma corticosteroid-binding globulin.

Binding, Competitive↗
PubMed1982

1,25-Dihydroxyvitamin D3 receptors and functions in cultured pig kidney cells (LLC PK1). Regulation of 24,25-dihydroxyvitamin D3 production.

Although 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) regulates the renal metabolism of 25-hydroxyvitamin D3 (25-OH-D3), the mechanism is not well understood. The established pig kidney cell line, LLC PK1, was used to study this feedback regulation. These cells possess a receptor for 1,25-(OH)2D3 with a sedimentation coefficient of 3.2 S. Scatchard analysis of 1,25-(OH)2D3 binding to cell cytosol yielded an apparent Kd of 0.12 nM and an Nmax of 26 fmol/mg of cytosol protein. LLC PK1 cells respond to 1,25-(OH)2D3 by changes in the metabolism of 25-OH-D3. When incubated with 25-OH-[3H]D3, homogenates of untreated cells did not produce detectable 1,25-(OH)2[3H]D3 or 24,25-(OH)2[3H]D3. However, after treatment of cell monolayers with 1,25-(OH)2D3 for 8 h, homogenates converted substantial 25-OH-[3H]D2 substrate to 24,25-(OH)2[3H]D3. The appearance of this 24-hydroxylase activity in response to 1,25-(OH)2D3 was time- and dose-dependent. Half-maximal levels of enzyme activity were achieved with 0.13 nM 1,25-(OH)2D3, a concentration almost identical to the Kd of the 1,25-(OH)2D3 receptor. The stimulation of 24-hydroxylase activity was shown to be an induction event; treatment of monolayers with 13 nM 1,25-(OH)2D3 for a 4-h pulse was sufficient to induce maximal activity assayed at h. The presence of the transcriptional inhibitor, actinomycin D, during the 4-h pulse abolished the induction of 24-hydroxylase activity. These results demonstrate for the first time the presence of both 1,25-(OH)2D3 receptors and stimulation of 24-hydroxylase activity within the same established mammalian kidney cell line.

Animals↗
PubMed1982

Glucocorticoid regulation of 1,25(OH)2vitamin D3 receptors: divergent effects on mouse and rat intestine.

We have investigated the possibility that glucocorticoids alter responsiveness to vitamin D by regulating the 1,25(OH)2D3 receptor in rat intestine. In contrast to the mouse where glucocorticoids caused receptors to decline, rats treated with glucocorticoids showed a substantial increase ( approximately 50%) in the number of intestinal 1,25(OH)2D3 receptors. This resulted from an increase in receptor content with no change in affinity for 1,25(OH)2D3. Receptor stimulation was even greater in vitamin D-deplete rats. Moreover, adrenalectomy led to a significant decline in receptor number. Although the properties of the receptor are similar in rat and mouse intestine, the divergent response to glucocorticoids emphasizes major differences between species in the regulation of 1,25(OH)2D3 receptor number.

Adrenalectomy↗
PubMed1982

Vitamin D resistant rickets with alopecia: cultured skin fibroblasts exhibit defective cytoplasmic receptors and unresponsiveness to 1,25(OH)2D3.

A new case of vitamin D dependent rickets (Type II) with alopecia in a 5 yr old child is reported. Skin fibroblasts were propagated in culture and analyzed for cytoplasmic 1,25(OH)2D3 receptors. The rachitic cells failed to exhibit specific, high-affinity binding sites by either Scatchard analysis or sucrose density gradient. Furthermore, a rise in 24-hydroxylase activity could not be elicited following incubation of the rachitig cells with 1,25(OH)2D3. Fibroblasts from a non-rachitic child examined in parallel experiments demonstrated high affinity binding sites (Kd = 0.1 nM, Nmax = 33 fmol/100 micrograms DNA) and the induction of 24-hydroxylase activity. The molecular basis of the unresponsiveness of the cells from the rachitic child appears to be due to defective or absent 1,25(OH)2D3 receptors.

Alopecia↗
PubMed1982

Ketoconazole blocks adrenal steroid synthesis.

Ketoconazole, a broad-spectrum, antifungal drug that is administered orally, has been shown to inhibit sterol synthesis in fungi. When gynecomastia developed in some patients taking this drug, we investigated the effects of ketoconazole on steroid synthesis in humans and in isolated adrenal cells from rats. In healthy humans, the cortisol response to adrenocorticotropic hormone was significantly blunted 4 hours after a 400-mg or 600-mg dose. The inhibition persisted for up to 8 hours and was absent by 16 hours. This finding indicated that adrenal androgen response was reduced. Easily achieved therapeutic concentrations of ketoconazole virtually eliminated corticosterone production by isolated adrenal cells from rats. Although ketoconazole at currently used doses has never been documented to cause clinical hypoadrenalism, caution is urged in high- or multiple-dose trials. The drug may prove useful as an agent to block steroid synthesis.

Adrenal Cortex↗
PubMed1982

1,25-dihydroxyvitamin D3 receptors in human epithelial cancer cell lines.

Specific, high-affinity cytosolic receptors for 1,25-dihydroxyvitamin D3 have been demonstrated in five human cancer cell lines. The cell lines were derived from tumors of breast, lung, cervix, and melanotic and amelanotic melanomas. Binding affinity (Kd) of the receptors for 1,25-dihydroxyvitamin D3 were all approximately 0.2 nM, and receptor content ranged from 21 to 174 fmol/mg cytosol protein. The receptors from all five cell lines sedimented at 3.2S on sucrose density gradients and exhibited preferential affinity for 1,25-dihydroxyvitamin D3 compared to other vitamin D metabolites.

Breast Neoplasms↗
PubMed1981

Effects of two albumins and two detergents on the activity of bovine milk lipoprotein lipase against very low density and high density lipoprotein lipids.

In this study we have determined the effects of two commercial albumin preparations (Sigma and Pentex albumins) and two detergents (sodium deoxycholate and Triton X-100) on the activity of lipoprotein lipase purified from bovine milk against biosynthetically labeled triacylglycerol in very low density lipoprotein and biosynthetically labeled phosphatidylcholine in very low density and high density lipoproteins. Pentex albumin decreased the activity of lipoprotein lipase in all assays to about one-fourth to one-third of that observed with Sigma albumin. Quantitative differences were observed in the distribution of labeled surface constituents (32P-labeled phospholipids, [3H]cholesterol and 125I-labeled apolipoprotein C) among density fractions during lipolysis of very low density lipoprotein carried out in the presence of Pentex or Sigma albumins. With Pentex albumin, more phospholipids and apolipoprotein C distributed to the density fraction of d 1.04-1.21 g/ml than with Sigma albumin. Sodium deoxycholate at a concentration of up to 2 mM had little effect in all assays. Triton X-100 decreased the activity of lipoprotein lipase against very low density lipoprotein lipids but increased the activity of the enzyme against high density lipoprotein lipids. The study has thus demonstrated marked quantitative differences of lipoprotein lipase activities when determined under slightly differing incubation conditions.

Animals↗
PubMed1981

1,25-dihydroxyvitamin D3 and malignant melanoma: the presence of receptors and inhibition of cell growth in culture.

In this study we demonstrate the presence of specific, high-affinity receptors for 1,25-dihydroxyvitamin D3 in malignant melanoma. Receptors are present both in cultured melanoma cells and in melanoma tumor tissue produced by inoculation of cells into athymic rats. The receptor sediments at 3.25 on sucrose density gradients, possesses a preferential affinity for 1,25-(OH)2D3 and has an apparent Kd of 0.18 nM by Scatchard analysis. We also demonstrate that human melanoma cells are responsive to 1,25-(OH)2D3 in vitro. Inclusion of 1,25-(OH)2D3 in the culture medium produced a marked increase in cell doubling time. This inhibitory effect of the hormone on melanoma cell proliferation was dose-related and represents the first demonstration of a 1,25-(OH)2D3 mediated action on tumor cells.

Calcitriol↗