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Biomedical subjects

E R Simpson

Publications and source records attributed to E R Simpson.

At least 253 records · Page 14Linked to original sources
PubMed1983

Aromatase activity of membrane fractions of human adipose tissue stromal cells and adipocytes.

Adipose tissue is the principal site of extraglandular estrogen formation in nonpregnant women. The importance of adipose tissue as a site of estrogen formation is emphasized by the finding that increased body weight is associated with an increased incidence of endometrial carcinoma. In the present study, the kinetics of estrogen formation from androstenedione by adipocytes and by stromal cells isolated from human adipose tissue as well as by membrane fractions prepared from these cells were investigated. Subcutaneous adipose tissue samples obtained from women were dispersed by collagenase treatment, and aromatase activity was assayed by the incorporation of tritium from [1-3H]androstenedione into [3H]water. As previously reported, aromatase activity was found in intact stromal cells of adipose tissue, whereas little aromatase activity was detected in intact adipocytes. When crude membrane fractions (100,000 X g pellet) of stromal cells and adipocytes were incubated in the presence of [1-3H]androstenedione and an NADPH-generating system, however, aromatase activity was found in membrane fractions of both stromal cells and adipocytes, and estrogen formation increased in a linear manner as a function of time and membrane protein concentration. The apparent Michaelis constant (Km) of aromatase for androstenedione of intact stromal cells was 0.03 microM, whereas intact adipocytes did not convert androstenedione to estrone at substrate concentrations up to 3.0 microM. In membrane fractions of stromal cells, the rate of aromatization as a function of androstenedione concentration did not follow simple Michaelis-Menten kinetics, and both low affinity (Km = 1.03 microM) and high affinity (Km = 0.10 microM) components were observed. The affinity of androstenedione for aromatase of adipocyte membrane fractions was low; the rate of aromatization was not saturable at concentrations of androstenedione up to 3.0 microM. When intact adipocytes were incubated with [1-3H]androstenedione, then homogenized, and the homogenate was treated by differential centrifugation, the radioactivity that was added to the medium was found almost entirely in the lipid fraction of the cells. This finding is indicative that the low aromatase activity in intact adipocytes is the result of sequestration of steroid in lipid droplets in the cells. We suggest that the stromal cells of adipose tissue are a major source of the increased estrogen production in obese persons; a role for adipocytes in the regulation of adipose tissue estrogen formation, however, cannot be excluded at this time.

Adipose Tissue↗
PubMed1983

Hypercholesterolemia due to elevated low density lipoprotein-cholesterol in newborns with anencephaly and adrenal atrophy.

In the present investigation, we evaluated the relationship between plasma lipoprotein-cholesterol and adrenal steroid production in abortuses and newborns in whom the adrenal was expected to be atrophic, i.e. in anencephalics. We found that umbilical cord plasma levels of dehydroepiandrosterone sulfate (DS) in 23 anencephalics delivered between 13.5 and 45.5 weeks of gestation (mean +/- SE, 176 +/- 37 ng/ml) were significantly lower than those in normal newborns of similar gestational ages; the umbilical cord plasma concentrations of cortisol in many anencephalics, however, were within normal limits. The levels of total cholesterol (134 +/- 10 mg/dl) and low density lipoprotein (LDL)-cholesterol (94 +/- 8 mg/dl) were substantially higher (up to 4-fold) in umbilical cord plasma of anencephalics than in umbilical cord plasma of normal newborns. The mean level of high density lipoprotein-cholesterol in umbilical cord plasma of anencephalic abortuses and newborns (38 +/- 4 mg/dl) was approximately 50% higher than that in normal newborns. The lowest plasma cholesterol level (56 mg/dl) and a concentration of DS (480 ng/ml) that was among the highest seen in the group of anencephalics were found in an anencephalic newborn in whom adrenals were of near-normal weight. Plasma cholesterol levels were inversely correlated to adrenal weights and plasma DS levels, and plasma DS levels were correlated to adrenal weight. Whereas the estimated plasma pool of DS in normal newborns increased to over 300 micrograms during the latter part of gestation, that of anencephalic newborns was much lower (less than 1 to 26 micrograms) and did not appear to increase as a function of gestational age. Conversely, the estimated plasma pool of cholesterol in normal newborns appeared to decline slightly during the last 10 weeks of gestation (80 mg at term), whereas that of anencephalic newborns expanded greatly near term; levels (approximately 200 mg) were attained that were about 3 times those in normal newborns. We conclude that the hypercholesterolemia in anencephalic newborns, due primarily to extremely elevated plasma levels of LDL-cholesterol, is a result of decreased uptake and utilization of plasma LDL-cholesterol for steroid biosynthesis by the adrenals. Since hypercholesterolemia is apparently early in gestation in anencephalic abortuses, we speculate that in normally developing fetuses, plasma LDL-cholesterol is used as substrate for adrenal steroidogenesis early in gestation as well as near term when the rates of growth and steroid production by the adrenals accelerate markedly.

Adrenal Glands↗
PubMed1983

Inhibition of prostaglandin biosynthesis in human adipose tissue by glucocorticosteroids.

Cultured stromal cells derived from human sc adipose tissue synthesized and secreted into the medium prostaglandin E2 (PGE2), PGF2 alpha, and 6-keto-PGF1 alpha, a metabolite of prostacyclin. PGE2 was quantitatively the major PG formed by these cells. Dexamethasone inhibited PG biosynthesis in a concentration- and time-dependent manner; dexamethasone (2.5 X 10(-10) M) caused greater than 50% inhibition of PGE2 synthesis after 24 h of incubation and 90% inhibition after 48 h. The effect of dexamethasone to inhibit PGE2 synthesis was blocked by simultaneous incubation of cells with cortisol-21-mesylate, an antagonist of the binding of glucocorticosteroids to cytosolic receptors. (Bu)2cAMP (1 mM), forskolin (10 microM), and 1-methyl-3-isobutylxanthine (0.1 mM) markedly stimulated PGE2 biosynthesis in cultured adipose stromal cells. The inhibitory effect of dexamethasone on PGE2 synthesis was attenuated by simultaneous incubation of cells with (Bu)2cAMP. The results of this study suggest that glucocorticosteroids inhibit PG biosynthesis in adipose tissue stromal cells, whereas cAMP and certain analogs thereof stimulate PG biosynthesis and overcome the inhibitory action of glucocorticosteroids.

6-Ketoprostaglandin F1 alpha↗
PubMed1983

Prostaglandin secretion by the neocortex and fetal zones of the human fetal adrenal gland.

Previously, we reported that the human fetal adrenal (HFA) gland secretes various prostaglandins (PGs) in vitro and that PG secretion is inhibited by endogenously synthesized glucocorticosteroids. In this investigation, the neocortex (NC) and fetal zone (FZ) of the HFA gland were separated by microdissection and maintained as tissue fragments in organ culture. The rate of PG secretion into the culture medium was determined by measuring various PGs using specific RIAs in media collected at 24-h intervals. During the first 24 h in culture, the secretion rates of PGF2 alpha and PGE2 were 6- and 7-fold greater by NC [14 +/- 5 and 9.9 +/- 3 ng mg protein-1 24 h-1 (mean +/- SE)], respectively, than by FZ tissue (2.5 and 1.4 ng mg protein-1 24 h-1). The secretion rates of PGFM and PGD2 were 2-fold greater in NC tissue than in FZ tissue, but the secretion rates of thromboxane B2 were similar in both zones of HFA tissue. In another study, the patterns of secretion of PGF2 alpha and PGE2 were determined as a function of days in culture. The secretion rates of PGF2 alpha and PGE2 fell rapidly in NC from 19.0 +/- 11 and 38.3 +/- 9.7 ng mg protein-1 24 h-1, respectively, to 1.3 +/- 7.2 and 4.8 +/- 3.3 by day 4. In contrast, the secretion rates of PGF2 alpha and PGE2 rose 8- and 3-fold in FZ tissue (from 0.7 +/- 0.2 and 0.9 +/- 0.6 ng mg protein-1 24 h-1, respectively, to 5.9 +/- 0.5 and 3.1 +/- 1.2 by day 4). The addition of ACTH or dexamethasone inhibited PG secretion in both zones, but to a greater degree in FZ tissue than in NC tissue. In summary, the NC secretes larger quantities of PG than the FZ, and the patterns of secretion are different in the two zones. The secretion of PGs is inhibited more in FZ than in NC tissue by ACTH and glucocorticosteroids.

Adrenal Glands↗
PubMed1982

Adrenodoxin biosynthesis by bovine adrenal cells in monolayer culture. Induction by adrenocorticotropin.

The long term effect of adrenocorticotropin (ACTH) on the synthesis of adrenodoxin in bovine adrenocortical cells was investigated. Primary, confluent monolayer cultures of adult bovine adrenocortical cells were incubated in the presence or absence of ACTH (10(-6) M) for periods up to 72 h. The amount of adrenodoxin precursor synthesized in a cell-free translation system programmed with RNA isolated from ACTH-treated cells increased to approximately 3 times the control level by 36 h. Similarly, ACTH increased the rate of incorporation of [35S]methionine into mature adrenodoxin in radiolabeled adrenocortical cells, an effect that was maximal 36 h after initiation of ACTH treatment. At longer times (48-72 h), the stimulatory effect of ACTH was not maintained, and adrenodoxin synthesis in both radiolabeled cells and cell-free translation systems declined to control levels. The content of adrenodoxin in cells treated with ACTH for 36 h, as measured by electron paramagnetic resonance spectroscopy, was approximately twice that in control cells. The results indicate that ACTH induces the synthesis of adrenodoxin in bovine adrenocortical cells. Based on the present results as well as those previously reported with respect to the induction of cholesterol side chain cleavage cytochrome P-450 by ACTH (DuBois, R. N., Simpson, E. R., Kramer, R. E., and Waterman, M. R. (1981) J. Biol. Chem. 256, 7000-7005), it is proposed that the synthesis of the mitochondrial components of the adrenocortical steroid hydroxylase system is controlled by ACTH in a coordinate fashion.

Adrenal Cortex↗
PubMed1982

Low density lipoprotein receptors in adrenal tissue of a human anencephalic fetus.

The binding of [125I]iodoLDL and [125I]iodoHDL to membrane fractions prepared from fresh adrenal tissue of an anencephalic newborn was examined and compared to the binding of these lipoproteins to membrane fractions prepared from normal human fetal adrenal (HFA) tissue. The apparent dissociation constant (KD) for the interaction of [125I]iodoLDL with its binding sites in membrane fractions prepared from HFA tissue of an anencephalic fetus (23.5 micrograms . ml-1) was similar to that in membranes from normal HFA tissue. However, the binding capacity for [125I]iodoLDL of membrane fractions prepared from adrenal tissue of an anencephalic fetus (600 ng . mg-1 protein) was only one-third of the binding capacity (1500 ng . mg-1 protein) of normal HFA membrane fractions. On the other hand, the binding capacity for [125I]iodoHDL of membrane fractions prepared from anencephalic and normal adrenal tissue was similar.

Adrenal Glands↗
PubMed1982

Inhibition of adenylate cyclase activity in human fat cells by alpha-adrenergic agonists.

The effects of the alpha 1-adrenergic agonist methoxamine and the alpha 2-adrenergic agonist clonidine on isoproterenol stimulated adenylate cyclase activity were examined in plasma membranes prepared from female human subcutaneous adipose tissue. It was found that in the presence of 10 microM GTP and 100 mM NaCl increasing concentrations of both agonists inhibited basal and isoproterenol-stimulated adenylate cyclase activity. The inhibitory action of 5 x 10(-7) M clonidine could not be overcome by increasing concentrations of isoproterenol. These results suggest both alpha 1- and alpha 2-adrenergic agonists inhibit beta-agonist-stimulated adenylate cyclase activity in human adipose tissue.

Adenylyl Cyclase Inhibitors↗
PubMed1982

Prostaglandin biosynthesis in the human fetal adrenal gland: regulation by glucocorticosteroids.

Human fetal adrenal (HFA) tissue was maintained in organ culture to evaluate the biosynthesis of prostaglandins and hormonal regulation of prostaglandin formation by this tissue. The HFA tissue secreted substantial amounts of prostaglandin E(2), prostaglandin F(2alpha), 13,14-dihydro-15-ketoprostaglandin F(2alpha), 6-ketoprostaglandin F(1alpha), and thromboxane B(2); secretion of prostaglandin D(2) could not be demonstrated. Prostaglandin biosynthesis in HFA tissue was inhibited in a time-dependent manner by corticotropin (ACTH; 0.4 muM); by the fourth day of culture, the extent of inhibition of biosynthesis of each prostaglandin was 60-90%. Progesterone (1 muM), cortisol (1 muM), and dexamethasone (1 muM) inhibited prostaglandin biosynthesis whereas estradiol (1 muM) did not. Of the compounds tested for inhibitory activity, dexamethasone was the most potent. An inhibitor of 11beta-hydroxylase activity (metyrapone; 0.1 mM) effectively eliminated the inhibition of prostaglandin biosynthesis caused by corticotropin and progesterone. Metyrapone treatment alone caused a 3-fold increase in prostaglandin biosynthesis by fetal adrenal tissues. Similar stimulatory effects resulted from treatment with inhibitors of (i) 3beta-hydroxysteroid dehydrogenase (cyanoketone; 15 muM), (ii) steroid 17alpha-hydroxylase (SU 10603; 19 muM), and (iii) cholesterol side-chain cleavage (aminoglutethimide; 1 mM). Inhibition of prostaglandin biosynthesis by dexamethasone in the presence or absence of metyrapone was concentration dependent and 50% inhibition could be demonstrated at 1 nM. A competitive inhibitor of the binding of glucocorticosteroids to cytoplasmic receptors (cortisol 21-mesylate; 1 muM) significantly reduced the inhibition of prostaglandin biosynthesis effected by dexamethasone (10 nM). These findings suggest that prostaglandin biosynthesis in the HFA gland is regulated by endogenously synthesized glucocorticosteroids, the actions of which are mediated by a glucocorticosteroid receptor. Such glucocorticosteroids induce the synthesis of a substance that inhibits prostaglandin biosynthesis.

Adrenal Glands↗
PubMed1982

Defective internalization of low density lipoprotein in epidermoid cervical cancer cells.

Cells of an epidermoid cancer cell line of human uterine cervix, which possessed a high-affinity, specific receptor for low density lipoprotein (LDL), internalized and degraded [125I]iodo-LDL at a very low rate. In these cells, LDL did not stimulate cholesteryl ester synthesis, nor did it suppress 3-hydroxy-3-methylglutaryl coenzyme A reductase to the same extent as in the control cells. The binding of [125I]iodo-LDL by these cells was not decreased by preincubation of the cells in medium containing LDL. Using ferritin-labeled LDL (F-LDL) and electron microscopy, it was determined that at 4 degrees C the cells bound F-LDL in the same way as other cancer cell lines that did not have a defect in internalization. When these cells were warmed to 37 degrees C the F-LDL remained on the surface, whereas in cells from control cancer cell lines the F-LDL was internalized and was no longer observed on the cell surface. On the basis of the results of these studies it is concluded that cells of this epidermoid cancer cell line have a defective ability to internalize LDL.

Binding Sites↗
PubMed1982

The levels of plasma cholesterol in the human fetus throughout gestation.

The present study was undertaken to define the umbilical cord plasma concentrations of cholesterol throughout human gestation. Mixed arterial and venous cord plasma samples obtained from abortuses of women undergoing elective abortion or from infants of women who underwent spontaneous premature vaginal delivery, and from infants of women who delivered vaginally at term were assayed for cholesterol by a micro-enzymatic method. No cases that involved any maternal or fetal complications (other than prematurity) were included in this study. Early in gestation (10-16 weeks post-conception), the total cholesterol level in cord plasma was 85.4 +/- 30.7 mg/dl (mean +/- SD), N = 68, with the cholesterol levels in some samples falling within the range of those of adults. Between 16.5 and 20 weeks post-conception, the umbilical cord plasma cholesterol level declined to 39.9 +/- 21.0 mg/dl, n = 19 (P less than 0.001). The cholesterol concentration in umbilical cord plasma then rose to 67.8 +/- 24.0 mg/dl, n = 17 (P less than 0.01) between 26.5 and 32 weeks of gestation. Thereafter, a second decline in the umbilical cord plasma cholesterol level occurred, with the values at 32.5-36 weeks being 58.4 +/- 13.6 mg/dl (n = 16), and at 36.5 to 40 weeks post-conception (term) being 51.4 +/- 11.5 mg/dl, n = 44 (P less than 0.01 vs. 26.5-32 wks). We suggest that the observed changes in fetal cholesterol levels could be related to alterations during development in the rates of lipoprotein-cholesterol biosynthesis and subsequent clearance from plasma by the fetal adrenals wherein cholesterol is used as substrate for steroid biosynthesis.

Abortion, Induced↗
PubMed1982

Lipoprotein-binding sites in human corpus luteum membrane fractions.

In a previous report evidence was presented that plasma low density lipoprotein (LDL), but not high density lipoprotein (HDL), is the major source of cholesterol used by the human corpus luteum for progesterone biosynthesis and, that plasma LDL is taken up by corpus luteum tissues via a receptor-mediated endocytotic process. Using membrane fractions prepared from fresh corpus luteum tissue obtained from nine women at various phases of the menstrual cycle, we conducted the present investigation to characterize lipoprotein-binding sites and changes in such binding sites that might occur throughout the ovarian cycle. High affinity, low capacity binding sites for [125I]iodo-LDL and also for [125I]iodo-HDL were detected in membrane fractions prepared from fresh corpus luteum tissue. The interaction of [125I]iodo-LDL with the high affinity binding sites in fresh corpus luteum membrane fractions was prevented by incubation with heparin. Also, the binding capacity of corpus luteum membrane fractions for [125I]iodo-LDL was reduced by treatment with pronase. On the other hand, the specific binding capacity for [125I]iodo-HDL was unaffected by pronase treatment. The binding capacity for [125I]iodo-LDL in membrane fractions prepared from midluteal phase corpus luteum was significantly greater than that of membrane fractions from tissue obtained at any other phase of the cycle, a finding that suggests that changes in progesterone biosynthesis throughout the cycle are positively correlated with changes in the numbers of binding sites for LDL in the corpus luteum.

Adult↗
PubMed1982

Low density lipoprotein binding and de novo synthesis of cholesterol in the neocortex and fetal zones of the human fetal adrenal gland.

The binding of low density lipoprotein (LDL) and the de novo synthesis of cholesterol in separated zones of human fetal adrenal (HFA) tissues were investigated. The number of LDL-binding sites was 2-fold greater in membrane fractions prepared from fresh fetal zone tissue than in those from neocortex tissue. The binding capacity for LDL in fetal zone and neocortex membrane preparations of HFA tissues maintained in culture in the presence of ACTH was 2-fold greater than that in membrane fractions of control tissues. The rates of de novo synthesis of cholesterol also were determined in separated zones of HFA tissue by measuring the specific activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase in microsomal fractions prepared from HFA tissues and by determining the rate of incorporation of tritium from [3H]water into cholesterol in HFA tissue fragments. The rate of de novo synthesis of cholesterol in fresh fetal zone tissue was twice that in neocortex tissue as estimated by these methods. When separated zones of HFA tissue were maintained in culture in the presence or absence of ACTH, the rates of de novo synthesis, as determined by the rate of incorporation of tritium from [3H]water into cholesterol, were stimulated to a similar extent by ACTH in both fetal zone and neocortex tissues. However, the specific activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase was increased to a greater extent by ACTH pretreatment in neocortex tissues than in fetal zone tissues. In summary, fetal zone tissues of the HFA gland have a larger number of LDL-binding sites and higher rates of de novo synthesis of cholesterol than do neocortex tissues, and ACTH stimulates LDL binding and de novo synthesis of cholesterol in both zones of the HFA gland.

Adrenal Cortex↗