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Biomedical subjects

E R Simpson

Publications and source records attributed to E R Simpson.

At least 289 records · Page 16Linked to original sources
PubMed1981

Deoxycorticosterone biosynthesis from progesterone in kidney tissue of the human fetus.

The levels of deoxycorticosterone (DOC) and DOC sulfate are extraordinarily high in the umbilical cord plasma of human newborns. However, the sources of DOC and DOC sulfate in the human fetus are not defined. Recently, it was shown that plasma progesterone was converted to DOC in extraadrenal tissues in pregnant, nonpregnant, and adrenalectomized women and in men. Thus, DOC formation from plasma progesterone constitutes another example of the formation of a biologically active steroid hormone from a circulating precursor. In addition, the conversion of [3H]progesterone to [3H]DOC in homogenates and microsome-enriched preparations of adult human kidney tissue was demonstrated. To investigate the origin of DOC in the human fetus, we sought 1) to ascertain whether steroid 21-hydroxylase activity was present in human fetal kidney tissue and 2) to establish the kinetics of the reaction catalyzed by this enzyme should it be present in fetal kidney. We employed microsome-enriched preparations of kidney tissue obtained from human abortuses of 9-20 weeks gestation. [3H]DOC formation from [3H]progesterone (1 microM) proceeded in a linear fashion for 3 h. and the rate of formation of [3H]DOC from [3H]progesterone (1 microM) was linear with a microsomal protein concentration between 0.026-0.512 mg X ml-1. The value computed for the apparent Km of steroid 21-hydroxylase in fetal kidney for progesterone was 0.146 microM. We conclude that the human fetal kidney may be an important site of DOC formation as well as a site of DOC action.

Desoxycorticosterone↗
PubMed1981

Cholesterol metabolism in human cancer cells in monolayer culture. V. The effect of progesterone on the regulation of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity by low density lipoprotein.

The effect of progesterone on the regulation of 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase activity by low density lipoprotein (LDL) was studied in eight gynecological cancer cell lines maintained in monolayer culture. In the absence of LDL and progesterone, the specific activity of HMG CoA reductase varied widely from one cell line to another (range, 200-2500 pmol mg microsomal protein-1 min-1). As in noncancerous cells, LDL suppressed the activity of HMG CoA reductase in all cell lines. Progesterone (40 microM) attenuated or abolished the decrease in specific enzyme activity normally brought about by LDL. Progesterone, in the absence of LDL, stimulated HMG CoA reductase activity in some of these cell lines. Other steroids (cortisol and estradiol) had no effect on the specific activity of HMG CoA reductase. The mechanism by which progesterone stimulates HMG CoA reductase activity is not clear, but since progesterone receptors were not detectable in these cells a mechanism independent of the classic hormone receptor-mediated phenomenon is likely to be involved.

Cells, Cultured↗
PubMed1981

Aromatization of androstenedione by human adipose tissue stromal cells in monolayer culture.

Stromal cells and adipocytes were separated after collagenase treatment of adipose tissue obtained from women undergoing elective surgery, and these cells were used to study aromatization of [3H]androstenedione in vitro. Aromatization activity was estimated either 1) by determining the incorporation of tritium from [1-3H]androstenedione into [3H]water or else 2) by determining the formation of [3H]estrone (E1) and [3H]estradiol (E2) from [1,2,6,7-3H]androstenedione. It was established that only 13% of the aromatase activity of adipose tissue resided in the adipocyte fraction, whereas 87% of the aromatase activity was in the stromal/vascular fraction. Subsequent studies of aromatization were conducted utilizing stromal/vascular cells grown to confluence in monolayer culture. In such cells, the formation of [3H]E2 was slower initially but increased with time, and after 48 h of incubation, the amount of [3H]E2 produced exceeded that of [3H]E1. The rate of [3H]E1 formation, as a function of [3H]androstenedione concentration, followed Michaelis-Menten kinetics. The Vmax ranged from 0.8-3.0 pmol and ranged from 0.16-0.67 pmol mg-1 cell protein 6 h-1 in cells from omental adipose tissue. The apparent Km for [3H]androstenedione in stromal cells derived from both omental and sc tissue was the same, i.e. about 25 nM. We conclude that the ability of human adipose tissue to form estrogen is not a function primarily of the adipocytes but rather resides principally in the cells of the stroma.

Adipose Tissue↗
PubMed1981

Synthesis of cholesterol in the human fetus: 3-hydroxy-3-methylglutaryl coenzyme A reductase activity of liver microsomes.

The specific activity of 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase in microsome-enriched fractions prepared from normal human fetal liver tissue was assessed. The mean specific activity was 0.58 +/- 0.18 nmol mevalonate formed min-1 mg-1 protein. The activity of the enzyme was inhibited by preincubation of microsomes with ATP (4 mM) and was greatly reduced when microsomes were prepared from tissue homogenized in the presence of NaF (50 mM). It can be computed that the activity of HMG CoA reductase in human fetal liver microsomes is adequate to provide cholesterol to meet the requirements of the fetal adrenal for steroid precursor provided that cholesterol synthesized in the fetal liver appears in the plasma in the form of low density lipoprotein.

Cholesterol↗
PubMed1981

Deoxycorticosterone sulfate biosynthesis in human fetal kidney.

The levels of deoxycorticosterone (DOC) and DOC-SO4 are extraordinarily high in umbilical cord plasma of human newborns compared to those in plasma of men and nonpregnant women. Progesterone is converted to DOC in minces, homogenates, and microsome-enriched preparations of human fetal kidney tissue. Since DOC, a mineralocorticosteroid, is formed in situ in kidney, its potential site of action, we sought to define whether sulfurylation of DOC also occurred in fetal kidney. We found that radiolabeled DOC-SO4 was formed from [3H]DOC and from [3H]progesterone in minces of human fetal kidneys. C-21 hydroxysteroid sulfotransferase activity was found to be localized principally in the cytosolic fraction of homogenates of human fetal kidney tissue and was present in both the cortical and medullary portions of fetal kidney. In cytosolic fractions prepared from homogenates of human fetal kidneys, we found that the apparent Km of the C-21 hydroxysteroid sulfotransferase for DOC was 4.7 microM; the reaction was linear with time for 60 min and was linear with protein concentration up to 1.2 mg X ml-1 incubation mixture. Thus, DOC-SO4, as well as DOC, is synthesized in the kidney of the human fetus, and kidney may be an important site of formation of these steroids that are found in fetal blood in very high concentrations.

Cytosol↗
PubMed1981

Inhibition of the lipolytic action of beta-adrenergic agonists in human adipocytes by alpha-adrenergic agonists.

The aim of this study was to define the role of the alpha-adrenergic receptor in the regulation of lipolysis by human adipocytes. Glycerol production by isolated human adipocytes was stimulated by the pure beta-adrenergic agonist isoproterenol in a dose-dependent fashion. This stimulation of lipolysis was inhibited by the alpha-adrenergic agonists methoxamine, phenylephrine, and clonidine. Epinephrine-stimulated lipolysis was potentiated by the alpha-adrenergic antagonists, dihydroergocryptine, phentolamine, phenoxybenzamine, and yohimbine. Whereas the attenuation of beta-adrenergic agonist-stimulated lipolysis by alpha-adrenergic agonists was reversed completely by the alpha 2-adrenergic antagonist yohimbine, the alpha 1-antagonist prazosin did not reverse such attenuation. It is concluded that alpha-adrenergic agonists act as antilipolytic agents in human adipocytes and that this action may result from the interaction of these compounds with a population of alpha 2-adrenergic receptors.

Adipose Tissue↗
PubMed1981

The role of receptor-mediated low-density lipoprotein uptake and degradation in the regulation of progesterone biosynthesis and cholesterol metabolism by human trophoblasts.

Dispersed human placental trophoblastic cells, maintained in primary culture, utilize principally cholesterol derived from low-density lipoprotein (LDL) for progesterone biosynthesis. The rate of secretion of progesterone by these cells increased (100 ng X mg-1 cell protein X 24 h-1 to 390 ng X mg-1 cell protein X 24 h-1) as the LDL concentration in the culture medium was increased (0 to 420 micrograms protein X ml-1). At an HDL-protein concentration of 1000 micrograms protein X ml-1, the rate of progesterone secretion by these cells was one-half that attained by cells maintained in medium containing LDL. The uptake of [125I]iodo-LDL by trophoblastic cells increased at [125I]iodo-LDL concentrations between 0 and 14 micrograms protein X ml-1 in the culture medium, was maximal by 5 to 6 h, and was diminished progressively as the concentration of non-radiolabelled LDL, but not HDL, was increased. Degradation of [125I]iodo-LDL increased as a function of the [125I]iodo-LDL concentration in the culture medium, was linear from 30 min to 32 h, was inhibited by the addition of choloroquine (40 mumol/1) to the culture medium; and was diminished progressively as the concentration of non-radiolabelled LDL, but not HDL, was increased. We observed a reduction in uptake and degradation of [125I]iodo-LDL by these cells as a function of the LDL concentration in the preincubation culture medium. Moreover, when the cells were preincubated with LDL for various times, a 90 per cent reduction in the rate of uptake and degradation of [125I]iodo-LDL was observed after 14 h. These findings are suggestive that trophoblastic cells are capable of down-regulating the number of LDL receptors on the cell surface. The incorporation of radiolabelled oleic acid into cholesteryl esters by these cells was linear for 6 h, increased as a function of the oleic acid concentration in the culture medium, and was stimulated when LDL was present in the culture medium. On the other hand, the synthesis of cholesteryl esters was inhibited half-maximally when the progesterone concentration in the culture medium was 20 mumol/1. Finally, the incorporation of [14C]acetate into cholesterol was inhibited as a function of the LDL concentration in the culture medium. Based on the results of these studies, our belief is that mechanism(s) have been elucidated to define the regulation of (1) progesterone biosynthesis; (2) de novo cholesterol synthesis; (3) intracellular cholesteryl ester storage; and (4) LDL uptake by normal human trophoblastic cells.

Cholesterol↗
PubMed1981

Effect of cell density and confluency on cholesterol metabolism in cancer cells in monolayer culture.

Cholesterol metabolism in four gynecological cancer cell lines in monolayer culture was evaluated as a function of cell density. The rate of uptake and degradation of [125I]iodinated low-density lipoprotein increased during the first 24 to 48 hr of culture, but decreased thereafter. Once the cells became confluent, the rate of metabolism of [125I]iodinated low-density lipoprotein was only one-tenth that in cells which were in the preconfluent state. The specific activity of 3-hydroxy-3-methylglutaryl coenzyme reductase increased during the first 24 to 48 hr of culture and subsequently declined, reaching a nadir after confluency was attained. The rate of incorporation of [14C]oleate into cholesteryl esters was low when the cells were in the log-exponential phase of replication but increased gradually as cell density increased. The highest specific activity of acylcoenzyme A: cholesterol acyltransferase was attained after the cells became confluent. Generally speaking, there was an inverse relationship between the specific activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase, on the one hand, and the rate of [125I]iodinated low-density lipoprotein metabolism and cholesteryl ester synthesis, on the other. It is concluded that cholesterol metabolism in cancer cells in monolayer culture is regulated, in part, by the rate of cell division. In the cancer cells utilized in this study, it is apparent that cholesterol metabolism was subject to the same regulatory mechanisms as are present in nonneoplastic cells.

Adenocarcinoma↗
PubMed1980

Inverse relation between low-density lipoprotein-cholesterol and dehydroisoandrosterone sulfate in human fetal plasma.

A striking inverse correlation was found in umbilical cord plasma between the concentrations of dehydroisoandrosterone sulfate and low-density lipoprotein (LDL)-cholesterol but not high-density lipoprotein-cholesterol or very low density lipoprotein-cholesterol. Dehydroisoandrosterone sulfate is a major secretory product of the human fetal adrenal and the principal precursor of placental estrogen production. The data suggest that the concentrations for LDL-cholesterol in fetal plasma are influenced by the rate of utilization of LDL-cholesterol by the fetal adrenal for steroidogenesis and are not necessarily related to a genetic predisposition for hypercholesterolemia or other lipoprotein disorders.

Adrenal Cortex↗
PubMed1980

Steroid secretion by ACTH-stimulated human fetal adrenal tissue during the first week in organ culture.

Fetal adrenal tissue has been reported to lose its in vivo secretory pattern by virtue of a loss of fetal zone cells after the first week in culture. Consequently, we studied the steroidogenic capacity and the responsiveness to ACTH of human fetal adrenal tissue during the first week in organ culture. The culture medium was removed daily and assayed for cortisol and dehydroisoandrosterone sulfate (DS). First, as the concentration of ACTH in the medium was increased from 0 to 1 micrograms/ml steroid secretion increased. When tissue fragments were maintained in the absence of ACTH for 3 to 4 days, there was a striking increase in steroid secretion upon addition of ACTH to the medium, with larger rates of secretion of cortisol than DS being observed. Second, the steroidogenic capacity of the separate zones of the fetal adrenal gland was assessed. Tissue from the fetal zone secreted large amounts of DS and small amounts of cortisol, whereas neocortex tissue secreted similar quantities of DS and cortisol. Third, fetal zone tissue was maintained the absence of ACTH for 4 days and thereafter ACTH was added to the media for an additional 6 days. In this experiment, there was a marked increase in DS secretion rate after the addition of ACTH and a smaller increase in cortisol secretion.

Adrenal Glands↗
PubMed1980

Deoxycorticosterone biosynthesis in human kidney: potential for formation of a potent mineralocorticosteroid in its site of action.

The extra-adrenal formation of deoxycorticosterone (DOC) from plasma progesterone has been demonstrated in humans. In those studies it was shown that in some persons the volume of plasma cleared of progesterone by DOC formation was great, namely, 75 liter/24 hr. Because steroid 231-hydroxylase activity [steroid 21-monooxygenase; steroid, hydrogen-donor:oxygen oxidoreductase (21-hydroxylating), EC 1.14.99.10] could not be demonstrated in homogenates or microsome-enriched preparations of human lung or liver tissue, we speculated that the 21-hydroxylation of plasma progesterone might take place in the kidney. Employing whole tissue homogenates and microsome-enriched preparations of human kidney tissue, we demonstrated the formation of [3H]DOC from kidney tissue, we demonstrated the formation of [3H]DOC from [3H]progesterone. The rate of formation of DOC from progesterone in microsomal preparations from kidney tissues of adult humans varied from 0 to 803 pmol per mg of microsomal protein per hr. The value computed for the apparent Km of the enzyme for progesterone was 0.140 microM. On the basis of these findings, we conclude that steroid 21-hydroxylase activity is present in human kidney tissue and that the kidney may be an important site of DOC formation as well as a site of DOC action.

Cell-Free System↗
PubMed1980

Conversion of plasma progesterone to deoxycorticosterone in men, nonpregnant and pregnant women, and adrenalectomized subjects.

During the third trimester of human pregnancy the concentrations of deoxycorticosterone (DOC) in maternal plasma are 4-50 times those in nonpregnant women and men. It has been suggested that the increased amount of DOC in maternal plasma originates in the fetal compartment. We considered an alternate explanation for the high levels of DOC in plasma or near-term pregnant women, viz., that DOC may be derived in part from 21-hydroxylation of maternal plama progesterone. To test this hyposthesis we measured the fractional conversion of plasma progesterone to DOC from the relationship between the 3H:14C ratio of the infused tracers, [3H]progesterone and [14C]-DOC, and the 3H:14C ratio or urinary 3 alpha,21-dihydroxy-5 beta-pregnan-20-one (tetrahydro-DOC). The fractional conversion of plasma progesterone to DOC ([rho](BU)P-DOC), measured in this manner, was 0.007 +/- 0.001 (mean +/- SEM, n = 26) in the subjects of this study. The values for [rho](BU)P-DOC varied widely among subjects (0.002-0.022) but the range of values for [rho](BU)P-DOC was similar among women pregnant with an anencephalic or dead fetus, nonpregnant and adrenalectomized women, and men. The transfer constant of conversion of progesterone to DOC in plasma, [rho](BB)P-DOC, remained constant in a nonpregnant woman during the infusion of nonradiolabeled progesterone at rates of 0-14 mg/h. Based on the results of these studied, we conclude that DOC is formed by extra-adrenal 21-hydroxylation of plasma progesterone and that the rate of formation of DOC by this pathway is proportional to the concentration of progesterone in plasma.

Adrenalectomy↗