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PubMedDate not supplied

Biophysical membrane interactions of steroid hormones: a potential complementary mechanism of steroid action.

There is strong evidence to support the hypothesis that the rapid effects of steroids on neurons are membrane-mediated. Rapid steroid effects have been demonstrated in the absence of intracellular receptors, in the presence of RNA or protein synthesis inhibitors, and in response to steroids coupled to large proteins that block access to intracellular receptors. This study selectively reviewed the emerging body of evidence suggesting that steroids have multiple sites of cellular actions, and explored in depth one possible membrane-mediated mechanism of action, the membrane-intercalation model.

Biophysical Phenomena↗
PubMed1983

Regulation of steroid-transforming enzymes by endogenous steroids.

The biosynthesis of C21, C19, and C18 steroids in adrenal, ovary, testis and foeto-placental unit is summarized and the properties of the enzymes involved are described, with particular reference to their substrate specificities and to the possible controlling effects of endogenous steroids. It is concluded that these may exert a considerable regulatory role in vivo in steroid hormone biosynthesis.

Adrenal Cortex↗
PubMed1999

[Steroid receptors and mechanism of action of sex steroids].

Steroid hormone receptors define a large family of proteins. Recently, a new estradiol receptor has been identified. This discovery suggests the existence of a previously unrecognized pathway of estrogen signalling. Moreover, it implies important pharmacological consequences. Receptors activation induces the modulation of transcription of specific genes. Proteins involved in this effect have been identified: coactivators, corepressors and cointegrators. Their mechanism of action have been characterized. They modify histone acetylation of the corresponding promotor. Sex steroid receptors are located in the nucleus. This nuclear localization is in fact a dynamic situation, resulting from a continuous shuttling of the receptor between the cytoplasm and the nucleus. Non genomic effects of steroids have also been described.

Animals↗
PubMed2005

Addition of inhaled long-acting beta2-agonists to inhaled steroids as first line therapy for persistent asthma in steroid-naive adults.

BACKGROUND: Consensus statements recommend the addition of long-acting inhaled beta2-agonists only in asthmatic patients who are inadequately controlled on inhaled corticosteroids. OBJECTIVES: To compare the efficacy of initiating anti-inflammatory therapy using the combination of inhaled corticosteroids and long-acting beta2-agonists (ICS+LABA) as compared to inhaled corticosteroids alone (ICS alone) in steroid-naive children and adults with persistent asthma. SEARCH STRATEGY: We identified randomised controlled trials (RCTs) through electronic database searches (Cochrane Airways Group Specialised Register, Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, EMBASE and CINAHL) until April 2004, bibliographies of identified RCTs and correspondence with manufacturers. SELECTION CRITERIA: RCTs comparing the combination of inhaled corticosteroids and long-acting beta2-agonists (ICS + LABA) to inhaled corticosteroids (ICS) alone in steroid-naive children and adults with asthma. DATA COLLECTION AND ANALYSIS: Studies were assessed independently by each reviewer for methodological quality and data extraction. Confirmation was obtained from the trialists when possible. The primary endpoint was rate of asthma exacerbations requiring systemic corticosteroids. Secondary endpoints included pulmonary function tests (PFTs), symptoms, use of other measures of asthma control, adverse events, and withdrawal rates. MAIN RESULTS: Eighteen trials met the inclusion criteria; nine (totaling 1061 adults) contributed sufficient data to be analysed. Baseline forced expiratory volume in one minute (FEV1) was less than 80% predicted value in four trials and equal to or greater than 80% in five trials. The long-acting beta2-agonists (LABA) formoterol (N=2) or salmeterol (N=7) were added to a dose of at least 800 microg/day of beclomethasone dipropionate (BDP) equivalent of inhaled corticosteroids (ICS) in three trials and to at least 400 microg/day in the six remaining trials. Treatment with ICS plus LABA was not associated with a lower risk of exacerbations requiring oral corticosteroids than ICS alone (relative risk (RR) 1.2; 95% confidence interval (CI) 0.8 to 1.9). FEV1 improved significantly with LABA (weighted mean difference (WMD) 210 ml; 95% CI 120 to 300), as did symptom-free days (WMD 10.74%; 95% CI 1.86 to 19.62), but the change in use of rescue fast-acting beta2-agonists was not significantly different between the groups (WMD -0.4 puff/day, 95% CI -0.9 to 0.1). There was no significant group difference in adverse events (RR 1.1; 95% CI 0.8 to 1.5), withdrawals (RR 0.9; 95% CI 0.6 to 1.2), or withdrawals due to poor asthma control (RR 1.3; 95% CI 0.5 to 3.4). AUTHORS' CONCLUSIONS: In steroid-naive patients with mild to moderate airway obstruction, the initiation of inhaled corticosteroids in combination with long-acting beta2-agonists does not significantly reduce the rate of exacerbations over that achieved with inhaled corticosteroids alone; it does improve lung function and symptom-free days but does not reduce rescue beta2-agonist use as compared to inhaled steroids alone. Both options appear safe. There is insufficient evidence at present to recommend use of combination therapy rather than ICS alone as a first-line treatment.

Administration, Inhalation↗
PubMed1987

Comparative study of plasma steroid and steroid glucuronide levels in normal men and in men with benign prostatic hyperplasia.

Plasma steroids were analyzed in 16 normal men and in 10 men with prostatic benign hyperplasia (BPH). The steroids measured by radioimmunoassay include pregnenolone, 17-OH-pregnenolone, dehydroepiandrosterone, androst-5-ene-3 beta, 17 beta-diol, testosterone, dihydrotestosterone, androstane-3 alpha, 17 beta-diol, androstane-3 beta, 17 beta-diol, estrone, and estradiol as well as their glucuronide derivatives. In addition, cortisol and the sulphoconjugated form of dehydroepiandrosterone were determined. Whereas the levels of pregnenolone, pregnenolone glucuronide and 17-OH-pregnenolone glucuronide are not different in the two groups, the levels of 17-OH-pregnenolone in the BPH group (0.87 +/- 0.07 ng/ml) exceed by two-fold (p less than 0.01) those observed in normal men. Plasma dehydroepiandrosterone and androst-5-ene-3 beta, 17 beta-diol concentrations are markedly elevated in the BPH group (1.49 +/- 0.23 and 0.55 +/- 0.08 ng/ml vs the control groups 0.43 +/- 0.11 and 0.31 +/- 0.05 ng/ml, respectively). Since the plasma cortisol and pregnenolone levels are comparable in these two groups, our data suggest that the elevation of plasma 17-OH-pregnenolone, dehydroepiandrosterone, and androst-5-ene-3 beta, 17 beta-diol reflects an increase of adrenal 17-hydroxylase activity in patients with BPH. A slight increase of the plasma dihydrotestosterone and androsterone glucuronide concentration is also observed in men with BPH, indicating an increase of 5 alpha-reduced androgen formation. We have also observed, in the BPH group, a 50% decrease (p less than 0.01) of plasma glucuronidated androst-5-ene-3 beta, 17 beta-diol, estrone, and estradiol levels, suggesting that the transformation of unconjugated estrogenic steroids into glucuronide derivative is inhibited in BPH patients. In summary, our data indicate that adrenal C-19 steroids might be involved in the process of BPH. Furthermore, whereas the estrogen glucuronide formation is diminished in men with BPH, the prostatic androgen metabolism as reflected by plasma dihydrotestosterone and androsterone glucuronide concentrations seems to be increased.

Aged↗
PubMed1990

Pharmacokinetics and pharmacodynamics of oral contraceptive steroids: factors influencing steroid metabolism.

The time-dependent alterations in the serum concentrations of ethinyl estradiol, gestodene, and 3-keto-desogestrel during treatment with 30 micrograms of ethinyl estradiol + 75 micrograms of gestodene or 30 micrograms of ethinyl estradiol + 150 micrograms of desogestrel were investigated during 12 months. The levels of gestodene and 3-keto-desogestrel increased between days 1 and 21 of each cycle, reaching maximal levels during the third and sixth cycles. The serum concentrations of gestodene were fourfold to fivefold higher than those of 3-keto-desogestrel. The ethinyl estradiol levels increased significantly between days 1 and 10 during each cycle and were significantly higher by 70% during intake of ethinyl estradiol/gestodene compared with ethinyl estradiol/desogestrel, although the dose was identical. Intake of gestodene, in addition to 35 micrograms of ethinyl estradiol + 2 mg of cyproterone acetate, caused a rise in ethinyl estradiol levels. During treatment with ethinyl estradiol/gestodene and an additional 150 micrograms of levonorgestrel, there was a continuous increase in gestodene levels, although sex hormone-binding globulin level did not change. During treatment with 30 or 35 micrograms of ethinyl estradiol and 75 micrograms of gestodene, 150 micrograms of desogestrel, or 2 mg of cyproterone acetate, there were large intraindividual and interindividual variations in the steroid levels and ratios of estrogen: progestogen levels. There was no correlation with the occurrence of intermenstrual bleedings. It is concluded that ethinyl estradiol and nortestosterone derivatives may inhibit steroid-metabolizing enzymes in the liver, which results in a rise in the serum levels of contraceptive steroids. The cause of the large intraindividual variations is as yet unknown, but it is probably from changes in steroid metabolism.

Adolescent↗
PubMed1981

Enzymic synthesis of steroid sulphates. XIV. Properties of human adrenal steroid alcohol sulphotransferase.

Pure steroid alcohol sulphotransferase (EC 2.8.2.-) has the property of sulphurylating hydroxyl groups on different positions of the steroid ring. It has now been established that although only monosulphates are formed from substrates such as 3,17-diols, the position of the sulphate group depends on the relative configuration of the hydroxyl groups. Androst-5-ene-3 beta,17 beta-diol, for example, is sulphurylated mainly at the 17-position. In addition, compounds such as epitestosterone and 17 alpha-estradiol are sulphurylated at much higher rates than their respective 17 beta-epimers. It is believed that the steroid can approach the sulphurylation site via (i) ring A with the beta-side upwards, and in this mode a 3 beta-hydroxyl is sulphurylated at a higher rate than a 3 alpha-hydroxyl, or (ii) ring D with the beta-side downwards, and in this mode a 17 alpha-hydroxyl group is oriented in an analogous fashion to the 3 beta-hydroxyl in (i). The enzyme exhibits non-Michaelis-Menten kinetics within physiological concentrations (0-2 micro M) of the substrate dehydroepiandrosterone and evidence was obtained for the presence of multiple interacting steroid-binding sites. A regulatory role for the enzyme in the secretion of dehydroepiandrosterone from the human adrenal gland is proposed.

Adrenal Glands↗
PubMed1992

Comparative affinity of steroidal and non-steroidal antioestrogens, cholesterol derivatives and compounds with a dialkylamino side chain for the rat liver antioestrogen binding site.

Steroidal and non-steroidal antioestrogens, steroidal compounds with (disubstituted) dialkyl amino side chain, cholesterol derivatives, histaminic and (anti)-progestational compounds were tested for their ability to compete with [3H]tamoxifen for the specific antioestrogen binding site (AEBS) in the post-mitochondrial fraction of rat liver homogenates. Relative binding affinity was highest for compounds with diethylamino or pyrrolidino ethoxy side chains. Affinity decreased with shortening of this side chain. No connection could be established between the carbon backbone of the compound and affinity, except for the presence of (sometimes aromatic) ring structures. Steroidal ring structures do not seem to be necessary for binding. The cholesterol derivatives, tested in our laboratory, showed very little affinity for the rat liver AEBS. Histamine, melatonin and the (anti)-progestational compounds showed no affinity for the AEBS; and we therefore conclude that the AEBS is not identical to receptors for these compounds. Results of these experiments could be useful in other investigations on the development of resistance of breast cancer to antioestrogens.

Animals↗
PubMed1992

Localization of the steroid-binding site of the human sex steroid-binding protein of plasma (SBP or SHBG) by site-directed mutagenesis.

The amino-terminal region of the human sex steroid-binding protein of plasma (SBP or SHBG) containing K134 and M139 was found to represent part of the steroid-binding site. This was accomplished by constructing and expressing site-directed mutants having the following replacements: M139L, M139K, M139S, K134A, H235S, and Y57F. The results indicated that M139L and H235S were fully-active, K134A and Y57F were 50 and 67% active, M139K was 7% active, and M139S was inactive. These results support affinity-labeling data indicating that both K134 and M139 are located in or near the site, and suggest that Y57 may play a role in steroid binding. The fully active H235S mutant reveals that H235 is not involved in the steroid-binding process.

Amino Acid Sequence↗
PubMed1985

Anti-inflammatory drugs in experimental atherosclerosis. Part 6. Combination therapy with steroid and non-steroid agents.

The pathological changes which accompany enhanced cholesterol deposition in atherosclerosis include inflammatory responses mediated by the prostaglandin cyclooxygenase and lipoxygenase-leukotriene metabolite of the arachidonic acid cascade. Cortisone suppresses arachidonic acid release, whereas non-steroid anti-inflammatory drugs inhibit principally the cyclooxygenase enzyme. Groups of New Zealand white rabbits were fed a 1% cholesterol diet for 12 weeks. Diets of selected groups were further supplemented daily with the non-steroid anti-inflammatory drugs phenylbutazone (100 mg), oxyphenylbutazone (240 mg), flufenamic acid (100 mg), either singly or in combination with cortisone acetate (10 mg or 5 mg), or 9-alpha-fluorohydrocortisone (30 micrograms or 200 micrograms). Serum lipid levels were measured at 0, 4, 8 and 12 weeks, and atherosclerotic plaque intensity in thoracic aorta was measured at 12 weeks using a planimetric technique: serum cholesterol levels in control groups increased from 38 +/- 5 to 1190 +/- 139 mg/100 ml. Neither the rate of increase nor the final lipid values attained were significantly changed by the non-steroid drugs. The non-steroid drugs reduced plaque coverage by about one third (phenylbutazone 34 +/- 10%, flufenamic acid 36 +/- 11%) compared to controls. In combination therapy, addition of cortisone acetate resulted in further plaque suppression. Cortisone 10 mg + phenylbutazone gave 100% suppression; cortisone 5 mg + phenylbutazone gave 82 +/- 18%, and cortisone 5 mg + flufenamic acid gave 84 +/- 3%.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1987

Contraceptive steroid treatment affects steroid binding proteins and the percentage of free 17 beta-estradiol and testosterone in the cynomolgus monkey (Macaca fascicularis).

The levels of steroid binding globulins were characterized in cynomolgus monkeys that were treated with contraceptive steroid preparations delivered either by intravaginal rings (CVR) or orally (OC) in the diet. Levonorgestrel (dNG) was the bioactive progestin and the estrogen was either 17 beta-estradiol (E2) in the CVR treatment or ethinyl estradiol (EE) in the OC treatment. Both contraceptive treatments lowered sex hormone binding globulin (SHBG) levels below those observed in males (P less than 0.05) and normal females (P less than 0.01). Corticosteroid binding globulin (CBG) was elevated (P less than 0.01) in the OC treatment, demonstrating the potency of EE. The distribution of E2 and testosterone (T) between binding to SHBG or albumin and the unbound fraction was calculated after the determination of the percentage of free steroid by centrifugal ultrafiltration. Both contraceptive treatments increased the percentage of free T and E2 (P less than 0.01) in the subset of monkeys that were evaluated, but the percentage bound to SHBG and albumin were different only for the CVR group (P less than 0.05). Decreased total T concentrations in the treatment groups offset any increase in free T concentrations associated with an increase in the percentage of free T. The differences in the distribution of binding to SHBG associated with these contraceptive steroid treatments was influenced more by the reduction in the binding capacity of SHBG than by the displacement of E2 and T from SHBG by dNG.

Administration, Oral↗
PubMed1991

Changes in the levels of protein and steroid hormones in the plasma and steroid hormone receptors in the uterus of normal cycling guinea pigs.

This study deals with the estrous cycle of guinea pigs in relation to sexual behavior, uterine weight, levels of gonadotropins, steroid hormones, and steroid hormone receptors in the uterus. The guinea pigs in this study showed cyclic changes in various reproductive functions broadly similar to other laboratory species studied. The increase in the uterine weight coincided with high concentration of steroid hormones (estradiol and progesterone) secreted during proestrus and estrus. The elevated levels of steroid hormone receptor concentrations in the uterus during these periods also confirm the role of these hormones. The rise in progesterone level from day 14 of the cycle was associated with lordosis and its related behavior. It was noted that the "estrus behavior" is the most accurate external marker for ovulation and sexual receptivity to males. It was also observed that there is an association between follicle-stimulating hormone and luteinizing hormone during the preovulatory period that was not demonstrated in previous studies.

Animals↗
PubMed1992

17-Epimerization of 17 alpha-methyl anabolic steroids in humans: metabolism and synthesis of 17 alpha-hydroxy-17 beta-methyl steroids.

The 17-epimers of the anabolic steroids bolasterone (I), 4-chlorodehydromethyltestosterone (II), fluoxymesterone (III), furazabol (IV), metandienone (V), mestanolone (VI), methyltestosterone (VII), methandriol (VIII), oxandrolone (IX), oxymesterone (X), oxymetholone (XI), stanozolol (XII), and the human metabolites 7 alpha,17 alpha-dimethyl-5 beta-androstane-3 alpha,17 beta-diol (XIII) (metabolite of I), 6 beta-hydroxymetandienone (XIV) (metabolite of V), 17 alpha-methyl-5 beta-androst-1-ene-3 alpha,17 beta-diol (XV) (metabolite of V), 3'-hydroxystanozolol (XVI) (metabolite of XII), as well as the reference substances 17 beta-hydroxy-17 alpha-methyl-5 beta-androstan-3-one (XVII), 17 beta-hydroxy-17 alpha-methyl-5 beta-androst-1-en-3-one (XVIII) (also a metabolite of V), the four isomers 17 alpha-methyl-5 alpha-androstane-3 alpha,17 beta-diol (XIX) (also a metabolite of VI, VII, and XI), 17 alpha-methyl-5 alpha-androstane-3 beta,17 beta-diol (XX), 17 alpha-methyl-5 beta-androstane-3 alpha,17 beta-diol (XXI) (also a metabolite of V, VII, and VIII), 17 alpha-methyl-5 beta-androstane-3 beta,17 beta-diol (XXII), and 17 beta-hydroxy-7 alpha,17 alpha-dimethyl-5 beta-androstan-3-one (XXIII) were synthesized via a 17 beta-sulfate that spontaneously hydrolyzed in water to several dehydration products, and to the 17 alpha-hydroxy-17 beta-methyl epimer. The 17 beta-sulfate was prepared by reaction of the 17 beta-hydroxy-17 alpha-methyl steroid with sulfur trioxide pyridine complex. The 17 beta-methyl epimers are eluted in gas chromatography as trimethylsilyl derivatives from a capillary SE-54 or OV-1 column 70-170 methylen units before the corresponding 17 alpha-methyl epimer. The electron impact mass spectra of the underivatized and trimethylsilylated epimers are in most cases identical and only for I, II, and V was a differentiation between the 17-epimers possible. 1H nuclear magnetic resonance (NMR) spectra show for the 17 beta-methyl epimer a chemical shift for the C-18 protons (singlet) of about 0.175 ppm (in deuterochloroform) to a lower field. 13C NMR spectra display differences for the 17-epimeric steroids in shielding effects for carbons 12-18 and 20. Excretion studies with I-XII with identification and quantification of 17-epimeric metabolites indicate that the extent of 17-epimerization depends on the A-ring structure and shows a great variation for the different 17 alpha-methyl anabolic steroids.

Adult↗
PubMed1990

Fetal fluid steroids and their relationship to gonadal steroid secretion in single and twin bovine fetuses.

Steroid concentrations in the fetal fluids of 153 single and 69 twin bovine pregnancies, ranging in age from 35 to 125 d of gestation, were studied to compare gonadal steroid secretions in vitro with the concentrations found in amniotic and allantoic fluids during the early stages of sex differentiation. Among the steroids measured in fetal fluids, only the testosterone level showed a correlation with the amount secreted by the gonads. Significantly higher concentrations of testosterone were associated with male fetuses than with female fetuses. The concentrations of androstenedione, estradiol and estrone in both fetal fluid compartments were generally correlated with age, reflecting the extra-gonadal source of steroids in these fluids. Androstenedione levels in fetal fluids were significantly higher in twins than in singletons, suggesting that this parameter may be useful for the diagnosis of fetal sex and/or type of pregnancy.

Journal Article↗
PubMed1983

Reduction of enzyme release from reperfused ischemic hearts by steroidal and non-steroidal prostaglandin synthesis inhibitors.

Exacerbation of heart tissue damage by reperfusion of the ischemic myocardium is a well documented phenomenon. The present study was undertaken to evaluate prostaglandin (PG) involvement in reperfusion-induced damage of isolated globally ischemic rat hearts. Reperfusion produced significant increases in creatinephosphokinase (CPK) and lactic dehydrogenase (LDH) efflux which was accompanied by enhanced PG release. Three non-steroidal antiinflammatory drugs; indomethacin, mefenamic acid and ASA, and the steroidal agents; dexamethasone, hydrocortisone and methylprednisolone significantly reduced both the release of CPK and PGs upon reperfusion whereas only indomethacin and mefenamic acid decreased LDH release. There was a significant correlation between the inhibition of PG synthesis and the attenuation of CPK leakage by both non-steroidal (P less than 0.001) and steroidal (P = 0.02) antiinflammatory agents. In spite of beneficial effects on enzyme release, drug treatment did not enhance recovery of mechanical function after reperfusion. The results suggest that inhibition of PG biosynthesis may be beneficial in preserving membrane, particularly mitochondrial integrity of the reperfused myocardium.

Animals↗
PubMed1994

Inhibition of steroid sulphatase activity by steroidal methylthiophosphonates: potential therapeutic agents in breast cancer.

The hydrolysis of steroid sulphates, by steroid sulphatase, is an important source of oestrogenic steroids (oestrone, oestradiol and 5-androstene-3 beta,17 beta-diol) which are found in tumours. In the present study, we have examined the effect of dehydroepiandrosterone-3-O-methylthiophosphonate (DHA-3-MTP), pregnenolone-3-O-methylthiophosphonate (pregnenolone-3-MTP) and cholesterol-3-O-methylthiophosphonate (cholesterol-3-MTP) on the inhibition of oestrone sulphatase as well as DHA sulphatase activities in intact MCF-7 breast cancer cells and in placental microsomes. All three methylthiophosphonates significantly (P < 0.01) inhibited the hydrolysis of oestrone sulphate (E1 S) in intact MCF-7 cells (31-85% inhibition at 1 microM and 53-97% inhibition at 10 microM). Significant inhibition of DHA sulphatase was also achieved. At a concentration of 50 microM, all three compounds inhibited the hydrolysis of dehydroepiandrosterone sulphate (DHAS) by > 95%. Using human placental microsomes, the Km and Vmax of E1S were determined to be 8.1 microM and 43 nmol/h/mg protein. The corresponding Ki values for DHA-3-MTP, pregnenolone-3-MTP and cholesterol-3-MTP were found to be 4.5, 1.4 and 6.2 microM, respectively. Such inhibitors which are resistant to metabolism may have considerable potential as therapeutic agents and may have additional advantage over aromatase inhibitors in also reducing tumour concentrations of the oestrogenic steroid, 5-androstene-3 beta,17 beta-diol, by inhibiting the hydrolysis of DHAS.

Antineoplastic Agents↗
PubMed1987

Genetic heterogeneity of steroid sulfatase deficiency revealed with cDNA for human steroid sulfatase.

Three cDNA clones with inserts of 1.2-1.6 kb that reacted both with antibodies and oligonucleotides specific for steroid sulfatase were isolated from a human placental library in lambda gt11. The 5'-end of one of the inserts, STS-3, was sequenced and colinearity with the amino acid sequence of 3 peptides of steroid sulfatase encompassing 64 amino acids was demonstrated. STS-3 hybridized with 2.5, 4.6 and 6.3 kb species in poly(A)+RNA and with 2.5, 4 and 9 kb fragments of EcoRI digested human DNA. The frequency of the EcoRI fragments in DNA from females was approximately twice that in DNA from males. DNA from two patients with steroid sulfatase deficiency and X-linked ichthyosis did not hybridize with STS-3. DNA from a third patient showed a normal hybridization pattern. It is concluded that steroid sulfatase deficiency is a genetically heterogenous disorder.

Amino Acid Sequence↗
PubMed1998

Microsomal steroid sulfatase: interactions with cytosolic steroid sulfotransferases.

Net sulfation of 4-methylumbelliferone in intact hepatocytes is regulated, in part, by substrate cycling between sulfotransferases (SULT) and arylsulfatases (ARS). Thus, ARS have the potential to influence rates of net sulfate conjugation of a variety of compounds in intact cells via interaction with SULT. Unlike ARSA and ARSB, which are lysosomal, steroid sulfate sulfatase (ARSC, also known as STS) is localized exclusively in the endoplasmic reticulum (ER). The present study was designed to assess the existence and extent of substrate cycling between steroids and their sulfate conjugates through ARSC and SULT, and also to initiate studies of the topology of the catalytic site of ARSC in the rat liver ER. Addition of rat liver microsomes to cytosol and 3'-phosphoadenosine 5'-phosphosulfate (PAPS) reduced rates of sulfation of dehydroepiandrosterone (DHEA) by SULT, and similarly hydrolysis of DHEA sulfate (DHEAS) was reduced when recombinant human hydroxysteroid SULT was added to rat liver microsomes in the presence of PAPS. There was no evidence for ARSC latency in the presence of detergent at either 4 or 37 degrees C, indicating that facilitated transport of steroid sulfates across the ER membrane may not be required for ARSC activity. The effect of proteases on ARSC activity in intact and disrupted microsomes was determined and compared with effects on components of the glucose-6-phosphatase system known to be localized on the lumenal and cytoplasmic surfaces of the ER. In contrast to the components of the glucose-6-phosphatase system, activity of ARSC in both intact and disrupted microsomes was substantially more resistant to protease inactivation. Our results indicate that substrate cycling of steroids and their sulfates does occur, and suggest that the active site of ARSC may be located within the ER membrane.

Animals↗