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R B Hochberg

Publications and source records attributed to R B Hochberg.

At least 37 records · Page 2Linked to original sources
PubMed1994

Androgen treatment decreases estrogen receptor binding in the ventromedial nucleus of the rat brain: a quantitative in vitro autoradiographic analysis.

Androgens oppose the actions of estrogen on a number of neuroendocrine functions in the rat including prolactin and gonadotropin secretion and the activation of the female pattern of sex behavior. Although in nonneural tissues antiestrogenic actions of androgens have been related to actions at the level of the estrogen receptor, previous attempts to demonstrate effects of nonaromatizable androgens on estrogen receptor levels in the brain have been unsuccessful, possibly because of the poor anatomical resolution of the methods used. We have used a new in vitro autoradiographic assay combined with an 125I-labeled estrogen receptor ligand to test the hypothesis that the nonaromatizable androgen, 5 alpha-dihydrotestosterone (5 alpha-DHT), may act to reduce estrogen binding in specific regions of the brain involved in reproductive neuroendocrine and behavioral responses. This in vitro autoradiographic method allows selective measurement of occupied estrogen receptors in tissue sections. Gonadectomized/adrenalectomized rats were divided into two groups per sex. All animals received daily injections of estradiol benzoate (EB: 40 micrograms/kg body wt) for 4 days. Animals in the 5 alpha-DHT treatment group received 5 alpha-DHT (10 mg/kg body wt) every 12 h for 4 days, while animals in the control group received vehicle injections. Animals were killed 4 h after the final EB/5 alpha-DHT injection and their brains processed for in vitro autoradiography. As previously reported, higher levels of estrogen binding were observed in the ventrolateral aspect of the ventromedial nucleus (vIVMN) and the periventricular and medial preoptic area of the female compared to the male.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenalectomy↗
PubMed1994

Partial demasculinization and feminization of sex behavior in male rats by in utero and lactational exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin is not associated with alterations in estrogen receptor binding or volumes of sexually differentiated brain nuclei.

In utero and lactational 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure partially demasculinizes and feminizes sexual behavior in adult male rats, presumably by causing incomplete sexual differentiation of the central nervous system (CNS). Our objective was to determine if TCDD exposure affects other aspects of sexual differentiation of the CNS. Because sex differences in the estrogen receptor system are thought to play a role in sexually dimorphic estrogen-mediated responses, and because estrogen is an important activator of both male and female sex behavior, the possible effect of TCDD exposure on estrogen binding in specific brain nuclei was examined. In addition, we investigated effects of in utero and lactational TCDD exposure on sex differences in the volumes of brain nuclei which are dependent on steroid hormone stimulation during the period of CNS sexual differentiation. Pregnant Holtzman rats were given TCDD (0.7 microgram/kg, po) or vehicle (control) on gestation Day 15. Offspring were exposed to TCDD in utero and via lactation and then assessed in adulthood. Demasculinized sexual behavior was evidenced in the TCDD-exposed males by increased intromission latencies and a greater number of intromissions prior to ejaculation. These males were then castrated, primed with ovarian steroids, and tested for feminine sexual behavior. In utero and lactational TCDD exposure increased both the frequency and intensity of lordotic behavior, indicating that the males were partially feminized. To determine if TCDD exposure had a generalized effect on estrogen receptor concentrations the arcuate nucleus, cortical and medical amygdala and the bed nucleus of the stria terminalis, previously found to have equivalent numbers of estrogen receptors in males and females, were evaluated in littermates of the rats whose sexual behavior had been assessed. TCDD had no effect in either sex. To determine if TCDD exposure had an effect specific to sexual differentiation of the brain, estrogen receptor concentrations in the medial preoptic nucleus (MPO), ventrolateral aspect of the ventro-medial nucleus, and periventricular preoptic area were assessed. As expected, females had higher estrogen receptor concentrations in these nuclei than did males, but TCDD exposure did not affect estrogen receptor concentrations in any of these sexually dimorphic brain nuclei. The volumes of sexually dimorphic brain nuclei were examined in additional littermates. In control rats, the volume of the sexually dimorphic nucleus of the preoptic area (SDN-POA) is larger in males than in females whereas the MPO is larger in females than in males. TCDD exposure had no effect on the volume of either the SDN-POA or MPO in either males or females.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1993

Rapid stimulation of rhodamine 123 efflux from multidrug-resistant KB cells by progesterone.

Rhodamine 123 is a mitochondrial dye that is retained for prolonged periods by carcinoma cells. While investigating causes of retention of this dye, we found that 10 microM progesterone caused a rapid stimulation of efflux of rhodamine 123 within 15 min from KB V20C cells, which overexpress the multidrug resistance pump. Progesterone did not stimulate efflux from KB cells that do not overexpress the pump, and verapamil blocked rhodamine 123 efflux in the presence or absence of progesterone, indicating that rhodamine 123 is removed from KB V20C cells by the multidrug resistance pump. Progesterone, however, is unlikely to stimulate rhodamine 123 efflux by simply increasing pump activity for two reasons: (1) progesterone inhibited the efflux of daunomycin from KB V20C cells, so it did not stimulate efflux of all drugs, and (2) progesterone inhibited efflux of rhodamine 123 from L1210/VMDRC cells and had little effect on Adr MCF7 cells; both overexpress the multidrug resistance pump. In the experiments with KB V20C cells, progesterone was the most active steroid tested. At 10 microM, progesterone caused a 70-fold stimulation, desoxycorticosterone, testosterone, promegestone and estradiol about 20-fold, and others had little or no effect. Progesterone may act by a non-genomic mechanism to decrease intracellular binding of rhodamine 123, making the dye accessible to the multidrug resistance pump.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1993

The isolation and characterization of estradiol-fatty acid esters in human ovarian follicular fluid. Identification of an endogenous long-lived and potent family of estrogens.

The estradiol-fatty acid esters are highly potent and long-lived estrogens that were first isolated and identified from in vitro biosynthetic experiments (Mellon-Nussbaum S., Ponticorvo, L., Schatz, F., and Hochberg, R. B. (1982) J. Biol. Chem. 257, 5678-5684). Other studies have indicated that these esters exist endogenously, but the evidence is indirect, resting solely on the presence of a nonpolar saponifiable metabolite. Although there are similar reports of other naturally occurring fatty acid esters of biologically active steroid hormones, likewise, none has been isolated and characterized. In this study we have found that follicular fluid from the ovaries of women stimulated with gonadotrophins contains relatively large amounts of a nonpolar saponifiable derivative of estradiol (approximately 10(-7)M), which we presumed to be fatty acid esters. Using a combination of chromatographic techniques we isolated these estradiol metabolites and identified them by mass spectral analysis. They are a mixture of five different estradiol-17 fatty acid esters. The amount of each present was determined by separating the esters by reversed phase high performance liquid chromatography and then quantifying each peak by radioimmunoassay and UV absorption. The esters of estradiol are predominantly unsaturated, with linoleate the most abundant comprising 43% of the total. The other esters are: palmitate (20%), arachidonate (19%), oleate (14%), and stearate (4%). Thus these studies conclusively demonstrate the existence of these unusually powerful estrogens.

Arachidonic Acid↗
PubMed1993

7 alpha-Methyl-17 alpha-(E-2'-[125I]iodovinyl)-19-nortestosterone: a new radioligand for the detection of androgen receptor.

We have synthesized two gamma-emitting, 125I-labeled steroids, E- and Z-7 alpha-methyl-17 alpha-(2'-[125I]iodovinyl)-19-nortestosterone [125I](E- and Z-MIVNT) for specific labeling of androgen receptors. [125I]E- and [125I]Z-MIVNT were synthesized stereospecifically from E- and Z-7 alpha-methyl-17 alpha-(2'-tri-n-butylstannyl-vinyl)-19-nortestosterone. The tin adducts were prepared by addition of tri-n-butyltin hydride to 7 alpha-methyl-17 alpha-ethynyl-19-nortestosterone, and after purification they were converted in high yield to the [125I]MIVNT isomers by reaction with 125I (generated in situ by oxidation of [125I]iodide with chloramine T). The 125I-labeled products were purified by high-performance liquid chromatography, and their mass determined with an ultraviolet detector (specific activity of both, approximately 2,200 Ci/mmol). In rat prostate cytosol, [125I]E-MIVNT bound with high affinity to a single class of binding sites. Nonspecific binding in the presence of 5 alpha-dihydrotestosterone was relatively low, and compared favorably with that obtained in parallel studies with [3H]methyltrienolone (R1881). The E-isomer bound prostate cytosol with at least twice the affinity of the Z-isomer; therefore, the interaction of the E-isomer with the androgen receptor as well as other steroid receptors was studied in greater detail. Complexes of the androgen receptor with [125I]E-MIVNT as well as [3H]R1881 dissociate very slowly at 4C (kdiss for both = 0.04 h-1). Displacement studies showed that the interaction of [125I]E-MIVNT with the androgen receptor is highly specific. Competition studies showed that unlabeled E-MIVNT binds poorly to other steroid receptors in rat tissue cytosols. These binding properties make [125I]E-MIVNT a promising ligand for study of the androgen receptor, and [123I]E-MIVNT a potential imaging agent for the detection of androgen-dependent tumors, such as prostate cancer.

Animals↗
PubMed1993

In vitro autoradiographic visualization of occupied estrogen receptors in the rat brain with an iodinated estrogen ligand.

Methods have been developed for the selective measurement of occupied estrogen receptors (ER) in brain tissue sections. Cryostat sections of unfixed tissue were incubated with radiolabeled estrogen at physiological temperatures, displacing endogenous receptor-bound estrogen by radioligand and thereby allowing the receptor complexes to be visualized autoradiographically after washing to remove nonspecifically bound steroid. The resultant autoradiographs were analyzed by computer-assisted densitometry. Synthetic 11 beta-methoxy-substituted radiolabeled estrogens gave the best autoradiographic images, as a result of reduced nonspecific labeling, although [3H]-estradiol was also used successfully. With the synthetic ER ligand 11 beta-methoxy 16 alpha-[125I]-iodo-estradiol, exposure times of less than 24 hr generated acceptable autoradiographs; with 3H-labeled estrogens, exposures of 3 months or more may be required. The method is sufficiently sensitive to detect physiological changes in ER occupation and to allow determination of receptor affinities and saturation binding capacities in discrete cell groups identified in sections from individual animals.

Animals↗
PubMed1992

Estrogen receptors colocalize with low-affinity nerve growth factor receptors in cholinergic neurons of the basal forebrain.

The rodent and primate basal forebrain is a target of a family of endogenous peptide signaling molecules, the neurotrophins--nerve growth factor, brain-derived neurotrophic factor, and neurotrophin 3--and of the gonadal steroid hormone estrogen, both of which have been implicated in cholinergic function. To investigate whether or not these ligands may act on the same neurons in the developing and adult rodent basal forebrain, we combined autoradiography with 125I-labeled estrogen and either nonisotopic in situ hybridization histochemistry or immunohistochemistry. We now report colocalization of intranuclear estrogen binding sites with the mRNA and immunoreactive protein for the low-affinity nerve growth factor receptor, which binds all three neurotrophins, and for the cholinergic marker enzyme choline acetyltransferase (acetyl-CoA:choline O-acetyltransferase, EC 2.3.1.6). Colocalization of estrogen and low-affinity nerve growth factor receptors implies that their ligands may act on the same neuron, perhaps synergistically, to regulate the expression of specific genes or gene networks that may influence neuronal survival, differentiation, regeneration, and plasticity. That cholinergic neurons in brain regions subserving cognitive functions may be regulated not only by the neurotrophins but also by estrogen may have considerable relevance for the development and maintenance of neural substrates of cognition. If estrogen-neurotrophin interactions are important for survival of target neurons, then clinical conditions associated with estrogen deficiency could contribute to the atrophy or death of these neurons. These findings have implications for the subsequent decline in those differentiated neural functions associated with aging and Alzheimer disease.

Animals↗
PubMed1992

Cellular variations in estrogen receptor mRNA translation in the developing brain: evidence from combined [125I]estrogen autoradiography and non-isotopic in situ hybridization histochemistry.

The spatial distribution of cells in the adult rodent forebrain which express estrogen receptor mRNA, as shown by in situ hybridization histochemistry with isotopically-labeled probes, has been reported to overlap with regions that are known targets of estrogen and which bind estrogen. The extent to which detection of estrogen receptor mRNA within developing forebrain neurons of the postnatal day 10-12 female rat is accompanied by translation into estrogen binding sites was investigated by combining [125I]estrogen autoradiography with non-isotopic (digoxigenin) in situ hybridization, using a 48-base oligodeoxyribonucleotide probe encoding a sequence of the estrogen-binding domain of rat uterine estrogen receptor cDNA. Estrogen receptor mRNA and estrogen binding sites appeared to be restricted to neurons. No mRNA or binding was seen in ependymal cells. Cells expressing estrogen receptor mRNA were widely distributed in the developing rat forebrain and were found in brain regions generally corresponding to those previously shown in the adult, with the addition of some regions not previously described, such as the medial habenula and dorsal endopiriform nucleus. Although there was widespread overlapping of estrogen receptor mRNA expression with known estrogen binding sites, there were regional and cellular variations in the extent of receptor mRNA translation. This pattern was true for developing forebrain regions previously defined as estrogen receptor-containing (hypothalamus, preoptic area, medial and lateral septum, vertical and horizontal nuclei of the diagonal band, cerebral cortex, hippocampus and amygdala) as well as for regions heretofore not considered estrogen targets (the thalamus, dorsal endopiriform nucleus, claustrum, ventral pallidum/substantia innominata and the basal nucleus of Meynert) or characterized as estrogen-responsive in the adult without previously documented estrogen binding [caudate-putamen (striatum)]. While estrogen binding and receptor mRNA expression always co-localized, neurons expressing estrogen receptor mRNA did not always exhibit ligand binding and there was no clear-cut relationship between the intensity of the hybridization signal and estrogen binding. Little, however, is known about translational control of estrogen receptor expression in the brain. Localization of estrogen binding sites to regions not generally considered targets of estrogen would appear to reflect the greater sensitivity of the iodinated ligand than the tritiated estrogens more commonly used for autoradiography. Non-isotopic in situ hybridization histochemistry combined with [125I]estrogen autoradiography represents a very powerful tool with which to study regulation of estrogen receptor gene expression at the single cell level with an exceptional degree of cellular and anatomical resolution.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1992

Aromatase and testosterone fatty acid esters: the search for a cryptic biosynthetic pathway to estradiol esters.

The estradiol fatty acid esters (lipoidal derivatives, LE2) are extremely potent estrogens that accumulate in fat, including fat of menopausal women. These steroidal esters are protected from metabolism and are converted to the free, biologically active steroid through the action of esterases. Previous studies have shown that biosynthetic pathways in the adrenal gland exist in which steroid fatty acid esters are substrates. This led us to determine whether a cryptic aromatase pathway exists in which testosterone esters could be converted directly into LE2. We tested a representative fatty acid ester, testosterone stearate, both as an inhibitor and as a substrate for the aromatase enzyme from human placental microsomes. This ester had neither activity. In addition, we tested [1 beta-3H]testosterone acetate as a substrate for this enzyme complex, measuring the production of 3H2O as evidence of aromatization. Although the rate of reaction was considerably slower than that of testosterone, 3H2O was produced. However, when [2, 4, 6, 7-3H]testosterone acetate was incubated and the steroidal products isolated, we found that hydrolysis of the substrate had occurred. Both [3H]-labeled testosterone and estradiol were found, and very little if any [3H]estradiol acetate was formed. Thus, we conclude that an aromatase pathway involving testosterone esters does not exist and that the sole source of LE2 is through direct esterification of estradiol.

Aromatase↗
PubMed1992

Intrinsic estrogenicity of some progestagenic drugs.

The intrinsic estrogenic activity of some progestins cannot be properly evaluated by using hormone responsive systems when the chosen end-points are also sensitive to progestagenic activity, usually antagonistic of estrogenic actions. We have therefore applied to the evaluation of some drugs commonly used in contraceptive and hormone replacement formulations a recently developed in vitro method to estimate estrogenic activities, which is based on measurements of the estrogen-stimulated alkaline phosphatase activity in cells of the Ishikawa-Var I human endometrial adenocarcinoma line, a response not influenced by progestins. Whereas progesterone, medroxyprogesterone acetate and danazol were found to be devoid of estrogenic activity in this assay, Org OD-14, norethynodrel, gestrinone (R 2323), norethindrone and dl-norgestrel provoked half maximal increases in alkaline phosphatase activity at concentrations (EC-50) of 7, 14, 140, 200 and 2900 nM, respectively, under conditions in which the corresponding value for estradiol was 8 pM. This intrinsic estrogenic activity can be inhibited by antiestrogens, as verified by reversing the effect of R 2323 with 4-hydroxytamoxifen. Since prostaglandin F2 alpha output by secretory endometrium is increased by estrogens and diminished by progestins, this end-point can serve to evaluate the net effect of drugs with intrinsic estrogenic and progestagenic activities. For instance, R 2323 showed estrogenic activity in this assay whereas Org OD-14 did not. The same in vitro system can be used to evaluate estrogen antagonistic activities of test compounds, using estradiol as the agonist. These in vitro systems are useful in establishing a profile of activities of a drug on a relevant human target tissue, in the screening of synthetic or natural compounds under investigation, and in studies on structure/action relationships.

Adenocarcinoma↗
PubMed1992

Measurement of estradiol-17-fatty acid esters in human tissues.

We have developed a gas chromatographic/mass spectral method for the sensitive and reproducible measurement of estradiol-17-fatty acid esters in human tissues and blood. To provide an internal standard for quantification, a trideuterated analog of a representative estradiol ester is added to the tissues. Estradiol (E2) released from the nonpolar ester fraction by alkaline hydrolysis is derivatized to form the ditrimethylsilyl ether and then analyzed by gas chromatographic/mass spectral, monitoring the molecular ions mass per U charge of the ditrimethylsilyl derivative of E2 and [2H3]E2. There are low but detectable levels of E2 ester in the blood of cycling females; there are none in urine. While the E2 ester is present in breast cyst fluid, its concentration, 77-140 pmol/L, is considerably less than E2, 110-2,863 pmol/L. But there is a large amount of E2 ester in fat. In premenopausal women the average E2 ester in fat (sc and omental) is 957 +/- 283 38 fmol/g (SEM); in women who are menopausal less than 12 yr, the E2 ester in fat is 669 +/- 158 fmol/g; in women who are menopausal at least 15 yr, the fat level is 399 +/- 146 fmol/g. Muscle from the same women have lower concentrations of the ester; in 8 out of 12 muscle specimens it was not detectable. The E2 esters are extremely potent estrogens. Although they are hormonally active they require enzymatic hydrolysis to exert their hormonal action. These studies show that these long chain esters of E2 are sequestered in fatty tissues, wherein they represent a protected store of preformed hormone. Under the proper stimulation, adipose tissue can activate the estrogenic signal through the action of hormonally sensitive esterases. Thus, through signaling between estrogen sensitive tissues and neighboring fat cells, a local paracrine loop may exist.

Adipose Tissue↗
PubMed1991

Distribution of estrogen target sites in the 2-day-old mouse forebrain and pituitary gland during the 'critical period' of sexual differentiation.

The present study provides a detailed anatomical description of estrogen target cells in the mouse forebrain and pituitary gland during the sexual imprinting stage of the brain. Six 2-day-old mice (3 males and 3 females) were s.c. injected with 16 alpha-[125I]iodo-11 beta-methoxy-17 beta-estradiol ([125I]MIE2) and two additional mice (one male and one female) were s.c. injected with 1000x unlabeled 17 beta-estradiol 1 h before [125I]MIE2 to check the specificity of estradiol binding. Two hours after injection the mice were decapitated, the brains dissected, frozen sectioned, and processed for thaw mount autoradiography. The highest intensity of nuclear labeling was observed in the preoptic-anterior hypothalamic area, amygdala and cortex entorhinalis. Strong labeling was present in the cerebral cortex and moderate to strong labeling in the lateral septum, bed nucleus of stria terminalis and pituitary gland. Weak to moderate labeling was observed in the bulbus olfactorius, circumventricular organs, basal ganglia, ventral striatum, thalamus, hippocampus and pineal gland. No sex differences were observed in the intensity of labeling and distribution of the estrogen target sites. The topographic distributions of estrogen-concentrating cells in the hypothalamus of the 2-day-old mouse forebrain was similar to the adult pattern but differed prominently in the cerebral cortex, entorhinal cortex and thalamus: the cerebral cortex showed an extensive and intensive labeling, the intensity of labeling in the entorhinal cortex greatly exceeded that observed in the adult and the nucleus anterior medialis thalami was distinctly labeled.

Animals↗
PubMed1991

Estrogen receptor binding in regions of the rat hypothalamus and preoptic area after inhibition of dopamine-beta-hydroxylase.

Previous studies have shown that administration of diethyldithiocarbamate (DDC), a dopamine-beta-hydroxylase inhibitor, results in a decreased concentration of estrogen receptors measured in the rodent hypothalamus and preoptic area. To determine if this modulation of receptor content is region-specific, in vitro estrogen binding assays were performed on cytosol and cell nuclear extracts of microdissected brain regions from female rats treated with DDC. For cytosol binding comparisons, ovariectomized (OVX) rats were treated with 550 mg DDC/kg b. wt. or the saline vehicle 12 h before sacrifice. For cell nuclear binding comparisons, OVX rats received a maximal dose of estradiol 12 h after DDC or saline treatment and 1 h before sacrifice. No region-specific decreases in estrogen binding were observed in either cytosol or nuclear extracts. To further examine possible regional specificity, quantitative autoradiographic analysis of the in vivo hypothalamic uptake of an iodinated analog of estradiol, 11 beta-methoxy-16 alpha-[125I]iodoestradiol (MIE2), in OVX rats treated with DDC was conducted. Animals received a saturating dose of [125I]MIE2 12 h after DDC or saline treatment and 1 h before sacrifice. DDC treatment resulted in higher background levels of radioactivity and a trend toward higher uptake levels in all brain regions, but with no evidence of marked regional specific effects in any area of the brain. In tissue uptake studies, DDC treatment resulted in higher levels of radioactivity recovered from serum and neural tissues of [125I]MIE2-injected rats, suggesting that DDC slows the clearance of MIE2.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1991

The biosynthesis of D-ring fatty acid esters of estriol.

Biological esterification with fatty acids is a feature that is now known to be common to most steroids. The esterification of estradiol in the D-ring at the 17 beta-hydroxyl leads to a family of extremely active estrogens. Similarly, esterification of the weaker estrogen, estriol (E3), has an even greater impact on its hormonal potency. We have recently shown that synthetic long chain esters of E3 at either 16 alpha- or 17 beta- are highly potent estrogens. The estrogenic activity of the synthetic E3 esters led us to determine whether E3 is biologically esterified, and if so, to characterize the resulting esters. Incubation of E3 with rat lung, a tissue which is highly active in esterifying estradiol, produces a nonpolar metabolite which upon saponification is converted back into E3. There was no evidence for the formation of a diester. Purification by high performance liquid chromatography separates the non-polar metabolite into two peaks, one the C-16 alpha- (approximately 60%) and the other the C-17 beta-ester (approximately 40%). The two fractions were further purified and characterized; each is a mixture of fatty acid esters of E3. The composition of the C-16 alpha- and the C-17 beta-fatty acid esters of E3 is identical. The predominant fatty acids are arachidonate, 34%, palmitate, 26%, followed by oleate 14%, linoleate 13%, stearate 8%, and palmitoleate 5%. The similarity of the esters at C-16 and C-17 may indicate that the fatty acid precursor for the acyltransferase is the same for both hydroxyl groups. It may also suggest that the same enzyme esterifies both positions in the D-ring. Since synthetic estriol fatty acid esters are extremely potent and long-lived estrogens, the enzymatic esterification of estriol produces powerful estrogens with considerable physiological potential.

Animals↗
PubMed1991

Steroidal fatty acid esters.

Several years ago we discovered an unexpected family of steroidal metabolites, steroidal fatty acid esters. We found that fatty acid esters of 5-ene-3 beta-hydroxysteroids, pregnenolone and dehydroisoandrosterone are present in the adrenal. Subsequently, others have shown the existence of these non-polar 5-ene-3 beta-hydroxysteroidal esters in blood, brain and ovaries. Currently, almost every family of steroid hormone is known to occur in esterified form. We have studied the esters of the estrogens and glucocorticoids in some detail, and have found that these two steroidal families are esterified by separate enzymes. In a biosynthetic experiment performed simultaneously with estradiol and corticosterone, we established that the fatty acid composition of the steroidal esters is quite different. The corticoid is composed predominantly of one fatty acid, oleate, while the estradiol esters are extremely heterogeneous. Our studies have demonstrated that the estrogens are extremely long-lived hormones, that they are protected by the fatty acid from metabolism. They are extremely potent estrogens, with prolonged activity. Esterification appears to be the only form of metabolism that does not deactivate the biological effects of estradiol. We have demonstrated the biosynthesis of fatty acid esters of estriol, monoesters at both C-16 alpha and C-17 beta. They too are very potent estrogens. These fatty acid esters of the estrogens are the endogenous analogs of estrogen esters, like benzoate, cypionate, etc., which have been used for decades, pharmacologically because of their prolonged therapeutic potency. We have found that the estradiol esters are located predominantly in hydrophobic tissues, such as fat. Sequestered in these tissues, they are an obvious reservoir of estrogenic reserve, requiring only an esterase for activation. To the contrary the biological activity of the fatty acid esters of the glucocorticoid, corticosterone, is not different from that of its free parent steroid. We have shown that the rapid kinetics of its induction of gluconeogenic responses is caused by its labile C-21 ester which is rapidly hydrolyzed by esterase enzymes. While it appears that the physiological role of the estrogen esters may be related to their long-lived hormonal activity, the role of the other families of steroidal esters is not yet apparent. They, and perhaps the estrogen esters as well, must serve other purposes. Indeed they may serve important biological functions beyond those which we ordinarily associate with steroid hormones.

Adrenal Glands↗
PubMed1991

Estrogenic action of estriol fatty acid esters.

Recent studies suggest that, estriol, like estradiol, is biosynthetically esterified with fatty acids. We have synthesized the stearate estriol, at C-16 alpha, C-17 beta and the diester, C-16 alpha,17 beta and tested these D-ring esters for their estrogenic action both in vivo and in vitro, comparing them to estradiol, estriol and estradiol-17-stearate. None of the estriol esters bind to the estrogen receptor. They are only weakly estrogenic in a microtiter plate estrogen bioassay: stimulation of alkaline phosphatase in the Ishikawa endometrial cells. However, both estriol monoesters are extremely potent estrogens when injected subcutaneously (in aqueous alcohol) into ovariectomized mice. Compared to the free steroids, they produced a dramatically increased uterine weight with a greatly prolonged duration of stimulation. The 16 alpha,17 beta-diester also induced a protracted uterotrophic response, but the stimulation of uterine weight was comparatively low. Since the esters of estradiol and estriol do not bind to the estrogen receptor, their estrogenic signal must be generated through the action of esterase enzymes. These naturally occurring esters have the potential of being extremely useful pharmacological agents for long-lived estrogenic stimulation.

Alkaline Phosphatase↗
PubMed1990

The synthesis and testing of E-17 alpha-(2-iodovinyl)-5 alpha-dihydrotestosterone and Z-17 alpha-(2-iodovinyl)-5 alpha-dihydrotestosterone as gamma-emitting ligands for the androgen receptor.

Two iodinated steroids, E-17 alpha-(2-iodovinyl)-5 alpha-dihydrotestosterone and Z-17 alpha-(2-iodovinyl)-5 alpha-dihydrotestosterone were synthesized in a search for a gamma-emitting androgen that binds with high affinity to the androgen receptor. Such compounds would be extremely useful research tools for studies of androgen responsive tissues and as in vivo probes of androgen responsive tumors such as prostate cancer. These 17 alpha-iodovinyl steroids were synthesized because many 17 alpha-substituents do not interfere markedly with binding to the androgen receptor and because similar analogs of other steroids, estrogens and progestins, have been shown to have the requisite properties for ligands to those receptors. Both of these potential ligands were tested for their ability to compete with [3H]R1881 for binding to the androgen receptor in cytosols from prostate, hypothalamus and pituitary. The relative binding affinities ranged between 5 and 20%, depending upon the tissue and steroid. In order to test the two ligands directly, they were both synthesized labelled with 125I and tested for binding to the androgen receptor in prostatic cytosol and in vivo for specific concentration in androgen responsive tissues. While there was considerable binding in the prostatic cytosol, it was not specific because 5 alpha-dihydrotestosterone did not compete. Likewise in the in vivo experiment there was no evidence for androgen receptor mediated concentration of the tracers. While on the basis of relative binding affinity, these 2 steroids appeared to be good candidates for androgen receptor ligands, neither were useful for this purpose. These results contribute new information which will be valuable in the design of other gamma-emitting androgens and emphasises that, in this process, other factors such as metabolism and nonspecific binding must be considered.

Animals↗