Combined effects of testosterone and estradiol on the ventral lobe of the rat postate in organ culture]

After organ culture without hormone, the epithelial gland cells of Rat veantral prostate undergo atrophic changes, whereas the interstitial stroma components tend to increase. Estradiol (1-1,000 nM),added to the culture medium, is ineffective. On the contrary, testosterone (1-100 nM) maintains epithelial cells and prevents the increase of interstitial stroma. When estradiol (1-1,000 nM) is combined with a physiological concentration of testosterone (1-4 nM), the epithelial cells are well maintained, but the inhibitory action of testosterone on the stroma is counteracted so that the glandular epithelium and the interstitial stroma are both stimulated. However, when testosterone is used at supraphysiological (10-100 nM) concentrations, estradiol is completely ineffective and the structure of the prostate is identical to the one given by the androgen alone.     

Binding of estradiol to synaptosomal mitochondria: physiological significance

The subsynaptosomal distribution and specific binding of 17beta-estradiol in vitro to mitochondria isolated from presynaptic nerve endings of female rat brain were examined. 17Beta-estradiol is (i) distributed unequally in synaptosomes and mitochondria posses the highest capacity to bind estradiol with respect to the available amount of the hormone. (ii) Estradiol binds specifically to isolated synaptosomal mitochondria. A Michaelis-Menten plot of specific binding was sigmoidal within a concentration range of 0.1-5 nM of added estradiol, with a saturation plateau at 3 nM. Binding of higher estradiol concentrations demonstrated an exponential Michaelis-Menten plot, indicating non-specific binding to mitochondria. Vmax and Km for the sigmoidal-shape range were estimated as 46 +/- 6 fmol of estradiol/mg of mitochondrial proteins and 0.46 +/- 0.07 nM free estradiol respectively. (iii) Estradiol binding is not affected by the removal of ovaries. The results show that inhibition of Na-dependent Ca2+ efflux from mitochondria by estradiol occurs according to an affinity change of the translocator for Na+, at the same estradiol concentrations that show specific binding to mitochondrial membranes. These data imply that physiological concentrations of estradiol, acting on mitochondrial membrane properties, extragenomically modulate the mitochondrial, and consequently the synaptosomal content of Ca2+, and in that way exert a significant change in nerve cell homeostasis.     

Fatty acid modulation of antiestrogen action and antiestrogen-binding protein in cultured lymphoid cells

Nonsteroidal antiestrogens reversibly and specifically inhibited the proliferation of two estrogen receptor-negative lymphoid cell lines (EL4 and Raji) in a dose-dependent manner. [3H]Thymidine incorporation of concanavalin A-stimulated primary splenocytes was also inhibited by 10(-6) M clomiphene (1-[4-(2-diethylaminoethoxy)phenyl]-1,2-diphenyl-2-chloroethylene). The antiproliferative effect could be prevented by the simultaneous presence in the growth medium of 10(-5) M linoleic acid or 10(-5) M arachidonic acid but not by 10(-6) M estradiol. Both lymphoid cell lines had high affinity antiestrogen-binding sites whose affinity could be altered by conditions of growth. Growth of EL4 cells in RPMI 1640 medium supplemented with charcoal-pretreated 5% fetal calf serum (charcoal-stripped medium) resulted in significantly higher affinity (Kd 0.54 nM +/- 0.11 nM; n = 6) than growth in medium supplemented with untreated serum (complete medium) (Kd = 1.68 nM +/- 0.48 nM; n = 6) (p less than 0.001). This change in affinity was partly due to removal of fatty acids from the growth medium by charcoal pretreatment, since addition of 10(-5) M linoleic acid or 10(-5) M gamma-linolenic to charcoal-stripped medium decreased the affinity of the antiestrogen-binding protein. In contrast, growth in 10(-5) M stearic acid or 10(-5) M oleic acid did not significantly alter the affinity of the antiestrogen-binding protein, whereas 10(-5) M palmitic acid significantly increased its affinity. The same fatty acids were also tested for their intrinsic effects on EL4 cell proliferation. Oleic, linoleic and gamma-linolenic acids were growth stimulatory while stearic and palmitic acids were not. Thus linoleic and gamma-linolenic acids whose presence in the growth medium was associated with decreased affinity of [3H]tamoxifen (1-[4-(2-dimethylaminoethoxy)phenyl]-1,2-diphenylbut-1(Z)-ene) binding to the intracellular antiestrogen-binding protein were also growth stimulatory. Unsaturated fatty acids have previously been shown to inhibit binding of [3H]tamoxifen to the antiestrogen-binding protein in a cell-free system. The present observations demonstrate that unsaturated fatty acids also modify the affinity of the antiestrogen-binding protein in intact cells.     

Aging and endothelin-1 induced vascular contractions

Summary Contractions produced by endothelin-1 (0.3–30 nM) have been investigated in aorta, renal arteries and mesenteric arteries from 2- and 24-month-old Sprague-Dawley rats. In senescent rats the EC₅₀ values of endothelin-1 for aorta and renal artery were significantly increased (aorta: from 6.2 to 12 nM; renal artery: from 5.2 to 7.8 nM). For mesenteric artery the EC₅₀ value (4.3 nM) was unchanged by aging, whereas the maximal contractile response to endothelin-1 was enhanced (from 8.3 to 11.7 mN). In contrast, there was no significant age-related difference in the maximal endothelin-1 response of aorta and renal artery. The present data demonstrate a reduced sensitivity for aorta and renal artery and an enhanced maximal response to endothelin-1 in the mesenteric artery in senescent rats.     

Aging and endothelin-1 induced vascular contractions

Contractions produced by endothelin-1 (0.3-30 nM) have been investigated in aorta, renal arteries and mesenteric arteries from 2- and 24-month-old Sprague-Dawley rats. In senescent rats the EC50 values of endothelin-1 for aorta and renal artery were significantly increased (aorta: from 6.2 to 12 nM; renal artery: from 5.2 to 7.8 nM). For mesenteric artery the EC50 value (4.3 nM) was unchanged by aging, whereas the maximal contractile response to endothelin-1 was enhanced (from 8.3 to 11.7 mN). In contrast, there was no significant age-related difference in the maximal endothelin-1 response of aorta and renal artery. The present data demonstrate a reduced sensitivity for aorta and renal artery and an enhanced maximal response to endothelin-1 in the mesenteric artery in senescent rats.     

Fatty acid modulation of antiestrogen action and antiestrogen-binding protein in cultured lymphoid cells

Summary Nonsteroidal antiestrogens reversibly and specifically inhibited the proliferation of two estrogen receptornegative lymphoid cell lines (EL4 and Raji) in a dose-dependent manner. [³H]Thymidine incorporation of concanavalin A-stimulated primary splenocytes was also inhibited by 10^–6 M clomiphene (1-[4-(2-diethylaminoethoxy)phenyl]-1,2-diphenyl-2-chloroethylene). The antiproliferative effect could be prevented by the simultaneous presence in the growth medium of 10^–5 M linoleic acid or 10^–5 M arachidonic acid but not by 10^–6 M estradiol. Both lymphoid cell lines had high affinity antiestrogen-binding sites whose affinity could be altered by conditions of growth. Growth of EL4 cells in RPMI 1640 medium supplemented with charcoal-pretreated 5% fetal calf serum (charcoal-stripped medium) resulted in significantly higher affinity (K_d 0.54 nM±0.11 nM; n=6) than growth in medium supplemented with untreated serum (complete medium) (K_d=1.68 nM±0.48 nM; n=6) (p<0.001). This change in affinity was partly due to removal of fatty acids from the growth medium by charcoal pretreatment, since addition of 10^–5 M linoleic acid or 10^–5 M gamma-linolenic to charcoal-stripped medium decreased the affinity of the antiestrogen-binding protein. In contrast, growth in 10^–5 M stearic acid or 10^–5 M oleic acid did not significantly alter the affinity of the antiestrogen-binding protein, whereas 10^–5 M palmitic acid significantly increased its affinity. The same fatty acids were also tested for their intrinsic effects on EL4 cell proliferation. Oleic, linoleic and gamma-linolenic acids were growth stimulatory while stearic and palmitic acids were not. Thus linoleic and gamma-linolenic acids whose presence in the growth medium was associated with decreased affinity of [³H]tamoxifen (1-[4-(2-dimethylaminoethoxy)phenyl]-1,2-diphenylbut-1(Z)-ene) binding to the intracellular antiestrogen-binding protein were also growth stimulatory. Unsaturated fatty acids have previously been shown to inhibit binding of [³H]tamoxifen to the antiestrogen-binding protein in a cell-free system. The present observations demonstrate that unsaturated fatty acids also modify the affinity of the antiestrogen-binding protein in intact cells.     

Thrombospondin co-localises with TGFβ and IGF-I in the extracellular matrix of human osteoblast-like cells and is modulated by 17β estradiol

Thrombospondin (TSP) is a multifunctional glycoprotein which is synthesised by several cell types including osteoblasts, and incorporated into the extracellular matrix (ECM) of these cells. The function and regulation of TSP in bone is not clear. In this study, using a long term culture model of human osteoblast-like cells, we examined the distribution of TSP in the ECM and its modulation by added estradiol. In this model the osteoblast-like cells form a regular multilayer which continues to increase in depth up to 50 days post confluence. In the ECM of these cultures and in 19-week fetal bone, the bone markers osteocalcin and alkaline phosphatase were diffusely distributed in the matrix. In contrast, labelling for TSP was concentrated, confined to the banded collagen and its immediately adjacent ECM. This pattern of labelling resembled that of the growth factors transforming growth factor-I (TGF), and insulin-like growth factor-I (IGF-I), with which TSP label co-localised. Labelling intensities were comparable between fetal bone and the in vitro material for TSP, TGF and IGF-I. TSP label was present by 10 days post confluence, reached a maximum by 20 days, and declined slowly thereafter, a time course which was similar to that of IGF-I. Incubation of osteoblast-like cell cultures with 17 estradiol resulted in an increase in multilayer depth and a maximal 3-fold increase in TSP labeling at 30 days as well as approximately 2-fold increases for TGF and IGF-I. The dose-response relationship for these responses to estradiol treatment was biphasic with maximal increases at 10^–10 M–10^–11 M of added estradiol. Treatment with 17 estradiol produced labelling intensities that were not significantly different from controls. Studies with other cell types have suggested that TSP may be involved in modulation of growth factor activity. The similarities between TSP, TGF and IGF-I, in terms of their distribution and regulation by 17 estradiol treatment, may indicate a role for TSP in modulating bone cell proliferation and function through interaction with local growth factors.     

Effect of sex hormones on plasma cholesterol in castrated and noncastrated male rats

Summary The administration of estradiol to both castrated and noncastrated male rats was associated with significantly increased plasma cholesterol levels as compared to controls, the estradiol in the noncastrated rats overriding the tendency of testosterone to lower plasma cholesterol.Acknowledgement. This work was supported by USPHS Grant NHLBI 17721.     

Peptide-stimulated release of prolactin from the fowl anterior pituitary gland

Summary Anterior pituitary glands from broiler fowl were preincubated for 24 h in either medium 199 only or medium containing estradiol 17, following which they were incubated in medium containing thyrotrophin releasing hormone (TRH), vasoactive intestinal polypeptide (VIP) or substance P (SP), alone or with the dopamine agonist, apomorphine. Estradiol priming stimulated release of prolactin and enhanced apomorphine-inhibition of prolactin release. TRH stimulated prolactin release, an effect reversed by apomorphine, and priming with estradiol potentiated both effects. VIP stimulated prolactin to a lesser degree and again this was inhibited by apomorphine and potentiated by estradiol. SP had little effect on the nonsteroid-primed pituitary, but stimulated release of prolactin after estradiol treatment, though less effectively than TRH or VIP.     

Effect of sex hormones on plasma cholesterol in castrated and noncastrated male rats

The administration of estradiol to both castrated and noncastrated male rats was associated with significantly increased plasma cholesterol levels as compared to controls, the estradiol in the noncastrated rats overriding the tendency of testosterone to lower plasma cholesterol.     

Plasma steroid and protein hormone concentrations in patients with benign prostatic hypertrophy and in normal men.

Serum levels of testosterone, FSH, LH and prolactin were measured by RIA in patients with BPH (benign prostatic hypertrophy) and in control men. The testosterone concentrations in the BPH group were significantly lower than that of the control group. Serum concentrations of FSH, LH, prolactin and estradiol, did not differ significantly in the 2 groups.     

Plasma steroid and protein hormone concentrations in patients with benign prostatic hypertrophy and in normal men

Summary Serum levels of testosterone, FSH, LH and prolactin were measured by RIA in patients with BPH (benign prostatic hypertrophy) and in control men. The testosterone concentrations in the BPH group were significantly lower than that of the control group. Serum concentrations of FSH, LH, prolactin and estradiol, did not differ significantly in the 2 groups.     

Inhibition of interleukin-1 beta release from cultured human peripheral blood mononuclear cells by prednisolone

Prednisolone, a water-soluble glucocorticoid hormone, suppressed the secretion of interleukin-1 beta from human peripheral blood mononuclear cells in culture. The prednisolone-induced suppression of the monokine release was dose-related and the half maximal response was observed at 0.1 nM.     

Reduced tuberoinfundibular dopaminergic neuronal function in rats after long-term withdrawal of estrogen treatment

Summary Hypothalamic fragments from female rats treated repeatedly with estradiol valerate (EV) and bearing prolactin (PRL)-secreting tumors contained, seven months after the last EV injection, lower concentrations of dopamine (DA) than age-matched controls. Depolarizing concentrations of K⁺ (35 mM) and amphetamine (50 M) evoked in PRL-secreting tumor bearing rats an endogenous DA release significantly lower than in controls.     

Characterization of the "4 S-trypsin" form of the cytosoluble estradiol receptor of calf uterus in a nondenaturing system]

The characterisation of the "4 S-trypsin" form of the estradiol-receptor from calf uterus cytosol was carried out by polyacrylamide gel electrophoresis under non-denaturing conditions. In a multiphasic buffer system (upper buffer Tris-glycine pH25degrees C = 8,6, lower buffer Tris HCl pH25degrees C = 7,4), three radioactive estradiol peaks were observed. The first was free estradiol, the second estradiol initially complexed with a macromolecule and dissociated during electrophoresis, and the third the hormone-receptor complex. The sedimentation coefficient (4 S) of this complex was the same before and after electrophoresis. Its mean geometric radius is R = 2.53 + 0.08 nm and its molecular weight was estimated to be 55,000.     

Effect of various doses of catecholestrogens on uterine eosinophilia in the immature rat

Summary This paper describes the induction of uterine eosinophilia as well as of deep endometrial edema and increase of uterine wet weight in the immature rat by the catecholestrogens 2-OH-estradiol and 4-OH-estradiol. These effects are thought to be mediated by eosinophils via a specific eosinophil receptor system. 4-OH-estradiol was equipotent with estradiol, whereas the effect of 2-OH-estradiol was significantly weaker.Acknowledgments. This work was supported by Grant B-1493-8435 from the University of Chile. We are indebted to Prof. R. Knuppen, Lübeck, for providing the CE.     

Demonstration of high affinity binding of estradiol in adrenal cortex tissue]

Evidence is presented demonstrating the presence of a high affinity (Ka10(8)M-1), limited capacity (3-4 pmoles/mg protein) estradiol binder in the soluble fraction of the Bovine, Rat and Human adrenal cortex. The binding appears specific to the estrogen structure whereas C19 and C21 steroids do not bind. Upon sucrose density gradient centrifugation, the estradiol binder sedimented at 9 S at low ionic strength and was shifted to 4.5 S in the presence of 0.5 M KCl. This demonstration of a receptor-like moiety for estradiol in the adrenal cortex lends biochemical support to previous observations suggesting that adrenal cortex functions may be modulated by a direct effect of gonadal steroid hormones.     

Variations of non conjugated steroids in the amniotic fluid during the last weeks of pregnancy]

Levels of cortisol, estradiol and progesterone were measured in 172 amniotic fluid samples collected by amniocentesis during late pregnancy. An increase in cortisol and estradiol levels was observed three weeks before vaginal delivery of term and premature infants; in both types of delivery, onset of labor was spontaneous. In contrast progesterone levels were only slightly decreased in these cases.     

Effect of various doses of catecholestrogens on uterine eosinophilia in the immature rat

This paper describes the induction of uterine eosinophilia as well as of deep endometrial edema and increase of uterine wet weight in the immature rat by the catecholestrogens 2-OH-estradiol and 4-OH-estradiol. These effects are thought to be mediated by eosinophils via a specific eosinophil receptor system. 4-OH-estradiol was equipotent with estradiol, whereas the effect of 2-OH-estradiol was significantly weaker.     

Chemical sympathectomy reveals pre-and postsynaptic effects of neuropeptide Y (NPY) in the cardiovascular system

Summary Intravenous injection of neuropeptide Y (NPY) caused short-lasting dose-dependent pressor responses in anesthetized rats. NPY was equipotent with noradrenaline in producing proportional pressor effects. Chemical sympathectomy, following the administration of 100 mg/kg 6-hydroxydopamine (6-OHDA), significantly potentiated the systemic pressor effects elicited by NPY or noradrenaline. Pretreatment with 2 nmol NPY enhanced the noradrenaline-induced pressor response in control rats. NPY did not change the basal tension of isolated rat aortic strips but significantly potentiated the contractile activity induced by 16 nM noradrenaline. This effect of NPY was not observed in aortic strips from rats pretreated with 6-OHDA. The presence of pre-and postsynaptic sites of action for NPY in the cardiovascular system of the rat is discussed.