Direct Effects Of Steroid Hormones Research Articles

The steroid hormone dydrogesterone inhibits myometrial contraction independently of the progesterone/progesterone receptor pathway

AimsIn this study, we aimed to investigate the direct effects of steroid hormones on pregnant myometrial contraction. Main methodsThe effect of steroids on oxytocin-induced contraction was examined in vitro using pregnant rat or human myometrium. Subsequently, we evaluated whether RU486, a potent progesterone antagonist, influenced the effects of progestin on myometrial contraction. Additionally, we evaluated the effects of progestin on high-concentration KCl-induced contraction caused by voltage-dependent calcium channels in order to investigate the mechanisms involved in this process. Key findingsOf the natural steroids examined, 17β-estradiol, progesterone, testosterone, cortisol, and aldosterone did not influence oxytocin-induced contraction at concentrations <10−6 M. Of the tested progestins, medroxyprogesterone acetate, norethisterone, chlormadinone acetate, levonorgesterol, 17α-hydroxyprogesterone capronate, and dienogest had no effect on contraction at <10−6 M. However, dydrogesterone showed rapid and direct inhibition of contraction at 10−6 M, and this inhibitory effect was dependent on dose and time. RU486 did not block the inhibitory effects of dydrogesterone on contraction. High-concentration KCl-induced contraction was also inhibited by dydrogesterone, and the inhibitory effects of dydrogesterone were observed at concentrations as low as 10−7 M. Additionally, oxytocin-induced contraction in pregnant human myometrium was inhibited by 10−6 M dydrogesterone. SignificanceThese results suggested that the rapid and direct effects of dydrogesterone on myometrial contraction were caused by a nongenomic pathway and that the progesterone receptor was not required for dydrogesterone action. Additionally, the mechanism of dydrogesterone action may involve voltage-dependent calcium channels.

Liver inflammation and regeneration in drug-induced liver injury: sex matters!

Drug-induced liver injury (DILI) remains a clinical challenge due to the poorly predictable outcomes. Accordingly, considerable efforts have been devoted to unravel the risk factors responsible for DILI worsening toward acute liver failure (ALF), liver transplantation (LT), and/or death. From a pathogenic point of view, exhaustion of drug metabolizing pathways, cell death mechanisms, activation of local immune cells, such as Kupffer cells, and recruitment of inflammatory leukocytes including monocytes and lymphocytes are key drivers of DILI progression. Taking into account that the liver is a sexually dimorphic organ, in the recent past several studies aimed to investigate the implications of gender differences in promoting DILI. While sex discrepancies in DILI include the hepatic drug metabolism or direct effects of steroid hormones (e.g. androgens and estrogens) on signaling pathways in the liver, relatively little is known on gender differences in modulating liver innate immune responses. In a previous issue of Clinical Science, Bizzaro and co-workers, analyzed sex-dependent differences in experimental acute liver injury and regeneration in mice. The authors observed a time-delay in the recovery process in male animals associated with a higher recruitment of monocytes expressing the androgen receptor (AR) as compared with females. Treatment of male mice with the pharmacological AR antagonist flutamide reduced monocyte recruitment in mice. Likewise, human male patients suffering from DILI displayed higher circulating immature and potentially more inflammatory monocytes. Altogether, these observations provide new insights into sex-dependent immune mechanisms in the context of acute liver injury, suggesting gender disparate inflammatory and regenerative responses following DILI.

Steroid hormone receptors and direct effects of steroid hormones on ram spermatozoa.

This study was based on the assumption that steroid hormones present in the female genital tract may have a rapid effect on ram spermatozoa by interaction with specific surface receptors. We demonstrate the presence of progesterone (PR) and estrogen (ER) receptors in ram spermatozoa, their localization changes during in vitro capacitation and the actions of progesterone (P4) and 17β-estradiol (E2) on ram sperm functionality. Immunolocalization assays revealed the presence of PR mainly at the equatorial region of ram spermatozoa. Western blot analyses showed three bands in ram sperm protein extracts of 40-45 kDa, compatible with those reported for PR in the human sperm membrane, and both classical estrogen receptors (66 kDa, ERα and 55 kDa, ERβ). ERα was located in the postacrosomal region of all the spermatozoa and ERβ on the apical region of 63.7% of the cells. The presence of ERβ was correlated with the percentage of non-capacitated spermatozoa evaluated by chlortetracycline staining (R = 0.848, P < 0.001). This significantly decreased after in vitro capacitation and nearly disappeared when acrosome reaction was induced. The addition of P4 and E2 before in vitro capacitation resulted in a higher (P < 0.001) acrosome-reacted sperm rate compared with the control (13.0%), noticeably greater after 3 h and when added to a high-cAMP medium (37.3% and 47.0% with E2 and P4, respectively). In conclusion, the results of this study demonstrate for the first time that ovine spermatozoa have progesterone and estrogen receptors and that both steroid hormones are related with the induction of the acrosome reaction.

Comparison of the Effects of the Selective Estrogen Receptor Modulators Ospemifene, Raloxifene, and Tamoxifen on Breast Tissue in Ex Vivo Culture.

Explant tissue culture provides a model for studying the direct effects of steroid hormones, their analogs, and novel hormonally active compounds on normal freshly isolated human breast tissues (HBTs). For this purpose, pre- and postmenopausal HBTs can be maintained in this culture system. The results demonstrate that the morphological integrity of HBT explants can be maintained in tissue culture up to 2 weeks and expression of differentiation markers, steroid hormone receptors, proliferation and apoptosis ratios can be evaluated as a response to hormonal stimulation. This chapter describes an ex vivo culture model that we have applied to study the effects of various hormonally active substances, including 17β-estradiol and selective estrogen receptor modulators (SERMs), on normal human breast tissues.

Expression of Ovarian Steroid Hormone Receptors in Tuberoinfundibular Dopaminergic Neurones During Pregnancy and Lactation

During late-pregnancy, tuberoinfundibular dopaminergic (TIDA) neurones, a critical component of the negative-feedback loop regulating prolactin secretion, become unresponsive to the stimulatory effects of prolactin. The change in TIDA responsiveness to prolactin at this time results in a decrease in dopamine secretion and a prolactin surge. As the onset of parturition and the antepartum prolactin surge depend on the withdrawal of progesterone in the presence of oestrogen, it is likely that ovarian steroid hormones mediate this change in TIDA responsiveness. To determine whether ovarian steroids can directly modulate TIDA activity, and whether changes of receptor numbers might contribute to overall steroid-regulation of these neurones, we investigated the level of oestrogen receptor alpha (ERalpha) and progesterone receptor (PR) expression within TIDA neurones during pregnancy and lactation. Animals were sacrificed on dioestrous, days 12, 19 and 21 of pregnancy and day 5 of lactation, and the proportion of TIDA neurones expressing ERalpha or PR, as well as the total number of PR expressing cells within the arcuate nucleus, was determined. Approximately 75% and 55% of tyrosine hydroxylase neurones expressed ERalpha and PR, respectively. Levels of steroid receptor expression within TIDA neurones remained fairly constant, except for an increase in ERalpha on days 12 and 19 of pregnancy compared to dioestrous and lactation day 5. The presence of steroid receptors on TIDA neurones during pregnancy and lactation supports the concept of a direct effect of steroid hormones on these neurones at this time. Thus, steroid hormones may directly act on TIDA neurones to regulate maternal prolactin secretion. The relatively stable level of expression during late pregnancy suggests that a shift in steroid receptor expression during late pregnancy does not contribute to the change in TIDA responsiveness to prolactin at this time.

Effects of estradiol and medroxyprogesterone acetate on morphology, proliferation and apoptosis of human breast tissue in organ cultures

BackgroundHuman breast tissue undergoes phases of proliferation, differentiation and regression regulated by changes of the levels of circulating sex hormones during the menstrual cycle or aging. Ovarian hormones also likely play a key role in the etiology and biology of breast cancer. Reports concerning the proliferative effects of steroid hormones on the normal epithelium of human breast have been conflicting. Some studies have shown that steroid hormones may predispose breast epithelial cells to malignant changes by stimulating their proliferation, which is known to be regulated tightly by stromal cells.The aim of this study was to investigate the effects of 17β-estradiol and medroxyprogesterone acetate on proliferation, apoptosis, expression of differentiation markers and steroid hormone receptors in breast epithelium using an in vitro model of freshly isolated human breast tissue, in which a proper interaction of breast epithelium and stroma has been maintained.MethodsHuman breast tissues were obtained from women undergoing surgery for breast tumours. Peritumoral tissues were excised and explants were cultured for 3 weeks in medium supplemented with E2 or MPA or with E2+MPA. Endpoints included histopathological, histomorphometric and immunohistochemical assessment of the breast explants.ResultsCulture of breast explants for 14 or 21 days with steroid hormones increased proliferative activity and the thickness of acinar and ductal epithelium. E2-treatment led to hyperplastic epithelial morphology, MPA to hypersecretory single-layered epithelium and E2+MPA to multilayered but organised epithelium.The proliferative response to E2 in comparison to control (p < 0.001) was more pronounced than to MPA (p < 0.05) or E2+MPA (p < 0.05) at 7 and 14 days for Ki-67 and PCNA. E2 treatment also decreased the proportion of apoptotic cells after 7 (p < 0.01) and 14 (p < 0.01) days. In addition, the relative number of ERα, ERβ and PR positive epithelial cells was decreased by all hormonal treatments.ConclusionOrgan culture system provides a model for studying the direct effects of steroid hormones and their analogues on postmenopausal human breast tissue.Addition of E2 or MPA or E2+MPA to breast explants caused characteristic changes in morphology, stimulated epithelial proliferation, lowered apoptosis ratio and decreased the relative number of epithelial cells expressing ERα, ERβ and PR.

Pituitary effects of steroid hormones on secretion of follicle-stimulating hormone and luteinizing hormone

Steroid hormones have a profound influence on the secretion of the gonadotropins, follicle-stimulating hormone (FSH) and luteinizing hormone (LH). These effects can occur as a result of steroid hormones modifying the secretion of gonadotropin-releasing hormone (GnRH) from the hypothalamus, or a direct effect of steroid hormones on gonadotropin secreting cells in the anterior pituitary gland. With respect to the latter, we have shown that estradiol increases pituitary sensitivity to GnRH by stimulating an increase in expression of the gene encoding the GnRH receptor. Since an estrogen response element (ERE) has not been identified in the GnRH receptor gene, this effect appears to be mediated by estradiol stimulating production of a yet to be identified factor that in turn enhances expression of the GnRH receptor gene. However, the importance of estradiol for enhancing pituitary sensitivity to GnRH during the periovulatory period is questioned because an increase in mRNA for the GnRH receptor precedes the pre-ovulatory rise in circulating concentrations of estradiol. In fact, it appears that the enhanced pituitary sensitivity during the periovulatory period may occur as a result of a decrease in concentrations of progesterone rather than due to an increase in concentrations of estradiol. Estradiol also is capable of altering secretion of FSH and LH in the absence of GnRH. In a recent study utilizing cultured pituitary cells from anestrous ewes, we demonstrated that estradiol induced a dose-dependent increase in secretion of LH, but resulted in a dose-dependent decrease in the secretion of FSH. We hypothesized that the discordant effects on secretion of LH and FSH might arise from estradiol altering the production of some of the intrapituitary factors involved in synthesis and secretion of FSH. To examine this hypothesis, we measured amounts of mRNA for activin B (a factor known to stimulate synthesis of FSH) and follistatin (an activin-binding protein). We found no change in the mRNA for follistatin after treatment of pituitary cells with estradiol, but noted a decrease in the amount of mRNA for activin B. Thus, the inhibitory effect of estradiol on secretion of FSH appears to be mediated by its ability to suppress the expression of the gene encoding activin.

Direct actions of gonadal steroid hormones on FSH secretion and expression in the infantile female rat

FSH-beta mRNA is dramatically regulated in the infantile female rat anterior pituitary. Elevated plasma levels of FSH correspond with increased FSH-beta mRNA levels which peak on PND 12. The source of this regulation does not appear to be GnRH, since the administration of a potent GnRH antagonist does not suppress FSH-beta mRNA levels. Consequently, we have examined the effects of the gonadal steroid hormones, estrogen and androgen, on the maintenance of gonadotropin secretion and gene expression both in vivo and in vitro. Androgen and estrogen action was blocked in vivo with the specific receptor antagonists, flutamide (150 μg) and tamoxifen (200 μg). Administration of antagonists during two different three day time-periods of infantile life [postnatal day (PND) 8–11 and PND 11–14] resulted in differing effects on both FSH and LH secretion as well as on FSH-beta and LH-beta mRNA levels. Flutamide and tamoxifen treatment both suppressed FSH secretion at either age examined ( p<0.01). LH secretion was suppressed by both treatments but only at the younger of the two ages ( p<0.01). In contrast to its effects on FSH secretion, tamoxifen suppressed FSH-beta mRNA levels in the later group only. LH-beta mRNA levels were suppressed by tamoxifen, but only in the younger age group ( p<0.05). The direct effects of steroid hormones on infantile pituitary gonadotrophs were examined in vitro by incubating cells with dihydrotestosterone propionate (DHTP; 10 −8 M) or 17 β-estradiol (E; 10 −8 M). Both DHT and E treatment stimulated FSH secretion when measured 48 h later ( p<0.01). There were no effects on LH secretion. FSH-beta mRNA levels were also stimulated by DHT at 48 h ( p<0.01). Estradiol treatment transiently increased FSH-beta mRNA levels at 2 and 6 h following treatment ( p<0.01) but not at 48 h. LH-beta levels were suppressed by DHT treatment ( p<0.05), and E transiently elevated LH-beta mRNA levels at 2 h ( p<0.05). Taken together these studies indicate that gonadotrophs from infantile female rats are capable of responding directly to steroid hormones, and may play a role in the selective stimulation of FSH secretion and expression in vivo.

Mitochondrial genes as sites of primary action of steroid hormones

Steroid and thyroid hormones act on nuclear gene transcription by activating protein receptors, which in turn bind to hormone response elements (HREs). Among the cell-specific processes regulated by steroid receptors is energy metabolism through increased synthesis of respiratory enzymes. As some of these enzymes are encoded by both nuclear and mitochondrial genes, coordination of their synthesis is probable, inter alia at the transcriptional level. We have postulated a direct effect of steroid hormones on mitochondrial gene transcription and here present the following evidence in support of this hypothesis. 1) The human and rodent mitochondrial genomes contain nucleotide sequences similar both to type I and type II HREs. 2) Glucocorticoid receptors (GR) rapidly translocate from the cytoplasm into mitochondria after administration of glucocorticoids. This process has been reproduced in vitro and deletion of the N-terminal part of the glucocorticoid receptor stops translocation into mitochondria. 3) Gel shift analysis has demonstrated binding of GR to putative mitochondrial GR elements. 4) In transfection experiments, mitochondrial HREs confer dexamethasone inducibility on hybrid reporter constructs, abolished in the presence of excess RU38486. 5) Similar results were obtained for thyroid hormone receptor (TR α) localization, import, and binding to TR elements. These findings, taken with the demonstrated effects of steroid (and thyroid) hormones on mitochondrial transcription and respiratory enzyme biosynthesis, strongly support the hypothesis of a direct effect of steroid (and thyroid) hormones on mitochondrial gene transcription.

Sexual dimorphism of blood pressure: Possible role of the renin-angiotensin system

The prevalence of hypertension in men is higher than in women and the onset of this disease is earlier in male than in female subjects. In spontaneously hypertensive rats, males also have higher blood pressures than females. Evidence from epidemiological, physiological, molecular biological and morphological studies concerning this sexual dimorphism is reviewed. We demonstrate that the gonadal steroids testosterone and estrogen have important effects on the gene regulation of the renin-angiotensin system. This may in part contribute to the sexual dimorphism in blood pressure control. The direct effect of steroid hormones on genes related to hypertension provides a suitable paradigm to improve our understanding of molecular and cellular mechanisms of cardiovascular control.

Update on the metabolic effects of oral contraceptives.

The pharmacokinetic properties of oral contraceptives (OCs) and the steroid action at cellular level are of fundamental significance to the side effects of these agents. As a result of variability from woman to woman in the pharmacokinetics of a particular steroid compound, metabolic side effects are heterogeneous. The direct effects of steroid hormones on target tissues such as liver, pancreas, kidney, and uterus usually decline within the first 3 months of OC use. Receptor mediated effects increase with dose, but level off when the dose becomes high. The cellular response to a hormone stimulus is in part dependent on the stability of the hormone/receptor complex and in part on the binding of the steroid/receptor complex to the DNA molecule. Thus, small changes in the steroid configuration can result in marked differences in the hormone response. In general, OCs decrease glucose tolerance and cause a rise in insulin levels. Both exogenous and endogenous sex steroids affect lipid metabolism and thus influence the risk of cardiovascular disease. There is epidemiological evidence that the risk of cardiovascular disease is dependent on the progestogen content and the estrogen/progestogen ratio of the contraceptive. OCs also have an effect on coagulation and the fibrinolytic enzyme system, with a dose-effect relationship related to the estrogen component of OCs.

The Sensitizing Effect of Estrogens and Catechol Estrogen on Cultured Pituitary Cells to Luteinizing Hormone-Releasing Hormone: Its Antagonism by Progestins*

The direct effect of steroid hormones upon LHRH-induced LH release was tested in cultured pituitary gonadotrophs. Anterior pituitary cells from adult ovariectomized rats were cultured for 2 days in the presence and absence of estrone (E1), 17β-estradiol (E2), estriol (E3), 2-hydroxyestrone (2-OHE1), 2-hydroxyestradiol (2-OHE2), progesterone (P), and 20α-dihydroprogesterone (20α-OHP). The cells were then washed and incubated for 3 h with graded doses of LHRH (10-12-10-6 M) and the LH released into the media was measured by RIA. A dose-related increase in LH secretion was observed with increasing concentrations of LHRH and, in control cultures, the half-maximal effective dose (ED50) of LHRH was 1.9 ± 0.6 × 10-9 M. Pretreatment with Ei or E2 (10-10-10-8 M) sensitized gonadotrophs to LHRH and caused a 10-fold decrease in the ED50. 2-OHE2 also exhibited a sensitizing effect, but it was less effective than E2 (2-OHE2: 10-10 M, no effect; 10-8 M, ED50, 2.5 ± 0.5 × 10-10 M). NO sensitizing effect was observed wit...

The ultrastructure of liver cells in women under steroid therapy. I. Normal pregnancy and trophoblastic growths.

ABSTRACT An electron microscopic study of liver biopsies of women under steroid therapy was carried out. Liver biopsies taken with a Menghini needle from five healthy young women during their last trimester of pregnancy revealed elongation, gigantism and lamellar osmiophilic matrical inclusions in approximately 10% of the population of mitochondria per cell examined. Liver biopsies taken from six women with hydatidiform mole and from one women with choriocarcinoma revealed mild focal dilatation and vesiculation of both the rough and smooth endoplasmic reticulum. Mitochondrial modifications were present and included gigantism, pleomorphism and also lamellar osmiophilic inclusions in 10 to 20% of the cases. In one case of hydatidiform mole this change was present in more than 60% of mitochondria examined. The findings are not interpreted as pathological but rather as an exaggeration of a physiological change brought about by an increased metabolic demand during pregnancy or else to a more direct effect of steroid hormones on liver cell organelles.

Effects of steroid hormones on the multiplication of cells

Since the action of hormones in the tissue is complicatedly related or combined, the pure action of steroid hormones is rarely investgated in vivo. The purpose of the present examinations was to determine the direct effects of steroid hormones on the multiplication of the following cells : Fibroblasts from chick embryo hearts, MK cells, PS cells, FL cells, HeLa cells and AH-130 cells. In order to determine accurately the effects, the tissue culture method was employed and a large quantity of the hormones was added in the medium.1) In 7 days' cultivation, paramethasone showed an inhibitory effect on the multiplication of Fibroblasts from chick embryo hearts proportional to the increase of concentration. Paramethasone in the concentration of 6.6 pg/cc had a 30% inhibitory effect in comparison with the control culture. In the concentration of 6.6 and 33.3μtg/cc, 4-chlorotestosterone acetate gave little effect on the multiplication of Fibroblasts, almost similar to the control culture, but in the concentration of 66.6 μg/cc, it gave slight inhibitory effects.2) The multiplication of MK cells and PS cells was inhibited by paramethasone in proportion to the increase of concentration : 6.6, 33.3 and 66.6 pg/cc of culture medium, respectively. The inhibitory effects were marked in the case of MK cells.3) The multiplication of strain FL cells was makedly inhibited by the addition of 66.6, ag/cc paramethasone, and paramethasone in the concentration of 6.6 μg/cc showed 46% inhibitory effect after 7 days' cultivation.Strain HeLa cells were much more sensitive to paramethasone than strain FL cells, namely, 33.3 μg/cc paramethasone showed 80% inhibitory effect in comparison with the control culture ; 4-chlorotestosterone acetate showed lower inhibitory effect than paramethasone in each concentration : 6.6, 33.3 and 66.6 μg/cc, respectively on the 7th day of cultivation.4) The inhibitory effects of adrenocorticosteroids on chick embryo pulmonary tissues and FL cells were reversible in a concentration lower than 100 μg per cc.Paramethasone in the concentration of 66.6 μg/cc had a marked inhibitory effect on the multiplication of FL cells tested, but cell growth was evident after removing paramethasone and thoroughly washing the culture with PBS solution, then adding YLE medium.5) On examination, the mixture of paramethasone and 4-chlorotestosterone acetate in the concentration of 10μg/cc a slight antagonistic effect was observed. When paramethasone was added alone to the medium, it gave a 50% inhibitory effect on the multiplication of FL cells. In the culture which 10μg/cc paramethasone and 4-chlorotestosterone acetate were added, the inhibitory effect was reduced to 38%.6) Paramethasone-treated FL cells became round in shape and tended to cohere themselves in sheets in which a great number of giant cells were often seen. These morphologic changes were also seen in high concentration of paramethasone, and FL cells turned into spindle-shaped ones in the concentration of 100 μg/cc.The nuclei of typical accites hepatoma 130 cells were of various sizes and irregularly shaped. To the contrary, when these cells were treated with paramethasone, they had small and nearly round nuclei in proportion to the increasing concentration, and the cytoplasm contained many vacuoles.7) In the concentration 50 /2g/cc paramethasone, the mitotic coefficiency decreased to two-thirds of control values in FL cells, five-eighths in MK cells, one-third in AH-130 cells. In 4-chlorotestosterone acetate-treated FL cells, it slightly increased.8) Prednisolone in the concentration of 10 μg/cc produced a delay and partial inhibition of cytopathic effect when introduced into HeLa cell cultures at the time of infection with HV J. The titer of virus as measured by hemagglutination was dereased slightly in prednisolone-treated groups after 72 hours of incubation.