Endogenous Progesterone Research Articles

Protein arginine methyltransferase 6 regulates multiple aspects of gene expression

It is well established that transcription and alternative splicing events are functionally coupled during gene expression. Here, we report that protein arginine N-methyltransferase 6 (PRMT6) may play a key role in this coupling process by functioning as a transcriptional coactivator that can also regulate alternative splicing. PRMT6 coactivates the progesterone, glucocorticoid and oestrogen receptors in luciferase reporter assays in a hormone-dependent manner. In addition, small interfering RNA (siRNA) oligonucleotide duplex knockdown of PRMT6 disrupts oestrogen-stimulated transcription of endogenous GREB1 and progesterone receptor in MCF-7 breast cancer cells, demonstrating the importance of PRMT6 in hormone-dependent transcription. In contrast, the regulation of alternative splicing by PRMT6 is hormone independent. siRNA knockdown of PRMT6 increases the exon inclusion:skipping ratio of alternatively spliced exons in endogenous vascular endothelial growth factor and spleen tyrosine kinase RNA transcripts in both the presence and absence of oestrogen. These results demonstrate that PRMT6 has a dual role in regulating gene expression and that these two functions can occur independently of each other.

G Protein Pathway Suppressor 2 (GPS2) Is a Transcriptional Corepressor Important for Estrogen Receptor α-mediated Transcriptional Regulation

We have identified G protein suppressor 2 (GPS2) as a stable component of the SMRT corepressor complexes. GPS2 potently represses basal transcription, with the repression domain mapped to the N-terminal silencing mediator of retinoic acid and thyroid hormone receptor (SMRT)-interacting domain. Knockdown of GPS2 abrogates, whereas overexpression potentiates, SMRT-mediated repression activity. The SMRT complexes are involved in 4-hydroxyl-tamoxifen (4OHT)-mediated gene repression by estrogen receptor alpha (ERalpha). We show that 4OHT recruits SMRT and GPS2 to the promoter of pS2, an ERalpha target gene, in a dynamic manner. Unexpectedly, we also found that estradiol (E2) promotes promoter recruitment of the SMRT complexes. While knockdown of GPS2 compromised 4OHT-mediated repression, it enhanced E2-induced expression of a reporter gene and several endogenous ERalpha target genes, including pS2, cyclin D1 (CCND1), progesterone receptor (PR), and c-MYC. Finally, we show that depletion of GPS2 or SMRT by siRNA promotes cell proliferation in MCF-7 breast cancer cells. Thus, we concluded that GPS2 is an integral component of the SMRT complexes, important for ligand-dependent gene regulations by ERalpha and a suppressor for MCF-7 cell proliferation.

Case of autoimmune progesterone dermatitis presenting as fixed drug eruption

Autoimmune progesterone dermatitis is a rare disorder that presents as a cyclical cutaneous eruption during the luteal phase of the menstrual cycle. It typically occurs in women due to an autoimmune phenomenon to endogenous progesterone production. We describe a 34-year-old woman with an erythematous round plaque with blistering, which recurred a few days before her menstrual cycle, at the identical site on the left arm. The diagnosis of autoimmune progesterone dermatitis is made with i.d. skin testing on the affected lesion with progesterone. After the beginning of oral prednisolone (40 mg daily) therapy during menstruation, although slight recurrence appeared, the severity was significantly improved.

Expression of Progesterone Receptor A form and Its Role in the Interaction of Progesterone with Cortisol on Cyclooxygenase-2 Expression in Amnionic Fibroblasts

Context: Human amnion fibroblasts produce abundant prostaglandins toward the end of gestation, which is believed to be one of the major events leading to parturition. Glucocorticoids have been shown to up-regulate cyclooxygenase-2 (COX-2) expression, the crucial enzyme catalyzing prostaglandin synthesis, in human amnion fibroblasts. Although a major propregnancy hormone, the effect of progesterone and the associated progesterone receptor subtypes in the regulation of both basal and glucocorticoid-induced COX-2 expression in human amnion fibroblasts have not been resolved. Methods and Results: Cultured human amnion fibroblasts prepared from the fetal membranes at term pregnancy without labor mainly expressed the progesterone receptor A form (PRA). Inhibition of endogenous progesterone production with trilostane or knockdown of PRA expression with small interfering RNA significantly enhanced the glucocorticoid receptor (GR)-mediated COX-2 induction by cortisol, whereas overexpression of PRA attenuated the i...

Expression of Progesterone Receptor A form and Its Role in the Interaction of Progesterone with Cortisol on Cyclooxygenase-2 Expression in Amnionic Fibroblasts

Human amnion fibroblasts produce abundant prostaglandins toward the end of gestation, which is believed to be one of the major events leading to parturition. Glucocorticoids have been shown to up-regulate cyclooxygenase-2 (COX-2) expression, the crucial enzyme catalyzing prostaglandin synthesis, in human amnion fibroblasts. Although a major propregnancy hormone, the effect of progesterone and the associated progesterone receptor subtypes in the regulation of both basal and glucocorticoid-induced COX-2 expression in human amnion fibroblasts have not been resolved. Cultured human amnion fibroblasts prepared from the fetal membranes at term pregnancy without labor mainly expressed the progesterone receptor A form (PRA). Inhibition of endogenous progesterone production with trilostane or knockdown of PRA expression with small interfering RNA significantly enhanced the glucocorticoid receptor (GR)-mediated COX-2 induction by cortisol, whereas overexpression of PRA attenuated the induction by cortisol. Co-immunoprecipitation assay revealed PRA in the GR protein complex. Although exogenous progesterone did not alter COX-2 expression under basal conditions, it attenuated cortisol-induced COX-2 expression at concentrations about 10- to 50-fold higher, which might be achieved by competition with cortisol for GR. We demonstrated in this study that endogenous progesterone might counteract the induction of prostaglandin synthesis by cortisol via PRA transdominant repression of GR function, whereas high levels of progesterone might further inhibit the induction by cortisol via competitive binding to GR in human amnion fibroblasts. These inhibitory actions of progesterone and PRA on glucocorticoids and GR may partly explain the inconsistent effects of glucocorticoids on parturition in humans.

A potential role of endogenous progesterone in modulation of GH, prolactin and thyrotrophin secretion during normal menstrual cycle

Previous studies investigating the fluctuations of endocrine secretion across the menstrual cycle yielded inconsistent results. Our objective was to evaluate during the menstrual cycle the potential role of endogenous oestradiol and progesterone in the regulation of hormones primarily controlled by the circadian clock and/or the sleep-wake cycle. Ten normally cycling young lean women were investigated once during follicular and once during luteal phase. Sleep was polygraphically recorded, and blood samples were obtained at 20-min intervals for 24 h. Sleep variables and diurnal melatonin and cortisol profiles (hormones primarily controlled by the circadian clock) were similar in both conditions. The TSH evening rise (a circadian marker) was similar in both conditions, but the sleep-related nocturnal TSH decrease occurred earlier during the luteal phase (P = 0.03) and tended to correlate positively with progesterone levels (r(s) = -0.64, P < 0.06). Daytime GH secretion and afternoon/evening PRL secretion (hormones primarily controlled by the sleep-wake homeostasis) were increased in the luteal phase compared with those of the follicular phase (GH: P = 0.04; PRL: P = 0.01). The increase in 24-h GH secretion was associated with higher progesterone levels (r(s) = 0.78, P = 0.02). In luteal phase, the evening PRL rise was associated with higher progesterone (r(s) = 0.70, P = 0.04) and oestradiol (r(s) = 0.72, P = 0.03) levels. The present data indicate that in normally cycling young women, daytime GH and PRL secretions are increased in luteal phase. These data also suggest that endogenous progesterone could play a modulation role on pituitary hormone secretion, stimulating GH and PRL release and enhancing the inhibitory action of sleep on TSH secretion.

Antimullerian hormone, but not follicle stimulating hormone, reflects decline in ovarian reserve associated with oral cyclophosphamide use in Wegener's granulomatosis for the WGET research group

OBJECTIVE: Our aim was to determine the effect of cyclophosphamide (CYC) on ovarian reserve markers antimullerian hormone (AMH) and follicle stimulating hormone (FSH) in patients with Wegener's Granulomatosis (WG). DESIGN: Case control study of stored plasma. MATERIALS AND METHODS: Patients in Wegener's Granulomatosis Etanercept Trial (WGET) received cyclophosphamide (CYC) for severe WG and methotrexate for limited WG. The first and last available plasma samples from the 42 women who were under age 50 at WGET enrollment were analyzed in duplicate for FSH (ELISA Calbiotech) and AMH (ELISA DSLabs). Linear regression analysis and Wilcoxon rank sum tests were performed. RESULTS: While the change in AMH during the study was closely correlated with the dose of CYC received (0.8 ng/mL decline per 10 gms CYC, p=0.009), change in FSH did not correlate with CYC dose. Furthermore, AMH, but not FSH, exhibited a significant decline in patients receiving CYC compared to no CYC.Table 1Change in AMH & FSH from baseline to end of study.CYC in WGETNo CYC in WGETParameterNMean ± std.NMean ± std.p-valueRaw Change of AMH (last visit - baseline)∗Baseline AMH<0.5 ng/mL excluded.11-2.15 ± 1.9460.36 ± 0.890.011Raw Change of FSH (last visit - baseline)†Oral contraceptive pill & GnRH agonist users excluded.227.40 ± 18.4410-1.73 ± 14.080.347∗ Baseline AMH<0.5 ng/mL excluded.† Oral contraceptive pill & GnRH agonist users excluded. Open table in a new tab CONCLUSIONS: In this study, AMH was more closely linked to CYC ovarian damage than FSH. FSH testing was not obtained in the early follicular phase, so was influenced by endogenous estradiol and progesterone during the ovarian cycle of ovulating women. In the oncofertility setting, where patients are not routinely seen in the early follicular phase and may be taking oral contraceptives, AMH is a superior marker of ovarian function.

Effect of endogenous progesterone and oestradiol-17β on the incidence of gastro-oesophageal reflux and on the barrier pressure during general anaesthesia in the female dog

ObjectiveTo investigate the potential effect of increased blood progesterone (P4) and oestradiol-17β (E2) concentrations on the barrier pressure (BrP) and the incidence of gastro-oesophageal reflux (GOR) in female dogs under general anaesthesia. Study designProspective, blinded experimental trial. AnimalsSeven female, adult, healthy dogs weighing 14–21 kg and aged 1–7 years. MethodsEach of the animals was studied under the influence of high blood E2 and basal P4 (study O), basal E2 and high P4 (study P) and basal E2 and P4 (study C) concentrations. Animals were premedicated with acepromazine and anaesthesia was induced with thiopental and maintained with halothane. Lower oesophageal pH was monitored continuously for 1 hour after induction. GOR was defined as oesophageal pH >7.5 or <4. Manometry of the posterior oesophageal sphincter (POS) was then performed using the slow pull-through technique. ResultsAcid GOR was detected in only one animal of study O. The three studies did not differ significantly in GOR. Mean BrP was 11.2 (study O), 9.1 (study P) and 11.6 mmHg (study C). No significant differences were detected with respect to mean BrP, intra-gastric pressure and POS pressure. ConclusionsIt is unlikely that the increased concentrations of E2 or P4 during the normal ovarian cycle influence the functional efficiency of the POS as a major barrier to GOR in healthy, female dogs under general anaesthesia. Clinical relevanceThe fact that female dogs undergoing obstetrical surgery represented a substantial sub-group of the animals which developed postoperative benign oesophageal stricture, should probably not be attributed to the effects of increased concentrations of female sex steroid hormones.

Total Laparoscopic Radical Hysterectomy in the Treatment of Early Cervical Cancer

Total laparoscopic radical hysterectomy (TLRH) has been reported since the early 1990 s. Although the acceptance of TLRH had been slow over the past 15 years, several teams throughout the world have recently reported promising results in the treatment of early cervical cancer with this procedure. Several modifications of the originally described technique have also been reported. From the currently existing data, these is no doubt that TLRH is technically feasible. Its operative safety profile is comparable to that of radical abdominal hysterectomy (RAH), and there exist sufficient data to suggest that the histopathologic outcome is also similar in terms of local radicality and lymph node yield. The duration of the procedure has become acceptable but remains still longer in comparison to RAH, in most series. It is now evident that with increasing experience, repetition, standardization, and incorporation of technological advances, duration can be reduced considerably and become similar to that of RAH. Total laparoscopic radical hysterectomy is associated with less blood loss, faster recovery and return of bowel function, reduced febrile morbidity, and a better cosmetic result. Nevertheless, shorter hospitalization in comparison to that observed after RAH is not consistently reported, and return of normal bladder activity is similar to that observed after RAH. It is also true that the currently existing recurrence and survival data are still immature to draw safe conclusions on its long-term oncological safety. Probably, the time has come for a multicenter randomized study between TLRH and RAH with participation of the institutions with significant experience in this procedure.

The progesterone receptor antagonist mifepristone abolishes carotid sinus nerve response to hypoxia in rat pups.

We tested the hypothesis that endogenous progesterone enhances peripheral chemoreceptors function in rat pups. Newborn rats have been treated with the progesterone receptor antagonist mifepristone (Mif) between postnatal days 3‐12 (70μg/g/day) and compared to control (Cont). We used the in‐vitro carotid body/carotid sinus nerve (CSN) preparation to record CSN activity (impulses/sec) in 12‐day‐old rats under normoxia (perfusion medium bubbled with 21%O2‐5%CO2), in response to hypoxia (5%O2‐5%CO2), or nicotine (mediator between carotid body type I cells and CSN terminals, 100μM) supefusion for 3‐4 minutes each. Baseline CSN firing frequency was similar between groups, at 2.7±0.3 in Cont and 2.6±0.3 Hz in Mif pups. At steady state hypoxic CSN activity increased to 7.5±0.9 Hz in Cont, but was left unchanged in Mif pups (3.0±0.5 Hz, p&lt;0.0002 between groups). In response to nicotine superfusion, CSN firing activity increased to 10.8±1.9 Hz in Cont and to 6.1±3.3 Hz in Mif (p&lt;0.04 between groups). We conclude that endogenous progesterone plays an important role in carotid body during postnatal development in rats. Founded by SickKids Foundation (XG07‐006)

Coincident frequencies and relative phases among brain activity and hormonal signals

BackgroundFourier transform is a basic tool for analyzing biological signals and is computed for a finite sequence of data sample. The electroencephalographic (EEG) signals analyzed with this method provide only information based on the frequency range, for short periods. In some cases, for long periods it can be useful to know whether EEG signals coincide or have a relative phase between them or with other biological signals. Some studies have evidenced that sex hormones and EEG signals show oscillations in their frequencies across a period of 28 days; so it seems of relevance to seek after possible patterns relating EEG signals and endogenous sex hormones, assumed as long time-periodic functions to determine their typical periods, frequencies and relative phases.MethodsIn this work we propose a method that can be used to analyze brain signals and hormonal levels and obtain frequencies and relative phases among them. This method involves the application of a discrete Fourier Transform on previously reported datasets of absolute power of brain signals delta, theta, alpha1, alpha2, beta1 and beta2 and the endogenous estrogen and progesterone levels along 28 days.ResultsApplying the proposed method to exemplary datasets and comparing each brain signal with both sex hormones signals, we found a characteristic profile of coincident periods and typical relative phases. For the corresponding coincident periods the progesterone seems to be essentially in phase with theta, alpha1, alpha2 and beta1, while delta and beta2 go oppositely. For the relevant coincident periods, the estrogen goes in phase with delta and theta and goes oppositely with alpha2.ConclusionFindings suggest that the procedure applied here provides a method to analyze typical frequencies, or periods and phases between signals with the same period. It generates specific patterns for brain signals and hormones and relations among them.

The effect of the neuroactive steroid 5β-pregnane-3β, 20(R)-diol on the time course of GABA evoked currents is different to that of pregnenolone sulphate

The endogenous progesterone metabolite allopregnanolone has a number of properties including anesthetic, sedative, antiepileptic, anxiolytic, impaired memory function and negative mood symptoms. Allopregnanolone is a potent positive GABA A receptor function modulators. In contrast, 3β-hydroxy-steroids (3β-steroids) usually modulate the GABA A receptor negatively. They have attracted some interest for their possible use as therapeutic agents that could counteract the negative symptoms induced by allopregnanolone. Two hypotheses for the action of 3β-steroids have been proposed: 1) 3β-steroids act in a similar way to pregnenolone sulphate, which non-competitively reduces GABA A receptor activity. 2) 3β-steroids specifically antagonize the effect of allopregnanolone. We have therefore tried to clarify this issue by comparing the effect of pregnenolone sulphate and 5β-pregnane-3β, 20(R)-diol on the GABA-evoked currents by the patch clamp technique on neurons from the medial preoptic nucleus. Both pregnenolone sulphate and 5β-pregnane-3β, 20(R)-diol increase the desensitization rate of the current response evoked by a 2 s GABA application. However, their effects on other parameters of the GABA evoked currents differed in degree and sometimes even in direction. The actions of pregnenolone sulphate and 5β-pregnane-3β, 20(R)-diol were not altered in the presence of allopregnanolone, which indicates that they do not directly interact with allopregnanolone. In addition, when 5β-pregnane-3β, 20(R)-diol was tested on spontaneous inhibitory postsynaptic currents (sIPSCs), it dramatically reduced the allopregnanolone-induced prolongation of the decay time constant but it had no effect on the decay under control conditions. In conclusion, the effect of 5β-pregnane-3β, 20(R)-diol on GABA-evoked currents is different to that of pregnenolone sulphate in medial preoptic nucleus neurons.

Artemisinin selectively decreases functional levels of estrogen receptor-alpha and ablates estrogen-induced proliferation in human breast cancer cells

MCF7 cells are an estrogen-responsive human breast cancer cell line that expresses both estrogen receptor (ER) alpha and ERbeta. Treatment of MCF7 cells with artemisinin, an antimalarial phytochemical from the sweet wormwood plant, effectively blocked estrogen-stimulated cell cycle progression induced by either 17beta-estradiol (E(2)), an agonist for both ERs, or by propyl pyrazole triol (PPT), a selective ERalpha agonist. Artemisinin strongly downregulated ERalpha protein and transcripts without altering expression or activity of ERbeta. Transfection of MCF7 cells with ERalpha promoter-linked luciferase reporter plasmids revealed that the artemisinin downregulation of ERalpha promoter activity accounted for the loss of ERalpha expression. Artemisinin treatment ablated the estrogenic induction of endogenous progesterone receptor (PR) transcripts by either E(2) or PPT and inhibited the estrogenic stimulation of a luciferase reporter plasmid driven by consensus estrogen response elements (EREs). Chromatin immunoprecipitation assays revealed that artemisinin significantly downregulated the level of endogeneous ERalpha bound to the PR promoter, whereas the level of bound endogeneous ERbeta was not altered. Treatment of MCF7 cells with artemisinin and the pure antiestrogen fulvestrant resulted in a cooperative reduction of ERalpha protein levels and enhanced G(1) cell cycle arrest compared with the effects of either compound alone. Our results show that artemisinin switches proliferative human breast cancer cells from expressing a high ERalpha:ERbeta ratio to a condition in which ERbeta predominates, which parallels the physiological state linked to antiproliferative events in normal mammary epithelium.

Effect of oral contraceptives on vascular endothelial growth factor, Cox-2 and aromatase expression in the endometrium of uteri affected by myomas and associated pathologies

Background The study was conducted to evaluate vascular endothelial growth factor (VEGF), Cox-2 and aromatase expression in the endometrium of uteri with myomas and other associated pathologies. Study Design Hysteroscopy was performed in 118 women of reproductive age with myomas and menorrhagia, 40 of whom were using a pill containing 75 mcg gestodene+30 mcg ethinylestradiol. Aromatase p450, VEGF and Cox-2 expression was detected using immunohistochemistry. Fisher's Exact Test and the Mann–Whitney test were used in the statistical analysis, with significance established at p<.05. Results In patients with myomas and menorrhagia, associated pathologies such as adenomyosis, endometrial polyps and endometriosis were found in 32%, 12% and 17% of cases, respectively. Aromatase, Cox-2 and VEGF expression was greater during the proliferative phase compared to the luteal phase of the cycle or following oral contraceptive use. Conclusion Endogenous progesterone or combined oral contraceptives are potent inhibitors of VEGF, aromatase and Cox-2 expression in the endometrium of patients with myomas and menorrhagia.

Estrous cycle regulates activation of hippocampal Akt, LIM kinase, and neurotrophin receptors in C57BL/6 mice

Estradiol modulates dendritic spine morphology and synaptic protein expression in the rodent hippocampus, as well as hippocampal-dependent learning and memory. In the rat, these effects may be mediated through nongenomic steroid signaling such as estradiol activation of the Akt and LIM kinase (LIMK) pathways, in addition to genomic signaling involving estradiol upregulation of brain-derived neurotrophic factor expression (BDNF). Due to the many species differences between mice and rats, including differences in the hippocampal response to estradiol, it is unclear whether estradiol modulates these pathways in the mouse hippocampus. Therefore, we investigated whether endogenous fluctuations of gonadal steroids modulate hippocampal activation of the Akt, LIMK, and the BDNF receptor TrkB in conjunction with spatial memory in female C57BL/6 mice. We found that Akt, LIMK, and TrkB were activated throughout the dorsal hippocampal formation during the high-estradiol phase, proestrus. Cycle phase also modulated expression of the pre- and post-synaptic markers synaptophysin and post-synaptic density 95. However, cycle phase did not influence performance on an object placement test of spatial memory, although this task is known to be sensitive to the complete absence of ovarian hormones. The findings suggest that endogenous estradiol and progesterone produced by the ovaries modulate specific signaling pathways governing actin remodeling, cell excitability, and synapse formation.

Rare Case of Autoimmune Progesterone Dermatitis Coexisting with Rheumatoid Nodules of the Ovary: Case Report and Review of the Literature

Autoimmune progesterone dermatitis (APD) represents a hypersensitivity reaction to endogenous progesterone. The condition presents with a variety of skin eruptions characterized by cyclical recurrent premenstrual exacerbations due to progesterone fluctuation during the menstrual cycle. We hereby describe the first case in modern literature of an association with a rheumatoid nodule of the ovary in a patient who presented with right-sided pelvic pain.

The effect of estrus synchronization treatments on somatic cell count of transitional-anestrus Awassi ewes’ milk

Fifty-three transitional-anestrus Awassi ewes, randomly assigned to three groups: fluorogestone acetate (FGA, n = 18), FGA-Prostaglandin (FGA-PGF, n = 18) and control (n = 17), were used to examine the effect of estrus synchronization protocols and steroid hormones concentrations on milk somatic cell count (SCC). Intravaginal FGA sponge was inserted for 13 days and 600 IU equine chorionic gonadotropin was administered for ewes of FGA and FGA-PGF groups at the time of sponge removal (day 0). In addition, 10 mg was administered to ewes of FGA-PGF group on day 0. Blood and milk samples were collected from all ewes on days -13, -6, 0, 1, 2, 7 and 14. Estradiol had significant positive correlation with the SCC during the periods of sponge insertion (P = 0.015, r = 0.235) and within two days (P = 0.063 r = 0.23) after sponge removal with no correlation with SCC of both udder halves during the luteal phase. Progesterone concentrations, on the other hand, had a significant positive correlation (P < 0.001; r = 0.420) with the SCC of both udder halves during the luteal phase of the experiment, but not during the periods of sponge insertion and expected estrus. SCC returned under the influence of endogenous progesterone on days 7 and 14 to pre-synchronization values. In conclusion, sheep milk SCC is affected significantly with induction of estrus and steroid hormones concentrations. However, peak SCC recorded during estrus was far below the upper limit of the current standard for normal milk. With the current standards for SCC of 1,000,000/ml as legal limit for abnormal milk control programs in sheep, estrus synchronization programs and the estrus status should not be considered when bulk-tank milk SCC is being investigated, but should be considered during the process of setting new standards.

Estrogen, medroxyprogesterone acetate, endothelial function, and biomarkers of cardiovascular risk in young women

Medroxyprogesterone acetate (MPA) is widely known for its use in combination hormone therapy for postmenopausal women. However, MPA is also commonly used in young women for contraception and treatment of a number of gynecological conditions. Despite its widespread use, the cardiovascular effects of MPA in young women are unclear. Therefore, the purpose of this study was to determine the acute effects of MPA when used in combination with estradiol on markers of cardiovascular risk in young women. We suppressed endogenous estrogens and progesterone in 10 premenopausal women using a gonadotropin-releasing hormone antagonist (GnRHa) for 10 days. On day 4 of GnRHa subjects received 0.1 mg of estradiol (GnRHa+E(2)), and on day 7 5 mg of MPA was added (GnRHa+E(2)+MPA). Endothelium-dependent vasodilation and endothelium-independent vasodilation of the brachial artery, lipids, homocysteine, high-sensitivity C-reactive protein, and endothelin-1 were assessed during treatment with GnRHa, GnRHa+E(2), and GnRHa+E(2)+MPA. Four additional subjects were tested to validate the efficacy of the GnRHa model and confirm the findings. Endothelium-dependent vasodilation was greater during GnRHa+E(2) than during GnRHa or GnRHa+E(2)+MPA (P = 0.006). Endothelin-1 was lower during GnRHa+E(2) than GnRHa alone (P = 0.039). Endothelin-1 increased with the addition of MPA and was not significantly different from GnRHa alone. There were no differences in the other markers of cardiovascular risk between hormone treatment days. These data suggest that acute MPA administration negates the beneficial effects of estradiol on endothelium-dependent vasodilation in young women. In addition, these data suggest that estradiol decreases endothelin-1 concentrations and the addition of MPA may counteract the effect of estradiol on endothelin-1.

Effects of Cu/Zn Superoxide Dismutase on Estrogen Responsiveness and Oxidative Stress in Human Breast Cancer Cells

The effects of estrogen on gene expression in mammary cells are mediated by interaction of the estrogen receptor (ER) with estrogen response elements in target DNA. Whereas the ER is the primary initiator of transcription, the recruitment of coregulatory proteins to the DNA-bound receptor influences estrogen responsiveness. To better understand how estrogen alters gene expression, we identified proteins associated with the DNA-bound ERalpha. Surprisingly, the antioxidant enzyme Cu/Zn superoxide dismutase (SOD1), which is known primarily as a scavenger of superoxide, was associated with the DNA-bound receptor. We have now demonstrated that SOD1 interacts with ERalpha from MCF-7 cell nuclear extracts and with purified ERalpha and that SOD1 enhances binding of ERalpha to estrogen response element-containing DNA. Although SOD1 decreases transcription of an estrogen-responsive reporter plasmid in transiently transfected U2 osteosarcoma cells, RNA interference assays demonstrate that SOD1 is required for effective estrogen responsiveness of the endogenous pS2, progesterone receptor, cyclin D1, and Cathepsin D genes in MCF-7 breast cancer cells. Furthermore, ERalpha and SOD1 are associated with regions of the pS2 and progesterone receptor genes involved in conferring estrogen-responsive gene expression. Interestingly, when MCF-7 cells are exposed to 17beta-estradiol and superoxide generated by addition of potassium superoxide (KO2) to the cell medium, SOD1 levels are increased and tyrosine nitration, which is an indicator of oxidative stress-induced protein damage, is significantly diminished. Our studies have identified a new role for SOD1 in regulating estrogen-responsive gene expression and suggest that the 17beta-estradiol- and KO2-induced increase in SOD1 may play a role in the survival of breast cancer cells and the progression of mammary tumors.

C‐reactive protein across the menstrual cycle

C-reactive protein (CRP) is a widely used, sensitive biomarker of inflammation. Studies conducted among users of exogenous hormones suggest that estrogen increases CRP, whereas progesterone decreases CRP. Examinations of CRP in normally cycling women suggest the opposite: CRP is negatively associated with endogenous estrogen and positively associated with endogenous progesterone. This work evaluates the association between menstrual cycle-related hormone changes and events (menstruation and ovulation) and CRP. Eight female subjects gave urine and blood samples from twelve days across the menstrual cycle, for a total of eleven cycles. Blood samples were assayed for CRP; urine samples for beta-follicle stimulating hormone (betaFSH), pregnanediol 3-glucuronide (PDG), and estrone glucuronide (E1G). Ovulation day was estimated using hormone levels. Presence or absence of menses was reported by subjects. Analyses were conducted with random-effects linear regression. All cycles were ovulatory; day of ovulation was identified for nine cycles. A ten-fold increase in progesterone was associated with a 23% increase in CRP (P = 0.01), a ten-fold increase in estrogen was associated with a 29% decrease in CRP (P = 0.05), and menses was associated with a 17% increase in CRP (P = 0.18); no association between ovulation or FSH and CRP was found. Hormone changes across the menstrual cycle should be controlled for in future studies of inflammation in reproductive-age women.