Browsing by Subject "Estrogens"
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Item Antagonistic Roles of miR-199a-3p/miR-214 and the miR-200 Family in the Regulation of Uterine Contractility During Pregnancy and Labor(2014-02-03) Williams, Koriand'r; Hammer, Robert; Mendelson, Carole; Mahendroo, Mala; Olson, EricProgesterone (P4) and estradiol-17β (E2) play critical and opposing roles in regulating myometrial quiescence and contractility during pregnancy and labor (Kamel et al., 2010). While these contrasting hormonal effects are likely mediated via differential regulation of inflammatory and contractile genes, the underlying mechanisms remain incompletely understood. Recently, we discovered that miR-200 family members, miR-200b and miR-429, and their target, transcription factor ZEB1, serve as P4/progesterone receptor (PR)-mediated regulators of uterine quiescence during pregnancy (Renthal et al., 2010). In the present study, we identified a novel role for another miR-200 family member, miR-200a, to enhance local metabolism of P4 in myometrium and, thus, decrease PR function during the progression towards labor (Williams et. al., 2012a). This occurs via miR-200a repression of signal transducer and activator of transcription (STAT)5b, a transcriptional repressor of the P4-metabolizing enzyme 20α-hydroxysteroid dehydrogenase (20α-HSD). We observed that miR-200a expression increased and STAT5b expression coordinately decreased in myometrium of mice as they progressed to labor and in laboring myometrium from pregnant women. These changes were associated with a dramatic increase in expression and activity of 20α-HSD in laboring myometrium from mouse and human. In a progesterone-withdrawal mouse model of preterm labor, preterm labor was associated with increased miR-200a, decreased STAT5b and enhanced 20α-HSD expression. In other studies, we also found that levels of the clustered miRNAs, miR-199a-3p and miR-214, were significantly decreased in laboring myometrium of pregnant mice and humans and in a inflammatory mouse model of preterm labor, while the miR-199a-3p/miR-214 target, cyclooxygenase-2 (COX-2), a critical enzyme in synthesis of pro-inflammatory prostaglandins, was coordinately increased (Williams et al., 2012b). The physiological relevance of the labor-associated increase in miR-199a-3p/214 expression was highlighted by the finding that overexpression of miR-199a-3p and miR-214 in cultured human myometrial cells inhibited COX-2 protein and blocked TNF-α-induced myometrial cell contractility. Notably, estrogen and P4 treatment of ovariectomized mice have opposing effects on uterine miR-199a-3p/214 expression that were mediated by ZEB1. Whereas, P4 stimulated ZEB1 and upregulated miR-199a/214 expression in mouse and human myometrium (Renthal et al., 2010), estrogen had an opposing inhibitory effect. Notably, ZEB1/2 inhibit miR-200 family expression. Together, our findings point to the key pivotal roles of myometrial ZEB1 and its miRNA targets as a hormonally-controlled regulators of inflammatory and contractile gene expression in the pregnant uterus during term and preterm labor.Item CHEMICAL MIXTURES OF XENOESTROGENS AND THEIR METABOLITES ALTER ESTRADIOL-INDUCED NON-GENOMIC SIGNALING(2013-06-03) Vinas, Rene; Watson, Cheryl S.; Cheng, Xiaodong; Laezza, Fernanda; Midoro-Horiuti, Terumi; Boonayaratanakornkit, VirojBACKGROUND: Xenoestrogens (XEs) pose a threat to human health by disrupting natural responses of physiologic estrogens. Bisphenol-A (BPA), a plastics monomer, is potently estrogenic via non-genomic signaling; however, bisphenol-S (BPS), a BPA substitute, has unknown endocrine actions. Nonylphenol is a surfactant and a ubiquitous contaminant. Our goals were to use a well-characterized estrogen-responsive cell model, the GH3/B6/F10 pituitary line, to characterize the potentially potent nongenomic signaling and functional responses to these compounds alone, as mixtures, as metabolites and congeners, and in combination with estradiol (E2), while also automating these assays to facilitate future screening of a potentially large number of XEs. METHODOLOGY: Environmentally relevant concentrations of XEs (10-15-10-7M) were assessed individually and as mixtures by challenging 1nM E2-induced responses. We quantified phospho-activation of extracellular signal-regulated kinases (ERKs) and c-Jun-N-terminal kinases (JNKs) by multi-well plate immunoassays. Cell proliferation was assessed by crystal violet assay, while apoptosis (caspase-8, -9) was assessed via the release of 7-amino-4-trifluoromethylcoumarin. Prolactin release was measured by radio-immunoassay after 1 min XE exposures. The BIOMEK FXP workstation was used to develop an automated screening system for changes in MAPKs activities due to XE exposures. RESULTS: XEs often activated MAPKs in a non-monotonic dose- and oscillating time-dependent (2.5-60 min) manner and attenuated 1nM E2 responses. While individual bisphenols did not activate JNK as NP did, the combination of all XEs with E2 generated an enhanced non-monotonic JNK dose-response. E2 and all XE compounds induced cell proliferation, while mixtures of these compounds with E2 suppressed proliferation. Caspase 8 activity was suppressed by E2, and elevated by BPS, while caspase 9 activity was inhibited by E2 and some XE combinations at later times. Mono- and di-chlorinated BPA activated, while tri-chlorinated BPA dephosphorylated ERK. Di- and tri-chlorinated BPAs caused JNK dephosphorylation. Phase II metabolites (sulfated and glucuronidated) were mostly unable to activate either kinase and in some cases severely inactivated them. CONCLUSIONS: Novel chemical analogues and conjugated forms of BPA individually or as mixtures with other known XEs had dramatic disrupting effects on physiologic estrogens, disrupting mechanisms of cell regulation and their downstream functional responses.Item Estrogen Therapy in a Viral Murine Model of Multiple Sclerosis(2012-10-19) Gomez, Francisco PascualMultiple sclerosis (MS) is an idiopathic neurodegenerative, demyelinating disease of the central nervous system (CNS). MS affects females more than males (3:1) and pregnancy reduces the number of relapses especially during the third trimester when 17-beta-estradiol (E2) and estriol (E3) are at their highest levels. In order to study the role of estrogens as potential therapeutic agents for MS we investigated their role in Theiler's murine encephalomyelitis virus (TMEV)-induced demyelination (TVID). SJL female mice were infected intracranially with Theiler's virus or PBS. The mice in the treatment groups were clinically scored and at week 20 they were ovarectomized (OVx) and given a subdermal pellet containing either 1) 0.1mg of E2, 2) 5mg of E3, or 3) placebo. Four weeks after treatment initiation, the mice were sacrificed and tissue samples were collected and vertebral columns and brains were fixed and placed in paraffin for histological analysis using either hematoxylin and eosin (H&E) stain for general anatomic features or Weil's stain for myelin. No signs of clinical disease developed in any of the sham-infected mice. Prior to ovariectomy, infected mice had developed significant clinical scores indicative of demyelination. Mice in the placebo and E3-treatment groups deteriorated rapidly whereas the E2-treated mice improved significantly during the course of the treatment. Uteri were used to assess hormonal effects post-ovariectomy. Hormone treated groups were significantly different from placebo, indicating hormones were present. Hormone treatment showed significant differences among treatment groups for both inflammation and demyelination. E2-treatment significantly decreased inflammation compared to placebo and E3. E2 was also effective in reducing demyelination compared to placebo groups but not E3. E3 treatment was effective in reducing inflammation compared to placebo, but no significance was found for demyelination. Both E3 and E2 treated mice developed lower antibody levels against TMEV. The improvement in clinical signs, inflammation, demyelination, and the reduction of antibody levels in 17-beta-estradiol-treated mice indicate a therapeutic potential for the treatment of MS.