Developmental Programming: Gestational Testosterone Treatment Alters Fetal Ovarian Gene Expression

Department of Molecular and Integrative Physiology, University of Kansas Medical Center, Kansas City, Kansas 66160, USA.
Endocrinology (Impact Factor: 4.5). 12/2011; 152(12):4974-83. DOI: 10.1210/en.2011-1182
Source: PubMed


Prenatal testosterone (T) treatment leads to polycystic ovarian morphology, enhanced follicular recruitment/depletion, and increased estradiol secretion. This study addresses whether expression of key ovarian genes and microRNA are altered by prenatal T excess and whether changes are mediated by androgenic or estrogenic actions of T. Pregnant Suffolk ewes were treated with T or T plus the androgen receptor antagonist, flutamide (T+F) from d 30 to 90 of gestation. Expression of steroidogenic enzymes, steroid/gonadotropin receptors, and key ovarian regulators were measured by RT-PCR using RNA obtained from fetal ovaries collected on d 65 [n = 4, 5, and 5 for T, T+F, and control groups, respectively] and d 90 (n = 5, 7, 4) of gestation. Additionally, fetal d 90 RNA were hybridized to multispecies microRNA microarrays. Prenatal T decreased (P < 0.05) Cyp11a1 expression (3.7-fold) in d 90 ovaries and increased Cyp19 (3.9-fold) and 5α-reductase (1.8-fold) expression in d 65 ovaries. Flutamide prevented the T-induced decrease in Cyp11a1 mRNA at d 90 but not the Cyp19 and 5α-reductase increase in d 65 ovaries. Cotreatment with T+F increased Cyp11a1 (3.0-fold) expression in d 65 ovaries, relative to control and T-treated ovaries. Prenatal T altered fetal ovarian microRNA expression, including miR-497 and miR-15b, members of the same family that have been implicated in insulin signaling. These studies demonstrate that maternal T treatment alters fetal ovarian steroidogenic gene and microRNA expression and implicate direct actions of estrogens in addition to androgens in the reprogramming of ovarian developmental trajectory leading up to adult reproductive pathologies.

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    • "Two of these miRNAs (miR-15b and miR-497) were examined by qRT-PCR, which confirmed their up-regulation in the ovaries of testosterone-exposed fetuses. Bioinformatics analysis identified members of the insulin-signaling pathway as potential targets of miR-15b and miR-497, a pathway that is known to be dysregulated in PCOS patients (Luense et al. 2011). Subsequent in vitro and in vivo studies are needed to identify potential targets of these miRNA as well as define the functional importance of these miRNAs. "
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    ABSTRACT: MicroRNAs (miRNAs) are posttranscriptional gene regulatory molecules that show regulated expression within ovarian tissue. Most research investigating miRNAs in the ovary has relied exclusively on in vitro analyses. In this review, we highlight those few studies in which investigators have illustrated an in vivo effect of miRNAs on ovarian function. We also provide a synopsis of how these small noncoding RNAs can impact ovarian disease. miRNAs have great potential as novel diagnostic biomarkers for the detection of ovarian disease and in the assisted reproductive technologies (ART) for selection of healthy viable oocytes and embryos. Copyright © 2015 Cold Spring Harbor Laboratory Press; all rights reserved.
    Cold Spring Harbor Perspectives in Medicine 05/2015; 5(9). DOI:10.1101/cshperspect.a022962 · 9.47 Impact Factor
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    • "s ( dot within the boxes ) and the 25 – 75 percentiles ( box sizes ) , and whiskers delineate the minimum and maximum values . Statistically significant p - value was obtained using the Mann – Whitney U - test . Scale bar represents 50 lm . n . s . = not significant 3b - HSD , CYP17 and 17b - HSD Expression in Foetal Gonads in Pig 7 et al . 2011 ; Luense et al . 2011 ) . In our study , we showed that prenatal flutamide treatment altered 3b - HSD mRNA expression at the same pattern as was observed for foetal testes . However , the immunostaining intensity of 3b - HSD was higher only in the GD90 group . On the other hand , flutamide treatment resulted in decreased CYP17 mRNA expression in the GD90 gro"
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    ABSTRACT: ContentsWe investigated whether the limited access to androgens during late prenatal period alters expression of steroidogenic enzymes involved in androgen production: 3β-hydroxysteroid dehydrogenase/Δ5-Δ4 isomerase (3β-HSD), cytochrome P450 17α-hydroxylase/17,20-lyase (CYP17) and 17β-hydroxysteroid dehydrogenase type 1 (17β-HSD1) or type 3 (17β-HSD3) in the foetal porcine gonads. Pregnant gilts were injected with anti-androgen flutamide (for seven days, 50 mg/day/kg bw) or corn oil (control) starting at 83 (GD90) or 101 (GD108) gestational day. To assess 3β-HSD, CYP17 and 17β-HSD1 or 17β-HSD3 expression, real-time PCR and immunohistochemistry were performed. In testes from flutamide-treated foetuses, increased 3β-HSD and CYP17 mRNA expression was observed in the GD90 group, while decreased 3β-HSD and 17β-HSD3 mRNA expression and increased CYP17 mRNA expression were found in the GD108 group. CYP17 and 17β-HSD3 were localized in Leydig cells. Following flutamide administration, the intensity of CYP17 immunostaining was higher in both treated groups, while 17β-HSD3 intensity was lower in the GD108 group. In ovaries from flutamide-treated foetuses in the GD90 group, mRNA level for 3β-HSD was elevated, but it was diminished for CYP17 and 17β-HSD1. In the GD108 group, flutamide treatment led to lower mRNA level for 3β-HSD but higher for CYP17. 3β-HSD was found in granulosa cells, while CYP17 was localized within egg nests and oocytes of forming follicles. Following flutamide treatment, the intensity of 3β-HSD and CYP17 immunostaining was higher in the GD90 and GD108 groups, respectively. Immunohistochemical staining for 3β-HSD was restricted to the ovary. Concluding, diminished androgen action in the porcine foetal gonads during late gestation induces changes in steroidogenic enzymes expression, which may led to changes in gonadal function. However, it seems that androgens exert diverse biological effects depending on the gestational period.
    Reproduction in Domestic Animals 08/2014; 49(5). DOI:10.1111/rda.12356 · 1.52 Impact Factor
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    • "In the past decade, there have been an increasing number of attempts to better understand the role of androgen signaling within the ovary during fetal and neonatal developmental periods. Studies using either prenatal androgen excess [1] [2] or deficiency [3] as a model both showed that disturbed androgen action led "
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    ABSTRACT: Recent studies suggest that disturbed androgen action during gestational and neonatal periods leads to reprogramming of the trajectory of ovarian development, manifested by altered follicular functioning in adulthood. In this study, we tested whether prenatal and neonatal exposure to antiandrogen flutamide affected ovarian 17β-estradiol (E(2)) synthesis and the associated gene expression in large antral follicles of adult pigs. Flutamide was injected into pregnant gilts between Days 80 and 88 of gestation and into female piglets between Days 2 and 10 postnatally. After animals reached sexual maturity, the ovaries were collected from treated and nontreated (control) pigs. The analysis of E(2) concentration in follicular tissues, as well as FSH and LH levels in plasma of control and flutamide-treated animals were conducted. In addition, the expression of mRNAs and proteins for FSH receptor (FSHR), cytochrome P450 aromatase (CYP19A1) and β-catenin (CTNNB1) was examined in large antral follicles of adult pigs. The E(2) concentration was greater in response to flutamide administered prenatally (P < 0.05) and neonatally (P < 0.01), whereas there was no changes in plasma gonadotropin concentration. Real-time polymerase chain reaction analysis revealed significant upregulation of FSHR, CYP19A1, and CTNNB1 at the mRNA level after maternal (P < 0.001, P < 0.01, P < 0.05, respectively) and neonatal (P < 0.001, P < 0.001, P < 0.01, respectively) flutamide exposure. The expression of FSHR protein was higher (P < 0.01) only after neonatal exposure to flutamide, whereas CYP19A1 and CTNNB1 proteins were upregulated in response to both prenatal (P < 0.01) and neonatal (P < 0.001) flutamide administration. Furthermore, membranous CTNNB1 immunolocalization indicates that it is not involved in regulation of FSH-mediated CYP19A1 activity as a transcription factor, but rather contributes to the intercellular adhesion. Concluding, it appears that the higher E(2) level in response to flutamide treatments is a result of the intensified aromatization and local E(2) action at the ovary level. The observed changes might influence the normal follicle development and pig fertility as a consequence.
    Theriogenology 10/2012; 78(9). DOI:10.1016/j.theriogenology.2012.07.026 · 1.80 Impact Factor
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