Estrogen receptor-alpha (Esr1) mediates estrogen action in regulating at all levels of the hypothalamic-pituitary-ovarian axis. Whereas the importance of Esr1 in hypothalamus and pituitary has been demonstrated by loss of fertility in the neuron- and pituitary-specific Esr1 knockout mice, whether Esr1 plays a critical role in the ovary remains to be determined. In the ovary, Esr1 is mainly expressed in the theca/interstitial cells and germinal epithelium and thus is believed to mediate estrogen action in these cells. In this study, we assessed the importance of Esr1 in the ovarian theca cells in regulating female reproduction. The Cre-LoxP approach was used to selectively delete the Esr1 gene in the theca cells, and the reproductive consequence of the deletion was measured. Adolescent theca-specific Esr1 knockout (thEsr1KO) mice (<4 months of age) are fertile and cycling. However, they begin to display an erratic pattern of estrous cycles and become infertile before they reach the age of 6 months. The ovaries of thEsr1KOmice (>or=4 months) have fewer corpora lutea but more antral follicles than the age-matching wild-type mice. The numbers of 17-hydroxylase-expressing cells are largely increased in the interstitium of the thEsr1KO mouse ovary. Interestingly, whereas basal levels of serum testosterone and FSH were mildly elevated, LH level was either markedly lower or undetectable in the thEsr1KO mice. When superstimulated by exogenous gonadotropins, thEsr1KO mice released significantly fewer oocytes that wild-type littermates and developed multiple hemorrhagic cysts. Taken together, this study demonstrates that theca Esr1 plays a critical role in regulating female reproduction.
A transgenic mouse line that expresses iCre under regulation of the Cytochrome P 450 17α-hydroxylase/17, 20-lyase (Cyp17) promoter was developed as a novel transgenic mouse model for the conditional deletion of genes specifically in the theca/interstitial cells of the ovary and Leydig cells of the testis. In this report we describe the development of Cyp17iCre mice and the application of these mice for conditional deletion of the estrogen receptor alpha (Esr1) gene in the theca/ interstitial and Leydig cells of the female and male gonad, respectively. These mice will prove a powerful tool to inactivate genes in the gonad in a cell-specific manner.Targeted gene deletion has become a powerful tool in the study of gene function with the utilization of this technology leading to marked progress in our understanding of both physiological and pathophysiological systems. The ovary is one organ where targeted genetic deletion has proven fruitful with the establishment of transgenic mice with Cre expression targeted to granulosa cells (Lécureuil et al., 2002), somatic cells (Bingham et al., 2006) and the oocyte (Lan et al., 2004;Lewandoski et al., 1997). However, our understanding of ovarian function cannot advance at optimal pace without a tool to specifically delete genes of interest from the other major endocrine cell population of the ovary, the theca/interstitial cells. Hence, our primary goal was to develop the mice necessary to allow the specific deletion of genes in the theca/interstitial cells of the ovary and in this report we describe the generation of such a line of transgenic mice, with codon-improved Cre (iCre) driven by the promoter to Cytochrome P 450 17α-hydroxylase/17, 20-lyase (Cyp17).Cytochrome P 450 17α-hydroxylase/17, 20-lyase, the product of Cyp17 gene expression, plays a major role in the control of sex steroid hormone synthesis by mediating the17α-hydroxylation of pregnenolone or progesterone to dehydroepiandrostenedione or androstenedione, respectively. In the female mouse, Cyp17 expression is primarily restricted to the ovary and placenta (Su et al., 2002) and within the ovary, Cyp17 is abundant in the gonadotropin-primed theca/interstitial cell population, but not the granulosa cells or oocyte (Zhang et al., 2001), making it ideal for iCre targeting. Furthermore, coincident to the need of a transgenic mouse line with iCre targeted to the theca/interstitial cells, is one also designed to allow the deletion of Leydig cell specific genes from the male gonad. Fortunately, the specificity of Cyp17 expression to the Leydig cells of the testis (Zhang et al., 2001) makes this line of mice also ideal as a tool to inactive genes specifically within that endocrine population of cells. Hence, this report was widened as a functional characterization of these mice inclusive to both the sexes.Three lines of Cyp17iCre founder mice were generated (A, B and C) by pronuclear injection of a KpnI/SalI DNA fragment derived from a Cyp17iCre expression plasmid (Figure 1). Then, to facilitate expression analys...
Estrogen acts to prime the pituitary prior to the GnRHinduced LH surge by undiscovered mechanisms. This study aimed to identify the key components that mediate estrogen action in priming the pituitary. RNA extracted from the pituitaries of metestrous (low estrogen) and proestrus (high estrogen) stage mice, as well as from ovariectomized wildtype and estrogen receptor a (ERa) knockout mice treated with 17b-estradiol (E 2 ) or vehicle, was used for gene expression microarray. Microarray data were then aggregated, built into a functional electronic database, and used for further characterization of E 2 /ERa-regulated genes. These data were used to compile a list of genes representing diverse biological pathways that are regulated by E 2 via an ERa-mediated pathway in the pituitary. This approach substantiates ERa regulation of membrane potential regulators and intracellular vesicle transporters, among others, but not the basic components of secretory machinery. Subsequent characterization of six selected genes (Cacna1a, Cacna1g, Cited1, Abep1, Opn3, and Kcne2) confirmed not only ERa dependency for their pituitary expression but also the significance of their expression in regulating GnRH-induced LH secretion. In conclusion, findings from this study suggest that estrogen primes the pituitary via ERa by equipping pituitary cells with critical cellular components that potentiate LH release on subsequent GnRH stimulations.
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