Hormonal Regulation of Human Follicle-Stimulating Hormone-β Subunit Gene Expression: GnRH Stimulation and GnRH-Independent Androgen Inhibition

Abstract

Previous studies from our laboratory demonstrated that chronic testosterone administration to castrated transgenic mice suppressed human follicle-stimulating hormone-β (FSHβ) mRNA levels transcribed from a human transgene to approximately 20% of control values. In the present study we used primary pituitary cultures prepared from the transgenic mice and in vivo experiments in hypogonadal (hpg) mice carrying the human transgene to assess the role of hypothalamic gonadotropin-releasing hormone (GnRH) in this inhibitory action. The levels of human FSHβ mRNA in monolayer cultures of pituitary cells were decreased by 24-hour treatments with 10 nM testosterone propionate or 5α-dihydrotestosterone to 13 and 26% of control values, respectively, in the absence of GnRH. For the in vivo experiments we introduced the 10-kb human FSHβ transgene into the hpg genetic background by selective crossbreeding. Daily injections of 1 µg GnRH for 14 days induced expression of the human FSHβ gene in male and female mice. Maximal effects were obtained by GnRH treatment of gonadectomized, hpg transgenic mice. Human FSHβ mRNA levels rose to approximately 4- or 10-fold that of control males and females, respectively. The stimulation was blocked completely by simultaneous administration of testosterone propionate in males and partially by estradiol in females. Pituitary content of immunoreactive FSH paralleled the mRNA changes. These data suggest that testosterone feedback inhibits the human FSHβ subunit gene directly at the pituitary gland in addition to the indirect mechanism of GnRH suppression. Furthermore, the in vitro data indicate that the suppression of human FSHβ gene expression is at least partly a direct androgen effect that does not require aromatization of testosterone to estradiol.

LbetaT2 gonadotroph cells secrete follicle stimulating hormone (FSH) in response to active A

“…Primary rat pituitary cultures have been a significant tool, but are not sustainable in culture. Further, although primary rat pituitary cultures in general demonstrate similar regulation to that observed in physiological studies in humans, we have observed some differences among the human FSHβ and mouse FSHβ genes in transgenic animals and the rat FSHβ gene, specifically with regard to androgen regulation (Kumar & Low 1995b). We postulate that the mouse gene may be more comparable to the human gene, making the LβT2 cell line, derived from a mouse pituitary adenoma, a more appropriate model.…”

Gonadotrope-specific Deletion of Dicer Results in Severely Suppressed Gonadotropins and Fertility Defects

“…Mice-Gender-specific regulation of gonadotropins has been well studied (68,70,74). To determine if loss of Dicer in gonadotropes affects gonadotropin synthesis and secretion in females similar to males, we first analyzed gonadotropin subunit mRNAs by real time qPCR assays using purified gonadotropes (GFP ϩ ) prepared from the pituitaries of adult female mice.…”

“…Isolation of Gonadotropes by Fluorescence-activated Cell Sorting-Pools of pituitaries from 3-4 adult mice of identical genotypes were surgically resected and enzymatically dispersed using collagenase treatment at 37°C as described (70). The final cell suspension was filtered through 100-m nylon cap-containing tubes (BD Biosciences) to remove cell clumps.…”

“…Gonadectomy and Superovulation Assays-Adult mice were surgically gonadectomized under isoflurane anesthesia and maintained for 1 week to totally deplete the endogenous steroids as described (70,73,74). At this point, blood was collected for hormone assays; one-half pituitary was used for RNA and the other half for Western blot analyses as described above.…”

Mechanisms of FSH synthesis: what we know, what we don't, and why you should care

“…In pituitary cultures from transgenic mice bearing a 10-kb human FSHB minigene (discussed above), androgens suppress both FSH release and human FSHB mRNA levels (the endogenous murine Fshb mRNA levels were too low to measure in the assays used at the time) (240). In vivo studies with GnRH-deficient men similarly suggest a suppressive effect of androgens on FSH release mediated at the pituitary level (241)(242)(243).…”