J. C. Mordacq

Transcriptional regulation of proliferin gene expression in response to serum in transfected mouse cells

The sequences upstream of a proliferin gene have been isolated, linked to a reporter gene, and transfected into mouse cell cultures. In low serum concentrations, transcription from the transfected DNA is very weak; transcriptional activity is induced 20‐ to 40‐fold in transfected cultures grown in high serum concentrations. Initiation of transcription occurs at the same site in the transfected DNA as in endogenous proliferin genes expressed in placental tissue and in cell cultures. Sequences within 578 nucleotides upstream of this initiation site are sufficient for complete serum‐inducible expression, but deletion of the upstream sequences to within 211 nucleotides of the start site abolishes promoter activity. In contrast, the upstream region from a second proliferin gene is only weakly inducible in transfected cell cultures, even though these two promoter regions share 97% nucleotide sequence homology.

Rat ovarian insulin-like growth factor binding protein-4 : A hormone-dependent granulosa cell-derived antigonadotropin

Objective: To assess the in vivo regulation of ovarian insulin-like growth factor binding protein-4 (GFBP-4) mRNA expression by gonadotropins and estrogen. Methods: Whole varian RNA, obtained from two models of follicular development, was extracted and analyzed by Northern blotting. Immature rats were treated with pregnant mare senum gonadotropin (PMSG) followed 48 hours later with hCG, or alternatively were hypophysectomized and treated with FSH and/or diethylstilbestrol (DES). Localization of IGFBP-4 expression was assessed in the former study by in situ hybridization. Finally, the ability of human IGFBP-4 to antagonize FSH-stimulated progesterone accumulation was assessed in vitro. Results: The ovarian content of IGFBP-4 transcripts increased threefold (P < .05) at 12 hours after PMSG but was near baseline at 24 and 48 hours. The abundance of IGFBP-4 mRNA increased (P < .05) again at 6 and 24 hours after hCG. The expression of IGFBP-4 was localized to granulosa cells of prenatral (untreated) and small antral (12 hours after PMSG) follicles. No IGFBP-4 expression was noted in large (gonadotropin-primed) antral follicles. Hypophysectomy increased (P < .05) the ovarian content of IGBP-4 mRNA by 1.5-fold, an effect further enhanced (1.8-fold; P < .05) by the provision of FSH and DES. In vitro studies revealed the ability of increasing concentrations (0.01-1 μg/mL) of recombinant human IGFBP-4 to inhibit the FSH-supported accumulation of progesterone. Conclusion: Increased expression after administration of PMSG, hCG, and FSH/DES suggests that IGFBP-4 is a dynamic and hormonally responsive component of the ovarian cycle. The lack of expression in preovulatory follicles and its antigonadotropic actions in vitro imply that the attenuated expression of IGFBP-4 may constitute a requirement for successful follicular maturation.

In vivo hormonal regulation of insulin-like growth factor binding protein-5 mRNA in the immature rat ovary.

Objective: Despite the potential importance of insulin-like growth factor binding protein-5 (IGFBP-5) to follicular development, the hormonal regulation of this antigonadotropic IGFBP has not been investigated. Therefore, it was the objective of this study to eludicate the role of gonadotropins and estrogen in the in vivo regulation of IGFBP-5 mRNA expression. Methods: Two models of follicular development in immature rats were used. Specifically, rats were hypophysectomized and treated with FSH and/or diethylstilbestrol (DES). Alternatively, terminal follicular development was induced in intact immature rats by pregnant mare serum gonadotropin (PMSG) and hCG. The IGFBP-5 mRNA in whole ovarian RNA was assayed by Northern blot hybridization. Localization of expression in PMSG and hCG-stimulated ovaries was further assessed by in situ hybridization. Results: Expression of IGFBP-5 mRNA was increased in ovaries from hypophysectomized rats. Treatment with FSH and/or DES did not alter the abundance of this mRNA. Treatment with PMSG induced a transient increase in IGFBP-5 expression that was localized in a subset of α-inhibin-negative follicles. At later times after PMSG, IGFBP-5 expression persisted in the surface epithelium but was not detected in large preovulatory follicles. In vitro studies affirmed the antigonadotropic action of IGFBP-5. Conclusion: In vivo expression of IGFBP-5 in the rat ovary is moderated by hormonal treatment both in terms of total expression and follicular localization.