Gonadotropin releasing hormone antagonists suppress aromatase and anti-Müllerian hormone expression in human granulosa cells

Winkler, Nurit; Bukulmez, Orhan; Hardy, Daniel B.; Carr, Bruce R.

doi:10.1016/j.fertnstert.2009.09.032

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Volume 94, Issue 5 , Pages 1832-1839, October 2010

Gonadotropin releasing hormone antagonists suppress aromatase and anti-Müllerian hormone expression in human granulosa cells

Nurit Winkler, M.D.
- Department of Obstetrics and Gynecology, the University of Texas Southwestern Medical Center at Dallas, Dallas, Texas
Orhan Bukulmez, M.D.
- Department of Obstetrics and Gynecology, Division of Reproductive Endocrinology and Infertility, University of Florida College of Medicine, Gainesville, Florida
Daniel B. Hardy, Ph.D.
- Departments of Obstetrics and Gynecology & Physiology and Pharmacology, The University of Western Ontario, London, Ontario, Canada
Bruce R. Carr, M.D.
- Department of Obstetrics and Gynecology, the University of Texas Southwestern Medical Center at Dallas, Dallas, Texas
- Reprint requests: Bruce R. Carr, M.D., Department of Obstetrics and Gynecology, Division of Reproductive Endocrinology and Infertility, University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Boulevard, Dallas, Texas 75235-9032 (FAX: 214-648-8066).

Received 9 May 2009; received in revised form 15 September 2009; accepted 16 September 2009. published online 06 November 2009.

Objective

To investigate the effects of a gonadotropin-releasing hormone antagonist (GnRH-ANT) on the expression of anti-Müllerian Hormone (AMH) and aromatase (via the exon CYP19IIa promoter), in cultured human granulosa cells (hGCs) and the human granulosa cell line (HGL5).

Design

Primary cell cultures of hGCs and culture of HGL5 cells.

Setting

Academic center.

Patient(s)

Women undergoing IVF because of male factor, tubal infertility, or donor eggs.

Intervention(s)

hGCs and HGL5 cells were treated with a GnRH-ANT (1 nM and 1 μM) alone or in combination with cAMP (1 mM). Media was collected and stored at −80°C until assayed.

Main Outcome Measure(s)

mRNA levels of CYP19 IIa, AMH, steroidogenic factor 1 (SF-1) and liver receptor homologue-1 (LRH-1) were determined by quantitative polymerase chain reaction. ELISA was used to determined estradiol (E₂) levels in the culture media. Pooled results from triplicate experiments were analyzed using one-way analysis of variance with Student–Newman–Keuls multiple-comparison methods.

Result(s)

The GnRH-ANT decreased the expressions of CYP19 IIa, AMH, SF-1, and LRH-1. cAMP induced aromatase and AMH expression. Cotreatment with cAMP and GnRH-ANT caused a dose-dependent suppression of AMH and CYP19 IIa mRNA. A GnRH agonist (GnRH-A) increased the mRNA expressions of CYP 19 IIa and AMH. The GnRH-ANT decreased E₂ production in cultured hGCs.

Conclusion(s)

GnRH-ANTs, in addition to their central suppressive effects on the pituitary, may have a direct effect on ovarian granulosa cells with inhibition of aromatase and AMH expression. Furthermore, the inhibitory effect could be mediated via suppression of SF-1 and LRH-1, and may play a role in estrogen-mediated ovarian folliculogenesis.

Key Words: AMH, MIS, aromatase, CYP 19 IIa, GnRH antagonist, GnRH agonist, GnRH analogues