Expression and regulation of gonadotropin-releasing hormone (GnRH) and GnRH receptor messenger ribonucleic acids in human granulosa-luteal cells.

Abstract

The present study investigated the expression and regulation of GnRH and GnRH receptor (GnRHR) messenger RNAs (mRNAs) in human granulosa-luteal cells using reverse transcription-polymerase chain reaction (RT-PCR). Granulosa-luteal cells were aspirated from preovulatory follicles obtained from women undergoing in vitro fertilization. Two sets of primers derived from human hypothalamic GnRHR complementary DNA (cDNA) were used to amplify cDNAs from granulosa-luteal cells. PCR products corresponding to the expected sizes of GnRH were obtained from granulosa-luteal cells as well as the brain, but not from skeletal muscle cDNA. The authenticity of the PCR products was confirmed by Southern blot hybridization with internal oligonucleotide probes and by subsequent cloning and sequencing. Similarly, using four sets of primers specific for the human pituitary GnRHR cDNA, PCR products with the expected sizes were detected from both brain and granulosa-luteal cells, but not from skeletal muscle. PCR products were subsequently confirmed by Southern blot hybridization using an internal oligonucleotide probe or a cDNA probe which was obtained from screening a human pituitary cDNA library. Cloning and sequencing of the PCR product in the 3'-untranslated region revealed identical sequence with the reported human pituitary GnRHR cDNA sequence. RNA samples obtained from cells immediately after dissociation or after 2, 5, and 8 days of culture were analyzed by RT-PCR, and in all cases, both GnRH and GnRHR mRNA were detected. To investigate how gene expression of GnRH and GnRHR is regulated, we examined the effect of GnRH and hCG on GnRH and GnRHR mRNA levels in cultured human granulosa-luteal cells. Treatment with different concentrations of GnRH induced biphasic responses. Both GnRH and GnRHR mRNA were significantly increased by 1 nM, but slightly decreased by 1 microM GnRH; 1 nM GnRH also significantly inhibited progesterone production, whereas higher doses had no effect. Treatment with hCG (1 IU/ml) decreased GnRHR mRNA levels without altering the expression of the GnRH gene. These results demonstrate for the first time that 1) both GnRH and GnRHR mRNAs are expressed in human granulosa-luteal cells; 2) GnRH mRNA levels are autoregulated by GnRH; and 3) GnRHR gene expression is up-regulated by GnRH, but down-regulated by hCG. These findings provide strong evidence that GnRH is an autocrine regulator in the human ovary.

Ovarian recovery after stem cell transplantation

“…15 It is also possible that GnRH analogs may exert direct protective effect on the ovary via peripheral GnRH receptors, which have been documented on human granulosa cells found in the developing and mature follicles. 16 However, there is lack of consistent reports from pilot human clinical studies and the benefit of ovarian protection by GnRHa is still unproven. There are limited prospective data and randomized controlled studies in humans, which are flawed by short-term follow-up and lack of controls.…”

Biology of the gonadotropin-releasing hormone system in gynecological cancers

“…In addition to this well-documented classic hypophysiotropic action, GnRH might have a role as a modulator of the activity of diverse systems in the brain and many peripheral organs (1, 4-10). An autocrine/paracrine function of GnRH has been suggested to exist, for instance, in the placenta (11 -14), granulosa cells (15)(16)(17), myometrium (18) and lymphoid cells (19 -21). In addition, it is probable that such GnRH-based autocrine systems are present in a number of human malignant tumors, including cancers of the breast, ovary, endometrium and prostate.…”

Genomic Structure and Expression Sites of Three Gonadotropin-Releasing Hormone Genes in One Species

“…By using the reverse transcription-polymerase chain reaction (RT-PCR) on human tissue, GnRH mRNA has been found in liver, heart, skeletal muscle, kidney, and placenta (e.g., Kakar and Jennes, 1995). GnRH gene expression has also been reported in the testis of several primates and rats (Dong et al, 1996), human ovary (Peng et al, 1994), and the rat prostate (Azad et al, 1993b). In H. burtoni we also found widespread GnRH expression by using RT-PCR, with heart, liver, spleen, kidney, testis, and brain of an adult male all containing 5Ser 8 6GnRH mRNA.…”