Effect of Progestin, Antiprogestin, and Relaxin on the Accumulation of Prolactin and Insulin-Like Growth Factor-Binding Protein-1 Messenger Ribonucleic Acid in Human Endometrial Stromal Cells1

Abstract

Prolactin (PRL) and insulin-like growth factor-binding protein (IGFBP-1) are two major secretory proteins of human endometrial/decidual cells. We have characterized the mRNA of PRL and IGFBP-1 and studied the effect of progestin, medroxyprogesterone acetate (MPA), anti-progestin (RU486), and relaxin (RLX) on the levels of these two mRNA transcripts in a long-term culture of human endometrial stromal cells. Northern blot analysis showed that the size of PRL mRNA was 1.15 kb and that of IGFBP-1 mRNA, 1.6 kb. Primer extension of endometrial/decidual IGFBP-1 mRNA showed two transcription initiation sites identical to those found in HepG2 human hepatoma cell line. The levels of mRNA in control samples remained low, approximately 2 pg PRL and approximately 5 pg IGFBP-1/microgram RNA at various times of culture. When stromal cells were treated with MPA for 28 days, PRL mRNA gradually increased 100-fold whereas IGFBP-1 mRNA exponentially increased approximately 1000-fold compared to control values and leveled after 25 days in culture. The timing of maximal stimulation was shortened by withdrawing MPA or by replacing MPA with RU486. After removal of MPA, levels of both mRNAs increased and each peaked after approximately 10 days, with PRL showing a 2-fold and IGFBP-1 a 20-fold increase compared to cells treated with MPA continuously. Replacing MPA by RU486 caused a rapid increase of PRL mRNA (2-3-fold) in 2-3 days followed by a gradual reduction to less than 20% of peak levels over the next 3 days. IGFBP-1 mRNA levels increased 30- and 100-fold in 1-2 days followed by a reduction to less than 20% of peak levels over the next 24 h. The reduction of mRNA levels by RU486 was reversed when cells were rechallenged with MPA. Relaxin alone caused a transient stimulation of PRL and IGFBP-1 mRNA. Maximal stimulation occurred between 10 and 20 days of culture and was 100-fold for PRL and 1000-fold for IGFBP-1 relative to control values. Cells treated with MPA and RLX in sequence had higher mRNA levels than cells treated with MPA continuously or cells subjected to MPA withdrawal. Maximal mRNA levels reached 0.4 ng PRL and approximately 8 ng IGFBP-1/microgram total RNA, approximately 0.04% and 0.8% of cellular RNA. The mRNA levels under various hormonal manipulations were similar to the previously published synthesis and secretion patterns of PRL and IGFBP-1 proteins in this system.(ABSTRACT TRUNCATED AT 400 WORDS)

Cyclic AMP and progesterone receptor cross-talk in human endometrium: a decidualizing affair

“…Peptide hormones and prostanoids implicated in promoting the decidual transformation include the gonadotrophins luteinizing hormone/ human chorionic gonadotrophin (LH/hCG), corticotrophin releasing hormone (CRH), relaxin (RLX), and prostaglandin E 2 (PGE 2 ) (Tseng et al 1992, Tang & Gurpide 1993, Frank et al 1994, Ferrari et al 1995. These molecules share the ability to provoke an increase in intracellular cAMP levels by binding to GPCR coupled to the stimulatory Gs protein (Gilchrist et al 1996, Herrlich et al 1996, Gravanis et al 1999, Narumiya et al 1999, Breyer et al 2001, Narumiya & FitzGerald 2001, Hillhouse & Grammatopoulos 2002, Hsu et al 2002, Sudo et al 2002.…”

“…In the context of decidualization, however, PR-A and LAP are the predominant isoforms and, hence, progesterone may be essential for maintaining and enhancing the expression of C/EBP -dependent genes in the decidua. Further support for the important role of PR-A in decidualization stems from the finding that the expression of IGFBP-1, another major decidual product which is regulated by progesterone and cAMP signalling (Tseng et al 1992, Frank et al 1994, Kim et al 1998, is more strongly induced by ligandactivated PR-A than PR-B (Gao et al 2000).…”

“…1). The decidualized endometrial stromal cell (ESC) becomes rounded, acquires myofibroblast characteristics, and secretes a variety of phenotypic antigens, including prolactin (PRL), insulin-like growth factor binding protein-1 (IGFBP-1) and tissue factor (TF) (Daly et al 1983, Irwin et al 1989, Tabanelli et al 1992, Tseng et al 1992, Lockwood et al 1993, Gellersen et al 1994, Oliver et al 1999b, Christian et al 2001b. At a molecular level, decidual transformation involves extensive reprogramming of many cell functions including altered steroid hormone receptor expression and steroid metabolism, remodelling of the extracellular matrix and cytoskeleton, altered expression of intracellular enzymes, growth factors and cytokines and their receptors, and induction of apoptosis modulators and decidua-specific transcription factors (Oliver et al 1999a, Popovici et al 2000, Brar et al 2001.…”

Effect of cryopreservation on the properties of human endometrial stromal cells used in embryo co-culture systems

“…Conditioned media from ESCs decidualized with MPA and β-Estradiol were assayed for secretion of PRL and IGFBP-1, which are wellestablished biochemical markers and quantitative indicators for decidualization [56]. Decreased PRL protein levels were measured in all 17 post-cryopreserved ESC cultures as compared to corresponding fresh ESCs at individual level (Fig.…”

Stimulation of a rat uterine stromal cell line in culture reveals a molecular switch for endocrine-dependent differentiation

“…PRL, IGFBP-1 and tissue factor are sensitive markers for differentiating human stromal cells (Irwin et al 1989, Tseng et al 1992, Lockwood et al 1993, Christian et al 2001. The tissue-specific expression of human PRL occurs by utilization of an alternative promoter region that becomes activated in differentiating cells (Berwaer et al 1994, Telgmann & Gellersen 1998.…”