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Local production and action of prolactin-releasing peptide (PrRP) in human decidua.
Materials/Methods: Human luteinized granulosa cells were obtained from the follicular fluid by transvaginal oocyte aspiration from infertile patients undergoing controlled ovarian hyperstimulation (COH) for in vitro fertilization (IVF). The cells were cultured for 72 hours with TNF-alpha concentrations of 1.0, 10.0, and 100.0 ng/ml, respectively. The cells not treated with TNF-alpha served as control. The concentrations of E2, P4, IGF-I, IGFBP-1, 2, and 3 were determined in conditioned culture media by immunoradiometric assay (IRMA) or radioimmunoassay (RIA). Results: The cell number in 100.0 ng/ml of TNF-alpha group was significantly higher than those in other groups, although the cell viabilities were similar in all groups. There were no statistically significant differences in the concentrations of E2 in all groups. However, the concentrations of P4 was seemed to be decreased as the concentrations of TNF-alpha were increased and the concentration of P4 in 100.0 ng/ml of TNF-alpha group was significantly lower than those in the control and other TNF-alpha groups. The concentrations of IGF-II, IGFBP-1, 2, and 3 were not different among the control and each TNF-alpha group. The secretion of E2 and P4 was not affected by IGF type I receptor antibody pretreatment. Conclusions: TNF-alpha might play a role as a regulator of ovarian physiology by modulating luteinized granulosa cellular proliferation and P4 secretion, and this mechanism might not be related to IGF system.
P-301 Expression of HOXA10 gene in human endometrium and of infertile and fertile women. S. Chen, H. Li, F. Xing. Southern Hosp, The First Military Medical Univ, Guangzhou, China. Objective: To study the expression of HOXA10 gene in the endometrium of fertile woman and unexplained infertile woman during different stages of the menstrual cycle. Design: Case-control study of two ethnic groups. Materials/Methods: Human endometrium samples were obtained by curettage during the different stages of the menstrual cycle. There are 52 fertile women in the control group and 38 unexplained infertile women in the study group. Expression of the HOXA10 gene was detected by in situ hybridization and reverse transcription polymerase chain reaction (RT-PCR) methods. Results: The HOXA10 mRNA gene can be detected in the epithelial cells and stromal cells of the endometrium of fertile women during the different stages of the menstrual cycle. HOXA10 mRNA levels were significantly higher in the mid- and late-secretory phases than that in the proliferative phase and early secretory phase (p ⬍ 0.01). HOXA10 mRNA levels in endometrial stromal cells of the mid-secretory and late-secretory phases were higher than that in the epithelial cells (p ⬍ 0.01). Patients with unexplained infertility have lower HOXA10 mRNA levels in the mid- and late-secretory endometrium as compared to control (p ⬍ 0.01). Conclusions: High expression of the HOXA10 gene during the window of implantation indicates the HOXA10 gene is involved in embryo implantation. Lower HOXA10 expression in patients with unexplained infertility suggests that change of development of endometrium at the molecular level may contribute to the etiology of infertility. The HOXA10 gene may be dominant within the decidua in early pregnancy in humans. Supported by: This research was supported by Scientific Research Fund from National Family Planning Committee and Guangdong Province Natural Science Fund in China.
P-302 Ovarian response to FSH stimulation in PCOS following pioglitazone administration. M. S. Coffler, K. S. Patel, M. H. Dahan, P. J. Malcom, R. J. Chang. Univ of CA, San Diego, La Jolla, CA. Objective: Women with polycystic ovary syndrome (PCOS) have been shown to be insulin resistant. Reduction of insulin resistance by insulin lowering agents has resulted in a resumption of follicular activity and in some cases ovulation. These data suggest that insulin resistance may inhibit follicular activity in PCOS. Our aim was to determine the role of insulin resistance on follicle function. Design: We examined granulosa cell (GC) responsiveness to recombinant human FSH (r-hFSH) in women with PCOS prior to and following treatment with pioglitazone.
FERTILITY & STERILITY威
Materials/Methods: Six women with PCOS were each given r-hFSH (Gonal-F, kindly provided by Serono), 75 IU, iv followed by frequent blood sampling over 24 hours. Subsequently, each was treated with pioglitazone, 30 mg per day for a period of three months. Administration of r-hFSH was then repeated as described. Serum levels of FSH and E2 were determined in each sample by radioimmunoassay. Data were analyzed by non-parametric analysis of variance for repeated measures. Results: Administration of r-hFSH stimulated a brisk increase of serum FSH from 4.1 ⫾ 1.2 mIU/ml (baseline) to a mean maximal value of 15 ⫾ 6.4 mIU/ml at 30 minutes. Corresponding E2 responses were characterized by a substantial increase from 229 ⫾ 28 pmol/ml to peak mean levels of 560 ⫾ 103 pmol/ml within 6 hours of r-hFSH administration. An effect of pioglitazone therapy on E2 responses to repeat r-hFSH stimulation was not apparent. E2 levels rose from 236 ⫾ 28 to 462 ⫾ 80 pmol/ml. There were no differences between groups as assessed by percent change (144 ⫾ 37 and 98 ⫾ 43% prior to and following 3 months of pioglitazone therapy, respectively). Conclusions: 1) Acute GC responsiveness may be achieved in PCOS women following the intravenous administration of r-hFSH; 2) The GC response to r-hFSH in PCOS is not influenced by the administration of pioglitazone. Supported by: This research was supported by NICHD/NIH through cooperative agreement [U54 HD12303-20] as part of the Specialized Cooperative Centers Program in Reproduction Research and in part by NIH grant M01 RR00827.
P-303 Local production and action of prolactin-releasing peptide (PrRP) in human decidua. F. M. Reis, B. Gaffuri, P. Vigano`, P. M. Spritzer, F. Petraglia, A. Di Blasio. UNIPAC and UFMG, Belo Horizonte, Brazil; Inst Auxologico Italy, Milan, Italy; UFRGS, Porto Alegre, Brazil; Univ of Siena, Siena, Italy. Objective: The human decidua is a source and target of prolactin (PRL), whose secretion is modulated by several unspecific molecular mechanisms. Prolactin-Releasing Peptide (PrRP) is a peptide recently isolated from the brain and characterized by a potent and specific stimulation of PRL release from pituitary cells. The present study investigated the expression of PrRP and its receptor in the decidua, and the effect of PrRP on decidual PRL release was evaluated as well. Design: Descriptive study of gene expression and controlled in vitro experiment. Materials/Methods: First trimester deciduas were obtained from women undergoing elective termination of pregnancy. Tissue specimens were stained by immunohistochemistry using a rabbit anti-human PrRP-31 antibody. Decidual cells were isolated from fragments of decidua compacta and maintained in culture for a minimum of 8 days. Cells were then counted and used for RNA extraction and mRNA analysis using reverse transcription polymerase chain reaction (RT-PCR). In a second experiment, cultured decidual cells were stimulated with serial concentrations of synthetic PrRP-31 for 24 hours and PRL levels were measured in culture medium by an enzyme immunoassay. Results: PrRP was localized in both epithelial cells of the decidual glands and in stromal cells. In primary cultures of decidual cells, PrRP and PrRP receptor gene expression were detected using RT-PCR, and the identity of the PCR products was further confirmed by restriction enzyme digestion. The addition of synthetic human PrRP-31 at the doses 10 (⫺12) to 10 (⫺9) M stimulated PRL release from cultured decidual cells. At the dose of 10 (⫺9) M, PrRP induced a 40% increase of PRL concentrations in the culture medium compared to vehicle-treated controls. Conclusions: PrRP and its receptor are expressed in human decidua of early pregnancy and PrRP stimulates PRL release from cultured decidual cells. These findings suggest that PrRP may have a role in the paracrine/ autocrine control of decidual PRL production. Supported by: Istituto Auxologico Italiano, CAPES and CNPq (Brazil).
P-304 Raloxifene effects on human endometrial carcinoma cell line. L. Staneva-Dobrovski, I. Christov, M. Glaeser, D. Niederacher. Ctr of Anatomy and Brain Research, H-Heine Univ of Duesseldorf, Duesseldorf, Ger-
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P-301 Expression of HOXA10 gene in human endometrium and of infertile and fertile women. S. Chen, H. Li, F. Xing. Southern Hosp, The First Military Medical Univ, Guangzhou, China. Objective: To study the expression of HOXA10 gene in the endometrium of fertile woman and unexplained infertile woman during different stages of the menstrual cycle. Design: Case-control study of two ethnic groups. Materials/Methods: Human endometrium samples were obtained by curettage during the different stages of the menstrual cycle. There are 52 fertile women in the control group and 38 unexplained infertile women in the study group. Expression of the HOXA10 gene was detected by in situ hybridization and reverse transcription polymerase chain reaction (RT-PCR) methods. Results: The HOXA10 mRNA gene can be detected in the epithelial cells and stromal cells of the endometrium of fertile women during the different stages of the menstrual cycle. HOXA10 mRNA levels were significantly higher in the mid- and late-secretory phases than that in the proliferative phase and early secretory phase (p ⬍ 0.01). HOXA10 mRNA levels in endometrial stromal cells of the mid-secretory and late-secretory phases were higher than that in the epithelial cells (p ⬍ 0.01). Patients with unexplained infertility have lower HOXA10 mRNA levels in the mid- and late-secretory endometrium as compared to control (p ⬍ 0.01). Conclusions: High expression of the HOXA10 gene during the window of implantation indicates the HOXA10 gene is involved in embryo implantation. Lower HOXA10 expression in patients with unexplained infertility suggests that change of development of endometrium at the molecular level may contribute to the etiology of infertility. The HOXA10 gene may be dominant within the decidua in early pregnancy in humans. Supported by: This research was supported by Scientific Research Fund from National Family Planning Committee and Guangdong Province Natural Science Fund in China.
P-302 Ovarian response to FSH stimulation in PCOS following pioglitazone administration. M. S. Coffler, K. S. Patel, M. H. Dahan, P. J. Malcom, R. J. Chang. Univ of CA, San Diego, La Jolla, CA. Objective: Women with polycystic ovary syndrome (PCOS) have been shown to be insulin resistant. Reduction of insulin resistance by insulin lowering agents has resulted in a resumption of follicular activity and in some cases ovulation. These data suggest that insulin resistance may inhibit follicular activity in PCOS. Our aim was to determine the role of insulin resistance on follicle function. Design: We examined granulosa cell (GC) responsiveness to recombinant human FSH (r-hFSH) in women with PCOS prior to and following treatment with pioglitazone.
FERTILITY & STERILITY威
Materials/Methods: Six women with PCOS were each given r-hFSH (Gonal-F, kindly provided by Serono), 75 IU, iv followed by frequent blood sampling over 24 hours. Subsequently, each was treated with pioglitazone, 30 mg per day for a period of three months. Administration of r-hFSH was then repeated as described. Serum levels of FSH and E2 were determined in each sample by radioimmunoassay. Data were analyzed by non-parametric analysis of variance for repeated measures. Results: Administration of r-hFSH stimulated a brisk increase of serum FSH from 4.1 ⫾ 1.2 mIU/ml (baseline) to a mean maximal value of 15 ⫾ 6.4 mIU/ml at 30 minutes. Corresponding E2 responses were characterized by a substantial increase from 229 ⫾ 28 pmol/ml to peak mean levels of 560 ⫾ 103 pmol/ml within 6 hours of r-hFSH administration. An effect of pioglitazone therapy on E2 responses to repeat r-hFSH stimulation was not apparent. E2 levels rose from 236 ⫾ 28 to 462 ⫾ 80 pmol/ml. There were no differences between groups as assessed by percent change (144 ⫾ 37 and 98 ⫾ 43% prior to and following 3 months of pioglitazone therapy, respectively). Conclusions: 1) Acute GC responsiveness may be achieved in PCOS women following the intravenous administration of r-hFSH; 2) The GC response to r-hFSH in PCOS is not influenced by the administration of pioglitazone. Supported by: This research was supported by NICHD/NIH through cooperative agreement [U54 HD12303-20] as part of the Specialized Cooperative Centers Program in Reproduction Research and in part by NIH grant M01 RR00827.
P-303 Local production and action of prolactin-releasing peptide (PrRP) in human decidua. F. M. Reis, B. Gaffuri, P. Vigano`, P. M. Spritzer, F. Petraglia, A. Di Blasio. UNIPAC and UFMG, Belo Horizonte, Brazil; Inst Auxologico Italy, Milan, Italy; UFRGS, Porto Alegre, Brazil; Univ of Siena, Siena, Italy. Objective: The human decidua is a source and target of prolactin (PRL), whose secretion is modulated by several unspecific molecular mechanisms. Prolactin-Releasing Peptide (PrRP) is a peptide recently isolated from the brain and characterized by a potent and specific stimulation of PRL release from pituitary cells. The present study investigated the expression of PrRP and its receptor in the decidua, and the effect of PrRP on decidual PRL release was evaluated as well. Design: Descriptive study of gene expression and controlled in vitro experiment. Materials/Methods: First trimester deciduas were obtained from women undergoing elective termination of pregnancy. Tissue specimens were stained by immunohistochemistry using a rabbit anti-human PrRP-31 antibody. Decidual cells were isolated from fragments of decidua compacta and maintained in culture for a minimum of 8 days. Cells were then counted and used for RNA extraction and mRNA analysis using reverse transcription polymerase chain reaction (RT-PCR). In a second experiment, cultured decidual cells were stimulated with serial concentrations of synthetic PrRP-31 for 24 hours and PRL levels were measured in culture medium by an enzyme immunoassay. Results: PrRP was localized in both epithelial cells of the decidual glands and in stromal cells. In primary cultures of decidual cells, PrRP and PrRP receptor gene expression were detected using RT-PCR, and the identity of the PCR products was further confirmed by restriction enzyme digestion. The addition of synthetic human PrRP-31 at the doses 10 (⫺12) to 10 (⫺9) M stimulated PRL release from cultured decidual cells. At the dose of 10 (⫺9) M, PrRP induced a 40% increase of PRL concentrations in the culture medium compared to vehicle-treated controls. Conclusions: PrRP and its receptor are expressed in human decidua of early pregnancy and PrRP stimulates PRL release from cultured decidual cells. These findings suggest that PrRP may have a role in the paracrine/ autocrine control of decidual PRL production. Supported by: Istituto Auxologico Italiano, CAPES and CNPq (Brazil).
P-304 Raloxifene effects on human endometrial carcinoma cell line. L. Staneva-Dobrovski, I. Christov, M. Glaeser, D. Niederacher. Ctr of Anatomy and Brain Research, H-Heine Univ of Duesseldorf, Duesseldorf, Ger-
S211