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Endometrium receptivity in in-vitro maturation (IVM) treatment
have identified hundreds of up- and down-regulated genes that play role in endometrial receptivity. The knowledge of transcription profiles of human embryos is much more limited, only a few studies have been reported, and the understanding of the complex dialogue between conceptus and endometrium is scarce. Our aim was to analyse and identify possible interactions between embryo and endometrium at the time of implantation. DESIGN: We compared whole genome expression between receptive endometrium and implanting embryo. MATERIALS AND METHODS: Affymetrix arrays were used for endometrial gene expression (mid-secretory endometrium n¼4, proliferative endometrium n¼4) and for embryonic gene expression analyses (30 8-cell embryos pooled n¼2, 30 blastocyst stage embryos pooled n¼2). Protein-protein interactions (PPI) from Human Protein Reference Database were mapped to genes up-regulated in embryos and genes up-regulated in endometrium, followed by filtering using Gene Ontology cell compartment annotations. The resulting PPI network was clustered into overlapping modules with a novel greedy algorithm that accounts for statistical enrichments between proteins and their interaction partners. RESULTS: In total we identified 442 interactions between studied genes, 91 interactions remained valid after the critical evaluation of the gene’s function and it’s location. A number of genes like laminins, collagens and fibulins, that are known to be involved in the implantation, were confirmed in our study. Meanwhile several new interactions were identified. CONCLUSION: To our knowledge, we are the first to demonstrate the interaction pattern between the human implanting embryo and the endometrium. Besides to the known molecules, we report a list of new genes involved in the embryo – endometrium cross-talk. Supported by: Enterprise Estonia, Swedish Research Council, Karolinska Institutet.
O-294 Wednesday, October 27, 2010 04:00 PM SHIFT IN APPEARANCE OF NUCLEOLAR CHANNEL SYSTEMS IN WOMEN FOLLOWING OVARIAN HYPERSTIMULATION WITH AND WITHOUT LUTEAL HORMONAL SUPPLEMENTATION. G. Zapantis, M. Szmyga, E. A. Rybak, D. Kreiner, N. Santoro, U. T. Meier. East Coast Fertility, Plainview, NY; Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, NY; Department of Obstetrics, Gynecology and Women’s Health, Albert Einstein College of Medicine, Bronx, NY; Department of Obstetrics and Gynecology, University of Colorado at Denver, Aurora, CO. OBJECTIVE: To evaluate the temporal appearance of nucleolar channel systems (NCSs) in the human secretory endometrium in natural cycles (NC) and following ovarian hyperstimulation with (OH+S) and without (OH) vaginal progesterone (P4) (Endometrin; Ferring Pharmaceuticals). We hypothesized that NCSs might appear earlier or later following ovarian hyperstimulation (OvH). DESIGN: Endometrial biopsies were obtained from 5 healthy, oocyte donor volunteers on cycle days 16 (CD16), 20 (CD20) and 26 (CD26) following random assignment to an NC, OH or OH+S group. Each subject consented to participate in all three cycle types, with at least one ‘wash-out’ cycle between study cycles. MATERIALS AND METHODS: NCS presence was assessed by indirect immunofluorescence using the nuclear pore complex antibody mAb414 as, none, low, or normal (in >10% of epithelial cell nuclei), and converted to a bimodal grade, i.e. no/low versus normal expression, for analysis (Sign-test). Estradiol and P4 levels were measured on the day of each biopsy. RESULTS: According to our previously determined CD19-24 NCS window of appearance, we confirmed NCS presence on CD20 and no/low abundance on CD16 and CD26 during NCs of individual women. Following OvH, however, NCSs appeared already on CD16, persisted through CD20 and disappeared on CD26. No other significant differences were found between NCs and OvH cycles, OH and OH+S cycles regarding NCS prevalence or hormone levels, or NCS presence and hormonal levels. CONCLUSION: In these healthy oocyte donors, NCS prevalence occurs when expected in the mid-luteal phase of NCs. In contrast, after OvH, NCSs appear earlier, either shifting the six-day NCS window up by at least three days or expanding it. Such an upshift of the NCS window (i.e., absence of NCSs after CD21) might be responsible for a reduced implantation rate after CD21 following embryo transfer in IVF cycles. Regardless, OvH protocols shift or expand the NCS window, a marker for secretory transformation. Supported by: Ferring Pharmaceuticals, March of Dimes Foundation.
FERTILITY & STERILITYÒ
O-295 Wednesday, October 27, 2010 04:15 PM THE ENDOCRINE PROFILE OF IN-VITRO MATURATION TREATMENT CYCLES. C. Ortega, L. Guzman, F. Albuz, P. Devroey, J. Smitz, M. De Vos. Centre for Reproductive Medicine, UZ Brussel, Brussels, Belgium; Laboratory of Follicular Biology, UZ Brussel, Brussels, Belgium. OBJECTIVE: In-vitro maturation treatment encompasses oocyte retrieval in the mid-follicular phase which impacts on luteal function and thus on endometrial competence. Here, we describe the dynamics of serum steroid levels in IVM-cycles and their influence on endometrial quality. DESIGN: A prospective cohort pilot study in 41 normo-ovulatory volunteers and 9 with PCOS. MATERIALS AND METHODS: 20 volunteers received a final oocyte maturation trigger using 5000 IU hCG; 30 received no such trigger. Endometrial priming consisted of 450 IU uFSH, followed by 6 mg daily transdermal estradiol gel (10 mg if endometrium thickness was < 8 mm) and 600 mg daily vaginal progesterone. Serum levels of progesterone (P) and estradiol (E2) were measured on cycle day 8, on the day of egg retrieval and during the luteal phase. Correlations were studied between endocrine serum levels and the endometrial histology. RESULTS: Serum progesterone levels did not differ between subgroups. Overall, hCG-triggering was correlated with decreasing E2 levels during the luteal phase. In PCOS, but not in normo-ovulatory volunteers, luteal phase E2 levels were significantly higher in volunteers with an in-phase endometrium than in those with a secretory delay. Normo-ovulatory volunteers with a competent endometrium who had not received an hCG-trigger had more than two-fold increased luteal phase E2 levels as compared to E2 levels at egg retrieval, whereas in those with a maturation delay E2 levels had dropped significantly after egg retrieval. CONCLUSION: The exact mechanisms underlying improper luteal function after egg retrieval in IVM-treatment are still open for speculation. The results of our pilot study indicate that distinct endocrine patterns may give clues to explain endometrial histological findings in IVM-cycles. Improper luteinisation of follicles may underlie rising estradiol levels after egg retrieval that are associated with a receptive endometrium after substitution therapy. Supported by: Besins Healthcare, Cook Medical, Mithra.
O-296 Wednesday, October 27, 2010 04:30 PM ENDOMETRIUM RECEPTIVITY IN IN-VITRO MATURATION (IVM) TREATMENT. M. De Vos, L. Guzman, F. Albuz, C. Bourgain, J. Smitz, P. Devroey. Centre for Reproductive Medicine, UZ Brussel, Brussels, Belgium; Laboratory of Follicular Biology, UZ Brussel, Brussels, Belgium; Department of Pathology, UZ Brussel, Brussels, Belgium. OBJECTIVE: Transforming endometrium from a mid-follicular to an effective luteal environment in IVM-cycles is challenging. Failure to obtain a receptive endometrium may underlie unsuccessful implantation in IVM. Research focussing on endometrium histology in IVM cycles is scarce. DESIGN: A prospective cohort pilot study in 41 normo-ovulatory volunteers and 9 with PCOS. MATERIALS AND METHODS: Oocytes were matured and fertilized for research purposes. No embryos were transferred, but endometrial development was investigated. Endometrial priming consisted of 450 IU uFSH, followed by 6 mg daily transdermal estradiol gel (10 mg if endometrium thickness was < 8 mm) and 600 mg daily vaginal progesterone. 20 volunteers received a final oocyte maturation trigger using 5000 IU hCG; 30 received no such trigger. Endometrium receptivity was assessed using vaginal ultrasound scanning and histological analysis of endometrium biopsies performed during the luteal phase of an IVM-cycle (on either day 4, 5, 6 or 7 after oocyte retrieval). RESULTS: Endometrium histology did not correlate with endometrium thickness and was not significantly influenced by hCG-triggering. According to Noyes’ criteria, 40,0 % (14/34) resp. 37,5 % (3/8) of representative samples from non-PCOS reps. PCOS volunteers showed a receptive endometrium. Overall, 56,3 % had an endometrium maturation delay exceeding 2 days, of which 33,3 % showed no secretory changes at all. CONCLUSION: A receptive endometrium is a prerequisite for successful implantation. In IVM-cycles, the results of our pilot study show a high incidence of secretory defects in spite of standard hormonal endometrial priming. Further research is needed to establish an optimal hormonal treatment protocol in IVM-cycles and to identify molecular markers that indicate a competent endometrium. Supported by: Besins Health Care, Cook Medical, Mithra.
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O-294 Wednesday, October 27, 2010 04:00 PM SHIFT IN APPEARANCE OF NUCLEOLAR CHANNEL SYSTEMS IN WOMEN FOLLOWING OVARIAN HYPERSTIMULATION WITH AND WITHOUT LUTEAL HORMONAL SUPPLEMENTATION. G. Zapantis, M. Szmyga, E. A. Rybak, D. Kreiner, N. Santoro, U. T. Meier. East Coast Fertility, Plainview, NY; Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, NY; Department of Obstetrics, Gynecology and Women’s Health, Albert Einstein College of Medicine, Bronx, NY; Department of Obstetrics and Gynecology, University of Colorado at Denver, Aurora, CO. OBJECTIVE: To evaluate the temporal appearance of nucleolar channel systems (NCSs) in the human secretory endometrium in natural cycles (NC) and following ovarian hyperstimulation with (OH+S) and without (OH) vaginal progesterone (P4) (Endometrin; Ferring Pharmaceuticals). We hypothesized that NCSs might appear earlier or later following ovarian hyperstimulation (OvH). DESIGN: Endometrial biopsies were obtained from 5 healthy, oocyte donor volunteers on cycle days 16 (CD16), 20 (CD20) and 26 (CD26) following random assignment to an NC, OH or OH+S group. Each subject consented to participate in all three cycle types, with at least one ‘wash-out’ cycle between study cycles. MATERIALS AND METHODS: NCS presence was assessed by indirect immunofluorescence using the nuclear pore complex antibody mAb414 as, none, low, or normal (in >10% of epithelial cell nuclei), and converted to a bimodal grade, i.e. no/low versus normal expression, for analysis (Sign-test). Estradiol and P4 levels were measured on the day of each biopsy. RESULTS: According to our previously determined CD19-24 NCS window of appearance, we confirmed NCS presence on CD20 and no/low abundance on CD16 and CD26 during NCs of individual women. Following OvH, however, NCSs appeared already on CD16, persisted through CD20 and disappeared on CD26. No other significant differences were found between NCs and OvH cycles, OH and OH+S cycles regarding NCS prevalence or hormone levels, or NCS presence and hormonal levels. CONCLUSION: In these healthy oocyte donors, NCS prevalence occurs when expected in the mid-luteal phase of NCs. In contrast, after OvH, NCSs appear earlier, either shifting the six-day NCS window up by at least three days or expanding it. Such an upshift of the NCS window (i.e., absence of NCSs after CD21) might be responsible for a reduced implantation rate after CD21 following embryo transfer in IVF cycles. Regardless, OvH protocols shift or expand the NCS window, a marker for secretory transformation. Supported by: Ferring Pharmaceuticals, March of Dimes Foundation.
FERTILITY & STERILITYÒ
O-295 Wednesday, October 27, 2010 04:15 PM THE ENDOCRINE PROFILE OF IN-VITRO MATURATION TREATMENT CYCLES. C. Ortega, L. Guzman, F. Albuz, P. Devroey, J. Smitz, M. De Vos. Centre for Reproductive Medicine, UZ Brussel, Brussels, Belgium; Laboratory of Follicular Biology, UZ Brussel, Brussels, Belgium. OBJECTIVE: In-vitro maturation treatment encompasses oocyte retrieval in the mid-follicular phase which impacts on luteal function and thus on endometrial competence. Here, we describe the dynamics of serum steroid levels in IVM-cycles and their influence on endometrial quality. DESIGN: A prospective cohort pilot study in 41 normo-ovulatory volunteers and 9 with PCOS. MATERIALS AND METHODS: 20 volunteers received a final oocyte maturation trigger using 5000 IU hCG; 30 received no such trigger. Endometrial priming consisted of 450 IU uFSH, followed by 6 mg daily transdermal estradiol gel (10 mg if endometrium thickness was < 8 mm) and 600 mg daily vaginal progesterone. Serum levels of progesterone (P) and estradiol (E2) were measured on cycle day 8, on the day of egg retrieval and during the luteal phase. Correlations were studied between endocrine serum levels and the endometrial histology. RESULTS: Serum progesterone levels did not differ between subgroups. Overall, hCG-triggering was correlated with decreasing E2 levels during the luteal phase. In PCOS, but not in normo-ovulatory volunteers, luteal phase E2 levels were significantly higher in volunteers with an in-phase endometrium than in those with a secretory delay. Normo-ovulatory volunteers with a competent endometrium who had not received an hCG-trigger had more than two-fold increased luteal phase E2 levels as compared to E2 levels at egg retrieval, whereas in those with a maturation delay E2 levels had dropped significantly after egg retrieval. CONCLUSION: The exact mechanisms underlying improper luteal function after egg retrieval in IVM-treatment are still open for speculation. The results of our pilot study indicate that distinct endocrine patterns may give clues to explain endometrial histological findings in IVM-cycles. Improper luteinisation of follicles may underlie rising estradiol levels after egg retrieval that are associated with a receptive endometrium after substitution therapy. Supported by: Besins Healthcare, Cook Medical, Mithra.
O-296 Wednesday, October 27, 2010 04:30 PM ENDOMETRIUM RECEPTIVITY IN IN-VITRO MATURATION (IVM) TREATMENT. M. De Vos, L. Guzman, F. Albuz, C. Bourgain, J. Smitz, P. Devroey. Centre for Reproductive Medicine, UZ Brussel, Brussels, Belgium; Laboratory of Follicular Biology, UZ Brussel, Brussels, Belgium; Department of Pathology, UZ Brussel, Brussels, Belgium. OBJECTIVE: Transforming endometrium from a mid-follicular to an effective luteal environment in IVM-cycles is challenging. Failure to obtain a receptive endometrium may underlie unsuccessful implantation in IVM. Research focussing on endometrium histology in IVM cycles is scarce. DESIGN: A prospective cohort pilot study in 41 normo-ovulatory volunteers and 9 with PCOS. MATERIALS AND METHODS: Oocytes were matured and fertilized for research purposes. No embryos were transferred, but endometrial development was investigated. Endometrial priming consisted of 450 IU uFSH, followed by 6 mg daily transdermal estradiol gel (10 mg if endometrium thickness was < 8 mm) and 600 mg daily vaginal progesterone. 20 volunteers received a final oocyte maturation trigger using 5000 IU hCG; 30 received no such trigger. Endometrium receptivity was assessed using vaginal ultrasound scanning and histological analysis of endometrium biopsies performed during the luteal phase of an IVM-cycle (on either day 4, 5, 6 or 7 after oocyte retrieval). RESULTS: Endometrium histology did not correlate with endometrium thickness and was not significantly influenced by hCG-triggering. According to Noyes’ criteria, 40,0 % (14/34) resp. 37,5 % (3/8) of representative samples from non-PCOS reps. PCOS volunteers showed a receptive endometrium. Overall, 56,3 % had an endometrium maturation delay exceeding 2 days, of which 33,3 % showed no secretory changes at all. CONCLUSION: A receptive endometrium is a prerequisite for successful implantation. In IVM-cycles, the results of our pilot study show a high incidence of secretory defects in spite of standard hormonal endometrial priming. Further research is needed to establish an optimal hormonal treatment protocol in IVM-cycles and to identify molecular markers that indicate a competent endometrium. Supported by: Besins Health Care, Cook Medical, Mithra.
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