- Email: [email protected]
Coculture using cumulus cells significantly improves day 3 embryo cleavage, but not fragmentation
TABLE 1. Comparison of sensitivity among assays Assay
TABLE 1. Comparison day 3 embryo morphology with coculture vs. regular culture
Dose¼ 24.4 mM
Dose¼ 48.8 mM
62% 48% 64% <0.001* 0.36 0.15
73% 71% 82% 0.044* 0.21 0.82
1-cell 2-cell HSSA 2-cell vs. HSSA 1-cell vs HSSA 1-cell vs 2-cell
* ¼ p< 0.05. TABLE 2. Within assay sensitivity with vs. without protein Assay
Oil
Protein
24.4 mM
48.8 mM
1 cell
No No Yes Yes No No Yes Yes No No Yes Yes
No Yes No Yes No Yes No Yes No Yes No Yes
62% p¼0.008* 38% 47% p<0.001* 13% 48% p¼0.002* 31% 72% NS 66% 64% NS 64%
72% NS 64% 75% p¼0.003* 48% 69% NS 63% 84% NS 80% 79% NS 86%
2 cell
HSSA
#Embryo Average Cell# Average Fragmentation score
Regular Culture
With Autologous
485 5.73 1.62 3.01 0.91
546 6.43 1.32 3.13 0.78
p 0.004 0.13
CONCLUSIONS: Our study suggests a cumulus cell coculture system may improve human embryo cleavage, but not fragmentation score. This result is consistent with our earlier data utilizing BRL cells. The mechanism of coculture needs to be further studied to explain this phenomenon. From a practical standpoint, autologous cumulus cell coculture is easy to perform, less time consuming and negligible risk for infection, and therefore, a feasible alternative to optimize human embryo culture system. Supported by: None.
OVARIAN STIMULATION P-727
* ¼p<0.05. CONCLUSIONS: The only statistically significant difference in sensitivity between the three assays is comparing the 2-cell vs HSSA. The addition of protein appears to have a negative effect on both the 1 and 2 cell assays but not the HSSA. These results indicate the HSSA is better at detecting formaldehyde than the 2 cell MEA and that protein has a significant negative effect on sensitivities on both MEA assays. Supported by: None.
P-726 COCULTURE USING CUMULUS CELLS SIGNIFICANTLY IMPROVES DAY 3 EMBRYO CLEAVAGE, BUT NOT FRAGMENTATION. X. Yang, M. P. Rosen, K. Ho, T. Choi, M. I. Cedars, S. Shen. OBGYN and Reproductive Sciences, University of California at San Francisco, San Francisco, CA. OBJECTIVE: The suggested beneficial effects of coculture include the secretion of embryotrophic factors and the elimination of potentially harmful substances, thereby detoxifying the culture medium. Multiple feeder cells have been recommended, such as human endometrium, granulosa cell, fibroblast, cumulus cell, and even non-human cells or cell lines. However, some of the coculture techniques are difficult to perform, time-consuming, or with risk of infection. In this study, we used autologous cumulus cell as the feeder to perform coculture in poor prognostic patients and evaluated whether this coculture system improves day3 embryo development within the same patient. DESIGN: Observational study. MATERIALS AND METHODS: In our center, coculture was performed in patients who had failed multiple cycles with low cleavage rates or high fragmentation. A total of 175 cycles of autologous cumulus cell coculture were evaluated and only the patients, who had two consecutive cycles, one with regular culture and one with coculture, were included in the study. In both cycles the same type of culture media was used, except in the second cycle, a monolayer of autologous cumulus cells were plated in the dish. Day 3 embryo morphology was compared within each patient. Cleavage was the cell number assessed at 48 hours after coculture. Fragmentation was scored by 1 to 5 with 1 as the best. Paired t-test was performed using STATA 7.0. RESULTS: Fifty three patients met the criteria and total 1,031 embryos were evaluated. Within the same patient in two consecutive cycles, day 3 cleavage rates were significantly improved after coculture with cumulus cells; however, fragmentation score remained unchanged (table).
S352
Abstracts
ARE PREGNANCY RATES AFFECTED BY USING HCG TO TRIGGER OVULATION IN CLOMIPHENE CITRATE-IUI SUPEROVULATION CYCLES? S. Smith, A. Priestley. Obstetrics and Gynecology, Franklin Square Hospital, Baltimore, MD; Franklin Square Hospital, Baltimore, MD. OBJECTIVE: To compare clinical pregnancy rates in patients undergoing superovulation with clomiphene citrate (CC) when human chorionic gonadotropin (hCG) was used to time intrauterine insemination (IUI) with CC cycles in which IUI was timed by detection of urinary LH surge. DESIGN: Retrospective analysis of an academic fertility center database. MATERIALS AND METHODS: One hundred and two ovulatory women who underwent 231 cycles of IUI after CC superovulation for unexplained, male factor or endometriosis associated infertility were identified from the database. All women received 100 mg CC from cycle days 3 to 7 and underwent a single intrauterine insemination 24-28 hours after the detection of a spontaneous urinary LH surge or 24-28 hours after administration of hCG to trigger ovulation. The primary outcome measure was clinical pregnancy per treatment cycle and secondary outcome measures were live birth per treatment cycle, and multiple pregnancies and miscarriage in those who became pregnant. To adjust for clinically significant covariates, a logistic regression analysis was conducted to determine possible statistically significant differences. Pearson’s Chi Square Test and Fisher Exact Test were used to compare rates between groups. Cumulative pregnancy rates for the first five treatment cycles were analyzed using Kaplan-Meier life table analysis, controlling for covariates to determine if pregnancy rates between groups were statistically significant. RESULTS: There was no statistically significant difference in clinical pregnancy rate based upon the manner in which an IUI was timed. The clinical pregnancy rate per cycle was 12.7% for IUI following hCG and 14.7% for IUI following urinary LH surge (p¼NS). The cumulative pregnancy rate after three or five treatment cycles for hCG timed IUIs was 30.8% and 51.1% and for urinary LH timed IUIs it was 36.9% and 52.5% (p¼NS). A multivariate regression analysis controlled for covariates of combined female-male factor, female age, mean number of mature follicles and mean total motile sperm. None of these variables had a statistically significant effect on primary or secondary outcome measures. CONCLUSIONS: The absence of a difference in pregnancy rates suggests that clinicians have practical options for timing IUIs after CC superovulation. As long as one or more dominant follicles are present, hCG may be administered to time an IUI without adversely affecting clinical pregnancy rates. Supported by: None.
Vol. 90, Suppl 1, September 2008
TABLE 1. Comparison day 3 embryo morphology with coculture vs. regular culture
Dose¼ 24.4 mM
Dose¼ 48.8 mM
62% 48% 64% <0.001* 0.36 0.15
73% 71% 82% 0.044* 0.21 0.82
1-cell 2-cell HSSA 2-cell vs. HSSA 1-cell vs HSSA 1-cell vs 2-cell
* ¼ p< 0.05. TABLE 2. Within assay sensitivity with vs. without protein Assay
Oil
Protein
24.4 mM
48.8 mM
1 cell
No No Yes Yes No No Yes Yes No No Yes Yes
No Yes No Yes No Yes No Yes No Yes No Yes
62% p¼0.008* 38% 47% p<0.001* 13% 48% p¼0.002* 31% 72% NS 66% 64% NS 64%
72% NS 64% 75% p¼0.003* 48% 69% NS 63% 84% NS 80% 79% NS 86%
2 cell
HSSA
#Embryo Average Cell# Average Fragmentation score
Regular Culture
With Autologous
485 5.73 1.62 3.01 0.91
546 6.43 1.32 3.13 0.78
p 0.004 0.13
CONCLUSIONS: Our study suggests a cumulus cell coculture system may improve human embryo cleavage, but not fragmentation score. This result is consistent with our earlier data utilizing BRL cells. The mechanism of coculture needs to be further studied to explain this phenomenon. From a practical standpoint, autologous cumulus cell coculture is easy to perform, less time consuming and negligible risk for infection, and therefore, a feasible alternative to optimize human embryo culture system. Supported by: None.
OVARIAN STIMULATION P-727
* ¼p<0.05. CONCLUSIONS: The only statistically significant difference in sensitivity between the three assays is comparing the 2-cell vs HSSA. The addition of protein appears to have a negative effect on both the 1 and 2 cell assays but not the HSSA. These results indicate the HSSA is better at detecting formaldehyde than the 2 cell MEA and that protein has a significant negative effect on sensitivities on both MEA assays. Supported by: None.
P-726 COCULTURE USING CUMULUS CELLS SIGNIFICANTLY IMPROVES DAY 3 EMBRYO CLEAVAGE, BUT NOT FRAGMENTATION. X. Yang, M. P. Rosen, K. Ho, T. Choi, M. I. Cedars, S. Shen. OBGYN and Reproductive Sciences, University of California at San Francisco, San Francisco, CA. OBJECTIVE: The suggested beneficial effects of coculture include the secretion of embryotrophic factors and the elimination of potentially harmful substances, thereby detoxifying the culture medium. Multiple feeder cells have been recommended, such as human endometrium, granulosa cell, fibroblast, cumulus cell, and even non-human cells or cell lines. However, some of the coculture techniques are difficult to perform, time-consuming, or with risk of infection. In this study, we used autologous cumulus cell as the feeder to perform coculture in poor prognostic patients and evaluated whether this coculture system improves day3 embryo development within the same patient. DESIGN: Observational study. MATERIALS AND METHODS: In our center, coculture was performed in patients who had failed multiple cycles with low cleavage rates or high fragmentation. A total of 175 cycles of autologous cumulus cell coculture were evaluated and only the patients, who had two consecutive cycles, one with regular culture and one with coculture, were included in the study. In both cycles the same type of culture media was used, except in the second cycle, a monolayer of autologous cumulus cells were plated in the dish. Day 3 embryo morphology was compared within each patient. Cleavage was the cell number assessed at 48 hours after coculture. Fragmentation was scored by 1 to 5 with 1 as the best. Paired t-test was performed using STATA 7.0. RESULTS: Fifty three patients met the criteria and total 1,031 embryos were evaluated. Within the same patient in two consecutive cycles, day 3 cleavage rates were significantly improved after coculture with cumulus cells; however, fragmentation score remained unchanged (table).
S352
Abstracts
ARE PREGNANCY RATES AFFECTED BY USING HCG TO TRIGGER OVULATION IN CLOMIPHENE CITRATE-IUI SUPEROVULATION CYCLES? S. Smith, A. Priestley. Obstetrics and Gynecology, Franklin Square Hospital, Baltimore, MD; Franklin Square Hospital, Baltimore, MD. OBJECTIVE: To compare clinical pregnancy rates in patients undergoing superovulation with clomiphene citrate (CC) when human chorionic gonadotropin (hCG) was used to time intrauterine insemination (IUI) with CC cycles in which IUI was timed by detection of urinary LH surge. DESIGN: Retrospective analysis of an academic fertility center database. MATERIALS AND METHODS: One hundred and two ovulatory women who underwent 231 cycles of IUI after CC superovulation for unexplained, male factor or endometriosis associated infertility were identified from the database. All women received 100 mg CC from cycle days 3 to 7 and underwent a single intrauterine insemination 24-28 hours after the detection of a spontaneous urinary LH surge or 24-28 hours after administration of hCG to trigger ovulation. The primary outcome measure was clinical pregnancy per treatment cycle and secondary outcome measures were live birth per treatment cycle, and multiple pregnancies and miscarriage in those who became pregnant. To adjust for clinically significant covariates, a logistic regression analysis was conducted to determine possible statistically significant differences. Pearson’s Chi Square Test and Fisher Exact Test were used to compare rates between groups. Cumulative pregnancy rates for the first five treatment cycles were analyzed using Kaplan-Meier life table analysis, controlling for covariates to determine if pregnancy rates between groups were statistically significant. RESULTS: There was no statistically significant difference in clinical pregnancy rate based upon the manner in which an IUI was timed. The clinical pregnancy rate per cycle was 12.7% for IUI following hCG and 14.7% for IUI following urinary LH surge (p¼NS). The cumulative pregnancy rate after three or five treatment cycles for hCG timed IUIs was 30.8% and 51.1% and for urinary LH timed IUIs it was 36.9% and 52.5% (p¼NS). A multivariate regression analysis controlled for covariates of combined female-male factor, female age, mean number of mature follicles and mean total motile sperm. None of these variables had a statistically significant effect on primary or secondary outcome measures. CONCLUSIONS: The absence of a difference in pregnancy rates suggests that clinicians have practical options for timing IUIs after CC superovulation. As long as one or more dominant follicles are present, hCG may be administered to time an IUI without adversely affecting clinical pregnancy rates. Supported by: None.
Vol. 90, Suppl 1, September 2008