- Email: [email protected]
Comparison of vitrification efficiency using thin plastic strip (TPS) and electron microscope (EM) grid on mouse blastocysts
was to evaluate the efficacy of two cryopreservation techniques for poor quality embryos. DESIGN: Retrospective. MATERIALS AND METHODS: In 2009 111 frozen-thawed embryo cycles were performed, twenty of which included only low quality embryos at cryopreservation on day 3 (classified using conventional criteria). These cycles were divided in two groups: Group 1 (n: 9) included cryopreserved embryos (slow freezing, Testart protocol) and Group 2 (n: 11) vitrified embryos (Vitrification-warming KIT, Kitazato Biopharma, Japan). Main outcome measures were embryo survival rate, blastomere loss, implantation and pregnancy rate. Statistical analysis was performed with chi square and student t test. P<0.05 was considered significant. RESULTS: Groups were comparable regarding age, endometrial type and thickness and number of embryos transferred. Eight embryo transfers were performed in Group 1 as all 3 embryos degenerated during one thawed cycle. Embryo survival rate was 84,4% (27/32) in Group 1 vs. 97.2% (35/36) in Group 2 (p¼0.048). Blastomere loss of viable embryos was 31,49% (57/ 181) vs.7,32% (15/205)(p¼0.0001). Implantation rate was 0% (0/21) for Group 1 vs. 23,33% (7/30) for Group 2 (p¼0.032) and clinical pregnancy rate was 0% (0/8) vs. 54,55% (6/11)(p¼0,018). Abortion rate was 33% (2/6) in Group 2. CONCLUSION: Vitrification may be less detrimental than slow freezing for the developmental ability of cryopreserved-thawed poor quality embryos. Further confirmation of these results could aid in the decision making process when considering the destination of surplus poor quality embryos.
P-57 Tuesday, October 26, 2010 VARIABLES INFLUENCING PREGNANCY RATE AFTER OOCYTE CRYOPRESERVATION. S. Ferrero, M. R. Privamera, S. Levi, A. J. Nicoletti, L. H. Abbamonte, P. Anserini. Department of Obstetrics and Gynecology, San Martino Hospital and University of Genoa, Genova, GE, Italy. OBJECTIVE: To evaluate the variables affecting clinical pregnancy rate after oocyte cryopreservation. DESIGN: Prospective study. MATERIALS AND METHODS: This study included 62 consecutive patients who underwent 74 cycles with cryopreservation of metaphase II (MII) oocytes. Oocytes from stimulated ovaries were cryopreserved using a slow freezing-rapid thawing method. Fertilization of thawed oocytes was achieved by ICSI. Embryo transfer (ET) was performed 40-72 hours after insemination. Clinical pregnancy was defined as evidence of a gestational sac on ultrasonography. RESULTS: The mean ( SD) patient age at retrieval was 34.3 ( 3.6) years. In 10 thawing cycles, none of the oocytes survived (13.5%). One to three oocytes were inseminated after a mean ( SD) post-thawing culture time of 210 minutes ( 50 minutes). The mean ( SD) number of inseminated oocytes was 2.7 ( 0.7). In 4 cycles, all inseminated oocytes failed to fertilise or to cleave; the mean ( SD) number of embryo obtained was 2.0 ( 1.0). 57 ET were performed (77.0% of all thaw cycles) and 14 clinical pregnancies (24.6% per ET) were observed. No significant difference was observed in the number of cryopreserved oocytes between women who had and did not have clinical pregnancies (p ¼ 0.619). Patients who had clinical pregnancies had a significantly higher number of oocytes that survived after thawing than patients who did not conceive (mean SD, 5.2 0.9 versus 3.1 2.0; p < 0.001). Patients with and without clinical pregnancies had no significant difference in age, cause of infertility, type of gonadotropin used for stimulation, dose of gonadotropin used for stimulation and endometrial thickness at ET. No significant difference was observed in the clinical pregnancy rate per ET between women < 35 years and those R 35 years (p ¼ 0.250). CONCLUSION: Patients with higher number of oocytes surviving postthawing have a higher clinical pregnancy rate. Age R 35 years does not decrease the clinical pregnancy rate after oocyte cryopreservation.
P-58 Tuesday, October 26, 2010 WHAT IS THE OPTIMAL STAGE FOR EMBRYO VITRIFICATIONA COMPARISON OF EMBRYO SURVIVAL AND CLINICAL OUTCOMES WITH DAY 3 CLEAVAGE VERSUS BLASTOCYST STAGE VITRIFICATION. N. Desai, T. Falcone, J. Goldberg, C. Austin, J. Goldfarb. OB-GYN/Women’s Health Institute, Cleveland Clinic Foundation, Beachwood, OH.
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Abstracts
OBJECTIVE: The optimal stage for embryo vitrification remains to be determined. Each stage has its own inherent advantages. This study compares our laboratory’s three year experience with embryo vitrification at either the 6-8 cell stage or at blastocyst. DESIGN: Retrospective examination of embryo vitrification. Patients R38 were excluded. MATERIALS AND METHODS: Embryos were vitrified using a two-step protocol: 7.5% DMSO/ethylene glycol (EG) (2-3 min) and then 15% DMSO/ EG for 45 seconds. Embryos were loaded on cryoloops and immersed in liquid nitrogen. Cryoprotectant was removed during warming by sequential washes (0.25 M, 0.125 M sucrose). Warmed embryos were cultured for 48 hours before transfer. Outcome parameters were embryo survival, morphology, positive hCG, intrauterine gestational sac and fetal cardiac activity on ultrasound, and implantation rate (fetal hearts/total embryos transferred. Data analyzed using the chi square and T-test. P values <0.05 considered statistically significant. RESULTS: Outomes shown below.
6-8 Cell Stage (Day3)
Blastocyst (Day 5-6)
33 2.8 223 2.38 0.73 87% 2.18 0.58
33.5 2.8 201 1.97 0.85 a 90% 1.83 0.60b
120/217 (55%) 102/217 (47%)
90/194 (46%) 80/194 (41%)
96/217 (44%) 122/473 (26%)
67/194 (35%)c 87/355 (25%)
Patient age Thaws Average embryos thawed Post-thaw survival Average embryos transferred Positive hCG/transfer(%) GestationalSacs/ transfer(%) Fetal hearts/transfer (%) Implantation rate (%)
Significantly different from cleavage thaw a,b P < 0.001; c P¼ 0.045. CONCLUSION: The implantation rates with vitrification at the 6-8 cell stage versus blastocyst were not different. Early stage vitrification is valuable for troubleshooting and gauging embryo potential on warming, whereas late stage vitrification ensures that embryonic genome activation has occurred. Tailoring vitrification protocols for specific cell stages and blastocyst morphologies may be necessary to optimize outcomes.
P-59 Tuesday, October 26, 2010 COMPARISON OF VITRIFICATION EFFICIENCY USING THIN PLASTIC STRIP (TPS) AND ELECTRON MICROSCOPE (EM) GRID ON MOUSE BLASTOCYSTS. J. Lee, Y. Hur, S. Yoon, C. Hur, W. Lee, J. Lim. Research, Maria Hospital, Seoul, Dongdaemun-gu, Korea; Maria Research Center, Maria Hospital, Seoul, Dongdaemun-gu, Korea; Maria Clinical Center, Maria Hospital, Seoul, Dongdaemun-gu, Korea. OBJECTIVE: As a vitrification tool, the EM grid has a strong point that has the highest cooling rate due to the extremely low volume of vitrification medium and directly plunging into liquid nitrogen(LN2). However, it also has a problem which is an expert embryo handling and an attaching embryo in grid after thawed. Accordingly, we attempted to apply a loading of plasticware which is everywhere made of polypropylene as a new vitrification tool, named thin plastic strip(TPS) for vitrification of mouse blastocysts. DESIGN: The aim of this study is to investigate the usefulness of TPS in vitrification of late stage mouse blastocysts comparing with an EM grid. MATERIALS AND METHODS: Blastocysts obtained from six weeks-old BDF1 mouse. The expanded blastocysts(EdB,n¼166) were vitrified respectively using TPS(n¼75) and EM grid(n¼91) after artificial shrinkage. And hatching blastocysts(HgB,n¼243) were vitrified respectively using TPS(n¼122) and EM grid(n¼121) following artificial shrinkage. Minimum volume of vitrification medium mixed with blastocysts loaded on TPS. Except loading method, other process of vitrification and thawing were prepared according to the previously described method (Son et al.,2003). The blastocysts were thawed a month later and cultured for a day. The viability was then assessed through survival and recovery. RESULTS: The recovery rate of the TPS(EdB,100%,75/75,HgB,100%,122/ 122) was significantly higher than EM grid(EdB,90.5%,83/91,HgB,75.2%,99/
Vol. 94., No. 4, Supplement, September 2010
121)(P<0.05). There was no significant difference in the survival rate of the TPS(EdB, 93.3%,67/75,HgB,88.3%,100/122) and EM grid(EdB,85.1%, 74/91,HgB, 68.6%,87/121)(P<0.05). CONCLUSION: This study showed that the vitrification using TPS has a potential usefulness on late stage mouse blastocysts. Comparison of survival rate of TPS is similar to EM grid. But, recovery rate of TPS is higher than EM grid. Therefore, the TPS would be useful as an alternative tool that overcomes the problems of EM grid. And also, it has the potential for clinical applications.
after ICSI was 75,7% and 67,0%, respectively. Embryo development was similar in both groups, as well as top quality embryo (29,2% and 31,9% respectively). Pregnancy and implantation rates were 47,1% and 24,02% in group I; 46,7% and 28,8% in group II. There were no statistically significant differences in none of the outcomes analyzed. CONCLUSION: Vitrification preserves oocyte potential to fertilize and further development. Our results show that oocyte vitrification procedure is equal to fresh egg donation with regard to embryo development and gestational results. This technique provides an excellent tool to establish an oocyte bank which in turn optimizes egg donation programs.
P-60 Tuesday, October 26, 2010
P-62 Tuesday, October 26, 2010
DEVELOPMENT OF A MURINE MODEL TO STUDY THE INFLUENCE OF MATERNAL BODY ON EMBRYO CYTOPLASMIC COMPOSITION. J. Weathers, N. Zimmerer, S. D. Prien. Animal and Food Sciences, Texas Tech University, Lubbock, TX; Ob/Gyn, Texas Tech University Health Sciences Center, Lubbock, TX.
THE OOCYTE CRYO-BANKING IS A PRACTICAL WAY TO BENEFIT MORE PATIENTS WHO NEED DONOR OOCYTES. C.-C. Chang, T. A. Elliott, G. Wright, D. B. Shapiro, H. I. Kort, Z. P. Nagy. Reproductive Biology Associates, Atlanta, GA.
OBJECTIVE: Cryopreservation is dependent on an equilibrium between cryoprotectants and the innercellular and extracellular water of embryos. It is recognized that the embryo component in this process, the cytoplasm, is primarily a product of maternal physiology.Previous research suggests than maternal body condition plays an important role in determining the biochemical makeup of the embryo. The present study describes a murine model that can be used to study the influence of maternal BMI on embryo composition. DESIGN: Use of an obese mouse strain to study BMI influence on embryo composition. MATERIALS AND METHODS: Embryos were obtained from three strains of mice; 1) a CB6F1 wild type (control), 2) a heterozygous C57BL/ 6NCrL-Leprdb-lb/Crl lean +/?, lean mouse and a homozygous C57BL/ 6NCrl-Leprdb-lb/Crl Obese +/+ mutation which produces obese animals, using standard superovulation techniques. Prior to embryo recovery, the animals were weighed and assessed for body condition. The relative weight of each embryo was then obtained from the zygote to the blastocyst stage using a modified specific gravity technique developed in this lab. Resulting data were analyzed using ANOVA and repeated measures. RESULTS: There was a difference in both weight (P <.001) and body condition score (P < .001) of the obese mice and the other two animal groups. Embryo weights changed over the 7 day window as embryos incorporated more water into their structures. However, embryos from the obese strain were significantly lighter than the other two groups at all time points (P <.001) and stages of embryo development (P <.001), indicating increased lipid content at all time points. CONCLUSION: Results indicate significant differences in relative embryo weight which had previously been demonstrated to be due to embryo lipid content. The data suggest the obese murine model can be used to study the influences of maternal body condition and BMI have on embryo biochemistry and how such changes affect ART success, especially in the area of cryopreservation.
P-61 Tuesday, October 26, 2010 IMPACT OF EMBRYO QUALITY IN FRESH VERSUS VITRIFIED OOCYTES IN AN EGG DONATION PROGRAM. S. M. Giordana, M. F. Insua, B. Lotti, N. Fernandez Peri, A. Pellicer, F. D. Neuspiller. IVI Buenos Aires, Ciudad Auto´noma de Buenos Aires, Buenos Aires, Argentina; IVI Valencia, Valencia, Spain. OBJECTIVE: The objective of this study was to compare the embryo quality and gestational results in an egg donation program using fresh and vitrified/thawed oocytes, previously stored in an oocyte bank. DESIGN: Retrospective, comparative, observational and transversal study. MATERIALS AND METHODS: A total of 43 patients who underwent 49 embryo transfers from January 1st to December 31st 2009 were analyzed. Group I involved 27 patients whose treatment was carried out using fresh oocytes, while group II involved 16 patients using vitrified/thawed oocytes by Cryotop method. Endometrial preparation consisted of increasing doses of estradiol valerate and micronized progesterone. Recipients with ovarian function were down-regulated with GnRH agonist in the lutheal phase of the previous cycle. Donor stimulation was performed with recombinant FSH (starting dose from 150 to 300 UI/day, according to antral follicle account) and GnRH antagonist. RESULTS: Mean age of the patients was 41 4,0 (group I) and 42,06 2,9 (group II). Survival rate of vitrified oocytes was 86,6%. Fertilization rate
FERTILITY & STERILITYÒ
OBJECTIVE: Our objective was to assess the efficacy and efficiency of oocyte donation cycles using oocyte cryo-banking in comparison to fresh oocyte donation cycles. DESIGN: Prospective, parallel-group study. MATERIALS AND METHODS: From Jan 2007 to Mar 2010, a total of 395 recipient cycles of donated oocytes were included in this study. Cryopreservation on oocytes was performed by minimum volume vitrification with 15% ethylene glycol, 15% DMSO, and 0.5M sucrose. Oocytes were fertilized by 40 hr after hCG administration or 2-3 hr after oocyte warming. Clinical pregnancy was confirmed by detecting fetal cardiac activity (FCA) with ultrasound. Results were analyzed by the One-way ANOVA or the Fisher’s exact tests, as appropriate (P<0.05). RESULTS: As shown below.
# of Recipients Total # of donor cycles Recipient age Donor age # of oocyte used/warmed (mean SD) # of oocyte remained # of oocyte survived (%) # of oocyte fertilized (mean SD) # of ET (mean SD) # of embryo vitrified # of (+)hCG (%) # of (+)FCA # of Implantation (%)
Fresh
Vitrified
P value
148 148 40.6 4.7 26.7 3.2 3107 (20.9 9.0)
247 139 41.0 4.6 26.3 2.7 1592 (6.4 2.0)
NA NA NS NS <0.0001
0 1823 (12.3 5.3)
1187 1386 (87.0) 1206 (4.8 1.5)
NA NA <0.0001
291 (1.9 0.3) 1435 (9.7 7.4) 113 (76.3) 89 (60.1) 146 (50.1)
494 (2.0 0.4) 290 (1.1 1.3) 175 (70.8) 147 (59.5) 219 (44.3)
NS <0.0001 NS NS NS
CONCLUSION: This study demonstrated that high pregnancy and implantation rates were obtained in both fresh and vitrified oocyte donation cycles. With the same clinical outcome as fresh donation cycles, it is possible to use oocyte cryo-banking to minimize the number of oocytes fertilized, thus reduce the number of excess embryos for cryopreservation. As the tremendous demand of oocyte donation remains, the less number of oocyte used per recipient means more recipients could acquire donated oocytes by the use of the oocyte cryo-banking.
P-63 Tuesday, October 26, 2010 THE TIME OF CRYOPRESERVATION DOES NOT INCREASE THE DAMAGE IN FROZEN OVARIAN TISSUE FOR FERTILITY PRESERVATION. J. R. Campos, B. R. Carvalho, J. C. Rosa-e-Silva, A. A. Vireque, R. A. Ferrani, A. C. J. S. Rosa-e-Silva. Obstetrics and Gynecology, Faculty of Medicine of Ribeir~ao Preto - University of S~ao Paulo, Ribeir~ao Preto, S~ao Paulo, Brazil. OBJECTIVE: To evaluate the time effect on ovarian tissue cryopreservation by analyzing the fresh and frozen-thawed samples according to its follicular density, follicular viability and steroidogenic capacity, comparing thawing after 30 and 180 days after the freezing procedure. DESIGN: A prospective study.
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P-57 Tuesday, October 26, 2010 VARIABLES INFLUENCING PREGNANCY RATE AFTER OOCYTE CRYOPRESERVATION. S. Ferrero, M. R. Privamera, S. Levi, A. J. Nicoletti, L. H. Abbamonte, P. Anserini. Department of Obstetrics and Gynecology, San Martino Hospital and University of Genoa, Genova, GE, Italy. OBJECTIVE: To evaluate the variables affecting clinical pregnancy rate after oocyte cryopreservation. DESIGN: Prospective study. MATERIALS AND METHODS: This study included 62 consecutive patients who underwent 74 cycles with cryopreservation of metaphase II (MII) oocytes. Oocytes from stimulated ovaries were cryopreserved using a slow freezing-rapid thawing method. Fertilization of thawed oocytes was achieved by ICSI. Embryo transfer (ET) was performed 40-72 hours after insemination. Clinical pregnancy was defined as evidence of a gestational sac on ultrasonography. RESULTS: The mean ( SD) patient age at retrieval was 34.3 ( 3.6) years. In 10 thawing cycles, none of the oocytes survived (13.5%). One to three oocytes were inseminated after a mean ( SD) post-thawing culture time of 210 minutes ( 50 minutes). The mean ( SD) number of inseminated oocytes was 2.7 ( 0.7). In 4 cycles, all inseminated oocytes failed to fertilise or to cleave; the mean ( SD) number of embryo obtained was 2.0 ( 1.0). 57 ET were performed (77.0% of all thaw cycles) and 14 clinical pregnancies (24.6% per ET) were observed. No significant difference was observed in the number of cryopreserved oocytes between women who had and did not have clinical pregnancies (p ¼ 0.619). Patients who had clinical pregnancies had a significantly higher number of oocytes that survived after thawing than patients who did not conceive (mean SD, 5.2 0.9 versus 3.1 2.0; p < 0.001). Patients with and without clinical pregnancies had no significant difference in age, cause of infertility, type of gonadotropin used for stimulation, dose of gonadotropin used for stimulation and endometrial thickness at ET. No significant difference was observed in the clinical pregnancy rate per ET between women < 35 years and those R 35 years (p ¼ 0.250). CONCLUSION: Patients with higher number of oocytes surviving postthawing have a higher clinical pregnancy rate. Age R 35 years does not decrease the clinical pregnancy rate after oocyte cryopreservation.
P-58 Tuesday, October 26, 2010 WHAT IS THE OPTIMAL STAGE FOR EMBRYO VITRIFICATIONA COMPARISON OF EMBRYO SURVIVAL AND CLINICAL OUTCOMES WITH DAY 3 CLEAVAGE VERSUS BLASTOCYST STAGE VITRIFICATION. N. Desai, T. Falcone, J. Goldberg, C. Austin, J. Goldfarb. OB-GYN/Women’s Health Institute, Cleveland Clinic Foundation, Beachwood, OH.
S110
Abstracts
OBJECTIVE: The optimal stage for embryo vitrification remains to be determined. Each stage has its own inherent advantages. This study compares our laboratory’s three year experience with embryo vitrification at either the 6-8 cell stage or at blastocyst. DESIGN: Retrospective examination of embryo vitrification. Patients R38 were excluded. MATERIALS AND METHODS: Embryos were vitrified using a two-step protocol: 7.5% DMSO/ethylene glycol (EG) (2-3 min) and then 15% DMSO/ EG for 45 seconds. Embryos were loaded on cryoloops and immersed in liquid nitrogen. Cryoprotectant was removed during warming by sequential washes (0.25 M, 0.125 M sucrose). Warmed embryos were cultured for 48 hours before transfer. Outcome parameters were embryo survival, morphology, positive hCG, intrauterine gestational sac and fetal cardiac activity on ultrasound, and implantation rate (fetal hearts/total embryos transferred. Data analyzed using the chi square and T-test. P values <0.05 considered statistically significant. RESULTS: Outomes shown below.
6-8 Cell Stage (Day3)
Blastocyst (Day 5-6)
33 2.8 223 2.38 0.73 87% 2.18 0.58
33.5 2.8 201 1.97 0.85 a 90% 1.83 0.60b
120/217 (55%) 102/217 (47%)
90/194 (46%) 80/194 (41%)
96/217 (44%) 122/473 (26%)
67/194 (35%)c 87/355 (25%)
Patient age Thaws Average embryos thawed Post-thaw survival Average embryos transferred Positive hCG/transfer(%) GestationalSacs/ transfer(%) Fetal hearts/transfer (%) Implantation rate (%)
Significantly different from cleavage thaw a,b P < 0.001; c P¼ 0.045. CONCLUSION: The implantation rates with vitrification at the 6-8 cell stage versus blastocyst were not different. Early stage vitrification is valuable for troubleshooting and gauging embryo potential on warming, whereas late stage vitrification ensures that embryonic genome activation has occurred. Tailoring vitrification protocols for specific cell stages and blastocyst morphologies may be necessary to optimize outcomes.
P-59 Tuesday, October 26, 2010 COMPARISON OF VITRIFICATION EFFICIENCY USING THIN PLASTIC STRIP (TPS) AND ELECTRON MICROSCOPE (EM) GRID ON MOUSE BLASTOCYSTS. J. Lee, Y. Hur, S. Yoon, C. Hur, W. Lee, J. Lim. Research, Maria Hospital, Seoul, Dongdaemun-gu, Korea; Maria Research Center, Maria Hospital, Seoul, Dongdaemun-gu, Korea; Maria Clinical Center, Maria Hospital, Seoul, Dongdaemun-gu, Korea. OBJECTIVE: As a vitrification tool, the EM grid has a strong point that has the highest cooling rate due to the extremely low volume of vitrification medium and directly plunging into liquid nitrogen(LN2). However, it also has a problem which is an expert embryo handling and an attaching embryo in grid after thawed. Accordingly, we attempted to apply a loading of plasticware which is everywhere made of polypropylene as a new vitrification tool, named thin plastic strip(TPS) for vitrification of mouse blastocysts. DESIGN: The aim of this study is to investigate the usefulness of TPS in vitrification of late stage mouse blastocysts comparing with an EM grid. MATERIALS AND METHODS: Blastocysts obtained from six weeks-old BDF1 mouse. The expanded blastocysts(EdB,n¼166) were vitrified respectively using TPS(n¼75) and EM grid(n¼91) after artificial shrinkage. And hatching blastocysts(HgB,n¼243) were vitrified respectively using TPS(n¼122) and EM grid(n¼121) following artificial shrinkage. Minimum volume of vitrification medium mixed with blastocysts loaded on TPS. Except loading method, other process of vitrification and thawing were prepared according to the previously described method (Son et al.,2003). The blastocysts were thawed a month later and cultured for a day. The viability was then assessed through survival and recovery. RESULTS: The recovery rate of the TPS(EdB,100%,75/75,HgB,100%,122/ 122) was significantly higher than EM grid(EdB,90.5%,83/91,HgB,75.2%,99/
Vol. 94., No. 4, Supplement, September 2010
121)(P<0.05). There was no significant difference in the survival rate of the TPS(EdB, 93.3%,67/75,HgB,88.3%,100/122) and EM grid(EdB,85.1%, 74/91,HgB, 68.6%,87/121)(P<0.05). CONCLUSION: This study showed that the vitrification using TPS has a potential usefulness on late stage mouse blastocysts. Comparison of survival rate of TPS is similar to EM grid. But, recovery rate of TPS is higher than EM grid. Therefore, the TPS would be useful as an alternative tool that overcomes the problems of EM grid. And also, it has the potential for clinical applications.
after ICSI was 75,7% and 67,0%, respectively. Embryo development was similar in both groups, as well as top quality embryo (29,2% and 31,9% respectively). Pregnancy and implantation rates were 47,1% and 24,02% in group I; 46,7% and 28,8% in group II. There were no statistically significant differences in none of the outcomes analyzed. CONCLUSION: Vitrification preserves oocyte potential to fertilize and further development. Our results show that oocyte vitrification procedure is equal to fresh egg donation with regard to embryo development and gestational results. This technique provides an excellent tool to establish an oocyte bank which in turn optimizes egg donation programs.
P-60 Tuesday, October 26, 2010
P-62 Tuesday, October 26, 2010
DEVELOPMENT OF A MURINE MODEL TO STUDY THE INFLUENCE OF MATERNAL BODY ON EMBRYO CYTOPLASMIC COMPOSITION. J. Weathers, N. Zimmerer, S. D. Prien. Animal and Food Sciences, Texas Tech University, Lubbock, TX; Ob/Gyn, Texas Tech University Health Sciences Center, Lubbock, TX.
THE OOCYTE CRYO-BANKING IS A PRACTICAL WAY TO BENEFIT MORE PATIENTS WHO NEED DONOR OOCYTES. C.-C. Chang, T. A. Elliott, G. Wright, D. B. Shapiro, H. I. Kort, Z. P. Nagy. Reproductive Biology Associates, Atlanta, GA.
OBJECTIVE: Cryopreservation is dependent on an equilibrium between cryoprotectants and the innercellular and extracellular water of embryos. It is recognized that the embryo component in this process, the cytoplasm, is primarily a product of maternal physiology.Previous research suggests than maternal body condition plays an important role in determining the biochemical makeup of the embryo. The present study describes a murine model that can be used to study the influence of maternal BMI on embryo composition. DESIGN: Use of an obese mouse strain to study BMI influence on embryo composition. MATERIALS AND METHODS: Embryos were obtained from three strains of mice; 1) a CB6F1 wild type (control), 2) a heterozygous C57BL/ 6NCrL-Leprdb-lb/Crl lean +/?, lean mouse and a homozygous C57BL/ 6NCrl-Leprdb-lb/Crl Obese +/+ mutation which produces obese animals, using standard superovulation techniques. Prior to embryo recovery, the animals were weighed and assessed for body condition. The relative weight of each embryo was then obtained from the zygote to the blastocyst stage using a modified specific gravity technique developed in this lab. Resulting data were analyzed using ANOVA and repeated measures. RESULTS: There was a difference in both weight (P <.001) and body condition score (P < .001) of the obese mice and the other two animal groups. Embryo weights changed over the 7 day window as embryos incorporated more water into their structures. However, embryos from the obese strain were significantly lighter than the other two groups at all time points (P <.001) and stages of embryo development (P <.001), indicating increased lipid content at all time points. CONCLUSION: Results indicate significant differences in relative embryo weight which had previously been demonstrated to be due to embryo lipid content. The data suggest the obese murine model can be used to study the influences of maternal body condition and BMI have on embryo biochemistry and how such changes affect ART success, especially in the area of cryopreservation.
P-61 Tuesday, October 26, 2010 IMPACT OF EMBRYO QUALITY IN FRESH VERSUS VITRIFIED OOCYTES IN AN EGG DONATION PROGRAM. S. M. Giordana, M. F. Insua, B. Lotti, N. Fernandez Peri, A. Pellicer, F. D. Neuspiller. IVI Buenos Aires, Ciudad Auto´noma de Buenos Aires, Buenos Aires, Argentina; IVI Valencia, Valencia, Spain. OBJECTIVE: The objective of this study was to compare the embryo quality and gestational results in an egg donation program using fresh and vitrified/thawed oocytes, previously stored in an oocyte bank. DESIGN: Retrospective, comparative, observational and transversal study. MATERIALS AND METHODS: A total of 43 patients who underwent 49 embryo transfers from January 1st to December 31st 2009 were analyzed. Group I involved 27 patients whose treatment was carried out using fresh oocytes, while group II involved 16 patients using vitrified/thawed oocytes by Cryotop method. Endometrial preparation consisted of increasing doses of estradiol valerate and micronized progesterone. Recipients with ovarian function were down-regulated with GnRH agonist in the lutheal phase of the previous cycle. Donor stimulation was performed with recombinant FSH (starting dose from 150 to 300 UI/day, according to antral follicle account) and GnRH antagonist. RESULTS: Mean age of the patients was 41 4,0 (group I) and 42,06 2,9 (group II). Survival rate of vitrified oocytes was 86,6%. Fertilization rate
FERTILITY & STERILITYÒ
OBJECTIVE: Our objective was to assess the efficacy and efficiency of oocyte donation cycles using oocyte cryo-banking in comparison to fresh oocyte donation cycles. DESIGN: Prospective, parallel-group study. MATERIALS AND METHODS: From Jan 2007 to Mar 2010, a total of 395 recipient cycles of donated oocytes were included in this study. Cryopreservation on oocytes was performed by minimum volume vitrification with 15% ethylene glycol, 15% DMSO, and 0.5M sucrose. Oocytes were fertilized by 40 hr after hCG administration or 2-3 hr after oocyte warming. Clinical pregnancy was confirmed by detecting fetal cardiac activity (FCA) with ultrasound. Results were analyzed by the One-way ANOVA or the Fisher’s exact tests, as appropriate (P<0.05). RESULTS: As shown below.
# of Recipients Total # of donor cycles Recipient age Donor age # of oocyte used/warmed (mean SD) # of oocyte remained # of oocyte survived (%) # of oocyte fertilized (mean SD) # of ET (mean SD) # of embryo vitrified # of (+)hCG (%) # of (+)FCA # of Implantation (%)
Fresh
Vitrified
P value
148 148 40.6 4.7 26.7 3.2 3107 (20.9 9.0)
247 139 41.0 4.6 26.3 2.7 1592 (6.4 2.0)
NA NA NS NS <0.0001
0 1823 (12.3 5.3)
1187 1386 (87.0) 1206 (4.8 1.5)
NA NA <0.0001
291 (1.9 0.3) 1435 (9.7 7.4) 113 (76.3) 89 (60.1) 146 (50.1)
494 (2.0 0.4) 290 (1.1 1.3) 175 (70.8) 147 (59.5) 219 (44.3)
NS <0.0001 NS NS NS
CONCLUSION: This study demonstrated that high pregnancy and implantation rates were obtained in both fresh and vitrified oocyte donation cycles. With the same clinical outcome as fresh donation cycles, it is possible to use oocyte cryo-banking to minimize the number of oocytes fertilized, thus reduce the number of excess embryos for cryopreservation. As the tremendous demand of oocyte donation remains, the less number of oocyte used per recipient means more recipients could acquire donated oocytes by the use of the oocyte cryo-banking.
P-63 Tuesday, October 26, 2010 THE TIME OF CRYOPRESERVATION DOES NOT INCREASE THE DAMAGE IN FROZEN OVARIAN TISSUE FOR FERTILITY PRESERVATION. J. R. Campos, B. R. Carvalho, J. C. Rosa-e-Silva, A. A. Vireque, R. A. Ferrani, A. C. J. S. Rosa-e-Silva. Obstetrics and Gynecology, Faculty of Medicine of Ribeir~ao Preto - University of S~ao Paulo, Ribeir~ao Preto, S~ao Paulo, Brazil. OBJECTIVE: To evaluate the time effect on ovarian tissue cryopreservation by analyzing the fresh and frozen-thawed samples according to its follicular density, follicular viability and steroidogenic capacity, comparing thawing after 30 and 180 days after the freezing procedure. DESIGN: A prospective study.
S111