Accepted Manuscript Title: Oocyte vitrification in the 21st century and post-warming fertility outcomes: a systematic review and meta-analysis Author: Neelam Potdar, Tarek A Gelbaya, Luciano G Nardo PII: DOI: Reference:
S1472-6483(14)00247-8 http://dx.doi.org/doi:10.1016/j.rbmo.2014.03.024 RBMO 1148
To appear in:
Reproductive BioMedicine Online
Received date: Revised date: Accepted date:
6-8-2013 20-3-2014 25-3-2014
Please cite this article as: Neelam Potdar, Tarek A Gelbaya, Luciano G Nardo, Oocyte vitrification in the 21st century and post-warming fertility outcomes: a systematic review and meta-analysis, Reproductive BioMedicine Online (2014), http://dx.doi.org/doi:10.1016/j.rbmo.2014.03.024. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Vitrified oocytes and fertility outcomes
Oocyte vitrification in the 21st century and post-warming fertility outcomes: a systematic review and meta-analysis
Neelam Potdar1, Tarek A Gelbaya1, Luciano G Nardo2 1 Leicester Fertility Centre, University Hospitals of Leicester, Leicester, LE1 5WW, UK. 2 Reproductive Medicine and Surgery Unit, Gynehealth, Manchester, M3 4DN, UK
Abstract Oocyte cryopreservation is a rapidly developing technology, which is increasingly being used for various medical, legal and social reasons. There are inconsistencies in information regarding survival rate and fertility outcomes. This systematic review and meta-analysis provides evidence-based information about oocyte survival and fertility outcomes post warming to help women to make informed choices. All randomized and non-randomized, controlled and prospective cohort studies using oocyte vitrification were included. The primary outcome measure was ongoing pregnancy rate/warmed oocyte. Sensitivity analysis for donor and non-donor oocyte studies was performed. Proportional meta-analysis of 17 studies, using a random-effects model, showed pooled ongoing pregnancy and clinical pregnancy rates per warmed oocyte of 7%. Oocyte survival, fertilization, cleavage, clinical pregnancy and ongoing pregnancy rates per warmed oocyte were higher in donor versus non-donor studies. Comparing vitrified with fresh oocytes, no statistically significant difference was observed in fertilization, cleavage and clinical pregnancy rates, but ongoing pregnancy rate was reduced in the vitrified group (odds ratio 0.74), with heterogeneity between studies. Considering the age of women and the reason for cryopreservation, reasonable information can be given to help women to make informed choices. Future studies with outcomes from oocytes cryopreserved for gonadotoxic treatment may provide more insight. Keywords: ICSI, IVF outcomes, oocyte cryopreservation, slow freezing, vitrification
Introduction Globally, various medical, legal and social reasons have emerged for oocyte cryopreservation. Traditionally, cryopreservation of oocytes has been considered for fertility preservation in women undergoing gonadotoxic treatment; other reasons are cryopreservation for oocyte donation programmes, and, in certain countries where the law prohibits embryo cryopreservation and gamete donation, excess oocytes have been cryopreserved for future use. Furthermore, with changing social cultures
Page 1 of 31
and role of women in the 21st century, social egg cryopreservation is gaining prevalence as a method of preserving reproductive potential. Since oocyte cryopreservation is a rapidly developing technology, there are inconsistencies in information provided to women with regard to survival rate and fertility outcome. Until recently, oocyte cryopreservation has been considered experimental; therefore clinicians and service providers have not been sure themselves of the actual success rate for oocyte cryopreservation. Many centres have cryopreserved oocytes for years, especially for oncology patients, before fertilization and transfer into women’s uteri. Many other oocytes have never been used because patients either regained their natural fertility or deceased. The first human birth from a cryopreserved oocyte was reported in 1986 (Chen 1986). The primary challenge with oocyte cryopreservation has been maintaining survival of the mature metaphase-II oocyte post warming, which is indirectly related to oocyte plasma membrane stability and permeability to water and cryoprotectants (Agca et al., 1998; Ford et al., 2000). In addition, the oocyte meiotic spindle, which is required for chromosomal segregation, is noted to be extremely sensitive to temperature changes and the dehydration–rehydration process (Baka et al., 1995; Chen et al., 2004; Bianchi et al., 2005; Larman et al., 2007; Cobo et al., 2008a). However, it has been demonstrated that the spindle disintegrates during the freeze–thaw process and that it reassembles again in most oocytes (Gook et al., 1994; Rienzi et al., 2004). Moreover, studies have shown that oocyte cryopreservation does not increase the frequency of developmental abnormalities and pregnancy complications (Chian et al., 2008; Noyes et al., 2009; Forman et al., 2012). The initial technique of oocyte cryopreservation began as slow freezing, followed by modifications to improve oocyte survival (Trad et al., 1999; Fabbri et al., 2001; Boldt et al., 2006; Parmegiani et al., 2008; Bianchi et al., 2012). In the last decade, vitrification techniques have been developed and modified to enhance survival and implantation rates for the oocytes and embryos, respectively (Fabbri et al., 2001; Borini et al., 2004; Kuwayama et al., 2005; Vajta and Nagy 2006; Cobo et al., 2010; Smith et al., 2010; Alpha Scientists in Reproductive Medicine., 2012). Previously, live birth rate per thawed–warmed oocyte was reported as 1.9% for slow freezing and 2.0% for vitrification (Oktay et al., 2006). Studies have compared fertility outcomes using slow-frozen versus vitrified oocytes and fresh versus vitrified oocytes (Cobo et al., 2008b; Rienzi et al., 2010; Ubaldi et al., 2010). The systematic review and metaanalysis by Oktay et al. (2006) compared outcomes of slow-frozen with fresh oocytes and concluded that fertilization, implantation and live birth rates were significantly better with fresh oocytes. Subsequently, Cobo and Diaz (2011) presented a meta-analysis for clinical application of oocyte vitrification and concluded that oocyte survival, fertilization and embryo cleavage rates per warmed oocyte were higher with vitrification compared with slow freezing. Since then, seven additional prospective studies have been conducted (Garcia et al., 2011; Parmegiani et al., 2011; Trokoudes et al., 2011; Cai et al., 2012; Forman et al., 2012; Rienzi et al., 2012;
Page 2 of 31
Chang et al., 2013) providing further evidence regarding outcomes using vitrified oocytes. To help women to make informed choices, this systematic review and meta-analysis was performed to compare fertility outcomes using vitrified oocytes with fresh oocytes and to provide evidence-based information about oocyte survival and fertility outcomes post warming (including ongoing pregnancy and clinical pregnancy rates per warmed oocyte).
Materials and methods Literature search Online searches of databases were performed in MEDLINE (1980–June 2013), EMBASE (1980–June 2013) and the Cochrane Library. The searches also included Conference Proceedings Citation Index and databases for registered and ongoing trials. A combination of Medical Subject Headings and words were used to generate a subset of citations for oocyte cryopreservation (‘oocyte’, ‘slow cooling’, ‘slow freeze’, ‘vitrification’ and ‘cryopreserv*’); for citations including outcomes after IVF and intracytoplasmic sperm injection (ICSI) (‘outcome’, ‘IVF’, ‘in-vitro fertilization’, ‘intracytoplasmic sperm injection’, ‘ICSI’ and ‘assisted reproduct*’). These subsets were combined using ‘AND’ to generate final citations addressing the research question. The reference lists of all published articles including review articles were examined to identify articles not noted by the electronic search of the databases. No language restrictions were placed on the searches so that relevant non-English studies were included. Authors were contacted to obtain further information, as appropriate.
Study eligibility criteria This study included randomized controlled trials (RCT), prospective nonrandomized controlled trials (NRCT) and prospective cohort studies that used vitrified oocytes for ICSI. The inclusion criteria were study population of women undergoing IVF treatment with exclusion of poor responders and intervention of vitrification and/or comparison with fresh or slowfrozen oocytes in the matched control group. In all studies, ICSI was performed because removal of cumulus has the potential to reduce fertilization and because there can be hardening of the zona pellucida post thawing/warming (Gook et al., 1994; Porcu et al., 1997). Observational studies were vigorously reviewed and prospective trials that met all other predefined criteria were included. These studies were included since their exclusion would have led to the omission of vital data and available evidence. The primary reasons for excluding studies were retrospective design or case series, no outcome measure reported and use of different intervention. Study selection and data extraction were performed by two authors (NP and TAG) independently. All articles, including abstracts from the electronic searches, were assessed and citations that met the predefined
Page 3 of 31
selection criteria were obtained. After quality assessment of full manuscripts, final inclusion decisions were made. Any disagreement between the two reviewers was resolved by consultation with the third author (LNG).
Data extraction The selected studies were assessed for the methodological quality. For RCT, quality was assessed using the domain-based risk for bias assessment tool recommended by the Cochrane Collaboration (Higgins et al., 2011). Information was sought on the method of randomization, blinding, allocation concealment, performance, detection, attrition and reporting bias. For non-randomized studies, quality assessment was performed using the Newcastle–Ottawa Quality Assessment Scale for selection, comparability at baseline and exposure or outcome measure and were reported on a scale of 0–9 stars (Wells et al., 2009). Further information was extracted as per the guidelines for Meta-analysis of Observational Studies in Epidemiology (MOOSE) (Stroup et al., 2000). For each study, information was obtained on participants (donor or nondonor sibling oocyte cycles, age and ovarian reserve), intervention used for cryopreservation (vitrification or slow freezing) and outcome measures. Authors were contacted for further details where there was lack of information.
Outcome measures The primary outcome measure was ongoing pregnancy rate (OPR) per thawed–warmed oocyte. Secondary outcome measures were oocyte survival rate, fertilization rate, cleavage rate and clinical pregnancy rate (CPR) per warmed oocyte and miscarriage rate. OPR was defined as ongoing pregnancy beyond 20 weeks of gestation; oocyte survival rate/warmed oocyte as the number of oocytes surviving after warming/thawing process; fertilization rate as the number of warmed/thawed oocytes fertilized after ICSI; cleavage rate as the number of embryos cleaved on day 3 of development (for most studies, the grading/scoring system used for classifying cleavage-stage embryos and blastocysts have been mentioned (Table 1; Alikani et al., 1999; Tomas et al., 1998; Rienzi et al., 2002; Veeck. 1999; Gardner et al., 1998; Alpha Scientists in Reproductive and Embryology 2011); and CPR was defined as gestational sac and fetal heart activity seen on transvaginal ultrasound scan after 6 weeks of gestation. Miscarriage rate was defined as pregnancy loss up to 20 weeks of gestation including ectopic pregnancies.
Search results The studies were selected and reported according to the PRISMA guidelines 2009 (Moher et al. 2009). A total of 3201 citations were identified, 48 of which were selected for detailed study. Finally 21 articles were included in the systematic review and meta-analysis (seven RCT, seven NRCT and seven prospective cohort studies; Figure 1). Twentyseven citations were excluded because they were review articles, case
Page 4 of 31
series or retrospective studies or because of differences in the intervention used; of these, 20 studies are described in detail (Table 2; Kuleshova et al., 1999; Yoon et al., 2000; Katayama et al., 2003; Chen et al., 2005; Kuwayama et al., 2005; Kyono et al., 2005; Lucena et al., 2006; Selman et al., 2006; Albani et al., 2008; Parmegiani et al., 2008; Fadini et al., 2009; Parmegiani et al., 2009; Borini et al., 2010; Noyes et al., 2010; Scaravelli et al., 2010; Selman et al., 2010; Hodes-Wertz et al., 2011; Bianchi et al., 2012; Tong et al., 2012; Garcia-Velasco et al., 2013).
Statistical analysis Study features, intervention used and outcomes were assembled in a tabular form, and formal meta-analysis was performed using Review Manager 5.1 (The Cochrane Collaboration 2011). To compare oocyte vitrification outcomes with slow-frozen or fresh oocytes, a fixed-effects model (Mantel–Haenszel method) were used where I2 statistic was 15 oocytes retrieved and surplus oocytes cryopreserved randomly
Intervention
Day of embryo transfer
Vitrification procedure
Outcomes reporteda
Vitrification versus fresh
Day 3 and blastocyst; fragmentation scoring Alikani et al. (1999); mean embryos transferred 2.1 1.2
Kuwayama et al. (2005); Cryotop method; vitrification for 1 h before warming
OSR, FR, CR, BR, IR, BPR; pregnancy outcomes of CPR, MR, MPR, OPR reported only for vitrified oocytes
Vitrification versus slow freezing
Days 3 and 5, scoring criteria Tomas et al. (1998)
Vitrification kit (Medicult, Jyllinge, Denmark); McGill Cryoleaf used; vitrification for minimum 1 month prior to warming
No transfer and outcomes reported; OSR, FR, CR, BR; oocytes with abnormalities
Comment [KMT8]: Author: please confirm edit or amend.
Page 17 of 31
Cobo et al. (2010)
Spain
Rienzi et al. (2010)
Italy
into two methods; GnRH antagonist cycle, stimulation with HMG, oocyte denudation using hyaluronidase and mechanical disruption Donor study: 600 women requiring donor oocytes recruited; 300 received vitrified and 300 fresh oocytes; age 2 miscarriages or other disorders; recipients’ inclusion criteria: age 18–49 years, BMI 18– 29 kg/m2, 6 normal-appearing MII oocytes undergoing ICSI, exclusion surgically retrieved spermatozoa and severe OAT; women’s age, cause of infertility similar in both groups; ovarian stimulation using GnRH agonist and antagonist regimes, HCG used for trigger Non-donor study: 230 infertility patients with >9 mature oocytes retrieved at ovarian stimulation; causes included tubal factor, severe male factor and unexplained infertility; follicular FSH 10 IU/l,
Vitrification versus slow freezing
al. (2002); mean embryos transferred 2.3 0.88
commercial kit (Kitazato BioPharma, Japan)
Day 3, graded by (Veeck, 1999); mean embryos transferred 3.1 0.1
Kuwayama et al. (2005); vitrificationto-warming interval 1 month
OSR, FR, CR, IR, BPR, CPR MR, MPR, LBR
age 32 1 years, BMI 30 kg/m2, regular cycles and presence of both ovaries and normal uterine cavity; laser-assisted hatching
Page 19 of 31
Parmegiani et al. (2011)
Forman et al. (2012)
Italy
USA
performed for all embryos Non-donor study: 31 patients, using sibling oocytes, >5 MII oocytes, age 35 0.8 years, ICSI with ejaculated spermatozoa; surgically retrieved and severe OAT excluded; liquid N2 sterilization via UV irradiation Non-donor study: 44 women, age 29.9 2.3 years, BMI 20 oocytes retrieved, consented for oocyte donation, age 28.1 3.2 years; 47 consented donation of surplus vitrified oocytes after healthy live birth Non-donor study: multicentric study, 450 couples, routine fertility
OSR, FR, CR, BR, IR, CPR, MPR, LBR
Page 25 of 31
programme, age
S1472-6483(14)00247-8 http://dx.doi.org/doi:10.1016/j.rbmo.2014.03.024 RBMO 1148
To appear in:
Reproductive BioMedicine Online
Received date: Revised date: Accepted date:
6-8-2013 20-3-2014 25-3-2014
Please cite this article as: Neelam Potdar, Tarek A Gelbaya, Luciano G Nardo, Oocyte vitrification in the 21st century and post-warming fertility outcomes: a systematic review and meta-analysis, Reproductive BioMedicine Online (2014), http://dx.doi.org/doi:10.1016/j.rbmo.2014.03.024. This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Vitrified oocytes and fertility outcomes
Oocyte vitrification in the 21st century and post-warming fertility outcomes: a systematic review and meta-analysis
Neelam Potdar1, Tarek A Gelbaya1, Luciano G Nardo2 1 Leicester Fertility Centre, University Hospitals of Leicester, Leicester, LE1 5WW, UK. 2 Reproductive Medicine and Surgery Unit, Gynehealth, Manchester, M3 4DN, UK
Abstract Oocyte cryopreservation is a rapidly developing technology, which is increasingly being used for various medical, legal and social reasons. There are inconsistencies in information regarding survival rate and fertility outcomes. This systematic review and meta-analysis provides evidence-based information about oocyte survival and fertility outcomes post warming to help women to make informed choices. All randomized and non-randomized, controlled and prospective cohort studies using oocyte vitrification were included. The primary outcome measure was ongoing pregnancy rate/warmed oocyte. Sensitivity analysis for donor and non-donor oocyte studies was performed. Proportional meta-analysis of 17 studies, using a random-effects model, showed pooled ongoing pregnancy and clinical pregnancy rates per warmed oocyte of 7%. Oocyte survival, fertilization, cleavage, clinical pregnancy and ongoing pregnancy rates per warmed oocyte were higher in donor versus non-donor studies. Comparing vitrified with fresh oocytes, no statistically significant difference was observed in fertilization, cleavage and clinical pregnancy rates, but ongoing pregnancy rate was reduced in the vitrified group (odds ratio 0.74), with heterogeneity between studies. Considering the age of women and the reason for cryopreservation, reasonable information can be given to help women to make informed choices. Future studies with outcomes from oocytes cryopreserved for gonadotoxic treatment may provide more insight. Keywords: ICSI, IVF outcomes, oocyte cryopreservation, slow freezing, vitrification
Introduction Globally, various medical, legal and social reasons have emerged for oocyte cryopreservation. Traditionally, cryopreservation of oocytes has been considered for fertility preservation in women undergoing gonadotoxic treatment; other reasons are cryopreservation for oocyte donation programmes, and, in certain countries where the law prohibits embryo cryopreservation and gamete donation, excess oocytes have been cryopreserved for future use. Furthermore, with changing social cultures
Page 1 of 31
and role of women in the 21st century, social egg cryopreservation is gaining prevalence as a method of preserving reproductive potential. Since oocyte cryopreservation is a rapidly developing technology, there are inconsistencies in information provided to women with regard to survival rate and fertility outcome. Until recently, oocyte cryopreservation has been considered experimental; therefore clinicians and service providers have not been sure themselves of the actual success rate for oocyte cryopreservation. Many centres have cryopreserved oocytes for years, especially for oncology patients, before fertilization and transfer into women’s uteri. Many other oocytes have never been used because patients either regained their natural fertility or deceased. The first human birth from a cryopreserved oocyte was reported in 1986 (Chen 1986). The primary challenge with oocyte cryopreservation has been maintaining survival of the mature metaphase-II oocyte post warming, which is indirectly related to oocyte plasma membrane stability and permeability to water and cryoprotectants (Agca et al., 1998; Ford et al., 2000). In addition, the oocyte meiotic spindle, which is required for chromosomal segregation, is noted to be extremely sensitive to temperature changes and the dehydration–rehydration process (Baka et al., 1995; Chen et al., 2004; Bianchi et al., 2005; Larman et al., 2007; Cobo et al., 2008a). However, it has been demonstrated that the spindle disintegrates during the freeze–thaw process and that it reassembles again in most oocytes (Gook et al., 1994; Rienzi et al., 2004). Moreover, studies have shown that oocyte cryopreservation does not increase the frequency of developmental abnormalities and pregnancy complications (Chian et al., 2008; Noyes et al., 2009; Forman et al., 2012). The initial technique of oocyte cryopreservation began as slow freezing, followed by modifications to improve oocyte survival (Trad et al., 1999; Fabbri et al., 2001; Boldt et al., 2006; Parmegiani et al., 2008; Bianchi et al., 2012). In the last decade, vitrification techniques have been developed and modified to enhance survival and implantation rates for the oocytes and embryos, respectively (Fabbri et al., 2001; Borini et al., 2004; Kuwayama et al., 2005; Vajta and Nagy 2006; Cobo et al., 2010; Smith et al., 2010; Alpha Scientists in Reproductive Medicine., 2012). Previously, live birth rate per thawed–warmed oocyte was reported as 1.9% for slow freezing and 2.0% for vitrification (Oktay et al., 2006). Studies have compared fertility outcomes using slow-frozen versus vitrified oocytes and fresh versus vitrified oocytes (Cobo et al., 2008b; Rienzi et al., 2010; Ubaldi et al., 2010). The systematic review and metaanalysis by Oktay et al. (2006) compared outcomes of slow-frozen with fresh oocytes and concluded that fertilization, implantation and live birth rates were significantly better with fresh oocytes. Subsequently, Cobo and Diaz (2011) presented a meta-analysis for clinical application of oocyte vitrification and concluded that oocyte survival, fertilization and embryo cleavage rates per warmed oocyte were higher with vitrification compared with slow freezing. Since then, seven additional prospective studies have been conducted (Garcia et al., 2011; Parmegiani et al., 2011; Trokoudes et al., 2011; Cai et al., 2012; Forman et al., 2012; Rienzi et al., 2012;
Page 2 of 31
Chang et al., 2013) providing further evidence regarding outcomes using vitrified oocytes. To help women to make informed choices, this systematic review and meta-analysis was performed to compare fertility outcomes using vitrified oocytes with fresh oocytes and to provide evidence-based information about oocyte survival and fertility outcomes post warming (including ongoing pregnancy and clinical pregnancy rates per warmed oocyte).
Materials and methods Literature search Online searches of databases were performed in MEDLINE (1980–June 2013), EMBASE (1980–June 2013) and the Cochrane Library. The searches also included Conference Proceedings Citation Index and databases for registered and ongoing trials. A combination of Medical Subject Headings and words were used to generate a subset of citations for oocyte cryopreservation (‘oocyte’, ‘slow cooling’, ‘slow freeze’, ‘vitrification’ and ‘cryopreserv*’); for citations including outcomes after IVF and intracytoplasmic sperm injection (ICSI) (‘outcome’, ‘IVF’, ‘in-vitro fertilization’, ‘intracytoplasmic sperm injection’, ‘ICSI’ and ‘assisted reproduct*’). These subsets were combined using ‘AND’ to generate final citations addressing the research question. The reference lists of all published articles including review articles were examined to identify articles not noted by the electronic search of the databases. No language restrictions were placed on the searches so that relevant non-English studies were included. Authors were contacted to obtain further information, as appropriate.
Study eligibility criteria This study included randomized controlled trials (RCT), prospective nonrandomized controlled trials (NRCT) and prospective cohort studies that used vitrified oocytes for ICSI. The inclusion criteria were study population of women undergoing IVF treatment with exclusion of poor responders and intervention of vitrification and/or comparison with fresh or slowfrozen oocytes in the matched control group. In all studies, ICSI was performed because removal of cumulus has the potential to reduce fertilization and because there can be hardening of the zona pellucida post thawing/warming (Gook et al., 1994; Porcu et al., 1997). Observational studies were vigorously reviewed and prospective trials that met all other predefined criteria were included. These studies were included since their exclusion would have led to the omission of vital data and available evidence. The primary reasons for excluding studies were retrospective design or case series, no outcome measure reported and use of different intervention. Study selection and data extraction were performed by two authors (NP and TAG) independently. All articles, including abstracts from the electronic searches, were assessed and citations that met the predefined
Page 3 of 31
selection criteria were obtained. After quality assessment of full manuscripts, final inclusion decisions were made. Any disagreement between the two reviewers was resolved by consultation with the third author (LNG).
Data extraction The selected studies were assessed for the methodological quality. For RCT, quality was assessed using the domain-based risk for bias assessment tool recommended by the Cochrane Collaboration (Higgins et al., 2011). Information was sought on the method of randomization, blinding, allocation concealment, performance, detection, attrition and reporting bias. For non-randomized studies, quality assessment was performed using the Newcastle–Ottawa Quality Assessment Scale for selection, comparability at baseline and exposure or outcome measure and were reported on a scale of 0–9 stars (Wells et al., 2009). Further information was extracted as per the guidelines for Meta-analysis of Observational Studies in Epidemiology (MOOSE) (Stroup et al., 2000). For each study, information was obtained on participants (donor or nondonor sibling oocyte cycles, age and ovarian reserve), intervention used for cryopreservation (vitrification or slow freezing) and outcome measures. Authors were contacted for further details where there was lack of information.
Outcome measures The primary outcome measure was ongoing pregnancy rate (OPR) per thawed–warmed oocyte. Secondary outcome measures were oocyte survival rate, fertilization rate, cleavage rate and clinical pregnancy rate (CPR) per warmed oocyte and miscarriage rate. OPR was defined as ongoing pregnancy beyond 20 weeks of gestation; oocyte survival rate/warmed oocyte as the number of oocytes surviving after warming/thawing process; fertilization rate as the number of warmed/thawed oocytes fertilized after ICSI; cleavage rate as the number of embryos cleaved on day 3 of development (for most studies, the grading/scoring system used for classifying cleavage-stage embryos and blastocysts have been mentioned (Table 1; Alikani et al., 1999; Tomas et al., 1998; Rienzi et al., 2002; Veeck. 1999; Gardner et al., 1998; Alpha Scientists in Reproductive and Embryology 2011); and CPR was defined as gestational sac and fetal heart activity seen on transvaginal ultrasound scan after 6 weeks of gestation. Miscarriage rate was defined as pregnancy loss up to 20 weeks of gestation including ectopic pregnancies.
Search results The studies were selected and reported according to the PRISMA guidelines 2009 (Moher et al. 2009). A total of 3201 citations were identified, 48 of which were selected for detailed study. Finally 21 articles were included in the systematic review and meta-analysis (seven RCT, seven NRCT and seven prospective cohort studies; Figure 1). Twentyseven citations were excluded because they were review articles, case
Page 4 of 31
series or retrospective studies or because of differences in the intervention used; of these, 20 studies are described in detail (Table 2; Kuleshova et al., 1999; Yoon et al., 2000; Katayama et al., 2003; Chen et al., 2005; Kuwayama et al., 2005; Kyono et al., 2005; Lucena et al., 2006; Selman et al., 2006; Albani et al., 2008; Parmegiani et al., 2008; Fadini et al., 2009; Parmegiani et al., 2009; Borini et al., 2010; Noyes et al., 2010; Scaravelli et al., 2010; Selman et al., 2010; Hodes-Wertz et al., 2011; Bianchi et al., 2012; Tong et al., 2012; Garcia-Velasco et al., 2013).
Statistical analysis Study features, intervention used and outcomes were assembled in a tabular form, and formal meta-analysis was performed using Review Manager 5.1 (The Cochrane Collaboration 2011). To compare oocyte vitrification outcomes with slow-frozen or fresh oocytes, a fixed-effects model (Mantel–Haenszel method) were used where I2 statistic was 15 oocytes retrieved and surplus oocytes cryopreserved randomly
Intervention
Day of embryo transfer
Vitrification procedure
Outcomes reporteda
Vitrification versus fresh
Day 3 and blastocyst; fragmentation scoring Alikani et al. (1999); mean embryos transferred 2.1 1.2
Kuwayama et al. (2005); Cryotop method; vitrification for 1 h before warming
OSR, FR, CR, BR, IR, BPR; pregnancy outcomes of CPR, MR, MPR, OPR reported only for vitrified oocytes
Vitrification versus slow freezing
Days 3 and 5, scoring criteria Tomas et al. (1998)
Vitrification kit (Medicult, Jyllinge, Denmark); McGill Cryoleaf used; vitrification for minimum 1 month prior to warming
No transfer and outcomes reported; OSR, FR, CR, BR; oocytes with abnormalities
Comment [KMT8]: Author: please confirm edit or amend.
Page 17 of 31
Cobo et al. (2010)
Spain
Rienzi et al. (2010)
Italy
into two methods; GnRH antagonist cycle, stimulation with HMG, oocyte denudation using hyaluronidase and mechanical disruption Donor study: 600 women requiring donor oocytes recruited; 300 received vitrified and 300 fresh oocytes; age 2 miscarriages or other disorders; recipients’ inclusion criteria: age 18–49 years, BMI 18– 29 kg/m2, 6 normal-appearing MII oocytes undergoing ICSI, exclusion surgically retrieved spermatozoa and severe OAT; women’s age, cause of infertility similar in both groups; ovarian stimulation using GnRH agonist and antagonist regimes, HCG used for trigger Non-donor study: 230 infertility patients with >9 mature oocytes retrieved at ovarian stimulation; causes included tubal factor, severe male factor and unexplained infertility; follicular FSH 10 IU/l,
Vitrification versus slow freezing
al. (2002); mean embryos transferred 2.3 0.88
commercial kit (Kitazato BioPharma, Japan)
Day 3, graded by (Veeck, 1999); mean embryos transferred 3.1 0.1
Kuwayama et al. (2005); vitrificationto-warming interval 1 month
OSR, FR, CR, IR, BPR, CPR MR, MPR, LBR
age 32 1 years, BMI 30 kg/m2, regular cycles and presence of both ovaries and normal uterine cavity; laser-assisted hatching
Page 19 of 31
Parmegiani et al. (2011)
Forman et al. (2012)
Italy
USA
performed for all embryos Non-donor study: 31 patients, using sibling oocytes, >5 MII oocytes, age 35 0.8 years, ICSI with ejaculated spermatozoa; surgically retrieved and severe OAT excluded; liquid N2 sterilization via UV irradiation Non-donor study: 44 women, age 29.9 2.3 years, BMI 20 oocytes retrieved, consented for oocyte donation, age 28.1 3.2 years; 47 consented donation of surplus vitrified oocytes after healthy live birth Non-donor study: multicentric study, 450 couples, routine fertility
OSR, FR, CR, BR, IR, CPR, MPR, LBR
Page 25 of 31
programme, age
