1 Academic Unit of Obstetrics, Gynaecology and Reproductive Healthcare, St Marys Hospital, Manchester and 2 Health Service Research Unit, Central Manchester Healthcare Trust and MANDEC, University Dental Hospital, Manchester, UK
3 To whom correspondence should be addressed at: Academic Unit of Obstetrics, Gynaecology and Reproductive Healthcare, St Marys Hospital, Manchester, M13 OJH, UK. e-mail: mwseif{at}man.ac.uk
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Abstract |
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Key words: assisted hatching/IVF/meta-analysis/randomized controlled trials/systematic review
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Introduction |
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Cultured embryos develop slowly and poorly in vitro, many fail to achieve blastocyst stage or hatch, and implant at lower rates than occur naturally (Harlow and Quinn, 1982; Hsu et al., 1999
; Mercader et al., 2001
). Hardening of the zona pellucida resulting from cross-linking of its constituent glycoproteins has been implicated in reduced hatching rates (Cohen, 1991
). Zona thickness (influenced by womens age, FSH levels and cause of infertility) has been correlated negatively with embryo implantation rates (Loret De Mola et al., 1997
). The combination of delayed embryo hatching and advanced endometrial development in assisted conception presents a highly unfavourable environment for implantation.
Assisted hatching (AH) is achieved by zona dissection, drilling or thinning, making use of acid solutions, proteinases, piezon vibrators and lasers (Al-Nuaim and Jenkins, 2002). In general, hatched embryos implant one day earlier than unhatched embryos (Rink et al, 1995
). The procedure is increasingly offered to older women, those with high FSH levels, higher risk of zona hardening, and following repeated implantation failure (Al-Nuaim and Jenkins, 2002
). Considerable uncertainty persists regarding the impact of AH, but reports suggest that it might be associated with higher rates of embryo damage and monozygotic twinning (Hershlag et al., 1999
). This review was undertaken to determine the impact of AH on live birth, clinical pregnancy and implantation.
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Materials and methods |
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Selection
All reviewers independently screened the identified abstracts for potential trials, which were retrieved and evaluated for inclusion. Criteria for inclusion were that participants were randomly allocated to AH (post-fertilization therapeutic disruption of the zona pellucida by the known method, including mechanical, chemical or laser) or no AH. The included women underwent only one cycle of treatment and had their own gametes. Trials utilizing mixed groups of hatched and un-hatched embryos were excluded. Primary outcomes included live births, clinical pregnancy and embryo implantation. The consensus definition of implantation (detection of a gestation sac on ultrasound scan) and clinical pregnancy (detection of fetal heart beats on ultrasound scan) were adopted (Human Fertilisation and Embryology Authority, 2000), and data from trials using definitions not conforming to these were excluded from the relevant analyses. Secondary outcomes included miscarriage, ectopic pregnancy, monozygotic twinning, and congenital or chromosomal abnormalities.
Validity and data extraction
Two reviewers (E.E. and L.H.) independently assessed trial quality (method of randomization, adequacy of allocation concealment, blinding, power calculation, intention to treat analysis, publication type and balanced age at baseline) and extracted trial data on forms designed for the review. Disagreements were resolved by discussion with the third reviewer (M.W.S.). Attempts were made to obtain additional information on trial methodology and outcomes from 11 principal authors of the 19 trials located before February 2002.
Quantitative data synthesis
For dichotomous data (such as live births), results for each trial were expressed as numbers of events per woman randomized, embryo transferred or clinical pregnancy, and pooled using random effects methodology (RevMan 4.1 software) where appropriate (discussed later). Heterogeneity between the results of different trials was examined using Cochrans test (assuming statistical significance at P < 0.1). It was intended that possible contributions of differences in participant characteristics to any heterogeneity would be investigated through the sub-grouping by age (< or 37 years), serum FSH (< or
8 IU/l), previous failed assisted conception cycles, and zona thickness (< or
12 µm). Where possible, these sub-groupings were directly extracted from included trials; otherwise the mean trial data were used to place the whole trial in one or other subgroup.
The STATA metareg command for random effects meta-regression and a funnel plot to investigate the presence of publication bias were used. Sensitivity analyses were undertaken to examine the stability of results in relation to adequacy of allocation concealment (removing trials with unclear or inadequate allocation concealment), adequacy of randomization (removing trials with an unclear method of randomization), baseline comparability [removing trials where the 95% confidence intervals (CI) for the difference in mean age between the two arms did not include zero or where insufficient information was provided to assess this].
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Results |
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The results of all meta-analyses, sub-groupings and sensitivity analyses undertaken are detailed in Table II.
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Miscarriage
Miscarriage was reported by 11 trials, with 69 miscarriages complicating 453 clinical pregnancies. Meta-analysis yielded an OR of 0.70 for miscarriage per clinical pregnancy in favour of AH and 1.07 for live birth per clinical pregnancy (the converse of miscarriage rates), again in favour of AH. The validity of the 95% CI and statistical significance of these meta-analyses was, however, questionable.
Other outcomes
Data on other outcomes were sparse, with only three trials reporting ectopic pregnancies, two monozygotic twinning, and two congenital and/or chromosomal abnormalities.
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Discussion |
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The reviewers felt it was of primary interest to determine the overall effect of AH in assisted conception to provide the prime statistic from both provider and consumer perspectives. The review pooled data from all suitable trials irrespective of the AH method employed, population of women studied or type of assisted conception procedure, before undertaking subgroup analysis in situations of particular interest. The overall methodological quality of included trials was sub-optimal as no trial met CONSORT criteria for the reporting of randomized controlled trials; no trials reported a power calculation, intention-to-treat analysis or adequate allocation concealment. This did not appear to improve with time over the 10-year span of the publications (1990 to 2000). Furthermore, only 10 trials described a valid method of randomization, thereby introducing the possibility of further bias to the results.
It was disappointing that the present review was severely limited by paucity of data on the impact of AH on the most sought after outcome of assisted conception, the take-home-baby rate. This probably reflects the gap that currently exists between the practice of assisted conception and clinical obstetrics, and the absence of an electronic database of records and outcomes that would facilitate follow-up of women by authorized agencies (such as the Human Fertilisation and Embryology Authority in the UK). That only six included trials reported live birth data suggested haste on the part of authors to disseminate information that was limited to short-term outcomes. For instance, none of the seven conference abstracts that was included reported live birth data. This should stimulate debate in the medical world about the criteria for reporting and publishing clinical trials of assisted conception. The small amount of existing data suggests that AH has no statistically significant impact on the odds of a live birth, increasing it by an average of 21% (the 95% CI include a reduction of 18% up to an increase of 78%), with significant heterogeneity between trials.
The odds of clinical pregnancy were significantly improved by AH, by an average of 63%, although statistical significance was lost on two of three sensitivity analyses. It was not possible to ascertain from the available evidence whether AH benefits women with high FSH levels, higher risk of zona hardening, or poor response to ovarian stimulation. Sub-group analysis suggested a stronger impact of AH on clinical pregnancy in women after failed cycle(s) of assisted conception (statistically significant OR of 2.33), compared with women undergoing first cycles (OR 1.40). Meta-regression suggested a stronger impact of AH in older women (although falling just short of statistical significance). Whilst additional trials are needed to further explore this trend, the evidence is that AH is most beneficial to women undergoing repeat cycles of assisted conception following previous treatment failure, and possibly the older ones.
Implantation is traditionally expressed per embryo transferred, but the pooling of these data for meta-analysis is statistically problematic. Transferring more than one embryo per woman results in an embryo cluster effect, necessitating an intra-cluster correlation coefficient to make the pooling of data meaningful. Otherwise, the larger number of transferred embryos than women randomized would narrow the 95% CI, suggesting statistical significance where there is none. A statistically more valid approach would be to report implantation per woman randomized. This approach is however based on the statistical assumption that not more than one gestation sac is counted in any particular woman, and this is clearly not the case in practice. The reported trials did not provide any information about the numbers of women in whom gestation sacs were detected, neither was any standard deviation presented for the number of gestation sacs detected per woman or per embryo transferred. In analysing implantation per woman randomized, the review made the statistical assumption that only one gestation sac was detected per woman, omitting the one trial (Isiklar et al., 1999) where the number of gestation sacs detected was greater than the number of women randomized in any arm. In the absence of an ideal way to analyse implantation data, both approaches were used to highlight their pitfalls. Assisted hatching appeared to improve the chance of embryo implantation in any particular woman by 97%, but the statistical significance of this remains questionable.
It was disappointing that the sub-optimal reporting of implantation data prevented investigation of the impact of AH on implantation in different subgroups, particularly women with high FSH levels. It was perplexing that this anomaly had previously not been mentioned in the medical literature despite the very obvious shortcomings of the current method of reporting and analysing implantation data. This problem will not exist if there is universal adherence to a one woman-one embryo strategy, allowing the collation and reporting of implantation data per woman randomized. A temporary solution would be the reporting of intra-cluster correlation coefficients for all trials replacing more than one embryo or counting more than one gestation sac per woman, allowing the collation and reporting of implantation per embryo transferred.
It is hoped that this systematic review generates debate in both provider and consumer groups to explore issues of quality and outcome recording in assisted conception treatment and research, and also leads to use of statistically valid methods of reporting and comparing implantation data.
In conclusion, the sub-optimal quality, flawed reporting and paucity of outcome data weaken the validity and strength of any recommendations from this systematic review and meta-analysis. On the available evidence, AH does not significantly improve the take-home-baby rate of assisted conception. However, it does improve the odds of clinical pregnancy in women undergoing repeat cycles of assisted conception following previous treatment failure, and probably in older women. It is probably justifiable, on the basis of this evidence, to recommend AH to this population. As miscarriage rates are not affected, it can be extrapolated that with an appropriate number of trials reporting usable outcome data, a positive impact would be expected on implantation and live births. This review highlights many unresolved issues that provide potential avenues for future research, including the actual effect of AH on live births, the method of reporting implantation data, the cost implications of AH on the take-home-baby rate, the place of AH in women with high FSH levels, thick or hardened zona pellucida, and the long-term consequences of the procedure on embryo damage, chromosomal abnormalities and congenital malformations. Trials are needed that are of high quality, of adequate size, conform to guidelines of reporting and provide data on live births.
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Acknowledgements |
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References |
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Submitted on March 13, 2003; resubmitted on April 23, 2003; accepted on May 2, 2003.