Zona thinning: ‘Laying it on thick’

Abstract

The zona pellucida is a multi-layered glycoprotein coat surrounding the mammalian egg and embryo. To implant successfully in the uterine wall, the embryo must shed this coat. During human IVF, two approaches have been used to facilitate this event in order to enhance implantation: (I) breaching the zona pellucida entirely (assisted hatching) and (II) removing a limited stretch of the outermost layer (zona thinning). It is important to distinguish between these two approaches since the outermost layer of the human zona pellucida appears less hardened than the inner layers. In this issue of Reproductive BioMedicine Online, Valojerdi and co-workers from Tehran (Iran) report on detrimental effects of a rather vigorous method of zona pellucida thinning using a laser along one quarter of the embryo’s circumference. This is the first report showing a detrimental effect after zona manipulation without blastomere biopsy. There are over 400 papers on assisted hatching and zona thinning in the literature with the majority having reported clinical results. Generally speaking, two types of outcomes have been reported: (I) an improvement in pregnancy and/or implantation rate, or (II) no effect of the intervention. The paper by Valojerdi et al. adds a new category to this list: a discernible detrimental effect of the intervention. In general, it is not surprising that outcomes have been so varied following zona pellucida manipulation, particularly considering the many different approaches used and the varied circumstances under which the investigations have been conducted. It is a considerable task to sort through all the studies and clinical trials. The latest Cochrane Review of the assisted hatching literature was published earlier this year (Das et al., 2009Das S. Blake D. Farquhar C. Seif M.M.W. Assisted hatching on assisted conception (IVF and ICSI).Cochrane Database System. Rev. 2009; https://doi.org/10.1002/14651858.CD001894.pub4Crossref PubMed Scopus (63) Google Scholar). The abstract of this study does not detail the long list of its findings and conclusions and anyone interested enough in this technique should perhaps read the entire document (which is nearly 80 pages long). The conclusion of this review is that in certain embryos assisted hatching has some clinical benefit, including those that were thawed or devitrified. Assisted hatching in the largest acceptable cohort of papers increased the chance of pregnancy, but not that of miscarriage. Yet, in the paper by Valojerdi et al., zona thinning appears detrimental in devitrified embryos. The authors provide a concise analysis of what could have caused this outcome, noting that similar studies do not confirm their findings. The message from this important work is that circumstances under which clinical trials involving methodology are conducted may not be comparable. Operator skill and experience and an understanding of the complexities of tools such as lasers play a critical role and, if compromised, can undermine a perfectly acceptable randomization protocol in a clinical trial. In the case of lasers, responsible manufacturers of certified instruments have made a significant effort to make their instruments sensitive and apparently fullproof for the inexperienced user. However, following manufacturer’s instructions carefully may not be enough. For instance embryologists usually calibrate the laser using ink painted on one side of a regular glass slide, but the Petri dish is not made of glass and the embryo is much nearer to the lens in a dish than the ink on a slide. Thinning the zona pellucida over one quarter of the embryo’s circumference involves a relatively large amount of energy, which may penetrate beyond the zona and into the embryo proper. Calibration errors may result in extensive release of energy into the embryo since the laser will not be properly focused. This is just one possible explanation for the observations in the study of Valojerdi et al. Although assisted hatching was introduced into clinical practice some 20 years ago, the method has not yet been standardized. This is unfortunate since it contributes to variability in results. Relying on publications and manufacturer guidelines alone is not always sufficient for proper application of complex technologies. Therefore careful (small) pilot studies should be conducted prior to large-scale trials in order to avoid unintended and unexpected detrimental effects in large numbers of cases.