An Overview of Temporal Trends in Multiple Births After Assisted Reproductive Technology in the USA

Judy E Stern


Expert Rev of Obstet Gynecol. 2013;8(4):357-368. 

In This Article

Increasing Implantation Rate

Changes in culture media formulations, as well as adjustments to laboratory conditions, have been at the heart of increasing ART effectiveness.[33–35] Culture media used for embryo growth in the early days of ART came from work with mouse and cattle embryos and we have come a long way from the media described with regard to an early, pre Louise Brown, IVF attempt as a "gloppy mixture in the test tube, which resembled a frothy chocolate milkshake".[36] Adjustments in media have included developing formulations that mimic human fallopian tube fluids[37] and media for which the components are changed in sequence over the course of several days in culture to mimic oviduct physiology.[38,39] Characteristics of these culture media and the affects on embryo development have been reviewed previously.[40]

In the early days of ART, embryos could not often be kept viable in culture for more than two days postfertilization. Now they are routinely cultured until day 5 or 6. This change has helped us select the embryo or embryos to transfer. On day 2 or 3 of culture, there will generally be more than one embryo that appears to be morphologically healthy and dividing normally, but only some of these embryos will remain viable and continue to develop with increasing time in culture. By extending an embryo's time in culture, we can observe that embryo as its continued development occurs. Culture to day 5 or 6 – also known as blastocyst culture – has improved live birth rates by allowing us to better choose embryos more capable of implantation and further development.[38] Choosing an embryo on day 5 is one way to better obtain a single embryo to transfer[41] and has been shown in at least one prospective randomized trial to be better than transfer of a single embryo on day 3.[42]

As with other ART advances, however, the increase in implantation potential of blastocysts has led to its own increased risk of multiple birth.[43,44] Kissin et al. compared multiple birth rate between day 3 and day 5 and found that the risk of multiples with transfer of two blastocysts (day 5) is greater.[44] They suggest that the only way to avoid this risk is to transfer no more than one blastocyst, although they demonstrate that for many patients, the live birth rate will be lower with transfer of one rather than two embryos. There may be other risks of extended culture to day 5. One difficulty that arises is that when embryos are held to day 5, some patients will not have an embryo transfer as a result of all embryos having arrested development.[43] There have been several reports of increased monozygotic twinning following blastocyst culture.[45,46] Of more concern, blastocyst culture might be associated with an increased rate of abnormalities in offspring and there are reports of imprinting disorders in animal models.[47,48] Whether these risks will turn out to be significant remains to be determined.

Other methods are being developed to assist in choosing the embryo with the greatest potential for implantation. One method is preimplantation screening in which biopsied cells of an embryo are assessed to screen for normal chromosome complement and thereby increase implantation and reduce pregnancy loss.[49] Embryos are biopsied by removing one or more cells on either day 3 or day 5 of culture in vitro. Assessment of the biopsied cells is made by FISH with five or more DNA probes. Embryos with normal genetic compliment, thought to have the most potential for continued development, are transferred: abnormal embryos are discarded. This procedure was used at many clinics for a number of years, particularly in women over 38 years where chromosomal abnormalities are common. Although early studies indicated that pregnancy loss rates were significantly improved with the technique,[49] a prospective randomized trial in 2007 suggested that pregnancy rates were actually lower in the biopsied group.[50] Nevertheless, recent studies using more complex gene analysis procedures have renewed hopes that these procedures will be useful in choosing the embryo most likely to implant.[51–53]

Noninvasive embryo screening methods are also being trialed. The morphological appearance of the embryo (cell number or stage, symmetry, fragmentation and other parameters) has long been used as a measure of an embryo's potential for implantation and, therefore, as a way of choosing embryos to transfer. These parameters, visually assessed under a light microscope, have not been sufficient to adequately choose only the high potential embryos. A different method for making this choice measures an embryo's metabolism through analysis of the culture media in which the embryo is grown. This method, known as metabolomics, involves assessment of substances such as glucose, pyruvate or amino acids.[54] Although preliminary studies suggest that this methodology may assist in choosing the best embryo, it is currently technically complex, expensive and not in general use in ART programs.

Another method for evaluating embryos through their morphologic appearance involves time lapse video monitoring of the embryo to measure developmental milestones with continued culture.[55] A video camera is set up, often within the incubator, and embryo development is measured in real time. Milestones that may ultimately provide cues to improved live birth rates include the time required for key mitotic steps to take place on different days of culture or the synchronicity with which the embryo divides.[56] As with metabolomics, time lapse monitoring requires equipment and expertise not currently available in the majority of ART laboratories.

Although the above methods may ultimately hold the key to our ability to choose a single embryo for transfer and eliminate the risk of multiples, none is currently in general use and their future promise has yet to be realized.