Single Step, Two-Steps, and Microfluidics in Embryo Culture. How Do They Measure up in Improving Embryo Quality?

Abstract

The issues surrounding the culture of the preimplantation human embryo are distinct from those associated with the propagation of somatic cells, given that the starting point of culture, the pronucleate oocyte, and the final stage of development, the blastocyst, are as distinct as any two cell types can be with regards to their nutrient requirements and metabolism. Furthermore, the cleavage stage period of embryo development is exquisitely sensitive to its environment and has limited capacity to regulate against disruptions in its surroundings, especially changes in pH. Historically the media used in human IVF, such as Human Tubal Fluid medium, were based on simple salt solutions but lacked key regulators of blastomere function, specifically amino acids. A resurgence of research in embryo metabolism and physiology in the 1980s paved the way for the development of Sequential media, designed not only to accommodate the changing requirements of the embryo itself (both in terms of carbohydrates and amino acids), but which were also based on available data on the dynamic/differing composition of the Fallopian tube and uterus. Sequential media, especially when used in combination with isolettes, were shown to be highly effective in supporting blastocyst development and significantly increased implantation rates, thereby facilitating the worldwide movement to single embryo transfer. However, while this approach partially satisfied the changing nutrient requirements of the embryo, and further facilitated the removal of toxins built up in the culture drop over 48h, such as ammonium, it necessitated moving embryos. Unless undertaken using warmed gassed isolettes, moving embryos inevitably resulted in shifts in temperature and pH. With the advent of time-lapse, there began a growing movement towards the use of one-step media in order to keep the embryo within an imaging incubator. To facilitate this, hybrid type media were formulated, and while they do not accommodate the dynamic requirements of the preimplantation period, their use meant embryos were not exposed to changes in either temperature or pH. To date, clinical data have reported similar outcomes using both media approaches, plausibly due to a trade-off of the inherent limitations of either approach. An alternative approach proposed decades ago, is to provide the embryo with a gradient of nutrients within a perfusion system, thereby facilitating a dynamic culture environment without the need to move embryos. Unfortunately, until recently, the technology to support this concept has not been available. However, the recent development of 3D printing using 2-photo lithography, has made it possible to create embryo-specific perfusion systems. Hence, we are now able to evaluate the potential of such dynamic systems and could find ourselves on the verge of a new era in embryo culture.