Cell fate regulation during gastrulation in humans and pigs

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To the renowned embryologist Lewis Wolpert is attributed the quote: "It is not birth, marriage or death, but gastrulation which is truly the most important time in your life". Gastrulation is the period of embryo development (around the second week of pregnancy in humans) when the major cell precursors that will form the organs in our body are determined. It is a critical time because if such development is disrupted, it can lead to irreversible abnormalities or have long term detrimental consequences for health in later life. How the gastrulation process occurs in humans is very poorly understood because human embryos cannot be directly studied during this period for ethical reasons. Mouse embryos have served as good models for understanding general principles of mammalian development, but we also know that mice and humans are very different in their development. Indeed, some of the key developmental differences correlate with distinct embryo morphology in mice and humans. Mouse embryos develop as a conical structure in which interactions with tissues contributing to the placenta are very different from other large mammals. Humans develop flat embryos, which establish different relationships with placental tissues. We have studied the significance of these differences in detail in pig embryos, which also develop as flat discs like humans. Our research showed that the genes that regulate the formation of sperm and egg precursor cells are different between pigs and mice. Importantly, we also showed that the mechanism used by pig is also used in human and other primate embryos. This landmark study showed for the first time that the molecular identity of a cell is different between mice and other larger mammals. Furthermore, it indicated that pig embryos can be used as a model for non-rodents mammals and be used to help understand human development. Further research is needed to find out whether differences also occur during the formation of other cells and organs, such as the muscle, liver, gut and pancreas. Such new understanding will ultimately help inform methods for driving human stem cells towards specific cell types with therapeutic potential. The first objective of the research is to determine the genetic program of different cell precursors during gastrulation in pig embryos. Using single cell molecular analysis we will build a map of how cells become restricted towards different cellular identities. This data will be then compared with available data from mouse and human stem cell differentiation studies that will allow us to establish similarities in developmental mechanisms between species. Next, we will validate these findings using new genome editing technologies to disrupt protein function in embryos and determine the morphology of mutant embryos. We will determine cell identity in mutant embryos and establish the roles played by these key proteins during this critical period of development. The final objective of the research is to determine the developmental pathway of human cells in pig/human chimeric embryos. Embryos generated from two species, known as chimeras, are excellent tools for assessing cell developmental potential in vivo. Here we will test the capacity of newly establish hESC and other progenitors combined with chemical methods to prevent cell death to enhance chimera integration. We will use labelling technologies to track cell development and determine cell fate in pig embryos. Understanding how pig/human chimeras develop will be an important step towards generating human organs in large animals for xenotransplantation. A better understanding of the biological barriers and technical limitations of these approaches will improve the safety and efficacy of these new transplantation technologies.