A Novel Large Animal Model for Studying the Developmental Potential and Function of LGR5 Stem Cells in Vivo and in Vitro

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Project Summary LGR5, a marker of adult and fetal stem cells in various tissues and organs, has been studied extensively for the past decades. Cells expressing LGR5 play key roles in organ/tissue development, homeostasis, regeneration, and disease, including cancer. Therefore, having the ability to identify, track, and manipulate these cells in vitro and in vivo, will allow their detailed role in normal and pathological organ function. To date, most of the research on LGR5 cells has been performed using mice, which have significant anatomical, physiological, and molecular differences from the human. Utilizing a newly developed LGR5-H2B-GFP pig line we have demonstrated its wide utility for studying the role of these unique cells and have now made seminal observations in a variety of tissues/organs. However, we have also been unable to address some key aspects due to the inability of the existing model to analyze the differentiated progeny (lineage tracking) of LGR5 cells during development, after transplantation, or during the process of injury and repair. In addition, while we have been able to inactivate both alleles of the LGR5 gene, we do not have the ability to delete the cell itself. Being able to delete the LGR5 expressing cell will increase our understanding of the importance of this unique cell (versus the LGR5 gene itself) in the phenotypes of interest. Thus, to overcome these two deficiencies, and further increase the value and impact of the original LGR5-H2B-GFP animal model, we propose to develop a highly improved LGR5 pig line that will allow labeling of the LGR5 cell, tracking of its progeny after injury, and delete it when desired. The high concordance between human and pig results to date with respect to LGR5 expression and function, not only increases the value of this line for basic research, but also for rapidly translating clinical findings to humans, further increasing the impact of this unique animal model. Successful completion of the proposed aims will generate a powerful animal model for study of translational aspects of LGR5 stem cells of the gut, lung, skin, liver, cochlea, and kidney among others.