Adult stem cells are present in many tissues and replenish the cellular constituents of those tissues throughout life. Recently, evidence has accumulated that at least under some conditions, progeny of adult stem cells can generate cells outside their tissue of origin. In our laboratory, we have examined this so-called stem cell """"""""plasticity"""""""" in the context of generation of blood by stem cells in the muscle, and generation of muscle by stem cells from bone marrow. Stem cells in muscle that generate blood are hematopoietic in origin, and were the focus of the previous funding period. We have also recently shown that the ability of hematopoietic stem cells (HSC) to generate muscle is most likely due to the direct incorporation of myeloid cells into regenerating muscle. Here, using liver regeneration as a model, we propose to examine the mechanisms of this non-autochthonous cell incorporation in much more detail, with the long-term view of improving the efficiency for its therapeutic use. In the first Aim, we will definitively identify the major hematopoietic cell types capable of participating in generation of hepatocytes. We will utilize cre-lox lineage marking strategies as well as cell transplantation into a genetically deficient liver model to assess hematohepatic conversion. We will also determine whether there is any evidence for a non-fusion hepatic regeneration mechanism in this model using multiple donor and recipient markers. In the second Aim, we will examine whether hemato-hepatic conversion can occur in the context of two distinct models of liver regeneration through either fusion-dependent or -independent mechanisms. This will reveal the applicability of this therapeutic strategy to different types of liver disease and modes of regeneration. Finally, in the third Aim, we will study the molecular requirements for hemato-hepatic conversion by a) Inducing fusion when cells are given in the absence of bone marrow transplantation, b) Increasing the efficiency of fusion using inducible viral fusion proteins, and c) Studying the nuclear reprogramming events occurring upon fusion using BAC-transgenics expressing tagged tissue-specific proteins. Together, these studies will improve our understanding of the factors that govern heterotypic cell fusion as well as the steps in the reprogramming of nuclei to alter cell identity. These insights will be essential for moving towards therapeutic applications of stem cell-based therapies.
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