Elucidating Mechanisms Guiding Enterocyte Migration

Cell migration sustains life. Throughout development, proteins regulate cell migration within the embryo to form layers and, eventually, specific organs. Post birth, controlled leukocyte migration ensures response to inflammation and infection in response to antigens. Within an injured tissue, cell migration is critical for healing.;Of particular interest to our group is Necrotizing Enterocolitis, NEC, a disease marked by pronounced inflammation and necrosis of the immature intestine. In this disease, a balance between injury and regeneration is required for health. Yet, mechanistically how this occurs remains incompletely understood. Thus, we explored the signaling mechanism regulating restitution using both in vitro cell culture-based as well as in vivo transgenic animal models. As we have long recognized importance of Platelet Activating Factor Receptor (PAFR) in pathophysiology of NEC, emphasis is paid downstream targets of PAFR, including FAK and PI3K, which, in many in vitro models, are indispensable for cell migration. Thus, we have untangled the mechanism by which activated PAFR inhibits enterocyte migration in a PI3K-AKT dependent manner, using a test medium more suitable for analysis of migration than the conventionally-used DMEM. By showing that upstream of PAFR enterocyte migration is regulated by betaArrestin proteins in an isoform dependent manner, our findings may explain reports of widely varying, and sometimes opposing, effects of PAF on migration in distinct cell lines. Our preliminary data suggests that treatment of enterocytes with TGFbeta1 prevents PAF-induced enterocyte inhibition of migration in a PI3K-independent, though FAK- and calmodulin kinase II (CaMK)-dependent mechanism. Finally, we analyzed the role of PAF receptor in aberrant enterocyte proliferation using murine models, showing that PAFR contributes to increased proliferation in response to stress in cross sections of ileum of wild-type, compared to PAFR knockout pups (as judging by the number of EdU-labeled cells along the crypt-villus axis), and to significant loss of the number of enterocytes in guts from wild-type littermates, underscoring the role of PAFR in apoptosis of cells of intestinal epithelial cell lineage.