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Dynamic studies of the wound-triggered inflammatory response and associated fibrosis and scarring


Wound repair is an important, highly-ordered process that aims for a rapid closure of the wound and regeneration of injured tissue. The wound-triggered inflammatory response, specifically invading neutrophils and macrophages, has long been considered essen tial for repair. Recently however, perfect scar-less healing has been observed in settings with limited or no inflammatory response. We therefore hypothesize that macrophages contribute to the negative side-effects of tissue repair, such as fibrosis and sc arring, and propose the following objectives to address this important biomedical question. Mice transgenic for GFP-tagged macrophages will be used to establish explant eye wound models in which we will dynamically image the migration of macrophages toward s a mammalian wound site, as little is currently known about the precise timing of recruitment and the underlying mechanisms of cell movement. Three-dimensional time-lapse movies will be generated, and we will quantify the number of macrophages recruited t o and dispersed from the wound, the speed and directionality of their migratory route, and the furthest distance from which they are drawn towards the wound. The second objective is to analyse the local effects of activated macrophages on wound fibroblasts with co-culture experiments. Physical interactions, timing and extent of macrophage actions, and fibroblast gene expression changes will be observed. Finally, the functional role of ¿inflammation-dependent genes¿ will be studied by anti-sense oligonucleot ide knockdown in vitro and at the wound site in vivo. These final experiments will begin to address whether knockdown of inflammation/fibrosis-associated genes has the potential to modulate the fibroblast response and quality of wound repair. By merging th e opportunities for live imaging of the inflammatory response with functional analysis of candidate ¿fibrosis genes¿, we hope to gain insight into the biology of the inflammatory response.

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