Endothelial cell-fibroblast communication in skin wound healing

PROJECT SUMMARY This application proposes a customized research training plan designed to promote the development of the applicant into an independent investigator. The plan includes advanced training in laboratory experimentation, along with tailored professional and career development opportunities. The training plan is supported by the outstanding availability of local and institutional resources at UIC. The proposed research will investigate the role of extracellular vesicles in mediating endothelial cell-fibroblast communication in the healing wound. In human skin, the process of wound repair involves angiogenesis, which includes the creation and then pruning of vessels. Simultaneously, fibroblasts in the wound are responsible for the deposition of the extracellular matrix components that surround the new vasculature. The goal of the proposed studies is to examine how endothelial cells communicate with surrounding fibroblasts in the wound. Several previous studies demonstrate that these two cell types may utilize paracrine methods of communication to influence or enhance their functional roles, but how this intercellular interaction occurs is largely unknown. One potential mechanism by which endothelial cells might communicate with fibroblasts is through extracellular vesicles and their microRNA cargo. Recent studies in our lab have shown that fibroblasts treated with extracellular vesicles from endothelial cells exhibit significant transcriptomic changes. Based on this preliminary data, our central hypothesis is that in the healing wound, endothelial cells secrete extracellular vesicles that influence the function of resident fibroblasts. The research plan utilizes methods for extracellular vesicle purification and application, traditional in vitro and in vivo wound healing assays, and advanced bioinformatics techniques to study extracellular vesicle-mediated communication between these two cells types. Aim 1 will determine how fibroblast phenotypes are altered by endothelial cell extracellular vesicle treatment, and will examine the role of cargo microRNAs in the observed changes. In vitro studies will elucidate the effect of endothelial cell extracellular vesicle treatment on the fibroblast transcriptome and key fibroblast functions, such as collagen contraction, deposition, and proliferation. Aim 2 will evaluate how endothelial cell extracellular vesicles affect fibroblast phenotypes in the wound in vivo using a splinted murine skin wound model. The effect of extracellular vesicle treatment will be examined for multiple parameters of healing, including collagen content and architecture, myofibroblast activation, angiogenesis, and wound closure. Finally, single cell-RNA sequencing of wound fibroblasts will be used to determine how endothelial cell extracellular vesicle uptake modifies the fibroblast transcriptional profile. Together, the aims will lead to a better understanding of the mechanisms by which endothelial cells might modulate fibroblast function, and may lead to the development of novel therapies to treat wounds or various skin and fibrotic diseases.