PROJECT SUMMARY / ABSTRACT Lymphedema is a chronic and debilitating condition currently without approved medical therapies. Lymphatic drainage insufficiency, caused by inflammation and lymphatic endothelial cell (LEC) dysfunction, is the main factor promoting the progression of lymphedema. LEC-derived hypoxia-inducible factor (HIF)-2α is required for proper lymphatic development and homeostasis; the decline of LEC HIF-2α, observed in lymphedema, may be directly involved with disease progression. In experimental models, reducing HIF-2α expression aggravates lymphatic drainage insufficiency and exacerbates accumulating interstitial fluid. Anti-inflammatory ketoprofen therapy was recently shown to be effective in lymphedema, and treated patients show increased LEC HIF-2α expression in their skin biopsies. The studies proposed in this grant are designed to address fundamental questions about why LEC HIF-2α is decreased in lymphedema and how this reduction influences lymphatic drainage. The grant hypothesis is that suppressed LEC HIF-2α expression, caused by inflammatory mediators, impairs lymphatic drainage by promoting LEC phenotypic transformation and abnormal cellular energetics. Specific Aim 1 is to explore the mechanisms by which inflammation inhibits LEC HIF-2α expression and map HIF-2α-regulated gene networks in LECs. Aim 2 will investigate how LEC HIF-2α reduction may cause LEC phenotypic transformation, promote lymphatic vascular remodeling, and aggravate lymphatic drainage insufficiency. Aim 3 will study how LEC HIF-2α reduction affects lymphatic structure and function by promoting abnormal cellular metabolism. Collectively, the proposed studies are expected to advance the understanding of the role that HIF-2α plays in lymphedema with the rationale that the knowledge gained can facilitate much- needed therapeutic development.