PROJECT SUMMARY/ABSTRACT Aging results in impaired lymphatic function in a variety of organ systems. This is important because the lymphatic system has many physiologic functions, and lymphatic abnormalities are implicated in many pathologic conditions that affect the elderly. However, while it is clear that aging impairs lymphatic function, the cellular mechanisms that regulate this response remain largely unknown. In addition, the effects of aging on lymphatic endothelial cell (LEC) response to vascular endothelial growth factor C (VEGFC), downstream pathways involving Pi3k and Akt signaling, and interactions with lymphatic muscle cells have not been analyzed. In previous and preliminary studies, we have found that chronic inflammation in a variety of settings, including obesity, lymphedema, and aging, results in decreased intracellular LEC Pi3K/Akt signaling. This is important since LEC Pi3K/Akt signaling is a key regulator of LEC proliferation, differentiation, and function. These findings are consistent with previous studies demonstrating that abnormalities in Pi3k/Akt signaling also contribute to age-related blood endothelial dysfunction. In this proposal, we aim to test the hypothesis that decreased LEC intracellular Pi3k/Akt signaling plays a causal role in age-related lymphatic dysfunction. Our study is innovative because previous studies have been largely observational and have not analyzed lymphangiogenic signaling in aging. We have also developed novel transgenic mice that enable us to selectively analyze the effects of changes in LEC intracellular Pi3k/Akt signaling, thus avoiding the limitations of previous studies that have relied on the systemic administration of recombinant VEGFC or VEGF-R3 antibodies. Using two aims, we will determine how aging changes LEC Pi3k/Akt signaling in different tissues and how these changes correlate with lymphatic function. These studies will help us determine how LECs respond to VEGFC in aging and how these changes modulate response to inflammation and nitrosative stress. In other studies, we will determine if inhibition of intracellular LEC Pi3k/Akt signaling in young mice can recapitulate the lymphatic phenotype of old mice. Finally, we will determine if increased LEC Pi3k/Akt signaling can improve lymphatic function in elderly mice. At the conclusion of our proposed research, we will have a detailed understanding of the cellular mechanisms that contribute to age-related lymphatic dysfunction. This understanding will provide the basis for future studies designed to improve lymphatic function.