Estrogen (E2) treatment is known to be neuroprotectant in spinal cord injury (SCI). This hormone is highly pleiotropic and has been shown to decrease apoptosis, modulate calcium signaling, regulate growth factor expression, act as an anti-inflammatory, and can drive angiogenesis. These beneficial effects were found at the low dose of 10μg/kg E2 (multiple i.v. injections; previous grant), which helped in obtaining IND approval for taking this into a small clinical safety trial. However, the dose remains non-physiologic and thereby poses a safety hurdle for clinical use. The emergence of smart drug delivery techniques, such as nanoparticles, may allow for increased drug safety and improved efficacy. Thus, the goal of this study is to investigate the effects of nanoparticle delivery of lower doses of E2 (2.5-5.0µg, single dose) that may avoid the high systemic exposures seen with traditional dose routes (i.v. or i.p.) and allow for enhanced protective and reparative effects on lesioned tissue. Preliminary data show that a single administration of rapid release formulated PLGA-PEG nanoparticles loaded with E2 can focally deliver E2 to the contused spinal cord with reduced plasma concentrations when compared with i.v. dosing and can drive estrogenic changes. Additionally, this approach has shown that a single 5.0µg dose of nanoparticle E2 (N-E2) can improve locomotor function recovery. Pilot data suggest that E2 supports the survival of oligodendrocyte precursor cells (OPC) when exposed to toxic factors from Th1 cells and microglia in vitro. Thus, using a cocktail of rapid and slow release nanoparticle formulations, an entirely novel approach, enhanced efficacy may be achieved after SCI. This approach may allow for rapid translation of this known neuroprotectant into clinical trials. We hypothesize that focal delivery of estrogen via nanoparticles will minimize plasma exposure and increase tissue concentrations thereby allowing for maximized therapeutic potential in SCI. To test the hypothesis, three specific aims are proposed: (1) Examine N-E2 release profile and estrogenic effects on inflammation, gliosis, and neuronal protection in acute SCI; (2) Determine the mechanisms of E2-driven protective effects on epithelial, glial and neuronal cells following slow release N-E2 in acute and chronic SCI; and (3) Investigate the potential additive effects of rapid and slow release N-E2 on reduced glial scarring, enhanced regeneration, and improved locomotor function in chronic SCI. Completion of this study will identify the optimal dosing strategy of E2 loaded nanoparticles for delivery of this therapeutic agent in SCI. Additionally, as novel targets of E2 signaling are explored, progress in understanding the mechanisms of estrogenic driven neuroprotection will be made. These data should provide sufficient evidence to support the translation of E2 into clinical trials, with the ultimate goal of providing a safe and effective therapeutic to treat both veterans...