Project Summary Glaucoma is a leading cause of irreversible blindness throughout the world and the second leading cause of blindness overall in the USA. Elevated intraocular pressure (IOP) and aging are the most important risk factors for most forms of glaucoma. IOP level is highly dependent on the rate at which the aqueous humor is filtered through the conventional outflow pathway containing the trabecular meshwork (TM). Reduced cellularity within the TM and abnormal extracellular matrix (ECM) turnover occur in glaucomatous conditions and correlate with increased outflow resistance, elevated IOP, and subsequent vision loss. The goal of this project is to define the mechanisms of stem cell homing and engrafting to the TM tissue, activating regeneration of the TM tissue, and hence restoring outflow facility, reducing IOP, and preventing vision loss. In our previous funding period, we have identified the mechanisms of stem cell homing and integration are partially associated with CXCR4/SDF1 chemokine pair and α5β1 integrin. We have also confirmed that TM stem cells (TMSCs), after intracamerally injection, can regenerate the TM tissue, reduce IOP, and preserve the retinal ganglion cell function in a mouse glaucoma model. This project is designed to test specific hypotheses about the mechanisms by which human TMSCs remodel the pathological TM tissue and restore the TM function. Specific Aim 1 tests the hypothesis that TMSCs and differentiated TM cells remodel the abnormal TM ECM via the COX2/PGE2/MMP pathway. We will utilize myocilin mutant TM cells and dexamethasone-treated TM cells as well as a mouse glaucoma model with myocilin mutation to test how TMSCs promote the ECM turnover and modify the TM segmental outflow pattern. Specific Aim 2 tests the hypothesis that transplanted TMSCs can promote endogenous TMSC activation, migration, and function via the SOX21/WNT signaling. We will unveil if the endogenous TMSCs are viable with a reduced number in aged and glaucomatous TM tissue in human and in mice and uncover how TMSCs awake endogenous TMSCs via SOX21/WNT signaling. The scientific impact of this study will be the elucidation of the cellular and molecular mechanisms of TM regeneration potential by stem cells. The results may also directly lead to the design of stem cell-based therapies or adjunctive treatments that prevent blindness from glaucoma clinically.