ABSTRACT Iron is an essential bio-metal, but requires tight regulation to minimize the generation of iron-catalyzed reactive oxygen species (ROS). This is accomplished by hepcidin, a mainly hepatic peptide hormone that regulates serum transferrin iron (Tf-iron) by downregulating ferroportin (Fpn), an iron export protein. The synthesis of local hepcidin in the ciliary epithelial, corneal endothelial, and trabecular meshwork (TM) cells suggests autonomous regulation of iron in the anterior segment, independent of the retina. However, hepcidin is upregulated by cytokines as well, the latter signal superseding the signal from Tf-iron. This raises the concern of cytokine- mediated upregulation of hepcidin, creating a toxic environment by increasing intracellular iron and ROS, known contributors of toxicity in primary open angle glaucoma (POAG) and other ocular conditions. Pertinent to this application is TGFβ2, a cytokine implicated in POAG. Data from my laboratory show that biologically active TGFβ2 upregulates hepcidin in primary human TM cells and ex-vivo human organ culture perfusion model of POAG. Remarkably, hepcidin upregulates full-length (FL) and biologically active TGFβ2, the latter probably through ROS, forming a self-sustained feed-forward loop that is disrupted by heparin, a hepcidin antagonist, and N-acetyl carnosine, an antioxidant. Based on these observations, we hypothesize that the anterior segment maintains autonomous regulation of iron by locally synthesized hepcidin, and cytokine-mediated upregulation of local hepcidin forms a self-sustained feed-forward loop that is disrupted by hepcidin antagonists and Fpn stabilizing agents. This hypothesis will be tested in three specific aims. In Aim 1,autonomous regulation of iron by the anterior segment will be explored in wild-type (wt) C57BL/6 and Balb/c mice at steady state and after systemic iron overload that is known to cause retinal iron accumulation. Transport of serum iron across the blood- aqueous barrier, and from the retina to the aqueous humor (AH) will be determinedin the absence or presence of exogenous synthetic hepcidin in the AH. The role of local hepcidin in corneal endothelial cells in regulating iron exchange between the AH and the cornea will be determined in the human corneal cup ex-vivo model. In Aim 2, levels of FL and bioactive TGFβ2 will be determined in hepcidin knock-out (hepc-/-) and littermate hepc+/+ controls at steady state, and after over-expressing FL TGFβ2 to evaluate whether absence of hepcidin and iron- mediated ROS decreases bioactive TGFβ2 and IOP . In addition, levels of hepcidin, Tf-iron, and TGFβ2 will be determined in the human AH of POAG cases and cataract controls collected at surgery. In Aim 3, ex-vivo human anterior segment organ culture perfusion model will be used to evaluate the efficacy of FDA approved Fpn stabilizing agent fursultiamine in reducing intracellular iron, ROS, and bioactive TGFβ2-mediated increase in IOP . Successful completi...