PROJECT DESCRIPTION During the progression of glaucoma, the retinal ganglion cells (RGC) and their axons degenerate. An important target to this damage occurs in the optic nerve head (ONH), where the RGC axons leave the globe to form the optic nerve. Although RGC axonal damage can be caused by different type of insults, it is well- established that elevated intraocular pressure (IOP) and stiffness in the peripapillary region (ppSC) are major contributors to this degeneration. It would seem logical to think that some kind of molecular regulation coordinating the anterior and posterior affected tissues would be of great benefit for a potential treatment of glaucoma. Previously, we had identified Matrix Gla (MGP) as one of most highly expressed genes in the human TM. We had also found that MGP was altered in the TM by elevated IOP, TGFβ and dexamethasone, and that calcification markers were increased in TM tissues from glaucoma patients and Mgp-KO mice. The gene transfer of a calcification inducer (BMP2) to the rat’s TM also elicited elevated IOP. Matrix Gla is a potent mineralization inhibitor secreted by cartilage chondrocytes and arteries’ vascular smooth muscle cells. Mgp KO mice die at 5-6 weeks due to massive arterial calcification. Arterial calcification results in arterial stiffness and higher systolic blood pressure. In order to investigate the abundance of Mgp in the eye and its contribution to a potential regulation of stiffness in glaucoma in a living animal, we used mouse genetics. To determine the Mgp spatial/ temporal expression in the eye, we generated an Mgp-Cre Knock-in (KI) mouse, containing Mgp DNA fused to an IRES-Cre-cassette. Crosses of this mouse with R26R-floxed reporters (lacZ and td.Tomato) revealed, as expected, Mgp’s high specific expression in the TM region, but also, and surprisingly, Mgp was highly and specifically expressed in the sclera, in particularly the ppSC. Based on these findings, we propose that MGP and its anti-calcification/ anti-stiffness function represents a sole mechanism that affects the source of two basic glaucomatous causes, elevated IOP and ONH damage. Thus, we hypothesize that MGP is a master key-mediator that prevents the occurrence of calcification/ stiffness in the targeted eye tissues, and as a consequence, controls the development and progression of glaucoma. To develop and prove this hypothesis, we propose to investigate the response of Mgp to glaucomatous insults in vivo using the newly generated Mgp-Cre-reporter mice (SA#1), to override the early death of the Mgp KO by creating TM and ppSC specific conditional Knock-outs (cKOs) (SA#2) and to evaluate the impact of the specific ablations on glaucoma phenotypes (SA#3 Results to be obtained with the execution of this proposal will provide the mechanistic understanding and the knowledge needed to develop a combined TM-ppSC therapy which could potentially lead to a totally new treatment of glaucoma.