Glaucoma is the second leading cause of blindness worldwide, and affects over 2 million people in the United States. Although elevated intraocular pressure (IOP) is a major risk factor for glaucoma, little is known about: (1) how alterations in IOP change tissue function within the optic nerve head (ONH), the most likely site of initial glaucomatous optic nerve damage, and (2) the specific cellular nature of these changes and the mechanisms by which they cause irreversible nerve injury. Our studies indicate that elevated IOP produces extensive changes in gene expression within the ONH. At least some of these changes are likely to contribute to early axonal injury, and can be termed lOP-injury gene changes. This proposal is based on the hypothesis that alterations in IOP cause changes in ONH scleral stress, which are sensed as cellular stretch by astrocytes in the ONH, and this in turn alters the expression of these genes. Because astrocytes are intimately connected to both the ONH connective tissues and axon bundles, these gene changes signal alterations in astrocyte function that either compromise their ability to support axons or induce them to injure axons directly. Specifically, this proposal will determine that: (1) lOP-injury gene changes occur within the unmyelinated portion of the ONH at the level of the sclera, and (2) these changes can be reproduced and their signaling pathways determined in cultured astrocytes exposed to physical stretch.
Two specific aims are proposed. (SA1) Altered gene pathways in the unmyelinated ONH will be identified by microarray analysis and quantitative real-time reverse transcriptase polymerase chain reaction using a rat model of focal nerve injury due to mild chronic IOP elevation. Then the expressed lOP-injury proteins will be localized by immunohistochemistry. By comparing this data with that from other experimental groups, we will isolate IOP injury gene expression from expression changes caused by secondary degeneration and physiological IOP fluctuation. (SA2) An in vitro model of ONH response to elevated IOP using astrocytes grown on distensible membranes and exposed to conditions of cellular stretch will be developed to reproduce the changes in IOP injury gene and protein expression identified in SA1. This culture model will be used to identify the signaling pathways by which stretch regulates the expression of the lOP-injury genes, with the ultimate goal of controlling their expression in vivo and minimizing axonal injury due to elevated IOP.
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