Adeno-associated virus-mediated expression of growth-associated protein-43 aggravates retinal ganglion cell death in experimental chronic glaucomatous injury

PURPOSE To examine whether adeno-associated virus (AAV) vector-mediated overexpression of growth-associated protein-43 (GAP-43) has protective or deleterious effects on retinal ganglion cell (RGC) survival in laser-induced chronic intraocular pressure (IOP) elevation injury. METHODS Adult Fischer 344 rats received unilateral intravitreal injection of either normal saline, AAV-green fluorescent protein (AAV-GFP), or a bicistronic AAV vector encoding GAP-43 and GFP (AAV-GAP-43). Two weeks later, experimental chronic glaucoma was induced in the injected eyes by scarring the trabecular meshwork with a diode laser. IOP was measured with an impact (rebound) tonometer. Survival of RGCs was estimated after 3 weeks of IOP elevation by quantifying β-III tubulin⁺ neurons in retinal whole mounts. The transfection efficiency of target genes was assessed with direct view of GFP and western blot analysis of GAP-43. RESULTS Quantification of β-III tubulin⁺ immunostaining revealed that, compared to uninjured eyes (1,172±80 cells/mm²), 3 weeks of laser-induced IOP elevation led to a 60% decline in RGC survival (496±136 cells/mm²). Transfection with control vector AAV-GFP by itself did not have a significant effect on RGC viability (468±124 cells/mm²). Overexpression of GAP-43 in RGC cell bodies and axons via bicistronic AAV-GAP-43 led to more severe RGC death (260±112 cells/mm²) in IOP elevated eyes, an 80% loss of the total RGC population. CONCLUSIONS Overexpression of GAP-43 aggravated RGC death in experimental chronic IOP elevation injury. GAP-43 was upregulated in RGCs regenerating after optic nerve injury. Thus, the finding that this same protein is deleterious to RGC viability after chronic IOP elevation may aid in understanding the mechanisms involved in RGC loss in glaucoma and how best to treat this condition.