Abstract
Purpose:
Our goal is to investigate the neuroprotective effects of systemic recombinant adeno-associated virus (rAAV) gene delivery of forms of erythropoietin (Epo) that have attenuated erythropoietic activity to provide long-term, sustained treatment in an induced mouse model of glaucoma.
Methods:
C57BL/6 mice were injected intramuscularly with 10^9 vgc rAAV2/8 delivering eGFP or Epo mutants R76E, R76E+S100E, or R76E+R103E. To induce ocular hypertension, anterior chambers were injected with 15-µm diameter microbeads or saline 1 month after rAAV administration. Elevation of intraocular pressure (IOP) was verified by tonometry. Baseline and endpoint visual function was assessed by flash visually evoked potential (fVEP) immediately prior and 1 month following induction. At 1 month post-induction, tissue was collected, and retinal ganglion cell (RGC) axons were evaluated by optic nerve histology and fluorescent cholera toxin B anterograde transport. Blood hematocrit was also assessed at collection.
Results:
Hematocrit was significantly elevated (86%) in mice treated with rAAV.EpoR76E but not elevated in mice treated with rAAV.EpoR76E+S100E (44%) or rAAV.EpoR76E+R103E (44%). Mean IOP of 18.7±2.8 mmHg was observed in microbead-injected eyes 1 week post-induction compared to 14.4±0.9 mmHg in saline controls. At 1 month post-induction, microbead-injected rAAV.eGFP controls developed 60% reduction in endpoint fVEP P1 amplitude from baseline, 30% reduction in RGC axon density, and impairment of RGC anterograde transport. Microbead-injected mice treated with rAAV.EpoR76E showed no significant decrease in endpoint fVEP P1 amplitude from baseline. RGC axon density in microbead-injected mice treated with rAAV.EpoR76E+S100E and rAAV.EpoR76E+R103E improved 25% and 20%, respectively, compared to rAAV.eGFP-treated mice. Microbead-injected mice treated with rAAV.EpoR76E showed improvement in RGC anterograde transport compared to rAAV.eGFP controls.
Conclusions:
Gene delivery of non-hematopoietic modified Epo appears to attenuate glaucomatous changes in the induced model of ocular hypertension, indicating that neuroprotection by Epo is not specific to the DBA/2J model of glaucoma.
Keywords: 538 gene transfer/gene therapy •
615 neuroprotection •
568 intraocular pressure