Abstract
Abstract: :
Purpose: In neonatal oxygen induced retinopathy (OIR) mouse model of retinal neovascularization (NV), AAV–K1K3(angiostatin) is reported to significantly reduce proliferating retinal vessels and increase levels of PEDF. We aimed to determine whether therapeutic effects of K1K3 are realized through induction of PEDF and whether appropriately timed injections of PEDF protein would be effective in the OIR model. Methods: AAV vectors expressing K1K3 were injected into one eye of postnatal day 0 (P0) mouse pups. Protein levels for K1K3 expression and subsequent up–regulation of PEDF were determined by ELISA daily for the 17–day period of this OIR model. Purified PEDF protein was also injected into one eye of P7 mouse pups just prior to hyperoxic exposure. FITC dextran angiography was used to follow the vaso–obliteration, vaso–proliferation effects of PEDF injection or K1K3 mediated upregulation of PEDF. Results: Protein levels measured by ELISA showed expression of K1K3 (15–113 ng/eye) and subsequent up–regulation of endogenous PEDF (3–15 ng/eye) are detectable as early as 1 day after vector injection and both persist at therapeutic levels for the 17–days in mice housed in normoxic conditions. When the K1K3 vector injected animals are exposed to hyperoxia in the OIR protocol, PEDF reaches levels of up to 32 ng/eye. This is comparable to PEDF levels obtained from a AAV–PEDF vector injection at P1. In untreated normoxic control eyes, endogenous expression of PEDF is measurable, but at levels lower than 1 ng/eye. In neonatal mice exposed to the 7–day period of hyperoxia in the OIR model, levels of PEDF increase to 1–2.5 ng/eye. Injection of PEDF protein at 100 ng/eye in P7 mice is measurable by ELISA at day 1 post–injection at levels of 3–15 ng/eye and leads to a reduction in vaso–proliferation at P17 similar to injection with the K1K3 vector. Conclusions: AAV–K1K3 vector may limit pathogenic retinal NV through induction of ocular PEDF levels above those normally seen in untreated eyes of OIR mice. This normal increase of PEDF during hyperoxia in the OIR protocol may trigger the observed vaso–obliteration that establishes the topographic ischemia and drives retinal NV. Data generated in this study indicate that PEDF levels must be elevated either before or during the period of hyperoxia to control retinal NV in the neonatal OIR mouse.
Keywords: retinopathy of prematurity • diabetic retinopathy • gene transfer/gene therapy