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AM Timmers, TB Nguyen, IA Elder, DR Saban; Lentiviral Delivery of DH-sCNTF to RPE in P23H Rats Rescues Photoreceptor Cells Morphologically but Not Functionally . Invest. Ophthalmol. Vis. Sci. 2002;43(13):2732.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Intraocular injection of ciliary neurotrophic factor (CNTF) transiently promotes survival of photoreceptors exposed to various types of insults. In this study, we tested whether long-term expression of a secretable form of CNTF by RPE cells in a rat model of retinitis pigmentosa (RP) could halt progression of photoreceptor cell degeneration. Methods: Four weeks post-natal, rats heterozygous for the P23H mutation in rhodopsin, were subretinally injected in the right eye with 5 * 106 particles of infectious lentivirus (LTV). The lentivirus carried a secretory form of CNTF with a stabilizing two amino acid base change (DH-sCNTF) under the control of the ubiquitous promoter, elongation factor-1α. CNTF expression was confirmed with RT-PCR. Retinal function of both eyes was assessed simultaneously using full-field scotopic ERG. Dark adapted rats were exposed to white light flashes with increasing intensity (-4.4 to 0.4 log cd.s.m-2). The ratio of A-wave and B-wave amplitudes of injected eyes over non-injected eyes was determined for evaluation of functional rescue. Photoreceptor cell number was determined by counting ONL nuclei in 15 µm cryostat sections of the eyecups stained with DAPI. Results: Full-field scotopic ERG analysis of P23H rat eyes injected subretinally with PBS showed that the injection itself does not accelerate retinal degeneration. Furthermore, the reduction of ERG signals correlates with the reduction of ONL thickness and the progression of retinal degeneration. Eyes injected with LTV-DH-sCNTF maintained the columnar ONL thickness at the time of injection while the columnar ONL thickness in non-injected contralateral eyes continued to decrease. A-waves and B-waves were affected differently by DH-sCNTF expression in RPE cells. The A-wave amplitudes in treated eyes and non-treated contralateral eyes were similar. In contrast, the B-wave amplitude in LTV-DH-sCNTF treated eyes decreased 50 to 60 % relative to non-treated contralateral eyes. This preferential loss of B-wave amplitudes is also observed in normal rat eyes injected with LTV-DH-sCNTF virus. Conclusion: The preservation of the ONL thickness at the time of injection indicates that LTV-DH-sCNTF treatment provides an immediate onset of photoreceptor cell rescue. Photoreceptor cell rescue was maintained for at least 2 months post-injection (the latest analysis at the time of submission of this abstract). ERG analysis revealed no effect on A-wave amplitudes but a strong reduction of B-wave amplitudes in P23H and normal rats.
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