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H. Ishikawa; Effect of GDNF Gene Transfer Into Axotomized Retinal Ganglion Cells Using in vivo Electroporation With a Contact Lens–Type Electrode . Invest. Ophthalmol. Vis. Sci. 2006;47(13):846.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal ganglion cells (RGCs) death is a major cause of blindness. Current study have suggested that neuroprotective factors have effects on the survival and regeneration of damaged RGCs. Therefore we newly developed an in vivo electroporation (ELP) method to introduce foreign genes into RGCs and electrointroduced glial cell line–derived neurotrophic factor (GDNF) into axotomized RGCs in adult rats.
After the intravitreous injection of the plasmid gene (20 µg), 5 electric pulses (6 V/cm, 100 ms duration) were each delivered twice with 5 min interval to the rat eye using a contact lens type electrode (cathodal) attached to the cornea and a needle electrode (anodal) inserted to the middle of the forehead. The efficiency of the genetic introduction into RGCs and tissue damage to the eyeball was evaluated using a green fluorescent protein (GFP) gene, TUNEL and histological observation. To confirm whether this method can actually rescue damaged RGCs, GDNF was electrointroduced into RGCs after optic nerve transection.
DiI retrograde labeling revealed that 24.4±4.7% of all RGCs was electrointroduced with the GFP gene. TUNEL and histological analysis showed a few tissue damages in the cornea, lens and retina. After the GDNF electrointroduction into axotomized RGCs, a significant increase in the number of surviving RGCs was observed 2 (47.0±7.4%) and 4 weeks (18.4±1.5%)after the optic nerve transection, and the decrease of caspase 3 and 9 was detected by RT–PCR.
This method may be useful for the delivery of genes into RGCs with simplicity and minimal tissue damage. Furthermore, The advantages of this newly–developed method is that molecules other than DNA, such as small proteins and peptides, can also be delivered into RGCs or other retinal cells by in vivo ELP. Therefore, this method can be applied to drug and gene delivery systems for RGCs disorder and other retinal desease such as retinoblastoma and choroidal neovascularization.
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