A selective method for transfection of retinal ganglion cells byretrograde transfer of antisense oligonucleotides against kynurenineaminotransferase II

[摘要] Purpose: Intravitreal administration of specific antisenseoligonucleotides (ODNs) effectively downregulates gene expression in theretina but does not modulate it exclusively in retinal ganglion cells(RGCs). Expression of kynurenine aminotransferase II (KAT II) in RGCshas been well described in the literature. We describe a new method fordownregulating cellular KAT II expression via transfection of RGC byretrograde transfer of ODN.Methods: Fluorescently labeled, specific ODNs against KAT II wereinjected into rats either intravitreally or into the superior colliculi.Fluorescence microscopy of retinal flat-mounts and radial sections wasused to compare the location, duration, and degree of transfection forboth methods of delivery. The effects of both methods on KAT IIexpression in RGCs were studied immunohistochemically with unlabeledODN. Retinal kynurenic acid (KYNA) contents were measured using highpressure liquid chromatography (HPLC).Results: After intravitreal injection, fluorescently labeled ODNreached all retinal layers, whereas injections into the superiorcolliculus resulted in transfection of the RGC layer alone.Immunohistochemistry showed that both methods of ODN application had asimilar effect on downregulation of KAT II expression in RGC. RetinalKYNA content decreased significantly 4 days after both types of ODNadministration.Conclusions: This study demonstrated that retrograde transfer ofspecific ODN into RGC is feasible and induces downregulation of KAT IIcellular expression. This may become a useful tool for modulating geneexpression in the retinal ganglion cell layer in vivo without directtransfer of ODN to other retinal cell layers.

[学科分类] 生物化学/生物物理