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In Vivo Delivery of Phosphorothioate Oligonucleotides Into Murine Retina
Masanori Hangai, MD, PhD;
Hidenobu Tanihara, MD, PhD;
Yoshihito Honda, MD, PhD;
Yasufumi Kaneda, MD, PhD
Arch Ophthalmol. 1998;116:342-348.
Objectives To determine the fate of phosphorothioate oligonucleotides (S-ODNs), which are commonly used for antisense strategy, in murine retina in vivo with the use of fluorescein isothiocyanate (FITC)–labeled S-ODNs, and to evaluate our fusogenic liposome system that may facilitate the delivery of S-ODNs.
Methods The FITC-labeled S-ODNs were encapsulated in liposomes, which were then coated with the envelope of inactivated hemagglutinating virus of Japan (HVJ; Sendai virus) by fusion (HVJ liposomes). Intravitreal injection of naked FITC-labeled S-ODNs or of the HVJ liposomes was done in ICR mice. After fixation, cryosections and flat-mounted retinas were prepared and examined by fluorescence microscopy.
Results Injection of naked FITC-labeled S-ODNs at 3 µmol/L exhibited weak fluorescence in 13% of the cells in the ganglion cell layer. When the concentration was increased to 30 µmol/L, high fluorescence was seen in 59% of cells in the ganglion cell layer at this time. This fluorescence diminished within a day. In contrast, injection of HVJ liposomes containing FITC-labeled S-ODNs at 3 µmol/L resulted in high fluorescence in 44% of the cells in the ganglion cell layer at 1 hour, and this fluorescence lasted for up to 3 days. This treatment also resulted in high fluorescence within retinal vessel walls, and weak fluorescence in photoreceptor cells.
Conclusions Intravitreally injected S-ODNs were rapidly eliminated from neural retina, and the use of HVJ liposomes could improve the delivery of S-ODNs. This method may be a potentially useful system for the antisense-based therapies for retinal diseases.
From the Department of Ophthalmology and Visual Science, Graduate School of Medicine, Kyoto University, Kyoto, Japan (Drs Hangai, Tanihara, and Honda), and Institute for Molecular and Cellular Biology, Osaka University, Osaka, Japan (Dr Kaneda). Dr Hangai is now with the Doheny Eye Institute, University of Southern California School of Medicine, Los Angeles.
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