Mol Ther. were not contaminated with adenovirus or rep-positive AAV.40 Another hypothesis is that the particles detected could be wild-type AAV particles following naturally acquired infection of the animals. However, the fact that these particles were systematically observed in the injected retina and not in the noninjected control retina is an important discussion against a naturally acquired illness with wt AAV. A very interesting getting was the presence of AAV vector particles inside and, to a lesser extent, outside of the injected area. In all studies published to day within the transfer of genes into the retina using known AAV serotypes, manifestation of the transgene was usually limited to the injected area. One exclusion was the recent getting by our group that AAV serotype 8 allows the manifestation of transgenes outside of the injected area in the retina of rats and dogs.41 The effects of the present study suggest that even though transgene expression following subretinal injection of AAV serotypes 2, 4, and 5 is limited to the injected area, some of the vector particles are transported outside of the injection zone. For those treated retinas, vector particles were found in cells that do not display expression PSI-7976 of the transgene. For rAAV2/4 that only transduces the RPE,42 vector particles were recognized in the OPL (P1) (Number 6a). For AAV2/5 (D1) and rAAV2/2 (D2) that display expression of the transgene in both the RPE and the photoreceptors,43 viral particles were found in the RPE, in the OPL and ganglion cell coating for D1 (Number 6b) and in the inner plexiform coating and OPL for D2 (Number 6c). These results suggest that manifestation of the transgene in a specific cell does not depend only on the ability of the vector to enter into this cell but may also be conditioned by the capacity of the vector to be processed from the endocytic pathway and to deliver PSI-7976 the viral genome into the nucleus of the cell. PSI-7976 The presence of AAV5 particles in ganglion cells and AAV2 particles in the inner plexiform coating, which are more distant from your injection site, is definitely somewhat hard to explain. One hypothesis would be a transsynaptic transport from photoreceptors to these cells. Open in a separate window Number 6 Schematic drawing of the retina and depiction of the localization PSI-7976 of the adeno-associated viral (AAV) particles found by electron microscopy. (a) Localization of AAV particles in the retina of P1, after subretinal injection of a rAAV2/4 vector. (b) Localization of AAV particles in the retina of D1 after subretinal injection of an AAV2/5 vector. (c) Localization of AAV particles in the retina of D2 after subretinal injection of an AAV2/2 vector. The reddish noticed circles indicate the location, where particles are found. The PSI-7976 green circles indicate the cells in which transgene expression can be observed. IPL, inner plexiform coating; OPL, outer plexiform coating; RPE, retinal pigment epithelium. It was previously observed that following Rabbit polyclonal to INPP1 intravitreal or subretinal injection of AAV2/2 vectors in dogs or primates, vector DNA was recognized along the visual pathway in the brain.44,45 More recently, we demonstrated that following subretinal injection of an AAV2/8 vector, we observed the expression of GFP in all the cells of the neuroretina and in distal neurons of the lateral geniculate nucleus contralateral to the injected eye.41 All together, these previous observations suggested that AAV particles can travel by axonal.