Differential transgene expression profiles in rat brain, using rAAV2/1 vectors with tetracycline-inducible and cytomegalovirus promoters.
The biodistribution of transgene expression in the CNS after localized stereotaxic vector delivery is an important issue for the safety of gene therapy for neurological diseases. The cellular specificity of transgene expression from rAAV2/1 vectors (recombinant adeno-associated viral vectors pseudotyped with viral capsids from serotype 1) using the tetracycline-inducible (TetON) expression cassette in comparison with the cytomegalovirus (CMV) promoter was investigated in the rat nigrostriatal pathway. After intrastriatal injection, although green fluorescent protein (GFP) was expressed mainly in neurons with both vectors, the relative proportions of DARPP-32-positive projection neurons and parvalbumin-positive interneurons were, respectively, 13:1 and 2:1 for the CMV and TetON vectors. DARP32-positive neurons projecting to the globus pallidus were strongly GFP positive with both vectors, whereas those projecting to the substantia nigra pars reticulata (SNpr) were efficiently labeled by the CMV vector but poorly by the TetON vector. Numerous GFP-positive cells were evidenced in the subventricular zone with both vectors. However, in the olfactory bulb (OB), GFP-positive neurons were observed with the CMV vector but not the TetON vector. We conclude that the absence of significant amounts of transgene product in distant regions (SN and OB) constitutes a safety advantage of the AAV2/1-TetON vector for striatal gene therapy. Midbrain injections resulted in selective GFP expression in tyrosine hydroxylase-positive neurons by the TetON vector whereas with the CMV vector, GFP-positive cells covered a widespread area of the midbrain. The biodistribution of GFP protein corresponded to that of the transcripts and not of the viral genomes. We conclude that the rAAV2/1-TetON vector constitutes an interesting tool for specific transgene expression in midbrain dopaminergic neurons.
Bockstael
Olivier
O
Multidisciplinary Research Institute (IRIBHM), Université Libre de Bruxelles, Hôpital Erasme, Brussels, Belgium.
Chtarto
Abdelwahed
A
Wakkinen
Janica
J
Yang
Xin
X
Melas
Catherine
C
Levivier
Marc
M
Brotchi
Jacques
J
Tenenbaum
Liliane
L
D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D003587:Cytomegalovirus / Q000235:genetics*; D000229:Dependovirus / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005455:Fluorescent Antibody Technique; D005786:Gene Expression Regulation / Q000502:physiology*; D015316:Genetic Therapy; D005822:Genetic Vectors; D051100:Glial Cell Line-Derived Neurotrophic Factor / Q000235:genetics / Q000378:metabolism; D007124:Immunoenzyme Techniques; D009474:Neurons / Q000187:drug effects / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013378:Substantia Nigra / Q000187:drug effects / Q000378:metabolism; D013752:Tetracycline / Q000494:pharmacology*; D014162:Transfection; D019076:Transgenes
D051100:Glial Cell Line-Derived Neurotrophic Factor; D011500:Protein Synthesis Inhibitors; D012333:RNA, Messenger; D013752:Tetracycline