Molecular neuroscienceAdeno-associated virus vectors serotyped with AAV8 capsid are more efficient than AAV-1 or -2 serotypes for widespread gene delivery to the neonatal mouse brain
Section snippets
Cell culture
The 293T human kidney cell line was obtained from Michele P. Calos (Department of Genetics, Stanford University School of Medicine, Stanford, CA, USA), and grown in Dulbecco’s Modified Eagle Medium (DMEM; Life Technologies/Invitrogen, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (Sigma, St. Louis, MO, USA and 100 units/ml penicillin and 100 μg/ml streptomycin (Life Technologies/Invitrogen) at 37°C in a 5% CO2 humidified atmosphere. Transfections of 293T cells for AAV vector
Results
In the present study, we compared the efficiency of AAV vectors pseudotyped with AAV8, AAV1 or AAV2 capsids for gene transfer to the mouse brain. The packaged genome was the same for all vectors, encoding GFP under control of a hybrid cytomegalovirus enhancer/chicken beta-actin promoter and carrying AAV2 inverted terminal repeats. At the day of birth, a total of 6.8×1010 genome copies were injected into both lateral ventricles of each mouse brain (n≥5 per vector). Mice were killed at postnatal
Discussion
The three AAV vectors used in this study carried the same genome with AAV2 ITR elements flanking the GFP expression cassette. Therefore it is reasonable to conclude that the differences between vectors in distribution, number of transduced cells, and intensity of gene expression in the brain are a direct result of different capsid properties. The distribution of transduced cells is most likely the result of different receptor tropism for each capsid. The receptor and co-receptors for AAV2 (Qing
Conclusion
In conclusion, AAV8 serotyped vectors can achieve widespread gene delivery to the mouse brain after neonatal i.c.v. injection. AAV8 transduced the cerebral cortex, hippocampus, olfactory bulb, and cerebellum much more efficiently than AAV1 or AAV2. Moreover the intensity of gene expression, as assessed here by a new method using a microarray reader, was considerably higher for AAV8 in all structures analyzed.
Acknowledgments
Supported by NIH AG08487 (B.T.H.), a Pioneer grant from the Alzheimer’s Association (B.T.H.), and a Fulbright Scholarship (M.L.D.B.). We would like to thank Igor Bagayev from the Confocal Microscopy Facility of the Neuroscience Center at Massachusetts General Hospital for his help. We would also like to thank Xandra Breakefield for helpful discussions during the course of this work and for her critical reading of the manuscript.
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