Elsevier

Neuroscience

Volume 138, Issue 2, 2006, Pages 501-510
Neuroscience

Molecular neuroscience
Adeno-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

https://doi.org/10.1016/j.neuroscience.2005.11.057Get rights and content

Abstract

Adeno-associated virus (AAV) vectors have gained a preeminent position in the field of gene delivery to the normal brain through their ability to achieve extensive transduction of neurons and to mediate long-term gene expression with no apparent toxicity. In adult animals direct infusion of AAV vectors into the brain parenchyma results in highly efficient transduction of target structures. However AAV-mediated global delivery to the adult brain has been an elusive goal. In contrast, widespread global gene delivery has been obtained by i.c.v. injection of AAV1 or AAV2 in neonates. Among the novel AAV serotypes cloned and engineered for production of recombinant vectors, AAV8 has shown a tremendous potential for in vivo gene delivery with nearly complete transduction of many tissues in rodents after intravascular infusion. Here we compare the efficiency of an AAV8 serotyped vector with that of AAV1 and AAV2 serotyped vectors for the extent of gene delivery to the brain after neonatal injection into the lateral ventricles. The vectors all encoded green fluorescent protein (GFP) under control of a hybrid CMV enhancer/chicken beta-actin promoter with AAV2 inverted terminal repeats, but differed from each other with respect to the capsid type. A total of 6.8×1010 genome copies were injected into the lateral ventricles of postnatal day 0 mice. Mice were killed at postnatal day 30 and brains analyzed for distribution of GFP-positive cells. AAV8 proved to be more efficient than AAV1 or AAV2 vectors for gene delivery to all of the structures analyzed, including the cerebral cortex, hippocampus, olfactory bulb, and cerebellum. Moreover the intensity of gene expression, assessed using a microarray reader, was considerably higher for AAV8 in all structures analyzed. In conclusion, the enhanced transduction achieved by AAV8 compared with AAV1 and AAV2 indicates that AAV8 is the superior serotype for gene delivery to the CNS.

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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