Adeno-associated viral vector (AAV)-mediated gene transfer in the red nucleus of the adult rat brain: Comparative analysis of the transduction properties of seven AAV serotypes and lentiviral vectors
Introduction
Adeno-associated viral vectors (AAVs) have evolved into one of the most attractive gene delivery systems for various tissues amenable for gene therapy. AAV vectors are of special interest for transduction of the central nervous system because of their ability to efficiently transduce neurons while inducing minimal immune responses in the host brain (Kaplitt et al., 1994, Kaplitt et al., 2007, McCown et al., 1996, Klein et al., 2002, Klein et al., 2008, Peel and Klein, 2000, Ruitenberg et al., 2004, Tenenbaum et al., 2004, Cearley and Wolfe, 2006, Cearley and Wolfe, 2007, Taymans et al., 2007, Tuszynski, 2007, Cearley et al., 2008, Lawlor et al., 2009). To date, the most widely used AAV vector has been AAV serotype 2.
One strategy to change tropism of the AAV vector entails cross packaging of the AAV2 genome in capsids of other AAV serotypes (Grimm et al., 2003, Burger et al., 2004). Different AAV serotypes transduced muscle cells of mice with differential efficiency (Louboutin et al., 2005, Zincarelli et al., 2008). The efficiencies of transduction in skeletal muscle with AAV7 and AAV8 were similar to AAV1 and higher than that seen with AAV2 and AAV5 (Louboutin et al., 2005, Hasbrouck and High, 2008). Various AAV serotypes have also been shown to efficiently and differentially transduce neuronal cells in the brain. For instance, AAV1, AAV5 and AAV8 are able to transduce neuronal cells in the hippocampus, substantia nigra and striatum (Kaplitt et al., 1994, Klein et al., 2002, Klein et al., 2008, Peel and Klein, 2000, Burger et al., 2004, Broekman et al., 2006). Non-neuronal transduction was observed when other serotypes were used, such as rh24 (Lawlor et al., 2009), whereas, hu.32, hu.11, pi.2, hu.48R3, and rh.8 resulted in preferential green fluorescent protein (GFP) expression in astrocytes or oligodendrocytes in the rodent brain (Cearley et al., 2008).
So far, the tropism and transduction properties of AAV serotypes for brain structures that have projections through the spinal cord (e.g. the red nucleus or corticospinal tract) has not been systematically evaluated, even though long lasting transduction of the red nucleus using AAV2 (Ruitenberg et al., 2002) and the red nucleus and corticospinal neurons with AAV5 vectors (Foust et al., 2008) has been documented. The red nucleus (RN) is an important target for gene therapeutic proof of concept studies because this relatively small nucleus in the brainstem contains the cell bodies of the rubrospinal tract (RST) that runs through the lateral white matter of the spinal cord. The RST is a well-defined spinal nerve tract and injury to the RST results in functional deficits of the fore and hind limbs (Kobayashi et al., 1997, Hendriks et al., 2006). The relatively small size of the nucleus makes this structure an interesting candidate for delivery of potential therapeutic genes (Ruitenberg et al., 2002, Ruitenberg et al., 2004, Blits and Bunge, 2006, Kwon et al., 2007). With this in mind, we compared the tropism, transduction efficiency and temporal transgene expression profile of 7 cross-packaged AAV serotypes and lentiviral vectors (LVs) following a single stereotactic injection into the RN (see Fig. 1). The results will facilitate the design and selection of AAV vectors to target the RN and advance the application of viral vectors for spinal cord injury research.
Section snippets
Production of AAV particles
AAV vectors encoding enhanced green fluorescent protein were all generated using the plasmid pTRCGW (Hermens et al., 1999, Blits et al., 2004). This vector contains a transgene expression cassette that carries the human cytomegalovirus (CMV) promoter to drive transgene expression, the transgenic cDNA encoding the marker protein GFP and a downstream SV40 polyadenylation (polyA) signal. Downstream of this sequence, the vector contained the cis-acting woodchuck posttranscriptional regulatory
Transduction efficiencies of AAV serotypes and LV in the red nucleus
Transduction efficiency was determined at 1 week and 1 month post-injection (Fig. 1). GFP staining of coronal sections of the brain stem revealed different expression patterns for the different serotypes at both time points. One week post-injection, serotypes 1, 2 and 6, as well as LV, all directed transgene expression in the red nucleus. The AAV-transduced cells had a neuronal phenotype based on co-localisation of GFP and NeuN expression. In contrast, LV transduced non-neuronal cell types as
Discussion
This study compared the transduction efficiency, temporal expression profile and cellular tropism of seven AAV serotype vectors and LV after a single stereotactic injection of viral vector in the RN of adult rats. We report differential transduction efficiencies, onset of transgene expression and cellular tropism. These observations will facilitate the design and selection of AAV vectors for gene transfer to the rubrospinal nucleus, an important and widely used target structure for studies on
Acknowledgements
We thank Corrinna Burger for supplying AAV vectors for pilot experiments, Eric Timmermans for technical assistance isolating AAV and Jurgen Kleischmidt and James Wilson for generously providing plasmids for cross packaging. These studies were supported by the International Spinal Research Trust (STR 090), SenterNovem, Amsterdam Molecular Therapeutics and the Netherlands Institute for Neuroscience.
References (56)
- et al.
Rescue and sprouting of motoneurons following ventral root avulsion and reimplantation combined with intraspinal adeno-associated viral vector-mediated expression of glial cell line-derived neurotrophic factor or brain-derived neurotrophic factor
Exp Neurol
(2004) - et al.
Adeno-associated virus vectors serotyped with AAV-8 capsid are more efficient than AAV-1 or -2 serotypes for widespread gene delivery to the neonatal mouse brain
Neuroscience
(2006) - et al.
Recombinant AAV viral vectors pseudotyped with viral capsids from serotypes 1, 2, and 5 display differential efficiency and cell tropism after delivery to different regions of the central nervous system
Mol Ther
(2004) - et al.
Transduction characteristics of adeno-associated virus vectors expressing cap serotypes 7, 8, 9, and Rh10 in the mouse brain
Mol Ther
(2006) - et al.
Expanded repertoire of AAV vector serotypes mediate unique patterns of transduction in mouse brain
Mol Ther
(2008) - et al.
Helper virus-free, optically controllable, and two-plasmid-based production of adeno-associated virus vectors of serotypes 1 to 6
Mol Ther
(2003) - et al.
Survival of chronically-injured neurons can be prolonged by treatment with neurotrophic factors
Neuroscience
(1999) - et al.
Safety and tolerability of gene therapy with an adeno-associated virus (AAV) borne GAD gene for Parkinson's disease: an open label, phase I trial
Lancet
(2007) - et al.
Measurements of vector-derived neurotrophic factor and green fluorescent protein levels in the brain
Methods
(2002) - et al.
AAV8, 9, Rh10, Rh43 vector gene transfer in the rat brain: effects of serotype, promoter and purification method
Mol Ther
(2008)
Rubrospinal neurons fail to respond to brain-derived neurotrophic factor applied to the spinal cord injury site 2 months after cervical axotomy
Exp Neurol
Efficient gene delivery and selective transduction of glial cells in the mammalian brain by AAV serotypes isolated from nonhuman primates
Mol Ther
Differential and persistent expression patterns of CNS gene transfer by an adeno-associated virus (AAV) vector
Brain Res
Adeno-associated virus vectors: activity and applications in the CNS
J Neurosci Methods
Dimerizer-regulated gene expression
Curr Opin Biotechnol
Transplantation of fetal spinal cord tissue into acute and chronic hemisection and contusion lesions of the adult rat spinal cord
Prog Brain Res
Adeno-associated viral vectors as agents for gene delivery: application in disorders and trauma of the central nervous system
Methods
Adeno-associated viral vector-mediated gene transfer of brain-derived neurotrophic factor reverses atrophy of rubrospinal neurons following both acute and chronic spinal cord injury
Neurobiol Dis
Delayed transplantation of fibroblasts genetically modified to secrete BDNF and NT-3 into a spinal cord injury site is associated with limited recovery of function
Exp Neurol
Analysis of AAV serotypes 1–9 mediated gene expression and tropism in mice after systemic injection
Mol Ther
Production and purification of serotype 1, 2, and 5 recombinant adeno-associated viral vectors
Methods
The 37/67-kilodalton laminin receptor is a receptor for adeno-associated virus serotypes 8, 2, 3, and 9
J Virol
Lentiviral vector-mediated transduction of neural progenitor cells before implantation into injured spinal cord and brain to detect their migration, deliver neurotrophic factors and repair tissue
Restor Neurol Neurosci
Direct gene therapy for repair of the spinal cord
J Neurotrauma
Adenovirus-mediated regulable target gene expression in vivo
Proc Natl Acad Sci U S A
A single injection of an adeno-associated virus vector into nuclei with divergent connections results in widespread vector distribution in the brain and global correction of a neurogenetic disease
J Neurosci
High-titer recombinant adeno-associated virus production utilizing a recombinant herpes simplex virus type I vector expressing AAV-2 Rep and Cap
Gene Ther
Axonal regeneration and functional recovery after complete spinal cord transection in rats by delayed treatment with transplants and neurotrophins
J Neurosci
Cited by (51)
Gene replacement therapy in two Golgi-retained CMT1X mutants before and after the onset of demyelinating neuropathy
2023, Molecular Therapy Methods and Clinical DevelopmentUsing genetically modified extracellular vesicles as a non-invasive strategy to evaluate brain-specific cargo
2022, BiomaterialsCitation Excerpt :NoMi-Nanoluc bioluminescence was used to monitored stable transduction of brain cells with lentivirus encoding the NoMi construct. Intracranial injection of lentivirus is typically well retained at the injection site in contrast to adenoassociated virus (AAV) vectors that can spread to other regions and cross the BBB [83,84]. Taking this into consideration, we detected NoMi-Nanoluc signal that peaked at the injection site and could still be detected 800 μm away from the injection site.
Viral Vectors for Studying Brain Mechanisms that Control Energy Homeostasis
2018, Cell Metabolism