In vivo adenovirus-mediated gene transfer and expression in ischemic rabbit spinal cord (original) (raw)
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Application of recombinant adenovirus for in vivo gene delivery to spinal cord
Brain Research, 1997
One strategy for treating spinal cord injury is to supply damaged neurons with the appropriate neurotrophins either by direct delivery or by transfer of the corresponding genes using viral vectors. Here we report the feasibility of using recombinant adenovirus for in vivo Ž . gene transfer in spinal cord. After injection of a recombinant adenovirus carrying a b-galactosidase b-gal reporter gene into the mid-thoracic spinal cord of adult rats, transgene expression occurred not only in several types of cells around the injection site but also in neurons whose axons project to this region from rostral or caudal to the injection site. Among labeled neurons were those of the red nucleus, the vestibular nuclei, reticular formation, locus coeruleus, and Clarke's nucleus. A non-specific immune reaction, which could be blocked by immunosuppression with Cyclosporin A, reduced the number of transduced cells surviving at the injection site by 1 month. In neurons away from the injection site, where the immune response was minimal, transgene expression lasted for at least 2 months. These results support the idea that recombinant adenovirus can be used in the spinal cord for in vivo delivery of therapeutic genes important for supporting neuron survival and axon regeneration. q 1997 Elsevier Science B.V.
Molecular neurobiology, 2015
The lack of methods to deliver transgene expression in spinal cord has hampered investigation of gene function and therapeutic targets for spinal cord diseases. Here, we report that a single intrathecal injection of recombinant adeno-associated virus rhesus-10 (rAAVrh10) into the lumbar cistern led to transgene expression in 60 to 90 % of the cells in the spinal cord. The transgene was expressed in all cell types, including neurons, glia, ependymal cells, and endothelial cells. Additionally, the transgene was expressed in some brain areas up to the frontal cortex and the olfactory bulb. The rAAV was distributed predominantly in the spinal cord, where its genome copy was over ten times that of the peripheral organs. Compared with intravenous injection, another method for rAAV delivery to the broad central nervous system (CNS), the intrathecal injection reduced the dosage of rAAV required to achieve similar or higher levels of transgene expression in the CNS by ~100-fold. Finally, the...
Adenovirus-Mediated Gene Transfer to Human Cerebral Arteries
Journal of Cerebral Blood Flow & Metabolism, 2000
Gene therapy is being investigated as a putative treatment option for cardiovascular diseases, including cerebral vasospasm. Because there is presently no information regarding gene transfer to human cerebral arteries, the principal objective of this study was to characterize adenovirus-mediated expression and function of recombinant endothelial nitric oxide synthase (eNOS) gene in human pial arteries. Pial arteries (outer diameter 500 to 1000 μm) were isolated from 30 patients undergoing temporal lobectomy for intractable seizures and were studied using histologic staining, histochemistry, electron microscopy, and isometric force recording. Gene transfer experiments were performed ex vivo using adenoviral vectors encoding genes for bovine eNOS (AdCMVeNOS) and Escherichia coli β-galactosidase (AdCMVLacZ). In transduced arteries, studied 24 hours after exposure to vectors, expression of recombinant β-galactosidase and eNOS was detected by histochemistry, localizing mainly to the adve...
Neurobiology of Disease, 2003
The current investigation tests whether adeno-associated viral vectors (rAAV) undergo remote delivery to the spinal cord via peripheral nerve injection as previously demonstrated with adenoviral vectors. The sciatic nerves of adult rats (n ϭ 10) were injected with either an rAAV (rAAVCMV-lacZ) or adenoviral (AdCMV-lacZ) vector (1.4 ϫ 10 7 particles/ml). After 21 days, the rAAV group demonstrated significantly higher spinal cord viral expression than the adenoviral group (P Ͻ 0.024). A second group of rats was injected with rAAV expressing the green fluorescence protein (GFP) reporter gene. GFP was detected 21 days after unilateral sciatic nerve injection in the neurons of the dorsal root ganglion and spinal cord. The codistribution of the viral genome and transgene in CNS neurons was confirmed with in situ hybridization. In summary, rAAV genes are expressed in CNS neurons following peripheral nerve injection at levels exceeding those seen following remote adenovirus injection.
Neuronal survival following remote adenovirus gene delivery
Journal of Neurosurgery: Spine, 2002
Object. Virus-mediated central nervous system gene delivery is a promising means of treating traumatized tissue or degenerative diseases. In the present study, the authors examined gene expression and neuronal survival in the spinal cord after sciatic nerve administration of an adenovirus vector expressing a LacZ reporter gene. Methods. The time course of adenovirus gene expression, DNA fragmentation, and neuronal density were quantified in rat lumbar spinal cord by staining for β-galactosidase (β-Gal), terminal deoxynucleotidyl transferase, and cresyl violet after microinjection of either saline or the reporter virus into rat sciatic nerve. The expression of β-Gal following remote vector delivery peaked at 7 days and declined thereafter but was not accompanied by neuronal cell death, as measured by DNA fragmentation. No significant difference in spinal motor neuron density was detected between virus-treated and control rats at any time point examined. Although the spinal cords remo...
Planet of the AAVs: The Spinal Cord Injury Episode
Biomedicines
The spinal cord injury (SCI) is a medical and life-disrupting condition with devastating consequences for the physical, social, and professional welfare of patients, and there is no adequate treatment for it. At the same time, gene therapy has been studied as a promising approach for the treatment of neurological and neurodegenerative disorders by delivering remedial genes to the central nervous system (CNS), of which the spinal cord is a part. For gene therapy, multiple vectors have been introduced, including integrating lentiviral vectors and non-integrating adeno-associated virus (AAV) vectors. AAV vectors are a promising system for transgene delivery into the CNS due to their safety profile as well as long-term gene expression. Gene therapy mediated by AAV vectors shows potential for treating SCI by delivering certain genetic information to specific cell types. This review has focused on a potential treatment of SCI by gene therapy using AAV vectors.
Adenoviral gene transfer to spinal cord neurons: intrathecal vs. intraparenchymal administration
Brain Research, 1998
The spinal cord is the site of many chronic, debilitating, neurological disorders that may be amenable to gene therapy. The present study, using quantitative and anatomical methods, examines the ability of replication deficient adenovirus to transfer a transcription Ž . cassette composed of the cytomegalovirus promoter driving the expression of the LacZ reporter gene AdCMVb gal to spinal-cord neurons. Rats were microinjected with AdCMVb gal into the spinal-cord parenchyma or subarachnoid space and sacrificed between 1 and Ž . Ž . 60 days post-infusion. The spinal cord was assayed for b-galactosidase b-gal activity fluorometrically MUG . Intraparenchymal Ž injection resulted in significant b-gal activity at day 1, which peaked at day 7, and decreased at day 14 21-, 57-and 9.8-fold of control . respectively . The spatial distribution of b-gal activity on day 7 was confined to the 1-cm section containing the injection site but was detected 2 cm caudal to this section by day 14. Histochemical staining and immunocytochemistry revealed a prominent reaction product in neurons, particularly motor neurons, and glia within the ventral grey matter bilaterally. Intrathecal viral injections showed Ž . comparatively modest, yet significant increases in b-gal activity throughout the spinal cord with the greatest activity 170% control closest to the catheter tip. This study demonstrates that AdCMVb gal injected into the ventral spinal cord results in extensive in vivo neuronal gene transfer with b-gal activity reaching a peak by day 7 and remaining detectable at 60 days. Intrathecal viral injections result in greater spatial distribution but a comparatively lower level of expression. q
An adenoviral vector can transfer lacZ expression into schwann cells in culture and in sciatic nerve
Annals of Neurology, 1995
Although a number of genetic defects in the PO, peripheral myelin protein-22, and connexin-32 genes recently were shown to cause the demyelinating forms of Charcot-Marie-Tooth disease, there is yet no effective treatment for these patients. Recent studies showed that replication defective adenoviral vectors can efficiently introduce genes into muscle, brain, lung, and other tissues, suggesting that this vector system may be useful for the treatment of a number of genetic diseases. In this work, we demonstrated that a replication deficient adenovirus expressing the Escherichia coli P-galactosidase gene (AdCMVLacZ) can introduce genes into Schwann cells, in culture as well as in sciatic nerve. Schwann cells cultured at a multiplicity of infection of 250: 1 did not demonstrate cytopathic effects. Following injection of AdCMVLacZ into sciatic nerve of rats, lacZ-expressing, myelinating Schwann cells could be detected for at least 45 days. These data suggest that in the future, these vectors may be useful both in perturbing Schwann cell gene expression and in designing therapies for the treatment of Charcot-Marie-Tooth disease. Shy ME, Tani M, Shi Y , Whyatt SA, Chbihi T, Scherer SS, Kamholz J. An adenoviral vector can transfer lacZ expression into Schwann cells in culture and in sciatic nerve. Ann Neurol 1995;38:429-436
Frontiers in pharmacology, 2017
The gene therapy has been successful in treatment of spinal cord injury (SCI) in several animal models, although it still remains unavailable for clinical practice. Surprisingly, regardless the fact that multiple reports showed motor recovery with gene therapy, little is known about molecular and cellular changes in the post-traumatic spinal cord following viral vector- or cell-mediated gene therapy. In this study we evaluated the therapeutic efficacy and changes in spinal cord after treatment with the genes encoding vascular endothelial growth factor (VEGF), glial cell-derived neurotrophic factor (GDNF), angiogenin (ANG), and neuronal cell adhesion molecule (NCAM) applied using both approaches. Therapeutic genes were used for viral vector- and cell-mediated gene therapy in two combinations: (1) VEGF+GDNF+NCAM and (2) VEGF+ANG+NCAM. For direct gene therapy adenoviral vectors based on serotype 5 (Ad5) were injected intrathecally and for cell-mediated gene delivery human umbilical cor...