Abstract
Treatment options for chronic spinal cord injury (SCI) remain limited due to unfavourable changes in the microenvironment. Gene therapy can overcome these barriers through continuous delivery of therapeutic gene products to the target tissue. In particular, adeno-associated virus (AAV) vectors are potential candidates for use in chronic SCI, considering their safety and stable gene expression in vivo. Given that different AAV serotypes display different cellular tropisms, it is extremely important to select an optimal serotype for establishing a gene transfer system during the chronic phase of SCI. Therefore, we generated multiple AAV serotypes expressing ffLuc-cp156, a fusion protein of firefly luciferase and Venus, a variant of yellow fluorescent protein with fast and efficient maturation, as a reporter, and we performed intraparenchymal injection in a chronic SCI mouse model. Among the various serotypes tested, AAVrh10 displayed the highest photon count on bioluminescence imaging. Immunohistological analysis revealed that AAVrh10 showed favourable tropism for neurons, astrocytes, and oligodendrocytes. Additionally, with AAVrh10, the area expressing Venus was larger in the injury epicentre and extended to the surrounding tissue. Furthermore, the fluorescence intensity was significantly higher with AAVrh10 than with the other vectors. These results indicate that AAVrh10 may be an appropriate serotype for gene delivery to the chronically injured spinal cord. This promising tool may be applied for research and development related to the treatment of chronic SCI.
Highlights
Traumatic spinal cord injury (SCI) results in severe functional deficits due to the limited regenerative capacity of the central nervous system (CNS)
To analyse the efficacy of transduction mediated by associated virus (AAV), we generated an AAV vector engineered to express ffLuc-cp[156], a fusion protein of firefly luciferase and the fluorescent protein Venus[41,42], as a reporter (Fig. 1A)
Since ffLuc-cp[156] is applicable to bioluminescence systems in addition to its use as a fluorescent protein, we could screen for desired AAV serotypes based on photon yield, as indicated by bioluminescence imaging, and cell-type specificity, as indicated by Venus expression in AAV-transduced tissues
Summary
Traumatic spinal cord injury (SCI) results in severe functional deficits due to the limited regenerative capacity of the central nervous system (CNS). There are few effective treatments for SCI in clinical settings; experimental interventions have been extensively examined in animal models Among these interventions, we and others have reported the efficacy of cell transplantation therapy using embryonic stem cell (ESC)-derived or inducible pluripotent stem cell (iPSC)-derived neural stem precursor cells[2,3,4] or mesenchymal stem cells[5]. We and others have reported the efficacy of cell transplantation therapy using embryonic stem cell (ESC)-derived or inducible pluripotent stem cell (iPSC)-derived neural stem precursor cells[2,3,4] or mesenchymal stem cells[5] In addition to those procedures, therapeutic intervention has been reported using chemical compounds and trophic factors, including riluzole[6,7,8], glibenclamide[9], granulocyte-colony stimulating factor (G-CSF)[10,11], hepatocyte growth factor (HGF)[12,13], an anti-Nogo antibody[14,15,16], chondroitinase ABC (c-ABC)[17,18,19], and a semaphorin 3A inhibitor[20].
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