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L. Sun, X. Qi, W.W. Hauswirth, J. Guy; AAV-Mediated Sod2 Gene Expression Driven by a Pro-Inflammatory Inducible Promoter: A Novel Method for Gene Therapy of Multiple Sclerosis . Invest. Ophthalmol. Vis. Sci. 2003;44(13):628.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose:To develop a treatment strategy for Multiple Sclerosis (MS). Currently available treatment options for MS consist of corticosteroids to treat relapses and immune modulators as prophylaxis to reduce the frequency and severity of relapses.We have previously shown the protective effect of increasing cellular defenses against reactive oxygen species (ROS) using adenoassociated virus (AAV) as a vector to deliver the genes for mitochondrial superoxide dismutase (Sod2) or catalase to SJL/J mice with experimental autoimmune encephalomyelitis (EAE), a frequently utilized animal model for MS. However, persistence of transgene expression after a single ocular inoculation diminished with time. Methods:To determine if expression of a protective gene, Sod2 in this case, could be triggered by pro-inflammatory mediators released during acute or relapsing EAE and MS we linked into AAV plasmid pTR-UF11 (1) an inducible enhancer element localized to a 280 nucleotide fragment in intron 2 of the rat Sod2 genome in place of the CMV enhancer and CßA promoter, (2) the enhancer element downstream of the CMV enhancer and CßA promoter (3) the inducible enhancer element in place of the CMV enhancer (downstream of the CßA promoter). In all constructs the gene for GFP was replaced by the human cDNA for Sod2. Murine NIH 3T3 cells were infected with rAAVs containing these 3 constructs as well as UF-11 containing the Sod2 gene driven by the CMV enhancer and CßA promoter, but lacking the inducible enhancer element. Cells infected with each of these 4 constructs were exposed to LPS (0.5 µg/ml) for 24 hours or grown in standard media without LPS. Cellular RNA was extracted and probed for human Sod2 gene expression using northern blot analysis. Human MnSOD mRNA was expressed as a ratio to an internal standard, GAPDH. Results:In the presence of LPS the enhancer element without any other promoters increased Sod2 gene expression 2.6 fold relative to cells infected with this rAAV but grown in the absence of the LPS pro-inflammatory mediator. This level of transgene expression was comparable to that achieved by the best promoters currently available to drive ocular gene expression (CMV and CßA). The addition of the CMV or CßA promoters to the inducible enhancer element did not further increase transgene expression. Conclusions:This pro-inflammatory inducible promoter may be useful to drive protective gene expression when most needed during relapses of EAE and perhaps MS.
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