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Chondroitinase activity can be transduced by a lentiviral vector in vitro and in vivo
Journal article   Open access   Peer reviewed

Chondroitinase activity can be transduced by a lentiviral vector in vitro and in vivo

Ying Jin, Andrea Ketschek, Zhilong Jiang, George Smith and Itzhak Fischer
Journal of neuroscience methods, v 199(2), pp 208-213
15 Aug 2011
DOI: https://doi.org/10.1016/j.jneumeth.2011.05.007
PMID: 21600922
Featured in Collection :   UN Sustainable Development Goals @ Drexel
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https://doi.org/10.1016/j.jneumeth.2011.05.007View
Published, Version of Record (VoR) Open

Abstract

Axons - enzymology Rats, Transgenic Chondroitin Lyases - metabolism Rats, Inbred F344 Cells, Cultured Rats Axons - physiology Chick Embryo Rats, Sprague-Dawley Genetic Vectors - genetics Chondroitin Lyases - genetics Animals Female Lentivirus - genetics Genetic Vectors - physiology Transduction, Genetic - methods
The bacterial enzyme chondroitinase ABC (ChABC), which cleaves chondroitin sulfate glycosaminoglycan chains, can degrade inhibitory scar tissue formed following spinal cord injury, thereby promoting axonal growth and regeneration. However, delivering the active enzyme for prolonged periods presents practical limitations. To overcome these problems, we prepared a lentiviral vector (LV) encoding chondroitinase AC (Chase) together with the green fluorescent protein (GFP) reporter (Chase/LV) and demonstrated its expression and enzymatic activity in vitro and in vivo. Neural precursor cells infected with Chase/LV expressed the GFP reporter at levels that increased dramatically with time in culture. Enzymatic activity from the supernatant of the infected cells was demonstrated by dot blot assay using an antibody that recognizes the digested form of CSPG and was compared with the bacterial ChABC enzyme. Chick DRG cultures plated adjacent to the CSPG border and incubated with supernatant from Chase/LV-infected cells showed neurites growing into the CSPG area, a response similar to that after treatment with ChABC. In contrast, in control cultures, the neurites turned to avoid the inhibitory CSPG interface. Degradation of CSPG in these cultures was confirmed by specific CSPG antibodies. A single injection of Chase/LV into the spinal cord resulted in sustained secretion of the enzyme, whose activity was detected for 8 weeks by expression of GFP and evidence of the digested form of CSPG. This study demonstrates the efficacy of the Chase/LV vector and its potential as a therapeutic tool to reduce scar inhibition and promote axonal growth and repair following central nervous system injury.

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30 citations in Web of Science
33 citations in Scopus

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Collaboration types
Domestic collaboration
Web of Science research areas
Biochemical Research Methods
Neurosciences
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