Lentiviral vectors enveloped with rabies virus glycoprotein can be used as a novel retrograde tracer to assess nerve recovery in rat sciatic nerve injury models
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Retrograde labeling has become the new “gold standard” technique to evaluate the recovery of injured peripheral nerves. In this study, lentiviral vectors with rabies virus glycoprotein envelop (RABV-G-LV) and RFP genes are injected into gastrocnemius muscle to determine the location of RFP in sciatic nerves. We then examine RFP expression in the L4-S1 spinal cord and sensory dorsal root ganglia and in the rat sciatic nerve, isolated Schwann cells, viral dose to expression relationship and the use of RABV-G-LV as a retrograde tracer for regeneration in the injured rat sciatic nerve. VSV-G-LV was used as control for viral envelope specificity. Results showed that RFP were positive in the myelin sheath and lumbar spinal motorneurons of the RABV-G-LV group. RFP gene could be detected both in myelinated Schwann cells and lumbar spinal motor neurons in the RABV-G-LV group. Schwann cells isolated from the RABV-G-LV injected postnatal Sprague Dawley rats were also RFP-gene positive. All the results obtained in the VSV-G-LV group were negative. Distribution of RFP was unaltered and the level of RFP expression increasing with time progressing. RABV-G-LV could assess the amount of functional regenerating nerve fibers two months post-operation in the four models. This method offers an easy-operated and consistent standardized approach for retrograde labeling regenerating peripheral nerves, which may be a significant supplement for the previous RABV-G-LV-related retrograde labeling study.
KeywordsLentiviral vectors Rabies virus glycoprotein Myelinated Schwann cells Retrograde labeling Peripheral nerve regeneration
This study was supported by the National Natural Science Foundation of China (81272787), Zhou Dafu Medical Research Fund (202836019-03) and the National Natural Science Foundation of Beijing (7122027).
- Burns JC, Friedmann T, Driever W, Burrascano M, Yee JK (1993) Vesicular stomatitis virus G glycoprotein pseudotyped retroviral vectors: concentration to very high titer and efficient gene transfer into mammalian and nonmammalian cells. Proc Natl Acad Sci U S A 90:8033–8037PubMedCentralPubMedCrossRefGoogle Scholar
- Kato S, Inoue K, Kobayashi K, Yasoshima Y, Miyachi S, Inoue S, Hanawa H, Shimada T, Takada M (2007) Efficient gene transfer via retrograde transport in rodent and primate brains using a human immunodeficiency virus type 1-based vector pseudotyped with rabies virus glycoprotein. Hum Gene Ther 18:1141–1151PubMedCrossRefGoogle Scholar
- Kato S, Kuramochi M, Takasumi K, Kobayashi K, Inoue K, Takahara D, Hitoshi S, Ikenaka K, Shimada T, Takada M (2011c) Neuron-specific gene transfer through retrograde transport of lentiviral vector pseudotyped with a novel type of fusion envelope glycoprotein. Hum Gene Ther 22:1511–1523PubMedCrossRefGoogle Scholar
- Mazarakis ND, Azzouz M, Rohll JB, Ellard FM, Wilkes FJ, Olsen AL, Carter EE, Barber RD, Baban DF, Kingsman SM, Kingsman AJ, O'Malley K, Mitrophanous KA (2001) Rabies virus glycoprotein pseudotyping of lentiviral vectors enables retrograde axonal transport and access to the nervous system after peripheral delivery. Hum Mol Genet 10:2109–2121PubMedCrossRefGoogle Scholar
- Muik A, Kneiske I, Werbizki M, Wilflingseder D, Giroglou T, Ebert O, Kraft A, Dietrich U, Zimmer G, Momma S, von Laer D (2011) Pseudotyping vesicular stomatitis virus with lymphocytic choriomeningitis virus glycoproteins enhances infectivity for glioma cells and minimizes neurotropism. J Virol 85:5679–5684PubMedCentralPubMedCrossRefGoogle Scholar
- Puigdellivol-Sanchez A, Prats-Galino A, Ruano-Gil D, Molander C (2000) Fast blue and diamidino yellow as retrograde tracers in peripheral nerves: efficacy of combined nerve injection and capsule application to transected nerves in the adult rat. J Neurosci Methods 95:103–110PubMedCrossRefGoogle Scholar
- Puigdellivol-Sanchez A, Valero-Cabre A, Prats-Galino A, Navarro X, Molander C (2002) On the use of fast blue, fluoro-gold and diamidino yellow for retrograde tracing after peripheral nerve injury: uptake, fading, dye interactions, and toxicity. J Neurosci Methods 115:115–127PubMedCrossRefGoogle Scholar
- van Neerven SG, Bozkurt A, O'Dey DM, Scheffel J, Boecker AH, Stromps JP, Dunda S, Brook GA, Pallua N (2012) Retrograde tracing and toe spreading after experimental autologous nerve transplantation and crush injury of the sciatic nerve: a descriptive methodological study. J Brachial Plexus Peripher Nerv Inj 7:5CrossRefGoogle Scholar