SHORT COMMUNICATION Incomplete Freund’s adjuvant enhances locomotor performance following spinal cord injury M. F. Azari a,c,d , T. Karnezis b , C. C. Bernard b , C. Profyris c,d , M. R. LeGrande e , D. W. Zang c,d , S. S. Cheema c,d and S. Petratos a a Molecular Neurobiology Laboratory, Department of Biochemistry and Molecular Biology, Monash University, Clayton Victoria; b Neuroimmunology Laboratory, Department of Biochemistry, School of Molecular Sciences, La Trobe University, Bundoora Victoria; c The Howard Florey Institute of Experimental Physiology and Medicine, The University of Melbourne, Parkville Victoria; d Department of Anatomy and Cell Biology, Monash University, Clayton Victoria; and e Heart Research Centre, The Royal Melbourne Hospital, Parkville Victoria, Australia Keywords: demyelination, incomplete Freund’s adjuvant, Nogo- A, spinal cord injury Received 17 August 2004 Accepted 30 December 2004 Following spinal cord injury (SCI), the pathological sequelae which ensue through the secondary mechanisms of degeneration produce myelin deposits which are potent inhibitors of endogenous neuroregeneration. We have enhanced the immune- mediated response following a hemisection lesion by immunizing adult C57Bl/6 female mice against the inhibitor of neurite outgrowth Nogo-A(623–640) peptide. Moderate anti-Nogo-A(623–640) antibody titre levels were obtained by using Montanide as the adjuvant. However, this antibody response was not obtained using incomplete Freund’s adjuvant (IFA). Significant benefit in locomotor per- formance was demonstrated only in animals which were vaccinated with IFA and not with Montanide. No further benefit could be demonstrated with the Nogo- A(623–640) peptide beyond that seen for adjuvant alone. These data imply that generating antibodies against Nogo-A(623–640) in vivo alone is not sufficient to enhance locomotor recovery and that subcutaneous injection of IFA prior to SCI can enhance locomotor performance. Introduction The pathological sequelae following acute spinal cord injury (SCI) include primary and secondary injurious mechanisms (Tator and Fehlings, 1991). The primary injury encompasses the focal destruction of neural tis- sue by trauma. A progressive wave of secondary injury mechanisms then ensues which leads to the destruction of axonal tracts left intact by the initial trauma becoming a major impediment to functional recovery after SCI (Tator and Fehlings, 1991; Tator, 1995; Beattie et al., 2002; Schwartz and Fehlings, 2002). The principal axonal re-growth inhibitor in the acute phase of SCI is central nervous system (CNS) myelin (Ng et al., 1996). The inhibitory components of myelin in- clude myelin-associated glycoprotein (McKerracher et al., 1994; Mukhopadhyay et al., 1994), Nogo-A (Chen et al., 2000; GrandPre et al., 2000; Prinjha et al., 2000) and oligodendrocyte-myelin glycoprotein (Kottis et al., 2002). Of the three myelin inhibitory molecules, Nogo-A has the most pronounced axonal re-growth inhibitory effects (Kim et al., 2003b; Simonen et al., 2003). Other secondary mechanisms include inflammatory responses which are of central importance in the pathogenesis of the acute and chronic phases of SCI. During these phases, the CNS recruits both the innate and adaptive arms of immunity (Hauben and Schwartz, 2003). Despite the success achieved by decreasing the immune-mediated mechanisms follow- ing SCI, a reduced presence of macrophages and microglia at the primary injury may be harmful. In addition to ridding the injured environment of myelin derived axon-growth inhibitory molecules, macroph- ages and microglia can potentiate the release of mediators that promote CNS repair (Nguyen et al., 2002). In fact, SCI performed on rats that are immunized against myelin proteins enhancing immune response, is associated with improved functional recovery (Hauben et al., 2000). Nevertheless, benefit from immunization is strain specific as recovery de- pends upon the individual strains ability to regulate the immune response (Kipnis et al., 2001). We have found that immunization of animals with incomplete Freund’s adjuvant (IFA) before SCI enhances their locomotor performance and may imply that phago- cytic monocytes are required for spinal cord regen- eration possibly through their ability to remove inhibitory myelin debris. Correspondence: Dr Steven Petratos, Department of Biochemistry and Molecular Biology, Monash University, Clayton 3800, Australia (tel.: +61 3 9905 3741; fax: +61 3 9905 3726; e-mail: steven. petratos@med.monash.edu.au). 1004 Ó 2005 EFNS European Journal of Neurology 2005, 12: 1004–1008