NCAM-mediated locomotor recovery from spinal cord contusion injury involves neuroprotection, axon regeneration, and synaptogenesis
Introduction
Spinal cord injury (SCI) causes serious loss of neural cells, tissue destruction and attenuation of neuronal function (reviewed by Kwon et al., 2004). A large variety of molecules are involved in the pathological developments. Some of them showed direct beneficial or detrimental effects on functional recovery (Profyris et al., 2004) but the mechanisms are poorly understood. From accumulated evidences it is known that spontaneous axonal growth occurs in experimental SCI animals (Okano et al., 2007, Hill et al., 2001). The axon outgrowth is believed to establish new circuits and contribute to the limited functional recovery (Bareyre et al., 2004, Bregman et al., 2002). Thus, elucidating influences of the activated neurotrophic factors on axon outgrow after SCI may provide clues to treatment.
Neural cell adhesion molecule (NCAM), which belongs to the immunoglobulin super-family, plays important roles in mediating cell migration, survival, neurite growth and synaptic plasticity (Rønn et al., 1998). There are three major isoforms of NCAM identified in most tissues with distinct molecular weight of 180, 140 and 120 kDa (Chuong and Edelman, 1984). All NCAM isoforms are transcribed from a single gene via mRNA alternative splicing mechanism (Santoni et al., 1987). Through homo- and heter-philic binding, NCAM associates with cell structural molecules and evokes several signaling cascades (Ditlevsen and Kolkova, 2008; Büttner et al., 2003). CAMP, Ca+, PKC and MAPK pathways play important roles in NCAM-stimulated neurite outgrowth (Maness and Schachner, 2007). Recent studies have revealed that NCAM actively participates in the regeneration of muscle (Dubois et al., 1994) and nervous system (Walsh and Doherty, 1996, Zhang et al., 2008). In completely transacted spinal cords, the expression level of NCAM was markedly changed (Tzeng et al., 2001), indicating its involvement in the pathological development after SCI. Moreover, polysialylated NCAM promotes selective targeting of regenerated motor neurons (Franz et al., 2005). These findings have aroused our interest to investigate the change in NCAM expression after SCI and its possible correlation with functional recovery of the spinal cord. NCAM-deficient (ND) mice and their wildtype (WT) littermates were employed in the present study of NYU weight-drop induced contusion spinal cord injury model. The locomotor ability of their hind limb was evaluated for up to 5 weeks after SCI. Apoptosis, astrocytosis, axon regeneration and synaptogenesis in the spinal cord were examined.
Section snippets
Surgical procedure and animal care
A total of 24 female B6.129P2-NCAM1tm1Cgn/J mice (ND) and 95 female wildtype (WT) littermates (Jackson Laboratory, Bar Harbor, ME, USA, 15–20 g) were used in this study. Animals were randomly selected for sham or injury groups. The New York University (NYU) model is favored in the present study because the lesions created seem to accurately mimic those seen in humans (Anderson and Stokes, 1992). Mice were anesthetized with pentobarbital (60 mg/kg) and their body temperatures were maintained by a
Expression and distribution of NCAM change dynamically following SCI
In the spinal cord, expression of NCAM was mainly demonstrated in the superficial dorsal horn of the sham animals (Fig. 1A), which is consistent with previous report (Seki and Arai, 1993). However, at 2 weeks after SCI, NCAM was highly expressed in gray and white matters of the injured spinal cord and hardly detectable in the scar area (Fig. 1A). To determine the overall NCAM expression in the damaged spinal cords, Western blot has been performed with spinal cord segment containing the injury
Discussion
Spinal cord injury was characterized by the drastic tissue destruction, cell death, scar formation and neural function loss. It has been widely accepted that the functional outcomes following SCI are determined by severity of the exogenous damage and capacity of the endogenerous recovery, which were attributed to the effects of tremendous number of factors (Profyris et al., 2004). Recent studies demonstrated that the endogenous function recovery is probably benefited from the limited axonal
Acknowledgments
We are grateful to Dr Yu Wei Ping in the National Neuroscience Institute of Singapore for his kind help with cryostat and thank Dr. Chew Sing Yian in the School of Chemical and Biomedical Engineering, Nanyang Technological University for critical reading the manuscript. This work was funded by Biomedical Research Council Grant (BMRC 06/1/33/19/479) of Singapore.
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