Chondroitinase ABC improves basic and skilled locomotion in spinal cord injured cats
Introduction
The inability of damaged axons to regrow following spinal cord injury (SCI) leads to permanent impairments in motor and sensory function below the level of the lesion. An injury-induced increase in chondroitin sulfate proteoglycans (CSPGs) at the lesion site is at least one factor that contributes to the growth inhibitory nature of the injured spinal cord (Lemons et al., 1999, Jones et al., 2002, Jones et al., 2003, Tang et al., 2003; reviewed by Morgenstern et al., 2002, Silver and Miller, 2004, Busch and Silver, 2007). The common feature among this large, diverse family of molecules is the presence of chondroitin sulfate glycosaminoglycans (CS GAGs), which are side chains attached to the core proteins of CSPGs.
Substantial evidence suggests that removal of CS GAGs, most commonly achieved using Chondroitinase ABC (Ch'ase ABC), substantially reduces the inhibitory nature of CSPGs during development and in vitro (Snow et al., 1990, McKeon et al., 1995, Zuo et al., 1998, Chung et al., 2000). More recently, disruption of CS GAGs with Ch'ase ABC in vivo has been shown to enhance axonal growth and behavioral recovery (Yick et al., 2000, Yick et al., 2003, Moon et al., 2001, Bradbury et al., 2002, Chau et al., 2003, Caggiano et al., 2005, Barritt et al., 2006, Houle et al., 2006), as well as synaptic plasticity in rodents (Tropea et al., 2003). These promising results have led our lab to investigate whether disruption of CS GAGs can promote functional recovery following SCI in a larger, more complex mammalian species, the cat.
The extension of potential therapeutic effects in the rat to the cat represents an important translational step. Ch'ase ABC continues to be studied, in part, because some think it may be a potential avenue to pursue for clinical studies of human SCI. One of the recommended strategies for translating preclinical-therapeutic candidates from the laboratory to clinical testing includes testing the efficacy of the therapy in multiple species (Anderson et al., 2005, Blight and Tuszynski, 2006). In addition to the cat's remarkable locomotor capacity and its importance as a model in providing the foundation for evolving locomotor rehabilitation strategies (Lovely et al., 1986, Hodgson et al., 1994, Behrman and Harkema, 2000, Wernig et al., 2000, de Leon et al., 2001, Behrman et al., 2006), the cat presents significant scale-up barriers that must be overcome to successfully treat the human central nervous system after injury. The translational impact of these findings is important due to several factors including the cat's sophisticated motor system and its size which presents physical challenges that are more similar to those that will be encountered in the human. To determine the effects of disrupting CS GAGs on motor recovery, the locomotor performances of three groups of cats with T10 spinal hemisections were compared: lesion-only, lesion + control, and lesion + Ch'ase ABC. The hemisection model was chosen as it permits substantial recovery, and thus enables evaluation of interventions on more skilled behaviors (Helgren and Goldberger, 1993). The control and Ch'ase ABC treatments were delivered intraspinally at the level of the lesion using subcutaneous ports with sub-dural tubing. Control or treatment injections into the ports were made every other day for 1 month (for a total of 15 injections). General recovery trends, as well as specific features of locomotion, were evaluated across a variety of basic and skilled locomotor tasks for 5 to 6 months. Our findings while confirming the work reported in rat models of SCI, identify novel effects of Ch'ase ABC including the accelerated onset of some locomotor recovery, as well as enhanced recovery of intralimb angular kinematic patterns and accuracy of limb trajectories, in the cat. Further, these findings suggest that Ch'ase ABC is effective in altering the extracellular matrix of the spinal cord and enhancing recovery of function across species.
Section snippets
Materials and methods
Fresh frozen, normal, adult, human spinal cord specimens were obtained from the University of Maryland's Brain and Tissue Bank for Developmental Disorders (Baltimore, MD). In compliance with HIPAA guidelines, a “Certificate of Research on Decedents” was acquired from the University of Florida Institutional Review Board and a “Report of Subcommittee on Human Studies” was acquired from the Malcom Randall VA Medical Center for the use of this tissue.
All animal procedures were conducted in
CS GAG similarity between human and cat
Chondroitinase ABC was used to cleave CS GAGs in spinal cord tissue samples from non-injured rat, cat, and human. The lyase products resulting from cleavage were assessed using FACE (Fig. 2). The Δ4S band density was similar in all three species; however, the Δ6S band was notably weaker in the rat compared to the cat and human. Although the Δ0S band could be seen in samples from all species, it was present in the smallest quantities (not shown). Thus, the cat and human are more similar to each
Discussion
The current study shows that degradation of CS GAGs after spinal cord injury in the cat can enhance behavioral recovery and promote axonal growth. Although all groups of cats recovered substantial locomotor function, the performance of the cats treated with intraspinal Ch'ase ABC was superior in multiple ways. During both bipedal treadmill and basic overground locomotion, the kinematic patterns of the Ch'ase ABC-treated cats were more similar than either control group to those seen prior to
Conclusions
The cat provides barriers that aptly challenge many of the demands set forth by human spinal cord injury. The data from this study demonstrate that administration of Ch'ase ABC in spinal cord injured cats may enhance axonal growth and significantly improve locomotor recovery. Most impressive in this model, is the increased rate and extent at which the ipsilateral hindlimb is effectively integrated by Ch'ase ABC-treated cats during skilled behaviors requiring balance and accuracy. These
Acknowledgments
This research was supported by NIH NINDS RO1 NS050699-01 and T32 HD043730, the Daniel Heumann Fund, the Department of Veterans Affairs, and the State of Florida Brain and Spinal Cord Injury Rehabilitation Trust Fund. Human tissue specimens were obtained from the NICHD Brain and Tissue Bank for Developmental Disorders at the University of Maryland (Baltimore, MD). We thank a variety of students and technicians including Stephanie Jefferson, Adele Blum, Sarah Sumner, David Kirby, Eric Neeley,
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2019, Experimental NeurologyCitation Excerpt :It is a lyase that degrades the chondroitin sulphate and dermatan sulphate chains of proteoglycan molecules; it also possesses hyaluronidase activity (Prabhakar et al., 2005; Yamagata et al., 1968). It has been widely used as a strategy to promote repair following spinal cord injury (SCI) (Bradbury et al., 2002; Garcia-Alias et al., 2009; Tester and Howland, 2008; Yick et al., 2000; Zuo et al., 1998c). Most of the therapeutic effects of ChABC can be attributed to its ability to degrade the sugar chains from a class of proteoglycan molecules, the chondroitin sulphate proteoglycans (CSPGs), (Asher et al., 2001; McKeon et al., 1991).
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