Chondroitinase treatment following spinal contusion injury increases migration of oligodendrocyte progenitor cells

Abstract

Following spinal cord injury (SCI), the demyelination of spared intact axons near the lesion site likely contributes to the loss of motor function. This demyelination occurs when oligodendrocytes, the myelinating cells of the central nervous system (CNS), are either destroyed during the initial trauma or die as a result of secondary pathology. In an attempt to remyelinate the affected axons, endogenous oligodendrocyte progenitor cells (OPCs) begin to accumulate at the border of demyelination. However, the differentiation of OPCs into fully myelinating cells is limited. While the reasons for this are unknown, it is well known that the injured spinal cord is rich in inhibitory molecules that block repair. One such family of molecules is the chondroitin sulfate proteoglycans (CSPGs), which are known to be highly inhibitory to the process of axonal elongation. Recent in vitro findings have demonstrated that CSPGs are also highly inhibitory to OPCs, affecting both their migration and differentiation. Treatment with the enzyme chondroitinase ABC (cABC), which removes the glycosaminoglycan side chains of CSPGs, reverses the inhibitory effects of CSPGs on these cells. In the present study, we examined the effects of cABC on the migratory behavior of endogenous OPCs in vivo following a moderate spinal contusion injury. The total number of OPCs surrounding the lesion site was significantly increased after cABC treatment as compared to controls. cABC treatment also enhanced axonal sprouting, but OPC migration occurs along a different time course and appears independent of new process outgrowth. These data suggest that CSPGs in the post-injury environment inhibit the migration of OPCs, as well as axonal regeneration. Therefore, cABC treatment may not only enhance regenerative axonal sprouting, but may also enhance remyelination after SCI.

Introduction

The loss of function that follows a spinal cord injury (SCI) is largely attributable to the damage of axonal tracts, but also results from the demyelination of spared intact axons. This demyelination is a result of the loss of oligodendrocytes, the myelin forming cell of the central nervous system (CNS). These cells are particularly sensitive to the homeostatic disruptions that accompany SCI (Bunge et al., 1993). Since a single oligodendrocyte can be responsible for myelinating up to 60 different axons (Remhal and Hildebrand, 1990), the death of one oligodendrocyte can result in significant areas of demyelination. The loss of oligodendrocytes occurs both acutely, as a result of the initial trauma, and at chronic times after injury at sites distant from the original lesion site (Crowe et al., 1997, Shuman et al., 1997, Totoiu and Keirstead, 2005, Arvanian et al., 2009). Prolonged demyelination of a spared axon may eventually lead to the degeneration of the axon itself, further contributing to the loss of nerve conduction in the spinal cord (Irvine and Blakemore, 2008). Ultimately the process of demyelination exacerbates the loss of motor function experienced post-SCI.

In an attempt to remyelinate axons, oligodendrocyte progenitor cells (OPCs), the cells that give rise to oligodendrocytes, that reside in the spinal cord begin to migrate towards the site of demyelination (Franklin et al., 1997, McTigue and Tripathi, 2008). However, their migration into the lesioned area and differentiation into fully myelinating cells are not sufficient to repair the chronic demyelination that occurs after injury (Shuman et al., 1997, Totoiu and Keirstead, 2005, Arvanian et al., 2009). The exact reasons why OPCs are unable to fully remyelinate uninjured axons remain unclear. However, one potential explanation is that the post-injury environment inhibits the migration of OPCs toward the lesion site (Fok-Seang et al., 1995, Levine et al., 2001).

A major inhibitory element in the post-injury environment is the glial scar, widely accepted to be a major reason for the abortive nature of axonal regeneration after SCI (reviewed by Fawcett and Asher, 1999, Profyris et al., 2007). Following SCI astrocytes near the injury proliferate, becoming hypertrophic, and secrete high levels of a family of extracellular matrix proteins known as chondroitin sulfate proteoglycans (CSPGs). Reactive astrogliosis ultimately leads to the formation of a dense CSPG rich glial scar. It has been well documented that treating CSPGs with the enzyme chondroitinase ABC (cABC), which cleaves the glycosaminoglycan (GAG) side chains from the central core protein, reverses this inhibition and creates a permissive environment for axonal sprouting (Zuo et al., 1998, Yick et al., 2000, Yick et al., 2003). Interestingly, while the effect of the glial scar and cABC treatment have been extensively studied in the context of axonal regeneration, little is known about how the glial scar and/or cABC treatment might affect OPC migration, differentiation, or the process of remyelination.

Previous work by our laboratory has demonstrated that specific CSPGs expressed within the glial scar exert a highly inhibitory influence on both OPC process outgrowth and differentiation using an in vitro culture model which mimics the glial scar (Siebert and Osterhout, unpublished data). OPCs actively avoid deposits of CSPGs and retract their processes when they come in contact with CSPGs. The effects of CSPGs can be neutralized by the application of the enzyme cABC. These data suggest that CSPGs are a major inhibitory presence at a spinal cord lesion, inhibiting not only axonal sprouting and regeneration, but OPCs as well.

In this study, we examined the effects of cABC treatment on the numbers of OPCs in and around the lesion following a moderate spinal contusion injury. Application of cABC resulted in a significantly greater number of OPCs near the lesion and within the lesion cavity itself at early times (1 and 2 weeks) post-injury. The increased number of cells appears to be a result of migration rather than proliferation. These findings suggest that cABC treatment may be beneficial for remyelination, as well as axonal sprouting and regeneration post-SCI.