Engineering Chondroitinase AC for Expression Within the Injured Spinal Cord

Neuronal damage due to spinal cord and brain injury can result in paralysis and/or loss of sensory function. A major contribution to loss of function is the inability of central nervous system neurons to regenerate to their original targets. One factor that confounds regeneration is the injury-induced upregulation of inhibitory proteoglycans (PGs), especially chondroitin sulfate proteoglycans (CSPGs). The majority of CSPGs produced after injury originate from reactive astrocytes surrounding the lesion site. CSPGs interact and prevent the growth of regenerating axons by as of yet an unknown mechanism; however, glycosaminoglycan side chains attached to the protein core are considered the responsible agent. This has been further demonstrated in a number of experiments in \vhich the digestion of chondroitin sulfate has increased regeneration. The enzyme used to digest these CSPGs are knovm as chondroitinases. These are bacterial enzymes with no know mammalian chondroitinases. Their administration presents some problems in that they require repeated injection (~ once every 3 days), function at alkaline pH, and may contain proteases that digest proteins as well. To overcome these difficults we have generated a form of chondroitinase (chase) AC that can be produced and secreted from mammalian cells. Using gene therapy we have been able to induce brain cells, in tissue culture, to produce and secret functional chaseAC. In this proposal we will use gene therapy to express chase AC at a lesion site in the spinal cord and examine regeneration of sensory axons. We will also examine the ability of nerve cells to produce chaseAC and secret it from their growth tips. This should aid motor nerves to gro\\1h through the injury site by digesting the inhibitory molecules as they grow