Overcoming the chondroitin sulphate barrier in injured nerve: the efficacy of chondroitinase ABC treatment and the potency of disaccharide products

Abstract

Chondroitin sulphate proteoglycans (CSPGs) up-regulated in the extracellular matrix of injured nerves, constitute barriers to nerve regeneration. Cleavage of CS using chondroitinase ABC (Proteus vulgaris) promises axon regrowth through the barrier. Albeit the occurrence of two subtypes (ChABCI, an endolyase, and ChABC II, and exolyase), only ChABC I is commercially available and tested for medical use. We produced recombinant (r) forms of the two subtypes and found that rChABC I was subject to competitive inhibition by the limit digestion product, CS-tetrasaccharides. Inclusion of rChABC II in the digestion mixture yielded essentially CS-disaccharides that were not inhibitors of rChABC I. Combinatorial use of both rChABC subtypes therefore improved efficiency of the CS-cleaving reaction and resulted in increased neurite lengths in a coculture model of cortical neurons with TGFβ1-stimulated astrocytes. Neurites growth-promoting effects of CS digestion products were revealed in cocultures with had been treated with rChABC combinations, washed of digestion products and then treated with graded supplements of CS-type ΔDi-4S or ΔDi-6S. By contrast, control treatments with sucrose or CS-type tetrasaccharides did not promote neurite growth . Evidence is thus provided for the dual role of combinatorial ChABC activities, both in cleaving the putative CS barrier to axon growth, and yielding CS-type disaccharide products that contribute to neurite growth.