Use of immobilized chondroitinase ABC in overcoming axon-restrictive chondroitin sulfates in the injured nerve environment

Abstract

Spinal cord injury causes physiological damage to nerve tissue and often lead to permanent paralysis. The trauma activates the reactive astrocytes to secret high level of chondroitin sulfate proteoglycans (CSPG) that act as physical barrier axonal regrowth, impedes repair and recovery of the damaged neurons. Chondroitinase ABC (ChABC) is an enzyme that can effectively digest the glycosaminoglycan (GAG) chain of CSPG and promote axonal regeneration. However, higher dosage and frequency are necessary for the treatment due to the instability of ChABC I. Here, we immobilized ChABC I on chitosan beads using genipin as crosslinking agent to provide a sustained activity at low dose. In vitro results confirmed that 5mU of immobilized functional ChABC I was significant degraded CS moieties and stimulated extension of rat cortical neurites in co-culture with astrocytes. In vivo experiment on rat model was performed by injecting 2mU of immobilized functional ChABC I into spinal cord at T9 incision. Immunofluorescence (IF) staining reveals the enzymes successfully cleared away the CS moieties and promoted regeneration of neurons into the lesion. The neurons were regenerated in a parallel manner towards the direction where the immobilized functional ChABC I was located. Therefore, low-dose immobilized ChABC I could provide effective axonal regeneration for further study.