Repair Rat Sciatic Nerve Defects With Multichannel Polycaprolactone/Chitosan Nerve Guide Conduits Modified With Neurotrophin-3

Purpose: To prepare a Polycaprolactone/Chitosan-multichannel nerve guide conduit by using a water-soluble Vylon textile thread template and load the conduits with neurotrophin-3(NT-3)by cross-linking grafting technique.The effect of using NT-3-modified PCL/CS nerve conduits to repair sciatic nerve defects in rats was evaluated。.Methods:In the first part of the study,the PCL/CS mixture was prepared by co-blending polycaprolactone(PCL)and chitosan(CS)through acetic acid as the solvent,and the PCL/CS hollow conduit was prepared by drying method as,and the PCL/CS multichannel conduit was prepared by sacrificial template method using water-soluble flexible Vylon textile thread as the template,and the two conduits were examined for material characterization.NT-3 was loaded onto the CS side chains in the PCL/CS material using a cross-linker and tested for slow-release properties.In the second part,material biocompatibility was tested by co-culture of Schwann cells,and then 10 SD rats were randomly selected from each group to make a 10 mm sciatic nerve defect model,and the nerve defects were repaired using PCL/CS-NT3 multichannel conduit,PCL/CS-NT-3hollow conduit and PCL/CS hollow conduit,respectively,with autologous nerve graft as the control group.Motor and sensory function tests were performed at 4 and 12 weeks postoperatively,and neurophysiology,gastrocnemius wet weight,cross-sectional myocyte area,collagen fiber ratio,and myelinated nerve fiber count of regenerated nerve tissue were examined at 12 weeks postoperatively to observe the effect of the conduit on the nerve defect is repair.Results: PCL/CS multichannel nerve conduit was successfully prepared,and it showed better properties than hollow conduit in tensile fracture test,bending resistance test,water absorption test and hydrophilic test,and it could maintain the lumen morphology and channel patency better than hollow conduit,and withstand bending load better than hollow conduit,which can meet the requirements as nerve bridging repair material.The release rate and concentration of NT-3 grafted to CS side chains were higher in the porous conduit group than in the hollow conduit.Cell proliferation assay showed no significant difference in the number of cell proliferation between the three groups at 24 h(P>0.05),and at 48 h of co-culture,the PCL/CS-NT-3 multi-channel conduit group showed a stronger proliferation trend than the PCL/CS-NT-3 hollow conduit group(P<0.05);at 72 h of culture,the PCL/CS-NT-3 multi-channel conduit group showed higher cell proliferation numbers than PCL/CS hollow conduit and blank control group with significant differences(P<0.05);while All four groups showed higher than normal warm retraction time at 4 weeks,and the retraction time of the autonomic group was significantly shorter than that of the three groups using artificial materials,and the retraction time of the PCL/CS-NT-3 multi-channel conduit group was shorter than that of the PCL/CS-NT-3 hollow conduit and the PCL/CS hollow conduit group,and the difference was significant(P<0.05).At 12 weeks postoperatively,all four groups of rats had different degrees of shortening of warm retraction time compared with 4 weeks postoperatively,while the shortest time was observed in the rats of the autonomic nerve group and the PCL/CS-NT-3 multichannel conduit group,and the retraction time was similar between the two groups with no significant difference(P>0.05),but all were significantly better than the PCL/CS-NT-3hollow conduit group and the PCL/CS hollow conduit group(P<0.05).At 4 weeks postoperatively,all four groups showed different degrees of hindfoot dysfunction,but there was no significant difference between the four groups(P>0.05).At 12 weeks postoperatively,the hindfoot function of the rats in each of the four groups recovered to different degrees compared with that at 4 weeks,among which the autonomic nerve group,PCL/CS-NT-3 multichannel conduit group and PCL/CS-NT-3 hollow conduit group all recovered better than the PCL/CS hollow conduit group(P>0.05).For neurophysiological testing,there was no significant difference in the latency of CMAP between the PCL/CSNT-3 multichannel conduit group and the autologous nerve group(P>0.05),but both were significantly shorter than the PCL/CS-NT-3 hollow conduit group and the PCL/CS hollow conduit group(P<0.05).The CMAP wave amplitude decreased sequentially in the autonomic nerve group,the PCL/CS-NT-3 multichannel conduit group,the PCL/CS-NT-3hollow conduit group and the PCL/CS hollow conduit group,while the differences between the groups were all significant(P<0.05).The autonomic nerve group had the highest rate of muscle wet weight recovery,which was significantly higher than the other three groups(P<0.05);while the PCL/CS-NT-3 multichannel conduit group was the second highest,which was significantly higher than the PCL/CS-NT-3 hollow conduit group and the PCL/CS hollow conduit group(P<0.05).Myocyte cross-sectional area: the recovery rates in the autonomic nerve group,PCL/CS-NT-3 multichannel conduit group,PCL/CS-NT-3hollow conduit group and PCL/CS hollow conduit group were decreasing in order,and the differences were statistically significant(P<0.05).The positive area of collagen fibers in the cross-section of gastrocnemius muscle,the proportion of collagen fibers in the autonomic nerve group,PCL/CS-NT-3 multichannel conduit group,PCL/CS-NT-3 hollow conduit group and PCL/CS hollow conduit group increased sequentially,and the differences were all statistically significant(P<0.05).The density of myelinated nerve fibers was higher in the autologous nerve group and the PCL/CS-NT-3 multichannel conduit group,than those in PCL/CS-NT-3 hollow conduit group and PCL/CS hollow conduit group(P<0.05).Immunofluorescence detection: the proportion of collagen fibers increased sequentially in the autologous nerve group,PCL/CS-NT-3 multichannel conduit group,PCL/CS-NT-3hollow conduit group and PCL/CS hollow conduit group,and the differences were all statistically significant higher than the PCL/CS-NT-3 hollow conduit group and the PCL/CS hollow conduit group(P<0.05).Conclusion: the PCL/CS-NT-3 multichannel catheter exhibited better mechanical properties,biocompatibility,and slow release ability to promote nerve regeneration and nerve function recovery,and showed better repair ability for larger peripheral nerve defects..