Study On Peripheral Nerve Regeneration With RGD Grafted Poly[LA-(Glc-Lys)]/PDLLA/β-TCP Composites

A novel composite film was investigated as nervous conduit materials to repair nerve defect.Firstly,RGD grafted Poly[LA-(Glc-Lys)](abbreviated as PRGD)was synthesized,then the copolymer was physical mixed with PDLLA,β-tricalcium phosphate(β-TCP)powder to prepare composite film which was expected to have good cytocompatibility.3S-[N^ε-(benzyloxycarbonyl)-L-lysyl]-2,5-morpholinedione was obtained by the intramolecular reaction of N^ε-romoacetyl-Lys(z),which was obtained by reactions of N^ε-(benzyloxy-carbonyl)-L-lysine with bromoacetyl bromide.Poly[LA-co-(Glc -alt-N^ε-benzyloxy-carbonyl-L-Lys)]was synthesized by ring opening copolymerization of D,L-lactide with 3S-[N^ε-(benzyloxycarbonyl)-L-lysyl]-2,5-morpholinedione. To obtain poly[LA-(Glc-Lys)],the deprotection of benzyl-oxycarbonyl group is accomplished with a Pd/C catalyst system.RGD grafted poly[LA-(Glc-Lys)]is obtained by the chemical attachment of the peptide RGD to the polymer's lysine residue.The copolymers were characterized with FT-IR,¹H-NMR and ¹³C-NMR.The results showed that N^ε-(Carbonylbenzoxy)-L-lysine was introduced into the main chain of the copolymer.The composition of the copolymer can be adjusted according to the ratio change in feed.¹H-NMR indicated that the protective group of the polymer has been removed.Animo acid analysis(AAA)showed that the amount of RGD peptide grafted onto the copolymer was 12.7～14.3μmol/g.PRGD and PDLLA with mass ratio 1:1 was dissolved in ethyl acetate, concentration of which was 10%.The solution was then mixed with proper quantities ofβ-TCP on a magnetic stirrer till the mixture was homogeneously.The composite film was then prepared by solvent pouring method.Water contact angle test was used to evaluate hydrophiIicity of RGD grafted poly[LA-(Glc-Lys)]/PDLLA/β-TCP composite film.The results showed that the hydrophilicity of the composite had been greatly improved compared to PDLLA.RSC96 cells were co-cultured with PRGD/PDLLA film and PDLLA film to evaluate their cell compatibility and potential to be used as nerve conduit material. The SEM photographs showed that PRGD/PDLLA film was superior in cell adhesion and cell growth to PDLLA film.RGD grafted Poly[LA-(Glc-Lys)] composite had good cytocompatibility and could be quite promising in nerve tissue engineering.In addition,animal experiments were conducted to evaluate the biocompatibility of the composites.The sciatic nerve defect model of adult rats was conducted in this experiment,then the defected nerves were bridged with PDLLA conduits,PRGD/PDLLA/β-TCP conduits respectively,with autograft group as the control group.The rats were anaesthetized and conducted with function recovery evaluation,morphological evaluation,histological evaluation,electrophysiological assay to evaluate the recovery of the regenerated nerve in 3 months and 6 months after the operation.The results showed that the composites had good biocompatibility,and could promote the regeneration of injured peripheral nerves effectively.The efficiency could compete with autograft group and was much more prominent than PDLLA group.