丝胶蛋白对仿生丝素蛋白神经导管的改性研究

摘要

目的 探索丝胶(SS)蛋白对仿生多通道丝素(SF)蛋白神经导管(NCs)的改性作用. 方法 在丝素蛋白水溶液中按比例加入丝胶蛋白,通过循序冷冻工艺制备丝胶/丝素共混神经导管(SS/SF-NCs),扫描电镜(SEM)观测其通道结构,X线衍射及红外光谱检测其内部分子结构,测量其孔隙率及机械性能,MTT实验量化分析PC12细胞与SS/SF-NCs共培养后的细胞活性,PC12细胞被用于检测从SS/SF-NCs中释放的NGF的细胞活性. 结果 SEM结果显示与SF-NCs相比,SS/SF-NCs具有周围神经束仿生结构的线性化导向的多通道,通道分布均匀呈片状纵向均匀排列,且通道微观结构上发生很大变化,机械性能得到很大提升,通道的壁间距、孔隙率、机械强度随着循序冷冻温度、丝胶比例的不同而变化.MTT试验结果显示SS/SF-NCs的PC12细胞的活性明显好于对照组(P＜0.05),从新型SS/SF-NCs中释放NGF释放时间长达4周并保持生物活性.结论通过丝胶蛋白的改性作用,新型SS/SF-NCs具有周围神经束的高度仿生结构,且具有更加优良的机械性能,可以作为人工神经导管的另一选择.%Objective To explore the modification of bionic silk fibroin nerve conduits (SF-NCs) by silk sencin.Methods The innovative SS/SF blended-NCs was fabricated by a vertical sequential cooling thermal induced phase separation (TIPS) processing with SF solution added sericin in proportion,its morphology was observed by Scanning electron microscopy (SEM),X-ray diffraction (XRD) and infrared spectroscopy (FTIR) were used to detect its internal molecular structure.MTT assay was used to quantitatively analyzed the PC12 cells viability co-cultured with the innovative SS/SF-NCs,SEM was used to observe the adhesion and morphology of PC12 cells seeded into the innovative SS/SF,PC12 cells were used to assess the NGF bioactivity released from the SS/SF.Results The SEM results showed that the new fabricated SS/SF-NCs had linearly oriented lamellar-like multiple-channel which distributed evenly,got great changes on the channel microstructure and their mechanical properties had been greatly improved,compared to SF-NCs.The XRD and FTIR results showed that the SS/SF-NCs had the similar internal molecular structure with natural silk.The spaces between parallel lamellar-like channels,porosities and compressive strengths of the SS/SF-NCs decreased with decreasing Sequential freezing temperature.MTT assay results showed that the viability of PC12 cell was better than the control group (P ＜ 0.05).The SEM observation indicated that PC12 cells showed good adhesion and differentiation with neuritis outgrowth during the period of co-culture with the SS/SF-NCs.NGF release from the innovative SS/SF-NCs was prolonged over 4 weeks,and remained bioactive.Conclusion The new fabricated SS/SF-NCs modified though silk sericin,which was highly bionic the structure of peripheral nerve fasciculus,had excellent mechanical properties and could be used as another alternative of artificial nerve conduits.