自组装IKVAV多肽纳米支架及其对背根神经节神经元细胞的作用

Effects of self-assembled IKVAV peptide nanofibers on dorsal root ganglion neurons

目的:应用自组装IKVAV多肽纳米支架与鼠背根神经节神经元细胞(DRGc)联合培养,观察其对DRGc的作用。方法:将多肽溶于0.1MNaOH溶液中,调整pH值为8.5,多肽浓度为0.01mg/μl,与等体积DMEM/F12混合触发多肽自组装为凝胶支架,透射电镜检测。采用原代分离培养方法获得DRGc单细胞悬液后分为实验组与对照组,实验组中DRGc接种于凝胶支架表面,对照组接种于多聚赖氨酸表面,倒置相差显微镜观察神经元生长情况,采用细胞计数结合免疫细胞化学染色方法,观察DRGc的存活和轴突生长情况,并行统计学分析。结果:透射电镜下显示自组装凝胶支架为编织状纳米纤维。实验组和对照组中DRGc培养1d时平均轴突长度分别为43.8±10.4μm、33.4±5.75μm;培养14d时实验组和对照组中神经元数目分别为36.50±1.78个/视野、19.70±3.71个/视野,神经元所占比例分别为(43.60±4.83)%、(26.97±4.90)%,两组间比较有显著性差异(P<0.05)。结论:自组装IKVAV多肽纳米支架能降低神经元的死亡率,并诱导轴突的发生和生长,具有支架及生物活性双重作用,可作为神经组织工程支架材料。

Objective：To observe the effects of self-assembled IKVAV peptide nanofibers on dorsal root ganglia neurons（DRGc） through co-culturing of DRGc and gel.Method：The peptide was dissolved in NaOH so- lution with pH value adjusted to 8.5 and the concentration of 0.01mg/ml,then the peptide was triggered to be self-organized into gel by the addition of DMEM/F12,which was detected under TEM.The DRGc suspension established by primary isolated culture methods,was divided into experimental group and control group.In experimental group（EG）：DRGc were transplanted on the surface of 2-D gels,while in control group,DRGc were transplanted on the surface of poly-lysine.Growth of DRGc was observed by IPCM.DRGc＇s survival time and the length of neurit in both groups were observed by cell counting and immunohistochemistry methods,and these datas were put into statistical analysis.Resuit：TEM showed that the self-assembled gel was composed of network nanofibers.The neurite length of DRGc at 1st day in EG and CG was （43.8±10.4）trm and （33.4± 5.75）μm respectively.The amount of neuron at 2st week for EG and CG were 36.50±1.78/EF and 19.70±3.71/EF respectively,with the percentage of （43.60±4.83）% and （39.04±3.87）% for EG and CG,which had significant difference （P〈0.05）.Conclusion：Self-assembled IKVAV peptide nanofibers,which can degrade the neuron mortality as well as promote neurite sprouting and growth,is a bioactivity scaffold,it can serve as a superordinary nerve tissue engineering scaffold.