摘要
目的制备Tenascin-R（TN．R）蛋白的EGF-L功能片段及该片段抗血清，联合TN-R蛋白研究其在体外对大鼠皮层神经元的作用，探讨该功能片段抗血清用于治疗中枢神经系统（CNSl损伤后轴突再生的可行性。方法制备人TN-R蛋白EGF-L片段抗血清，再以该抗血清联合TN-R蛋白包被多孔板形成不同的铺板状态，分EGF．L抗血清组、TN-R蛋白组、TN-R蛋白＋EGF-L抗血清组，仅包被有多聚赖氨酸（PLL）处为对照组。单细胞悬液法原代培养大鼠皮层神经元24h后免疫荧光染色检测β-Ⅲ-tubulin，比较不同的铺板状态下所黏附的细胞数；7d后免疫荧光染色检测β-Ⅲ-tubulin，观察神经元的形态并比较神经元突起主干、分支点数量及突起总长度。小组织块法培养大鼠皮层神经元7d后观察包被的蛋白与PLL之间形成的边界对神经元突起生长的影响。结果成功制备高浓度的TN-R蛋白EGF-L片段抗血清。4组所黏附的细胞密度不同，比较差异有统计学意义（P〈0．05）。与EGF-L抗血清组和对照组比较，TN-R蛋白组所黏附的细胞密度、神经元的突起主干、分支点数量及突起总长度较低，TN-R蛋白＋EGF-L抗血清组上述指标均较高，差异有统计学意义（P〈0．05）；TN-R蛋白组从组织块中迁徙出的细胞及细胞突起均几乎不能越过边界，TN-R蛋白＋EGF-L抗血清组可见有少量的细胞突起越过边界生长。结论TN-R蛋白EGF-L片段抗血清在体外能削弱TN-R蛋白对于神经元生长的抑制功能，有望将该抗血清用于体内，为CNS损伤后轴突再生创造有利的内环境。
Objective To study the effect of the EGF-L functional region containing human TN-R protein in prokaryocytes and anti-EGF-L serum on the cortical neurons in vitro in rats. Methods The anti-EGF-L serum was obtained from the rabbits immunized with the protein of EGF-L, then combine with TN-R coat petri dish to prepare various culture substrate. The effect on adhere, migrate and neurite outgrowth of neuron by differ culture substrate was observed. Results High concentration of antiserum against protein of EGF-Lwas successfully obtained f^om the rabbits, and it has no effect on the neuron when coated on petri dish as culture substrate with PLL in vitro, but can add the adhesive to the neuron, partly neutralize the inhibitory on the neuron neurite of TN-R. The antiserum can also allow the neuron with its neurite to migrate to the area coated with TN-R. Conclusion The anti-EGF-L serum can weaken the inhibitory on the neuron of TN-R in vitro, the function in vivo need further research,
出处
《中华神经医学杂志》
CAS
CSCD
北大核心
2012年第3期217-221,共5页
Chinese Journal of Neuromedicine
基金
国家自然科学基金一广东省联合基金（U0632008）
国家临床重点专科建设项目
