搭载紫杉醇脂质体的聚乳酸-羟基乙酸共聚物静电纺丝膜对神经干细胞增殖分化的影响

目的：探讨搭载不同浓度紫杉醇脂质体的聚乳酸-羟基乙酸共聚物（PLGA）静电纺丝膜对神经干细胞（NSCs）增殖和分化的影响，评价载有紫杉醇脂质体的PLGA静电纺丝膜在脊髓损伤组织工程修复中应用的可行性。方法：将紫杉醇脂质体和PLGA以不同比例混合，利用静电纺丝技术分别构建1、5和10 μg·L^-1紫杉醇脂质体PLGA静电纺丝膜。从胎鼠大脑组织中分离纯化NSCs。扫描电镜（SEM）检测各组紫杉醇脂质体PLGA静电纺丝膜纤维直径，免疫荧光染色鉴定分离纯化所得的NSCs。将NSCs分别培养在含有不同浓度（0、1、5和10 μg·L^-1）紫杉醇脂质体PLGA静电纺丝膜的培养基中（作为纯PLGA静电纺丝膜组和1、5及10 μg·L^-1紫杉醇脂质体PLGA静电纺丝膜组），MTT法检测各组NSCs增殖活性，RT-PCR法检测各组NSCs中Tuj-1和GFAP mRNA表达水平。结果：紫杉醇脂质体PLGA静电纺丝膜呈白色薄膜状。SEM检测，各组紫杉醇脂质体PLGA静电纺丝膜纤维直径比较差异无统计学意义（P>0.05）。免疫荧光染色，从胎鼠脑组织中分离出的细胞呈球状，所有细胞均有Nestin蛋白阳性表达。MTT法检测，5 μg·L^-1紫杉醇脂质体PLGA静电纺丝膜组NSCs增殖活性高于其他3组（P<0.05）。RT-PCR检测，与纯PLGA静电纺丝膜组比较，5 μg·L^-1紫杉醇脂质体PLGA静电纺丝膜组NSCs中Tuj-1 mRNA表达水平升高（P<0.05），GFAP mRNA表达水平降低（P<0.05）。结论：中等浓度紫杉醇能够促进NSCs增殖，诱导NSCs向神经元分化。载有中等浓度紫杉醇脂质体的PLGA静电纺丝膜在脊髓损伤修复中有一定的应用价值。

Effect of PLGA electrospinning membrane carrying paclitaxel-liposomes on proliferation and differentiation of neural stem cells

Objective: To investigate the effects of polyactic-co-glycolic(PLGA) electrospinning membrane carrying paclitaxel-liposomes with different concentrations on the proliferation and differentiation of the neural stem cells (NSCs), and to evaluate the feasibility of PLGA electrospinning membrane carrying paclitaxel liposomes in the tissue engineering repair of spinal cord injury. Methods: The paclitaxel-liposomes and PLGA were mixed in different proportions and the electrospinning technique was used to construct 1, 5 and 10μg·L^-1electrospinning membrane carrying paclitaxel-liposomes. The NSCs were isolated and purified from the fetal rat brain tissue. The diameters of PLGA electrospinning membrane carrying paclitaxel-liposomes in various groups were detected by scanning electron microscope (SEM), and the isolated and purified NSCs were identified by immunofluorescence staining. The NSCs were cultured in PLGA electrospinning membrane carrying paclitaxel-liposomes with different concentrations (0,1,5 and 10μ g·L^-1)as PLGA electrospinning membrane group and PLGA electrospinning membrane carrying 1,5 and 10μ g·L^-1paclitaxel-liposomes groups. The proliferation levels of the NSCs in various groups was detected by MTT method, and the expression levels of Tuj-1 and GFAP mRNA in the NSCs in various groups were detected by RT-PCR method. Results: The PLGA electrospinning membrane carrying paclitaxel-liposomes were white thin films. The SEM results showed that there was no statistical differences in the fiber diameters of PLGA electrospinning membrane carrying paclitaxel-liposomes between various groups (P>0.05).The immunofluorescence staining results showed that the cells isolated from fetal rat brain tissue were spherical and positive expression of Nestin protein. The MTT assay results showed that the proliferation level of NSCs in PLGA electrospinning membrane carrying 5μg·L^-1paclitaxel-liposomes group was higher than those in the other three groups(P<0.05).The RT-PCR results showed that compared with PLGA electrospinning membrane group, the expression level of Tuj-1 mRNA in the NSCs in PLGA electrospinning membrane carrying 5μg·L^-1 paclitaxel-liposomes group was increased(P<0.05) and the expression level of GFAP mRNA in the NSCs in PLGA electrospinning membrane carrying 5μg·L^-1 paclitaxel-liposomes group was decreased(P<0.05). Conclusion: Medium concentration of paclitaxel can promote the proliferation of NSCs and induce the differentiation of NSCs into the neurons. PLGA electrospinning membrane carrying medium concentration of paclitaxel-liposomes has certain application value in the repair of spinal cord injury.