模拟脑组织生物力学环境下温敏脐带间充质干细胞分化特性探究.doc

模拟脑组织生物力学环境下温敏脐带间充质干细胞分化特性探究 　　[摘要] 目的 模?M脑组织弹性模量制备相应二维培养基，比较亚低温联合温敏脐带间充质干细胞（tsUC）与常温下脐带间充质干细胞（UC）的分化特性。 方法 应用单、双丙烯酰胺的聚合作用，制备弹性模量为0.5 kPa的聚丙烯酰胺（PA）水凝胶，用于模拟脑组织的生物力学环境，并测其弹性模量。从新生儿脐带中分离培养脐带间充质干细胞，通过感染携温度敏感型猿猴病毒40大T抗原（ts-SV40LT）基因的逆转录病毒来制备tsUC。实验分为3组：UC+常温+玻片组（A组）、UC+常温+0.5 kPa组（B组）、tsUC+亚低温+0.5 kPa组（C组）。动态观察各组细胞的生长情况和形态变化，并于7 d后行细胞免疫荧光检测各组细胞的分化水平并计算分化神经元的轴突长度。 结果 PA水凝胶弹性模量的检测结果为（0.50±0.03）kPa。B、C两组部分细胞出现细长的胞突，并存在β-tubulin Ⅲ阳性细胞，A组细胞镜下无明显神经元形态，也无β-tubulin Ⅲ阳性表达。B、C两组的神经元分化率以及荧光下轴突长度均明显高于A组，但组间差异无统计学意义（P 0.05）。 结论 在模拟脑组织弹性模量的环境中，tsUC具有向神经元分化的能力，可应用于亚低温治疗下脑损伤修复的细胞移植研究 [关键词] 温敏脐带间充质干细胞；聚丙烯酰胺水凝胶；弹性模量；生物力学 [中图分类号] R651.15 [文献标识码] A [文章编号] 1673-7210（2016）12（c）-0014-04 [Abstract] Objective To manufacture two-dimensional culture medium of brain tissue elastic modulus， and to compare the differentiate property of temperature-sensitive umbilical cord mesenchymal stem cells （tsUC） under mild hypothermia treatment （MHT） and umbilical cord mesenchymal stem cells （UC） under normal temperature （NT）. Methods Acrylamide and bis-acrylamide were applied to synthesise polyacrylamide hydrogel of 0.5 kPa in elasticity modulus. UC were isolated from umbilical cord of the newborn， and then the tsUC were obtained through the infection of temperature-sensitive Simian 40 Large T-antigen （ts-SV40LT） retrovirus. There were three experiment groups in this study， including UC+NT+glass （group A）， UC+NT+0.5 kPa PA （group B）， and tsUC+MHT+0.5 kPa PA （group C）. Cell growth and morphological changes in each group were given dynamic observation. 7 days later， cell immunofluorescence were implemented， with the differentiation level of each group estimated and the axon length of the differentiated neurons calculated by using Image J software. Results The elasticity modulus of PA gels was （0.50±0.03） kPa. A small amount of cells resembling the neurons were presented in group B and C， with small soma and long， slender protuberances. Cell immunofluorescence statistics demonstrated that the cells diffe