毛囊来源的神经嵴干细胞修复大鼠坐骨神经缺损  

Transplanted epidermal neural crest stem cell in a peripheral nerve gap

作　　者：张路[1,2] 张洁元[3] 李兵仓[3] 刘争[4] 刘彬[1,2] 

机构地区：[1]西南大学生命科学学院,重庆400715 [2]淡水鱼类资源与生殖发育教育部重点实验室,重庆400715 [3]第三军医大学大坪医院野战外科研究所创伤,烧伤与复合伤国家重点实验室,重庆400042 [4]重庆医科大学生理学教研室神经科学研究中心,重庆400016

出　　处：《生物工程学报》2014年第4期605-614,共10页Chinese Journal of Biotechnology

基　　金：国家重点基础研究发展计划(973计划)(No.2012CB518106);创伤、烧伤与复合伤国家重点实验室Ⅰ类研究课题(No.SKLZZ201003);国家自然科学基金(Nos.31271036,81371341)资助~~

摘　　要：毛囊来源的神经嵴干细胞(Epidermal Neural Crest Stem Cell,EPI-NCSC)由于取材方便,具有多种分化潜能,是一种具有良好应用前景的组织工程种子细胞。目前,在神经损伤修复领域中,EPI-NCSC主要被应用于脊髓损伤的修复。为了探讨EPI-NCSC对周围神经缺损的修复作用,对原代培养的GFP-SD大鼠来源的EPI-NCSC的体外性质进行了考察,并以其为种子细胞,将其等量与细胞外基质(Extracellular matrix,ECM)混合后,预置入聚乳酸-聚羟基乙酸共聚物(Poly lactic acid co glycolic acid copolymer,PLGA)导管中,同时,以等量的达尔伯克(氏)改良伊格尔(氏)培养基(Dulbecco's Modified Eagle's medium,DMEM)代替EPI-NCSC作为对照,以用于修复大鼠坐骨神经10 mm距离的缺失。噻唑蓝(Methyl thiazolyl tetrazolium,MTT)比色分析结果显示,EPI-NCSC在PLGA膜上的初期粘附率为89.7%。在第1、3、5、7天细胞相对增殖率分别为89.3%、87.6%、85.6%和96.6%。细胞周期与DNA倍体分析表明,与PLGA共培养的EPI-NCSC与单独培养的EPI-NCSC相比较,二者的细胞周期变化趋势相同,增殖指数变化趋势也相同。在神经导管移植4周,术部实现了组织学水平的修复。大鼠手术一侧后肢感觉功能有所恢复,坐骨神经指数有所提高。研究结果表明,在PLGA导管中预置EPI-NCSC,有望实现较好的周围神经缺损的修复效果。Neural crest stem cells originated from hair follicle （epidermal neural crest stem cell, EPI-NCSC） are easy to obtain and have potentials to differentiate into various tissues, which make them eminent seed cells for tissue engineering. EPI-NCSC is now used to repair nerve injury, especially, the spinal cord injury, To investigate their effects on repairing peripheral nerve injury, EPI-NCSC from a GFP-SD rat were primarily cultured on coated dishes and on a poly lactic acid coglycolic acid copolymer （PLGA） membrane. Methyl thiazolyl tetrazolium （MTT） assay showed that the initial adhesion rate of EPI-NCSC was 89.7% on PLGA membrane, and the relative growth rates were 89.3%, 87.6%, 85.6%, and 96.6% on the 1st, 3rd, 5th, 7th day respectively. Cell cycles and DNA ploidy analysis demonstrated that cell cycles and proliferation indexes of cultured EPI-NCSC had the same variation pattern on coated dishes and PLGA membrane. Then cultured EPI-NCSC were mixed with equal amount of extracellular matrix and injected into a PLGA conduit to connect a 10 mm surgery excision gap of rat sciatic nerve, Dulbecco＇s Modified Eagle＇s medium（DMEM） was used to substitute EPI-NCSC in the control group, After four weeks of transplantation, the defected sciatic nerve achieved a histological restoration, the sensory function of rat hind limb was partly recovered and the sciatic nerve index was also improved. The above results showed that a PLGA conduit filled with EPI-NCSC has a good repair effect on the peripheral nerve injury.

关 键 词：表皮神经嵴干细胞 周围神经 细胞外基质(ECM) 聚乳酸-聚羟基乙酸共聚物(PLGA) 细胞周期 

分 类 号：R318[医药卫生—生物医学工程] Q813[医药卫生—基础医学]