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

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。

Abstract:

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.