《中国康复理论与实践》 ›› 2010, Vol. 16 ›› Issue (4): 335-338.

• 论文 • 上一篇    下一篇

脂肪干细胞与温敏型支架构建人工髓核的体外研究

文天用1,2,李放2,叶超群2,刘连江1,2   

  1. 1.第四军医大学,陕西西安市 710032;2.北京军区总医院骨科,北京市 100700。
  • 收稿日期:2010-03-09 修回日期:1900-01-01 出版日期:2010-04-25 发布日期:2010-04-25
  • 通讯作者: 李放

Construction of Tissue Engineered Nucleus Pulposus with Adipose-derived Stromal Cells and Thermo-Sensitive Scaffold

WEN Tian-yong, LI Fang, YE Chao-qun, et al.   

  1. the Forth Military Medical University, Xi'an 710032, Shaanxi, China
  • Received:2010-03-09 Revised:1900-01-01 Published:2010-04-25 Online:2010-04-25

摘要: 目的评测壳聚糖基温敏型支架与诱导后脂肪干细胞的相容性及对其功能状态的影响。方法将兔脂肪干细胞体外培养3代、诱导2周,RT-PCR检测细胞内RNA表达水平,证明向髓核分化后,与制备好的支架复合,37 ℃成胶后,培养2周,行AO/PI检测细胞存活情况,并以扫描电镜观察超微结构。结果脂肪干细胞在单层诱导培养后有Ⅱ型胶原(Col Ⅱ)和蛋白聚糖(Aggrecan)表达;与支架复合后,细胞90%以上存活,复合培养2周后无去分化现象,且Col Ⅱ、Aggrecan mRNA表达水平升高(P<0-05)。结论温敏型壳聚糖支架与诱导后脂肪干细胞相容性良好,一定程度上利于干细胞表型、功能的维持与进一步分化,可作为干细胞植入的可注射性载体材料。

关键词: 组织工程, 椎间盘, 脂肪干细胞, 可注射支架, 温敏型

Abstract: ObjectiveTo evaluate the feasibility of using injective chitosan scaffold and induced- adipose-derived stromal cells(ADSCs) to construct tissue engineered injectable nucleus pulposus (NP).MethodsADSCs were harvested from rabbits to culture 3 passage and induce 2 weeks to NP-like cells. The injective chitosan hydrogel scaffold was made of chitosan and disodium β-glycerophosphate. Its physical properties and gross condition were observed. The tissue engineered NP was constructed by compounding the scaffold and induced-ADSCs. Then, the viability of ADSCs in the scaffold was observed 2 days after compound culture and the growth condition of ADSCs on the scaffold was observed by scanning electron microscope (SEM) 14 days after compound culture. Expression of aggrecan and Col Ⅱ mRNA in ADSCs were analyzed by RT-PCR 14 days after inductive culture and compound culture.ResultsThe injective chitosan hydrogel was liquid at room temperature and solidified into gel at 37 ℃ (10~15 minutes) due to crosslinking reaction. Acridine orange/propidiumiodide staining showed that the viability rate of induced-ADSCs in chitosan scaffoldl was above 90%. Scanning electron microscope observation demonstrated that the ADSCs were distributed in the reticulate scaffold. RT-PCR results showed that the expression of Col Ⅱ and aggrecan mRNA in induced-ADSCs demonstrated differentiation of ADSCs to a phenotype which showed similarities to NP cells, and the co-culture NP-like cells with scaffold didn`t cause dedifferentiation.ConclusionWith good cellular compatibilities, C/Gp scaffold may be a potential NP cells carrier for tissue engineered NP.

Key words: tissue engineere, intervertebral disc, adipose-derived stromal cells(ADSCs), injective scaffold, thermo-sensitive