诱导hUC-MSCs和VECs转化为角膜内皮样细胞的实验研究

发布时间：2018-01-21 00:36

  本文关键词： 脐带间充质干细胞 猪角膜基质后板层 纱布 冻存 血管内皮细胞 组织工程　出处：《暨南大学》2012年硕士论文　论文类型：学位论文

【摘要】：1.目的（1）提高体外分离hUC-MCs的成功率 （2）探讨hUC-MCs作为组织工程角膜种子细胞的可能性 （3）改良冻存VECs的方法 （4）探讨VECs作为组织工程角膜种子细胞的可能性 2.方法（1）用多片脐带组织片悬浮法联合Ⅳ型胶原酶消化法、单纯多片脐带组织片悬浮法及脐带组织片悬浮法这三种方法体外分离hUC-MCs，并比较其成功率，鉴定细胞表面标记并诱导其分化 （2）以猪角膜基质后板层为载体培养hUC-MCs构建组织工程角膜后板层并行动物移植，观察角膜透明情况，并行hUC-MCs表面特异内皮标志物NSE和ZO-1的检测 （3）采用改良及传统方法冻存VECs，4周后复苏分别比较其存活率、形态、生长曲线，流式细胞术检测其凋亡率 （4）以猪角膜基质后板层为载体培养VECs构建组织工程角膜后板层，扫描电镜观察其表面结构 3.结果（1）多片悬浮脐带片法联合Ⅳ型胶原酶消化法及单纯多片悬浮脐带片法成功率高于悬浮脐带片法 （2）种植hUC-MCs移植组角膜较非种植组透明，hUC-MCs表面特异内皮标志物NSE和ZO-1表达阳性 （3）改良的冻存法VECs复苏后形态上较传统组好，存活率、生长曲线及凋亡率与传统组无明显差异 (4）扫描电镜可见hUC-MCs在猪角膜基质后板层表面生长良好，，连接成片 4.结论（1）本研究探讨了hUC-MCs体外分离的几种方法，提供了一种较为高效的培养方法 （2）hUC-MCs可向CECs方向转化，为组织工程后板层的构建提供了新的种子细胞来源 （3）改良的VECs冻存法能大幅度缩短冻存所需时间达到与传统方法类似的效果 (4）VECs或能为组织工程后板层的构建提供了新的种子细胞来源
[Abstract]:1. Objective 1) to improve the success rate of hUC-MCs isolation in vitro. To explore the possibility of hUC-MCs as a seed cell of tissue engineering cornea Improved method of VECs cryopreservation To explore the possibility of VECs as a seed cell of tissue engineering cornea 2. Methods hUC-MCs was isolated in vitro by multi-cord tissue suspension method combined with collagenase IV digestion method, simple multi-cord tissue suspension method and umbilical cord tissue suspension method in vitro. Compared with the success rate, the cell surface markers were identified and induced to differentiate. (2) using porcine corneal stromal posterior lamina (hUC-MCs) as the carrier to construct tissue engineered corneal posterior lamina and animal transplantation, the corneal transparency was observed. Detection of NSE and ZO-1 on hUC-MCs Surface specific Endothelial markers The survival rate, morphology, growth curve and apoptosis rate of VECs were measured by flow cytometry after 4 weeks resuscitation. VECs was used as the carrier to construct the tissue engineered posterior lamina of cornea, and the surface structure was observed by scanning electron microscope (SEM). 3. Results: the success rate of multislice suspension cord method combined with type 鈪

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