iPSCs定向分化的内耳毛细胞与支持细胞间相互作用的研究

发布时间：2018-06-09 12:41

本文选题：耳聋 + 诱导多能性干细胞(iPSCs)　；参考：《中华耳科学杂志》2017年04期

【摘要】：目的利用诱导多能性干细胞定向分化的内耳毛细胞和支持细胞探究这两种体外诱导分化细胞之间的相互作用。方法首先,利用细胞单层贴壁两步诱导法将三株i PS细胞(野生株、MYO7A缺陷株、MYO7A校正株)向内耳祖细胞及内耳毛细胞诱导分化,探究i PSCs定向分化内耳毛细胞的过程中是否有支持细胞的产生;其次,通过细胞免疫化学的方法探究体外分化的内耳毛细胞和支持细胞间的相互作用;最后,将表达绿色荧光蛋白(EGFP)的上皮样内耳祖细胞以圆窗膜穿刺的方法移植到白化荣昌猪的内耳中观察分析移植细胞在体内的迁移、分化以及在体内形成的联系。结果三株i PS细胞诱导分化为内耳毛细胞的过程中均有一部分细胞分化为支持细胞;对分化细胞进行E-cadherin、N-cadherin和ZO-1的免疫荧光检测结果显示,E-cadherin、N-cadherin和ZO-1在支持细胞-支持细胞连接和支持细胞-毛细胞连接间都有表达;移植4周后,耳蜗免疫组织化学结果显示,三株不同来源的移植细胞均有少量细胞成功迁移到了毛细胞受损部位—柯底氏器,并表达毛细胞标志性蛋白MYO7A。移植细胞之间以及移植细胞与宿主细胞之间有E-cadherin、N-cadherin和ZO-1的表达。结论 i PSCs诱导分化的内耳毛细胞和支持细胞在体内、外均能形成钙粘连接和紧密连接。这些研究结果对毛细胞取代法治疗耳聋策略的完善与发展有一定的科学意义。
[Abstract]:Aim to explore the interaction between differentiated inner ear hair cells and Sertoli cells induced by pluripotent stem cells in vitro. Methods first, three iPS cells (wild strain MYO7A defective strain) were induced to differentiate into inner ear progenitor cells and inner ear hair cells by two steps of cell monolayer adherence. To explore whether there are Sertoli cells in the process of I PSCs directional differentiation of inner ear hair cells. Secondly, to explore the interaction between differentiated inner ear hair cells and Sertoli cells by means of cellular immunocytochemistry. The epithelioid inner ear progenitor cells expressing green fluorescent protein (EGFP) were transplanted into the inner ear of Rongchang albino pig by circular window membrane puncture. The migration, differentiation and association of the transplanted cells in vivo were observed and analyzed. Results some of the three iPS cells differentiated into Sertoli cells during the process of differentiation into inner ear hair cells. Immunofluorescence analysis of E-cadherin and ZO-1 showed that E-cadherin and ZO-1 expressed between Sertoli cell junctions and Sertoli cells junctions. A few of the three transplanted cells from different sources migrated successfully to the damaged part of the hair cell, the Cordy organ, and expressed the marker protein MYO7A of the hair cell. E-cadherin N-cadherin and ZO-1 were expressed between transplanted cells and host cells. Conclusion I PSCs induced differentiation of hair cells and Sertoli cells in vivo and in vitro can form cadherin connection and tight junction. These results are of scientific significance for the improvement and development of the strategy of hair cell replacement for deafness.
【作者单位】：浙江大学紫金港校区生命科学学院细胞与发育研究所;解放军总医院耳鼻咽喉头颈外科解放军耳鼻喉研究所;
【基金】：国家重大基础研究(973)项目,编号:2014CB541705~~
【分类号】：R764.43

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