腺病毒转染新生及成年鼠内耳细胞的研究

发布时间：2018-01-12 11:36

本文关键词：腺病毒转染新生及成年鼠内耳细胞的研究　出处：《中华细胞与干细胞杂志(电子版)》2016年05期 　论文类型：期刊论文

【摘要】：目的探讨腺病毒携带目的基因Cre和Oct4导入新生和成年小鼠转染内耳细胞的可行性,为内耳基因治疗提供可行性研究。方法采用纳升级显微操作系统,经耳蜗中阶显微注射携带基因编辑基因Cre和绿色荧光蛋白(GFP)基因的5型重组腺病毒悬液于新生鼠(P1)和成年鼠,并以腺病毒携带Oct4基因转染入成年鼠内耳,注射4d后取双侧耳蜗标本做基底膜铺片,观察GFP和Oct4表达情况。结果新生鼠组:术后第4d见耳蜗底圈62%和中圈54%支持细胞表达GFP。成年鼠组:术后第4天可见耳蜗底圈96%和中圈51%内毛细胞表达GFP,底圈72%和中圈40%支持细胞表达GFP。31%内毛细胞和43%支持细胞表达Oct4。未注射耳未见GFP和Oct4表达。结论显微注射腺病毒携带目的基因Cre和Oct4可高效导入新生鼠及成年鼠内耳并表达,可用来研究内耳基因功能和基因治疗。
[Abstract]:Objective to investigate the feasibility of transfection of adenovirus carrying target genes Cre and Oct4 into neonate and adult mouse inner ear cells and to provide a feasibility study for gene therapy of inner ear. Methods Nano-upgrade microoperating system was used. The recombinant adenovirus type 5 carrying the gene editing gene Cre and the green fluorescent protein (GFP) gene was injected into the cochlea by microinjection into the cochlea, and the recombinant adenovirus type 5 was suspended in the newborn mouse (P1) and the adult mouse. The adenovirus carrying Oct4 gene was transfected into the inner ear of adult rats and the bilateral cochlea specimens were taken for basement membrane preparation 4 days after injection. Results the expression of GFP and Oct4 was observed in neonatal rats on the 4th day after operation, cochlear basal ring 62% and middle ring 54% Sertoli cells were observed. On the 4th day after operation, GFP was expressed in the hair cells of cochlear basal ring 96% and middle circle 51%. Basal ring 72% and middle ring 40% Sertoli cells expressed GFP.31% inner hair cells and 43% Sertoli cells expressed Oct4.The expression of GFP and Oct4 was not found in ear injection. Conclusion Microinjection of adenovirus-carrying cells can not be seen. Cre and Oct4 with target genes can be efficiently introduced into the inner ear of newborn and adult rats and expressed. It can be used to study the gene function and gene therapy of inner ear.
【作者单位】：复旦大学附属眼耳鼻喉科医院耳鼻喉头颈外科国家计委听觉医学重点实验室;哈佛大学医学院麻省眼耳医院耳鼻喉头颈外科;
【基金】：国家自然科学基金(NSFC81300824) 上海市浦江人才资助项目(15PJ1401000)
【分类号】：R764
【正文快照】： 据2006?我国第?次残疾人抽样调查显示?我国听力残疾人达2 780万?并且还在不断增加[1]。耳聋可?致分为3种类型?传导性耳聋、感音神经性耳聋和混合性耳聋。感音神经性耳聋是由于耳蜗Corti氏器毛细胞、听神经、听传导径?或各级神经元受损害?致声音的感受与神经冲动传递障碍。因噪

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