中华耳鼻咽喉头颈外科杂志
中華耳鼻嚥喉頭頸外科雜誌
중화이비인후두경외과잡지
CHINESE JOURNAL OF OTORHINOLARYNGOLOGY HEAD AND NECK SURGERY
2014年
8期
680-686
,共7页
管利娜%陈燕红%朱恒涛%陈静%江红群
管利娜%陳燕紅%硃恆濤%陳靜%江紅群
관리나%진연홍%주항도%진정%강홍군
诱导性多潜能干细胞%Corti器%毛细胞%螺旋神经节%细胞分化%小鼠
誘導性多潛能榦細胞%Corti器%毛細胞%螺鏇神經節%細胞分化%小鼠
유도성다잠능간세포%Corti기%모세포%라선신경절%세포분화%소서
Induced pluripotent stem cells%Organ of Corti%Hair cells%Spiral ganglion%Cell differentiation%Mice
目的 探讨体外定向诱导小鼠诱导性多潜能干细胞(induced pluripotent stem cell,iPSC)向耳蜗毛细胞及螺旋神经节细胞分化的可行性.方法 建立小鼠iPSC培养体系并对iPSC加以鉴定.分离出生3d小鼠的耳蜗Corti器,将小鼠iPSC与耳蜗Corti器共培养.对产生的分化细胞用毛细胞标志物肌球蛋白Ⅶa(Myosin Ⅶa),Math1(碱性螺旋-环-螺旋转录因子家族成员),钙视网膜蛋白(Calretinin),Espin(F肌动蛋白的结合蛋白)进行免疫细胞化学鉴定.分离出生3d小鼠的耳蜗蜗轴,将小鼠iPSC与耳蜗蜗轴共培养,对分化细胞用神经元的标志物神经上皮干细胞蛋白(Nestin)、中等分子量神经丝蛋白(Neurofilament-M)、β-Ⅲ微管蛋白(β-ⅢTubulin),Ⅰ型囊泡膜谷氨酸转运体(vesicular glutamate transporter 1,VGluT1)进行免疫细胞化学鉴定.结果 免疫细胞化学染色结果显示,小鼠iPSC与耳蜗Corti器共培养18 d后可得到表达毛细胞标志蛋白MyosinⅦa、Math1、Calretinin 及Espin的细胞;小鼠iPSC与耳蜗蜗轴共培养18 d后可得到表达神经元标志蛋白Nestin、Neurofilament-M、β-ⅢTubulin,VGluT1的细胞.结论 共培养方法可成功将小鼠iPSC诱导分化为表达毛细胞及螺旋神经节细胞分子标志物的细胞.
目的 探討體外定嚮誘導小鼠誘導性多潛能榦細胞(induced pluripotent stem cell,iPSC)嚮耳蝸毛細胞及螺鏇神經節細胞分化的可行性.方法 建立小鼠iPSC培養體繫併對iPSC加以鑒定.分離齣生3d小鼠的耳蝸Corti器,將小鼠iPSC與耳蝸Corti器共培養.對產生的分化細胞用毛細胞標誌物肌毬蛋白Ⅶa(Myosin Ⅶa),Math1(堿性螺鏇-環-螺鏇轉錄因子傢族成員),鈣視網膜蛋白(Calretinin),Espin(F肌動蛋白的結閤蛋白)進行免疫細胞化學鑒定.分離齣生3d小鼠的耳蝸蝸軸,將小鼠iPSC與耳蝸蝸軸共培養,對分化細胞用神經元的標誌物神經上皮榦細胞蛋白(Nestin)、中等分子量神經絲蛋白(Neurofilament-M)、β-Ⅲ微管蛋白(β-ⅢTubulin),Ⅰ型囊泡膜穀氨痠轉運體(vesicular glutamate transporter 1,VGluT1)進行免疫細胞化學鑒定.結果 免疫細胞化學染色結果顯示,小鼠iPSC與耳蝸Corti器共培養18 d後可得到錶達毛細胞標誌蛋白MyosinⅦa、Math1、Calretinin 及Espin的細胞;小鼠iPSC與耳蝸蝸軸共培養18 d後可得到錶達神經元標誌蛋白Nestin、Neurofilament-M、β-ⅢTubulin,VGluT1的細胞.結論 共培養方法可成功將小鼠iPSC誘導分化為錶達毛細胞及螺鏇神經節細胞分子標誌物的細胞.
목적 탐토체외정향유도소서유도성다잠능간세포(induced pluripotent stem cell,iPSC)향이와모세포급라선신경절세포분화적가행성.방법 건립소서iPSC배양체계병대iPSC가이감정.분리출생3d소서적이와Corti기,장소서iPSC여이와Corti기공배양.대산생적분화세포용모세포표지물기구단백Ⅶa(Myosin Ⅶa),Math1(감성라선-배-라선전록인자가족성원),개시망막단백(Calretinin),Espin(F기동단백적결합단백)진행면역세포화학감정.분리출생3d소서적이와와축,장소서iPSC여이와와축공배양,대분화세포용신경원적표지물신경상피간세포단백(Nestin)、중등분자량신경사단백(Neurofilament-M)、β-Ⅲ미관단백(β-ⅢTubulin),Ⅰ형낭포막곡안산전운체(vesicular glutamate transporter 1,VGluT1)진행면역세포화학감정.결과 면역세포화학염색결과현시,소서iPSC여이와Corti기공배양18 d후가득도표체모세포표지단백MyosinⅦa、Math1、Calretinin 급Espin적세포;소서iPSC여이와와축공배양18 d후가득도표체신경원표지단백Nestin、Neurofilament-M、β-ⅢTubulin,VGluT1적세포.결론 공배양방법가성공장소서iPSC유도분화위표체모세포급라선신경절세포분자표지물적세포.
Objective In this study,we investigated the potential of mouse induced pluripotent stem cells(iPSC) for use as a source of transplants for the restoration of auditory hair cells and spiral ganglion neurons.Methods We co-cultured the mouse iPSC with the cells of the cochlear organ of Corti or the modiolus in vitro.The cochlear organ of Corti (which contains cochlear hair cells) and the modiolus (which contains auditory spiral ganglion neurons) were obtained from postnatal day 3 (P3) CD-1 ICR mice.After 18 days of coculture with the cells of newborn mouse cochleae.The expressions of hair cell markers (Myosin Ⅶ a,Math1,Calretinin,Espin) and Spiral ganglion neuron markers [Nestin,Neurofilament-M,β-Ⅲ Tubulin,Vesicular glutamate transporter 1 (VGluT1)] were detected by immunocytochemical analysis.Results Immunocytochemical analysis results indicated that the differentiated iPSC expressed auditory hair cell markers (Myosin Ⅶ a,Math1,Calretinin,Espin) and spiral ganglion markers (Nestin,Neurofilament-M,β-Ⅲ Tubulin,VGluT1).Conclusion Mouse iPSC in virto cultured could successfully be induced to differentiate into hair cell-like cells and spiral ganglion-like cells with hair cell and spiral ganglion molecular markers.
