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
2011年
4期
288-292
,共5页
王家佳%刘天津%魏春生%李诗敏
王傢佳%劉天津%魏春生%李詩敏
왕가가%류천진%위춘생%리시민
声带%创伤和损伤%羊膜%上皮细胞%移植,异种%细胞移植%兔
聲帶%創傷和損傷%羊膜%上皮細胞%移植,異種%細胞移植%兔
성대%창상화손상%양막%상피세포%이식,이충%세포이식%토
Vocal cords%Wounds and injuries%Amnion%Epithelial cells%Transplantation,heterologous%Cell transplantation%Rabbits
目的 研究人羊膜上皮细胞(human amniotic epithelial cells,hAEC)移植入兔声带损伤组织内的生长分布特点,探索hAEC促进声带损伤后修复再生的潜能.方法 分离和培养hAEC,慢病毒增强型绿色荧光蛋白(1entivirus enhanced green fluorescent protein,Lenti-EGFP)基因转染作标记.建立深及声韧带的兔声带损伤模型,设hAEC移植组(13侧声带)、损伤对照组(13侧声带)及正常对照组(4侧声带).荧光显微镜下连续观察hAEC在声带内的存活、分布情况,应用HE染色和免疫组化染色分析胶原、纤维连接蛋白等主要细胞外基质在损伤后3个月时的含量及分布.结果 hAEC原代培养6 d后呈铺路石样生长,植入声带损伤组织后可在固有层内持续存活2个月,细胞呈纵向排列,有趋向性.声带损伤2个月时免疫荧光显示hAEC移植组兔肌细胞标志结蛋白荧光阳性,提示hAEC向肌细胞分化;同时Ⅲ型胶原荧光阳性,提示hAEC植入后具有分泌Ⅲ型胶原功能.光镜观察见hAEC移植后3个月兔胶原纤维密度和排序较损伤对照组改善,但未及正常;免疫组化染色示hAEC移植组纤维连接蛋白含量和分布介于损伤对照组和正常对照组之间.结论 hAEC可在异种动物声带损伤组织内持续存活、生长,并有向声带组织分化和分泌部分细胞外基质的潜能,可能促进声带损伤后的修复再生.
目的 研究人羊膜上皮細胞(human amniotic epithelial cells,hAEC)移植入兔聲帶損傷組織內的生長分佈特點,探索hAEC促進聲帶損傷後脩複再生的潛能.方法 分離和培養hAEC,慢病毒增彊型綠色熒光蛋白(1entivirus enhanced green fluorescent protein,Lenti-EGFP)基因轉染作標記.建立深及聲韌帶的兔聲帶損傷模型,設hAEC移植組(13側聲帶)、損傷對照組(13側聲帶)及正常對照組(4側聲帶).熒光顯微鏡下連續觀察hAEC在聲帶內的存活、分佈情況,應用HE染色和免疫組化染色分析膠原、纖維連接蛋白等主要細胞外基質在損傷後3箇月時的含量及分佈.結果 hAEC原代培養6 d後呈鋪路石樣生長,植入聲帶損傷組織後可在固有層內持續存活2箇月,細胞呈縱嚮排列,有趨嚮性.聲帶損傷2箇月時免疫熒光顯示hAEC移植組兔肌細胞標誌結蛋白熒光暘性,提示hAEC嚮肌細胞分化;同時Ⅲ型膠原熒光暘性,提示hAEC植入後具有分泌Ⅲ型膠原功能.光鏡觀察見hAEC移植後3箇月兔膠原纖維密度和排序較損傷對照組改善,但未及正常;免疫組化染色示hAEC移植組纖維連接蛋白含量和分佈介于損傷對照組和正常對照組之間.結論 hAEC可在異種動物聲帶損傷組織內持續存活、生長,併有嚮聲帶組織分化和分泌部分細胞外基質的潛能,可能促進聲帶損傷後的脩複再生.
목적 연구인양막상피세포(human amniotic epithelial cells,hAEC)이식입토성대손상조직내적생장분포특점,탐색hAEC촉진성대손상후수복재생적잠능.방법 분리화배양hAEC,만병독증강형록색형광단백(1entivirus enhanced green fluorescent protein,Lenti-EGFP)기인전염작표기.건립심급성인대적토성대손상모형,설hAEC이식조(13측성대)、손상대조조(13측성대)급정상대조조(4측성대).형광현미경하련속관찰hAEC재성대내적존활、분포정황,응용HE염색화면역조화염색분석효원、섬유련접단백등주요세포외기질재손상후3개월시적함량급분포.결과 hAEC원대배양6 d후정포로석양생장,식입성대손상조직후가재고유층내지속존활2개월,세포정종향배렬,유추향성.성대손상2개월시면역형광현시hAEC이식조토기세포표지결단백형광양성,제시hAEC향기세포분화;동시Ⅲ형효원형광양성,제시hAEC식입후구유분비Ⅲ형효원공능.광경관찰견hAEC이식후3개월토효원섬유밀도화배서교손상대조조개선,단미급정상;면역조화염색시hAEC이식조섬유련접단백함량화분포개우손상대조조화정상대조조지간.결론 hAEC가재이충동물성대손상조직내지속존활、생장,병유향성대조직분화화분비부분세포외기질적잠능,가능촉진성대손상후적수복재생.
Objective To investigate the survival, growth and distribution of human amniotic epithelial cells (hAEC) after injected into injured rabbit vocal folds, in addition, to assess the ability of hAEC to affect the components of lamina propria extracellular matrix (ECM) and prevent vocal fold scarring.Methods hAEC were isolated from human amnion and marked by Lenti-EGFP. Fifteen New Zealand rabbits were used for this experiment. EGFP-hAEC was injected into the left injured vocal folds in thirteen rabbits, and the contralateral thirteen vocal folds experienced an injured procedure only (" injured untreated control"), and four vocal folds were left as untreated controls. The survival, distribution, differentiation potential and secretion function of hAEC were examined by immunofluorescence method. HE staining and immunohistochemical staining were performed for the evaluation of collagen and fibronectin respectively.Results hAEC showed a cobblestone-like growth. After implanted into the injured vocal folds, hAEC could survive in vocal fold lamina propria for 2 months. The immunofluorescence analysis showed the evidence of hAEC differentiation into muscle cells as well as secretion the ECM protein. Three months postoperatively, the density of collagen was higher in the injured untreated control folds than that in the injured vocal folds injected with hAEC and the untreated controls. Besides, the content of fibronectin in the injured untreated control group was significantly increased. Conclusions hAEC survived in the vocal folds lamina propria,and had the potentiality to differentiate into vocal folds tissue and secret some ECM components. The histological improvement caused by the injected cells demonstrate that hAEC had the ability to promote the repairment and regeneration of injured vocal folds.
