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
2010年
7期
587-591
,共5页
雷霏%孙建军%刘阳%姜丹泉
雷霏%孫建軍%劉暘%薑丹泉
뢰비%손건군%류양%강단천
间质干细胞%成纤维细胞%鼓膜%细胞,培养的%大鼠
間質榦細胞%成纖維細胞%鼓膜%細胞,培養的%大鼠
간질간세포%성섬유세포%고막%세포,배양적%대서
Mesenchymal stem cells%Fibroblasts%Tympanic membrane%Cells,cultured%Rats
目的 观察大鼠脂肪干细胞(adipose-derived stem cell,ASC)对鼓膜成纤维细胞增殖、迁移的作用,初步探讨脂肪干细胞用于鼓膜损伤修复治疗的可行性及意义.方法 以Wistar大鼠为研究对象,分别分离、培养及鉴定ASC和鼓膜成纤维细胞.通过transwell共培养方法检测ASC对鼓膜成纤维细胞增殖的影响;通过transwell细胞迁移实验检测ASC对鼓膜成纤维细胞迁移速度的影响.激光共聚焦显微镜下评估各项检测指标.结果 大鼠ASC表面标志免疫荧光鉴定结果显示CD44、CD29表达阳性,CD34表达阴性;鼓膜成纤维细胞表面标志鉴定结果显示波形蛋白(vimentin)表达阳性.细胞增殖测量结果显示,与ASC共培养的鼓膜成纤维细胞增殖速度比对照组明显加快,细胞计数差异具有统计学意义(t=6.75,P=0.003).细胞迁移测量结果显示,在ASC条件培养基作用下,鼓膜成纤维细胞于1、2.5和4 h在Matrigel胶内位置距底部聚碳酸酯膜孔的平均距离比对照组均明显缩小,即迁移速度明显加快,差异具有统计学意义(P值均<0.05);5 h穿越至聚碳酸酯膜下表面的细胞数明显多于对照组,组间差异具有统计学意义(P<0.05).结论 ASC可能通过旁分泌作用促进鼓膜成纤维细胞增殖和迁移,将有利于鼓膜纤维层的修复.
目的 觀察大鼠脂肪榦細胞(adipose-derived stem cell,ASC)對鼓膜成纖維細胞增殖、遷移的作用,初步探討脂肪榦細胞用于鼓膜損傷脩複治療的可行性及意義.方法 以Wistar大鼠為研究對象,分彆分離、培養及鑒定ASC和鼓膜成纖維細胞.通過transwell共培養方法檢測ASC對鼓膜成纖維細胞增殖的影響;通過transwell細胞遷移實驗檢測ASC對鼓膜成纖維細胞遷移速度的影響.激光共聚焦顯微鏡下評估各項檢測指標.結果 大鼠ASC錶麵標誌免疫熒光鑒定結果顯示CD44、CD29錶達暘性,CD34錶達陰性;鼓膜成纖維細胞錶麵標誌鑒定結果顯示波形蛋白(vimentin)錶達暘性.細胞增殖測量結果顯示,與ASC共培養的鼓膜成纖維細胞增殖速度比對照組明顯加快,細胞計數差異具有統計學意義(t=6.75,P=0.003).細胞遷移測量結果顯示,在ASC條件培養基作用下,鼓膜成纖維細胞于1、2.5和4 h在Matrigel膠內位置距底部聚碳痠酯膜孔的平均距離比對照組均明顯縮小,即遷移速度明顯加快,差異具有統計學意義(P值均<0.05);5 h穿越至聚碳痠酯膜下錶麵的細胞數明顯多于對照組,組間差異具有統計學意義(P<0.05).結論 ASC可能通過徬分泌作用促進鼓膜成纖維細胞增殖和遷移,將有利于鼓膜纖維層的脩複.
목적 관찰대서지방간세포(adipose-derived stem cell,ASC)대고막성섬유세포증식、천이적작용,초보탐토지방간세포용우고막손상수복치료적가행성급의의.방법 이Wistar대서위연구대상,분별분리、배양급감정ASC화고막성섬유세포.통과transwell공배양방법검측ASC대고막성섬유세포증식적영향;통과transwell세포천이실험검측ASC대고막성섬유세포천이속도적영향.격광공취초현미경하평고각항검측지표.결과 대서ASC표면표지면역형광감정결과현시CD44、CD29표체양성,CD34표체음성;고막성섬유세포표면표지감정결과현시파형단백(vimentin)표체양성.세포증식측량결과현시,여ASC공배양적고막성섬유세포증식속도비대조조명현가쾌,세포계수차이구유통계학의의(t=6.75,P=0.003).세포천이측량결과현시,재ASC조건배양기작용하,고막성섬유세포우1、2.5화4 h재Matrigel효내위치거저부취탄산지막공적평균거리비대조조균명현축소,즉천이속도명현가쾌,차이구유통계학의의(P치균<0.05);5 h천월지취탄산지막하표면적세포수명현다우대조조,조간차이구유통계학의의(P<0.05).결론 ASC가능통과방분비작용촉진고막성섬유세포증식화천이,장유리우고막섬유층적수복.
Objective To explore the effects of rats adipose-derived stem cell (ASC) on the proliferation and migration of fibroblasts from the tympanic membrane, and to discuss the possibility and significance of therapy with ASC for tympanic membrane (TM) healing and regeneration. Methods Wistar rats were sacrificed, and then the isolation, culture and identification of both ASC and the TM fibroblasts were performed respectively. To verify the effect of ASC on fibroblasts proliferation, transwell coculture system was used. To examine the effect of ASC on fibroblasts migration, cell migration assay with transwell was also applied. All the data were analyzed under a confocal laser scan microscopy system.Results Immunofluorescence of cell surface markers indicated that rats ASC were positive for both of CD44 and CD29, but negative for CD34. The rat TM fibroblasts were positive for vimentin. The fibroblasts cocultured with ASC proliferated faster than the fibroblasts of control group, and the difference of the cell counting number between the two groups was significant ( t = 6. 75, P = 0. 003 ). Compared with the control group, the fibroblasts cultured with ASC conditioned culture medium migrated significantly faster, and the space between the fibroblasts and the polycarbonate membrane pore was significantly shortened at different time point ( 1, 2. 5 and 4 h, P ＜ 0. 05 ). The cell number of the fibroblasts that had migrated through the polycarbonate membrane had been significantly increased 5 h after migration ( P ＜ 0. 05 ). Conclusion ASC might promote TM fibroblasts proliferation and migration by paracrine activation, and it will faciliate the regeneration of TM fibrous layer.
