CAJ | 학술논문

背景:羊水干细胞的分离培养及生物学特性相关研究取得了明显进展,但未见45,X/46,XX(Turner综合征)羊水细胞建系的报道.目的:建立体外培养人羊水来源干细胞的方法,初步探讨羊水干细胞的生物学特性.方法:孕中期羊膜腔穿刺获得1例染色体核型异常(45,X/46,XX)羊水标本,采用梯度稀释后低密度种植的方法获得羊水干细胞,体外传代培养,倒置显微镜观察细胞的贴壁生长情况,细胞染色体检查确认核型,流式细胞仪检测羊水干细胞特异性表面标志和细胞周期,进行成骨诱导分化及碱性磷酸酶染色和茜素红染色鉴定.结果与结论:羊水干细胞在体外培养体系中增殖迅速,核型为45,X/46,XX,流式细胞仪检测羊水干细胞表达CD29、CD44、CD90和CD105间充质干细胞表面标志,不表达造血干细胞标志CD45和CD34;大部分细胞处于 G1期,增殖能力强.羊水干细胞经成骨诱导分化后,碱性磷酸酶染色和茜素红染色阳性.结果可见该实验成功分离获得45,X/46,XX (Turner 综合征)羊水干细胞,其增殖能力强,表现间充质干细胞的特性.
배경:양수간세포적분리배양급생물학특성상관연구취득료명현진전,단미견45,X/46,XX(Turner종합정)양수세포건계적보도.목적:건입체외배양인양수래원간세포적방법,초보탐토양수간세포적생물학특성.방법:잉중기양막강천자획득1례염색체핵형이상(45,X/46,XX)양수표본,채용제도희석후저밀도충식적방법획득양수간세포,체외전대배양,도치현미경관찰세포적첩벽생장정황,세포염색체검사학인핵형,류식세포의검측양수간세포특이성표면표지화세포주기,진행성골유도분화급감성린산매염색화천소홍염색감정.결과여결론:양수간세포재체외배양체계중증식신속,핵형위45,X/46,XX,류식세포의검측양수간세포표체CD29、CD44、CD90화CD105간충질간세포표면표지,불표체조혈간세포표지CD45화CD34;대부분세포처우 G1기,증식능력강.양수간세포경성골유도분화후,감성린산매염색화천소홍염색양성.결과가견해실험성공분리획득45,X/46,XX (Turner 종합정)양수간세포,기증식능력강,표현간충질간세포적특성.
@@@@BACKGROUND: Great progress has been achieved on studies on isolation, culture and biological characteristics of amniotic fluid-derived mesenchymal stem cel s. However, few studies are reported on amniotic fluid-derived mesenchymal stem cel s in 45, X/46, XX (Turner’s syndrome). OBJECTIVE: To develop a simple culture protocol to isolate amniotic fluid-derived mesenchymal stem cel s and investigate the biological characteristics of amniotic fluid-derived mesenchymal stem cel s. METHODS: We developed a gradient dilution culture protocol to isolate a population of 45, X/46, XX (Turner’s syndrome) amniotic fluid-derived mesenchymal stem cel s from second-trimester amniocentesis. The morphology of amniotic fluid-derived mesenchymal stem cel s was observed by light microscope. The karyotype was analyzed. Specific cel surface antigens and cel cycle of the clonal amniotic fluid-derived mesenchymal stem cel s at passage 4 were characterized by flow cytometry. Osteogenic differentiation of amniotic fluid-derived mesenchymal stem cel s was confirmed by alkaline phosphatase staining and alizarin red staining. RESULTS AND CONCLUSION: The cultured human amniotic fluid-derived mesenchymal stem cel s proliferated rapidly after passage. Karyotype mapping showed abnormal female chromosome type with 45, X/46, XX observed. The amniotic fluid-derived mesenchymal stem cel s had an immunophenotype similar to that of common mesenchymal stem cel s and were positive for CD29, CD44, CD90 and CD105, but negative for CD34 and CD45. The cel cycle measurement showed that amniotic fluid-derived mesenchymal stem cel s cultured in vitro could maintain strong proliferation ability. Alkaline phosphatase staining and alizarin red staining results confirmed that amniotic fluid-derived mesenchymal stem cel s could be successful y induced to differentiate into osteocytes under specific culture media. These results demonstrated that amniotic fluid-derived mesenchymal stem cel s in 45, X/46, XX (Turner’s syndrome) were successful y isolated and the cel s had a great potential of proliferation and showed the characteristics of mesenchymal stem cel s.