人AGM区基质细胞对脐血LTC推荐.doc

人AGM区基质细胞对脐血LTC 人AGM区基质细胞对脐血LTC 【摘要】 为探讨人主动脉-性腺-中肾（AGM）区基质细胞对脐血长期培养启动细胞（LTC-IC）的造血支持作用，建立了人AGM区基质细胞与脐血CD34+细胞体外长期共培养体系。采用免疫磁珠方法分离人脐血CD34+细胞，接种于已制备好人AGM区基质细胞（hAGMS1-S5）饲养层的24孔板中共培养，同时设无饲养层组作为对照，分别于共培养 5、6、7、8周时收获细胞行造血细胞集落培养，并采用极限稀释法（LDA）检测与人AGM区基质细胞共培养后的脐血CD34＋ 细胞的LTC-IC含量。结果表明，无饲养层的对照组培养5周后不再产生造血细胞集落，而以hAGMS1-S5作为饲养层，共培养5周后造血细胞仍具有集落形成能力。hAGMS1-S5维持LTC-IC的作用组间比较有显著性差异（P 0.05），其中以hAGMS3与S4组支持作用最强。LDA检测结果显示，hAGMS3和S4维持LTC-IC的能力无显著性差异（P 0.05）；与hAGMS3和S4共培养14天后脐血CD34＋ 细胞中的LTC-IC扩增，分别是达到ere plated on 24-ell plates as feeder cells. CD34+ cells ere positively selected from human umbilical cord blood through immunomagnetic bead selection method，seeded on the feeder cells，and co-cultured for 8 eeks. The hematopoietic cells ere collected at 5，6，7 and 8 eeks for CFC analysis. Frequencies of LTC-IC in umbilical cord blood CD34+ cells after co-culture ith AGM stromal cells ere detected through limiting dilute analysis. The results shoed that there as no any hematopoietic CFC in the feeder cell-free culture system after 5 eeks of co-culture. Hoever，in AGM feeder cells culture systems，there ere still CFCs after 5 eeks of co-culture，hich indicated that human AGM stromal cells could maintain LTC-IC in vitro. In groups of hAGM feeders，hAGMS3 and S4 had better supportive effects than other AGM groups . The absolute number of LTC-IC in hAGM S3 and S4 culture systems got expansion up to % and % respectively ithout significant difference beteen hAGMS3 and S4 . It is concluded that human AGM stromal cells S1-S5 support the maintenance of umbilical blood LTC-IC in vitro，hile hAGMS3 and S4 cells have better effects on maintaining LTC-IC and expansion of LTC-IC. Key ords AGM region；stromal cell；LTC-IC；cord blood 主动脉-性腺-中肾（aorta-gonad-mesonephros，AGM）区是胚胎期永久造血的起源部位，正成为国际上造血发生、分化机理研究中的热点。AGM区造血微环境对多能造血干细胞的发生及发育起重要作用。基质细胞是造血微环境的主要成分，通过研究基质细胞的体外造血支持功能是分析造血微环境的重要途径。本研究探讨人AGM区基质细胞对人脐血来源的长期培养启动细胞（long-term cultur-initiating cell，LTC-IC）的造血支持作用，为阐明AGM 区基质细胞在胚胎造血发生中的作用提供理论和实验依据。 材料和方法 主要