脐血干/祖细胞短期体外扩增后归巢相关粘附特性的研究

Studies on the homing-related adhesion activities of UCB HSPC in short-term ex vivo expansion

目的 探讨体外扩增对脐血 (UCB)造血干 祖细胞 (HSPC)原有粘附功能的影响。方法 将从新鲜UCB标本中纯化的CD34+ 细胞接种于无血清、无基质的悬浮扩增体系 ,分别于培养第 7天、第10天和第 14天从扩增产物中再次纯化CD34+ 细胞 ,比较扩增前后CD34+ 细胞表面VLA 4 (CD4 9d)、VLA 5(CD4 9e)、LFA 1(CD1 1a)、L selectin (CD6 2L)、PECAM 1(CD31 )、ICAM 1(CD54)和HCAM(CD4 4)等归巢相关粘附分子 (CAMs)的表达情况 ,并检测CD34+ 细胞与纤连蛋白 (FN)间的自发粘附率和基质细胞来源因子 1(SDF 1)诱导粘附率。结果 ①在第 14天的培养扩增中 ,表达上述CAMs的各CD34+ 细胞亚群均有不同程度 (15～ 72倍 )的扩增 ;②扩增后CD34+ 细胞表面粘附分子CD4 4、CD1 1a、CD4 9e和CD4 9d的表达与原代CD34+ 细胞持平或高于培养开始时水平 ,而CD6 2L、CD54和CD31 的表达则有不同程度下调 ;③在前 10d的扩增中 ,CD34+ 细胞与FN间的自发粘附率和SDF 1诱导粘附率均呈上升趋势。结论 所建立的短期培养体系不仅可以支持表达重要CAMs的UCBCD34+ 细胞亚群的有效扩增 ,而且扩增后的HSPC总体上保持原有的粘附功能。

Objective To study the effect of ex vivo expansion on the adhesion activities of umbilical cord blood hematopoietic stem and progenitor cells (HSPC). Methods Fresh UCB CD 34 + cells were cultured in a serum and stroma free culture system. At day 7, day 10 and day 14, CD 34 + cells were re selected from the expanded products. The expression of adhesion molecules (CAMs) such as VLA 4, VLA 5, LFA 1, ICAM 1, HCAM, L selectin and PECAM 1, and the adhesion activity of the expanded CD 34 + cells were evaluated and compared with those of precultured fresh CD 34 + cells. Results ① The CD 34 + cells expressing homing related CAMs were increased (from 15 fold increase for CD 34 + CD 54 + subset to 72 fold increase for CD 34 + CD 49e + subset at day 14). ②The expressions of CD 49d , CD 44 , CD 11a and CD 49e on the expanded CD 34 + cells were increased or sustained the same levels as those on fresh UCB CD 34 + cells, while the expression of CD 62L , CD 54 and CD 31 on expanded CD 34 + cells declined with the cultivating. ③Spontaneous adhesion and SDF 1 induced adhesion tended to be increased in the course of the first 10 day's culture. Conclusions The culture system used in this study could substantially support the expansion of HSPCs expressing the above CAMs, and the expanded HSPCs would sustain their intrinsic adhesion potentials.