Alt Rüdiger, Wilhelm Franziska, Pelz-Ackermann Oliver, Egger Dietmar, Niederwieser Dietger, Cross Michael
Division of Hematology and Oncology, University of Leipzig, Leipzig, Germany.
Exp Hematol. 2009 Feb;37(2):294-301. doi: 10.1016/j.exphem.2008.09.015. Epub 2008 Dec 19.
The ABCG2 transporter has been identified as a determinant of the side population (SP) in murine bone marrow and a potential marker for primitive stem cells. To assess the potential of the SP phenotype and ABCG2 expression to identify stem cells in human umbilical cord blood (hUCB), we have examined directly the relationship between SP; expression of ABCG2, CD133, and CD34; and hematopoietic potential in UCB samples.
Multicolor fluorescence activated cell sorting analysis was combined with the Hoechst SP procedure to allow the simultaneous detection of the SP phenotype together with surface markers in cells from fresh and cryopreserved UCB. Sorted populations were analyzed for cobblestone area-forming cell (CAFC) activity and by quantitative reverse transcriptase polymerase chain reaction for expression of mRNA from the ABC transporters ABCG2, MDR1, and MRP1. ABCG2(+) cells were enriched by magnetic-activated cell sorting for stringent analysis.
hUCB-derived SP cells were negative for ABCG2, but comprise approximately 20% CD133(+)/CD34(+) cells. Sorted SP cells from UCB were enriched 20-fold for week 13 CAFC activity, while magnetic-activated cell sorting-enriched ABCG2(+) cells retained no hematopoietic activity either in CAFC or liquid cultures. There was no significant difference in the SP frequency, immunophenotype, or CAFC potential of fresh and cryopreserved UCB. ABCG2 mRNA was not enriched in the SP and was specifically diminished ninefold in CD133 cells, which were eightfold enriched for MDR1 mRNA.
We find no evidence for an association of ABCG2 with SP activity or hematopoietic progenitor function in hUCB.
ABCG2转运蛋白已被确定为小鼠骨髓中侧群细胞（SP）的决定因素以及原始干细胞的潜在标志物。为了评估SP表型和ABCG2表达在鉴定人脐带血（hUCB）干细胞方面的潜力，我们直接研究了SP、ABCG2、CD133和CD34的表达与脐带血样本中造血潜力之间的关系。
将多色荧光激活细胞分选分析与Hoechst SP程序相结合，以便同时检测新鲜和冷冻保存的脐带血中细胞的SP表型和表面标志物。对分选的细胞群体进行鹅卵石区域形成细胞（CAFC）活性分析，并通过定量逆转录聚合酶链反应检测ABCG2、MDR1和MRP1等ABC转运蛋白的mRNA表达。通过磁激活细胞分选富集ABCG2(+)细胞以进行严格分析。
hUCB来源的SP细胞ABCG2呈阴性，但约20%为CD133(+)/CD34(+)细胞。从脐带血中分选的SP细胞在第13周CAFC活性方面富集了20倍，而磁激活细胞分选富集的ABCG2(+)细胞在CAFC或液体培养中均无造血活性。新鲜和冷冻保存的脐带血在SP频率、免疫表型或CAFC潜力方面没有显著差异。ABCG2 mRNA在SP中未富集，在CD133细胞中特异性减少了9倍，而CD133细胞中MDR1 mRNA富集了8倍。
我们没有发现ABCG2与hUCB中的SP活性或造血祖细胞功能相关的证据。
