Anthony R S, McKelvie N D, Cunningham A J, Craig J I, Rogers S Y, Parker A C
University of Edinburgh, John Hughes Bennett Laboratory, Western General Hospital, UK.
Bone Marrow Transplant. 1998 Mar;21(5):441-6. doi: 10.1038/sj.bmt.1701134.
Quantifying progenitor cells in peripheral blood stem cell (PBSC) harvests by flow cytometric enumeration of CD34+ cells does not account for cell viability. Cell membrane asymmetry in early apoptosis exposes phosphatidylserine on the cell surface. This can be detected by staining with annexin V FITC. Apoptosis in 30 autologous PBSC harvests mobilised by cyclophosphamide + G-CSF or standard chemotherapy + G-CSF was analysed immediately after collection by dual-colour flow cytometry with CD34 PE and annexin V FITC. Harvests contained a median of 3.4 x 10(6)/kg (range 0.3-91.8) CD34+ cells. Of these 87.6% (range 30-96.5) were annexin V-. In 10% of harvests more than 50% of CD34+ cells were apoptotic. Differences in PBSC mobilisation or collection could not explain the variation in annexin V binding. Cyclophosphamide + G-CSF significantly increased the yield of CD34+ cells but did not increase apoptosis. Comparison of consecutive harvests showed no difference in the numbers of CD34+ cells collected but found a significant decrease in apoptotic CD34+ cells through multiple collections. Analysis of annexin V binding in PBSC harvests is a simple flow cytometry technique which gives additional information on the status of CD34+ progenitor cells.
通过流式细胞术对CD34+细胞进行计数来定量外周血干细胞（PBSC）采集物中的祖细胞，并未考虑细胞活力。早期凋亡过程中的细胞膜不对称性会使磷脂酰丝氨酸暴露于细胞表面。这可以通过用膜联蛋白V FITC染色来检测。在收集后，立即采用CD34 PE和膜联蛋白V FITC双色流式细胞术分析了30份由环磷酰胺+粒细胞集落刺激因子（G-CSF）或标准化疗+G-CSF动员的自体PBSC采集物中的细胞凋亡情况。采集物中CD34+细胞的中位数为3.4×10⁶/kg（范围为0.3 - 91.8）。其中87.6%（范围为30% - 96.5%）的细胞不与膜联蛋白V结合。在10%的采集物中，超过50%的CD34+细胞发生凋亡。PBSC动员或采集的差异无法解释膜联蛋白V结合情况的变化。环磷酰胺+G-CSF显著提高了CD34+细胞的产量，但并未增加细胞凋亡。对连续采集物的比较显示，所采集的CD34+细胞数量没有差异，但发现多次采集后凋亡的CD34+细胞数量显著减少。分析PBSC采集物中膜联蛋白V的结合情况是一种简单的流式细胞术技术，它能提供有关CD34+祖细胞状态的额外信息。
