阵发性睡眠性血红蛋白尿症患者骨髓造血干/祖细胞体外单个细胞培养

目的 研究阵发性睡眠性血红蛋白尿症（PNH）患者单个不同表型骨髓造血干／祖细胞体外生长特性。方法 用免疫磁珠富集C34^ 细胞，用流式细胞仪自动分选系统将患者CD34^ CD59^ 和CD34^ CD59^-造血干／祖细胞单个分选入96孔培养板，进行单个细胞培养，观察不同细胞各个生长指标，并与正常对照细胞比较。结果 ①单个细胞培养条件下，患者CD34^ CD59^-细胞在细胞发生分裂（细胞数≥2个）的比率、形成集落（细胞数≥50）的比率及扩增的总数（所有细胞发生分裂的孔的细胞数的总和）都超过了其CD34^ CD59^ 细胞（P＜0．05）。②PNH CD34^ CD59^-细胞单个细胞扩增的数目及细胞扩增的总数代于正常对照（P＜0．05）。③PNH CD34^ CD59^ 细胞在较大集落（细胞数≥500）形成、单个细胞扩增的数目及细胞扩增总数均低于正常对照（P＜0．01）。④患者异常与正常表型细胞单个细胞次级集落的形成能力差异无显著性（P＞0．05），但都明显低于正常对照。（P＜0．001）。结论 在单个细胞培养条件下，患者异常表型的造血干／祖细胞较其正常表型的造血干／祖细胞具有一定的生长优势。但它们与正常骨髓细胞相比，都存在一定的生长缺陷，正常表型细胞的缺陷尤为明显。

Proliferative capacity of the isolated single CD34+ glycosylphosphatidylinesitol-anchored (GPI) protein negative and positive hematopoietic cells in paroxysmal nocturnal hemoglobinuria

OBJECTIVES: To investigate the stroma-independent growth ability, multilineage differentiation and expansion of the single hematopoietic stem/progenitor cell from patients with paroxysmal nocturnal hematoglobinuria (PNH). METHOD: The CD(34)(+) CD(59)(+) and CD(34)(+) CD(59)(-) cells from PNH patients and CD(34)(+) CD(59)(+) cells from normal volunteers were sorted as each single cell into a well of 96 well culture plates containing culture medium supplemented with SCF, IL-3, Epo, GM-CSF, G-CSF, IL-6, Tpo and Flt-3 ligand. RESULTS: (1) Single PNH CD(34)(+) CD(59)(-) cell had a higher capacities for plating efficiency, colony (>/= 50 cells) formation and cell expansion than that of the PNH CD(34)(+) CD(59)(+) cells (P < 0.05); (2) Both the single CD(34)(+) CD(59)(-) cells from PNH patients and the single CD(34)(+) CD(59)(+) cells from normal controls had similar capacities for cell plating efficiency and colony and large colony formation. The PNH CD(34)(+) CD(59)(-) cells had a lower average cell production and cell expansion capacity. (3) The single CD(34)(+) CD(59)(+) cells from both PNH patients and normal controls showed the same capacities for cell plating efficiency and colony formation. The PNH CD(34)(+) CD(59)(+) cells exhibited much lower capacity for large colony formation, average cell production and total cell expansion. (4) A diminished secondary colony formation ability was also observed in the PNH CD(34)(+) CD(59)(+) and CD(34)(-) CD(59)(-) clones. CONCLUSION: The single PNH CD(34)(+) CD(59)(-) cells had growth advantage over the single PNH CD(34)(+) CD(59)(+) cells to some extent, but they both had impaired growth abilities as compared with CD(34)(+) cells from normal volunteers.