【摘要】 目的　探讨阵发性睡眠性血红蛋白尿症 (PNH)患者CD34+ CD59+ 细胞的特性及PNH克隆呈优势造血的可能原因 ,以探索PNH发病的内在机制。方法　用免疫磁珠吸附法富集纯化CD34+ 细胞 ,再用流式细胞仪分选出PNH患者的CD34+ CD59+ 细胞、CD34+ CD59- 细胞及正常对照CD34+ 细胞。分别进行体外扩增液体培养 2周 ,并对扩增前、后的细胞进行半固体培养。结果　①PNH患者CD34+CD59+ 细胞与正常对照CD34+ 细胞形成集落形成单位 (CFU)均在第 7天达到扩增高峰 ,并且扩增后的细胞仍能保持CD59抗原 ,无GPI锚连蛋白的丢失。②正常对照的CD34+ 细胞在生存、增殖、形成CFU及扩增能力上均明显强于PNH患者的CD34+ CD59+ 细胞及CD34+ CD59- 细胞。③PNH患者CD34+ CD59+ 细胞及CD34+ CD59- 细胞体外半固体培养 ,其形成CFU的能力无明显差异。④PNH患者CD34+ CD59+ 细胞及CD34+ CD59- 细胞在SCF +IL 3 +IL 6 +FL +Tpo及SCF +IL 3+IL 6 +FL +Tpo +Epo组合下液体培养 ,其生存、增殖、扩增等能力上均无明显差异。但在SCF +IL 3+IL 6 +FL +Tpo +Epo +GM CSF组合下液体培养 ,CD34+ CD59- 细胞的生存、增殖、扩增能力均明显强于CD34+ CD59+ 细胞。结论　①正常对照的CD34+ 细胞在生存、增殖、形成CFU及扩增能力上均明显强于PNH患者的CD34更多还原

【Abstract】 Objective To explore the characteristics of CD 34+CD 59+ cells from paroxysmal nocturnal hemoglobinuria(PNH) patients’ bone marrow and the possible reasons of hematopoietic clonal dominance of PNH clones. Methods CD 34+CD 59+ and CD 34+CD 59- cells from PNH patients and CD 34+ cells from normal control were selected from the bone marrow mononuclear cells by means of immunomagnetic microbead-flow cytometry two step sorting method undergone ex vivo expansion in liquid culture for two weeks and performed semisolid cultures before and after expansion. Results ① Cultivation for seven days was the optimum for ex vivo expansion of PNH CD 34+CD 59+ cells and normal CD 34+ cells, both cell populations remained CD 59 positive after expansion. ②Normal CD 34+ cells had higher capacities of proliferation and expansion, and stronger potential to survival than that of both PNH CD 34+CD 59+ and PNH CD 34+CD 59- cells. ③In terms of semisolid culture, there was no significant difference in the yields of CFU formation between CD 34+CD 59+ and CD 34+CD 59- cells. ④In liquid culture with combinations of hematopoietic factors SCF+IL-3+IL-6+FL+Tpo or SCF+IL-3+IL-6+FL+Tpo+Epo, there was no significant difference in the capabilities of survival, proliferation and expansion between CD 34+CD 59+ and CD 34+CD 59- cells; but with combination of SCF+IL-3+IL-6+FL+Tpo+Epo+GM-CSF, CD 34+CD 59- cells had better proliferation and expansion capacities and stronger potential to survival than that of CD 34+CD 59+ cells. Conclusions ① Normal CD 34+ cells had better proliferation, expansion capacities and stronger potential tosurvival than that of PNH CD 34+CD 59+ cells. ②In semisolid and liquid culture with hematopoietic factor combinations, there was no significant difference in the capabilities of survival, proliferation and expansion between CD 34+CD 59+ and CD 34+CD 59- cells. It was suggested that CD 34+CD 59- cells had no clonal hemotopoiesis dominance. GM-CSF might be one of the reasons for PNH clones to possess clonal hematopoiesis dominance.更多还原