Stec Malgorzata, Weglarczyk Kazimierz, Baran Jaroslaw, Zuba Ewa, Mytar Bozenna, Pryjma Juliusz, Zembala Marek
Department of Clinical Immunology and Transplantation, Polish-American Institute of Pediatrics, Jagiellonian University Medical College, Wielicka str. 265, 30-663 Cracow, Poland.
J Leukoc Biol. 2007 Sep;82(3):594-602. doi: 10.1189/jlb.0207117. Epub 2007 Jun 26.
To determine whether monocytes can be generated from CD34+ hematopoietic progenitors in large numbers, cord blood CD34+ cells were first expanded for 3-10 days in X-VIVO 10 medium supplemented with FCS, stem cell factor (SCF), thrombopoietin (TPO), and Flt-3 Ligand (Flt-3L), and then differentiated in IMDM medium supplemented with FCS, SCF, Flt-3L, IL-3 and M-CSF for 7-14 days. These two step cultures resulted in up to a 600-fold mean increase of total CD14+ cells. Using this approach, two subpopulations of monocytes were obtained: CD14+CD16(-) and CD14++CD16+ occurring at 2:1 ratio. 1.25(OH)2 Vitamin D3 added to the differentiation medium altered this ratio by decreasing proportion of CD14++CD16+ monocytes. In comparison to CD14+CD16(-), the CD14++CD16+ cells showed different morphology and an enhanced expression of CD11b, CD33, CD40, CD64, CD86, CD163, HLA-DR, and CCR5. Both subpopulations secreted TNF and IL-12p40 but little or no IL-10. CD14++CD16+ monocytes released significantly more IL-12p40, were better stimulators of MLR but showed less S. aureus phagocytosis. These subpopulations are clearly different from those present in the blood and may be novel monocyte subsets that represent different stages in monocyte differentiation with distinct biological function.
为了确定CD34 +造血祖细胞能否大量生成单核细胞，首先将脐血CD34 +细胞在补充有胎牛血清（FCS）、干细胞因子（SCF）、血小板生成素（TPO）和Flt-3配体（Flt-3L）的X-VIVO 10培养基中扩增3 - 10天，然后在补充有FCS、SCF、Flt-3L、白细胞介素-3（IL-3）和巨噬细胞集落刺激因子（M-CSF）的IMDM培养基中分化7 - 14天。这两步培养使总CD14 +细胞平均增加了600倍。采用这种方法，获得了两个单核细胞亚群：CD14 + CD16（-）和CD14 ++ CD16 +，其比例为2:1。添加到分化培养基中的1,25（OH）2维生素D3通过降低CD14 ++ CD16 +单核细胞的比例改变了这一比例。与CD14 + CD16（-）相比，CD14 ++ CD16 +细胞表现出不同的形态，且CD11b、CD33、CD40、CD64、CD86、CD163、HLA-DR和CCR5的表达增强。两个亚群均分泌肿瘤坏死因子（TNF）和白细胞介素-12p40，但很少或不分泌白细胞介素-10。CD14 ++ CD16 +单核细胞释放的白细胞介素-12p40明显更多，是混合淋巴细胞反应（MLR）的更好刺激物，但对金黄色葡萄球菌的吞噬作用较弱。这些亚群与血液中存在的亚群明显不同，可能是代表单核细胞分化不同阶段且具有不同生物学功能的新型单核细胞亚群。
