Payuhakrit Witchuda, Panichakul Tasanee, Charoenphon Natthawut, Chalermsaenyakorn Panus, Jaovisidha Adithep, Wongborisuth Chokdee, Udomsangpetch Rachanee
Department of Pathobiology, Faculty of Science, Mahidol University, Thailand.
Faculty of Science and Technology, Suan Dusit Rajabhat University, Thailand.
EXCLI J. 2015 Sep 9;14:1031-9. doi: 10.17179/excli2015-506. eCollection 2015.
Hematopoietic stem cells (HSC) from cord blood are potentially high sources for transplantation due to their low immunogenicity and the presence of the multipotent cells. These cells are capable of differentiating to produce various lineages of blood cells under specific conditions. We have enriched highly purified CD34(+) cells from cord blood, determined in vitro growth of the cells in culture systems in the absence (condition A) or presence of GM-CSF and G-CSF (condition B), and determined the profile of immune cells during the period of cultivation by using flow cytometry. PhytohemagglutininA (PHA) was used as a mitogen to stimulate T lymphocytes derived from hematopoietic stem cells. GM-CSF and G-CSF prolonged the survival of the growing cells and also maintained expansion of cells in blastic stage. By day 12 of cultivation, when cell numbers peaked, various types of immune cells had appeared (CD14(+) cells, CD40(+)HLA-DR(+) cells, CD3(+)CD56(+) cells, CD19(+) cells, CD3(+)CD4(+) cells, CD3(+)CD8(+)cells and CD3-CD56(+)). A significantly higher percentage of monocytes (p = 0.002) were observed under culture with GM-CSF, G-CSF when compared with culture without GM-CSF, G-CSF. In addition, T lymphocytes derived from HSC responded to 50 µg/ml of PHA. This is the first report showing the complete differentiation and proliferation of immune cells derived from CD34(+) HSC under in vitro culture conditions. Lymphocytes, monocytes, dendritic cells and polymorph nuclear cells derived from HSC in vitro are unique, and thus may benefit various studies such as innate immunity and pathophysiology of immune disorders.
由于脐血造血干细胞（HSC）免疫原性低且存在多能细胞，它们潜在地是移植的高来源。这些细胞能够在特定条件下分化产生各种血细胞谱系。我们从脐血中富集了高度纯化的CD34(+)细胞，确定了这些细胞在无GM-CSF和G-CSF（条件A）或有GM-CSF和G-CSF（条件B）的培养系统中的体外生长情况，并通过流式细胞术确定了培养期间免疫细胞的概况。植物血凝素A（PHA）用作有丝分裂原以刺激源自造血干细胞的T淋巴细胞。GM-CSF和G-CSF延长了生长细胞的存活时间，还维持了处于母细胞阶段的细胞扩增。到培养第12天，细胞数量达到峰值时，出现了各种类型的免疫细胞（CD14(+)细胞、CD40(+)HLA-DR(+)细胞、CD3(+)CD56(+)细胞、CD19(+)细胞、CD3(+)CD4(+)细胞、CD3(+)CD8(+)细胞和CD3-CD56(+)）。与无GM-CSF、G-CSF的培养相比，在有GM-CSF、G-CSF的培养下观察到单核细胞百分比显著更高（p = 0.002）。此外，源自HSC的T淋巴细胞对50μg/ml的PHA有反应。这是第一份显示在体外培养条件下源自CD34(+) HSC的免疫细胞完全分化和增殖的报告。体外源自HSC的淋巴细胞、单核细胞、树突状细胞和多形核细胞是独特的，因此可能有益于各种研究，如先天性免疫和免疫疾病的病理生理学研究。
