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从胚胎干细胞诱导发育为浆细胞的体外实验研究 |
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乔春平 李树浓 黄绍良 徐 令 梁晓燕 张秀明
摘 要 目的：建立胚胎干细胞体外定向诱导分化为浆细胞的体系，为研究造血发生的基因调控及B淋巴细胞发育过程奠定基础。方法：分阶段培养，第一、二阶段使用半固体培养，分组加入细胞因子、2－巯基乙醇和胚胎细胞条件培养液，用流式细胞仪、免疫组化及Wright s染色鉴定细胞，第三阶段移入骨髓基质细胞滋养层。结果：第一阶段：加入SCF、TPO、IL－6、IL－3、Epo、和G－CSF，4 d后细胞大片坏死，仅加入2－巯基乙醇可形成胚胎体。第二阶段：加入SCF和TPO，细胞增殖缓慢，再加入条件培养液，发育3～4 d可形成CD34＋的细胞集落。第三阶段：移至骨髓基质细胞饲养层后，发育为浆细胞。结论：胚胎干细胞在体外可定向分化为浆细胞。胚胎干细胞在早期发育不需要细胞因子，胚胎细胞的条件培养液可促使其向造血细胞发育，骨髓基质细胞饲养层可使其向浆细胞分化。
主题词 胚胎；干细胞；细胞活素类；造血干细胞；浆细胞
Induction of plasma cells from embryonic stem cells in vitro culture
QIAO Chun-Ping, LI Shu-Nong,HUANG Shao-Liang, XU Ling, LIANG Xiao-Yan, ZHANG Xiu-Ming
Department of Pathophysiology, Sun-Yat-sen University of Medical Sciences, Guangzhou(510089)
Abstract AIM：To find a suitable microenviroment for improved induction of plasma cells from murine embryonic stem(ES)cells,we explored the roles of cytokines and primary bone marrow stromal cells in the development of ES－D3 cells.METHOD and RESULTS：The process was divided into three stages.First stage(0－4 day):adding SCF(stem cell-factor) ，TPO(thrombopoietin)，IL－6，IL－3，erythropoietin，G－CSF into methycellulose culture,ES－D3 cells died four days later,ES－D3 cells developed into embryoid bodies and they grew well in the presence of 2－Mercaptoethanol (2-ME) only.Second stage(4－7 dyas)：the embryoid bodies were trypsined,then transferred into secondary methylcellulose culture,cells grew slowly when adding SCF and TPO, but they(86%)developed quickly into the colonies of CD34＋ cells in the presence of conditional medium(first stage of liquid culture,adding 2ME only).Third stage:CD34＋ cells developed into CD34－ CD38＋ cells when transferred onto prepared bone marrow stromal cells in the presence of IL－6 and SCF,they looked like plasma cells by cell morphology,and immunoglobulin test was positive .CONCLUSIONS：Cultured in a suitable microenviroment,ES－D3 cells can developed into plasma cells.cytokines were not needed in the early develo[1] [2] 下一页
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