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Cocultivation of umbilical cord blood CD34^+ cells with retro-transduced hMSCs leads to effective amplification of long-term culture-initiating cells
作者姓名:Xie CG  Wang JF  Xiang Y  Qiu LY  Jia BB  Wang LJ  Wang GZ  Huang GP
作者单位:Chun-Gang Xie Jin-Fu Wang Ying Xiang Li-Yan Qiu Bing-Bing Jia Li-Juan Wang Guo-Zhong Wang Guo-Ping Huang,College of Life Sciences,Zhejiang University,Hangzhou 310012,Zhejiang Province,China Institute of Life Science,Jiangsu University,Zhenjiang 212013,Jiangsu Province,China
摘    要:AIM:To establish a novel coculture system for ex vivoexpansion of umbilical cord blood(UCB)hematopoieticprogenitors using thrombopoietin(TPO)/Flt-3 ligand(FL)-transduced human marrow-derived mesenchymalstem cells(tfhMSCs)as feeder.METHODS:UCB CD34~ cells were isolated and culturedusing four culture systems in serum-containing or serum-free medium.Suitable aliquots of cultured cells wereused to monitor cell production,clonogenic activity,and long-term culture-initiating culture(LTC-IC)output.Finally,the severe-combined immunodeficient(SCID)mouse-repopulating cell(SRC)assay was performed toconfirm ability of the cultured cells to reconstitute long-term hematopoiesis.RESULTS:There were no significant differences in thenumber of total nucleated cells among different culturesystems in serum-containing medium during 21-dculture.However,on d 14,the outputs of CD34~ cells,CFU-C and CFU-GEMM in ffhMSCs coculture system weresignificantly enhanced.LTC-IC assay demonstrated thatthe tfhMSCs coculture system had the most powerfulactivity.The severe-combined immunodeficient(SCID)mouse repopulating cell(SRC)assay confirmed extensiveability of the expanded cells to reconstitute long-termhematopoiesis.Furthermore,PCR analysis demonstratedthe presence of human hematopoietic cells in the bonemarrow and peripheral blood cells of NOD/SCID mice. CONCLUSION:The TPO/FL-transduced hMSCs,incombination with additive cytokines,can effectivelyexpand hematopoietic progenitors from UCB in vitro andthe tfhMSCs coculture system may be a suitable systemfor ex vivo manipulation of primitive progenitor cellsunder contact culture conditions.

关 键 词:血栓形成  红细胞  细胞培养  脐带血  hMSCs
收稿时间:2005 Apr 22

Cocultivation of umbilical cord blood CD34+ cells with retro-transduced hMSCs leads to effective amplification of long-term culture-initiating cells
Xie CG,Wang JF,Xiang Y,Qiu LY,Jia BB,Wang LJ,Wang GZ,Huang GP.Cocultivation of umbilical cord blood CD34+ cells with retro-transduced hMSCs leads to effective amplification of long-term culture-initiating cells[J].World Journal of Gastroenterology,2006,12(3):393-402.
Authors:Xie Chun-Gang  Wang Jin-Fu  Xiang Ying  Qiu Li-Yan  Jia Bing-Bing  Wang Li-Juan  Wang Guo-Zhong  Huang Guo-Ping
Institution:1. College of Life Sciences, Zhejiang University, Hangzhou 310012,Zhejiang Province, China;Institute of Life Science, Jiangsu University,Zhenjiang 212013, Jiangsu Province, China
2. College of Life Sciences, Zhejiang University, Hangzhou 310012,Zhejiang Province, China
Abstract:AIM:To establish a novel coculture system for ex vivo expansion of umbilical cord blood(UCB)hematopoietic progenitors using thrombopoietin(TPO)/Flt-3 ligand (FL)-transduced human marrow-derived mesenchymal stem cells(tfhMSCs)as feeder. METHODS:UCB CD34~ cells were isolated and cultured using four culture systems in serum-containing or serum- free medium.Suitable aliquots of cultured cells were used to monitor cell production,clonogenic activity, and long-term culture-initiating culture(LTC-IC)output. Finally,the severe-combined immunodeficient(SCID) mouse-repopulating cell(SRC)assay was performed to confirm ability of the cultured cells to reconstitute long- term hematopoiesis. RESULTS:There were no significant differences in the number of total nucleated cells among different culture systems in serum-containing medium during 21-d culture.However,on d 14,the outputs of CD34~ cells, CFU-C and CFU-GEMM in ffhMSCs coculture system were significantly enhanced.LTC-IC assay demonstrated that the tfhMSCs coculture system had the most powerful activity.The severe-combined immunodeficient(SCID) mouse repopulating cell(SRC)assay confirmed extensive ability of the expanded cells to reconstitute long-term hematopoiesis.Furthermore,PCR analysis demonstrated the presence of human hematopoietic cells in the bone marrow and peripheral blood cells of NOD/SCID mice. CONCLUSION:The TPO/FL-transduced hMSCs,in combination with additive cytokines,can effectively expand hematopoietic progenitors from UCB in vitro and the tfhMSCs coculture system may be a suitable system for ex vivo manipulation of primitive progenitor cells under contact culture conditions.
Keywords:Mesenchymal stem cells  Thrombopoietin  Fit-3 ligand  Hematopoiesis
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