Fernandez Irina, Ooi Tracy P, Roy Krishnendu
Department of Biomedical Engineering, The University of Texas at Austin, Austin, Texas, USA; Dell Pediatric Research Institute and, The University of Texas at Austin, Austin, Texas, USA.
Stem Cells. 2014 Jan;32(1):93-104. doi: 10.1002/stem.1512.
In vitro differentiation of mouse and human stem cells into early T cells has been successfully demonstrated using artificial Notch signaling systems. However, generation of mature, antigen-specific, functional T cells, directly from human stem cells has remained elusive, except when using stromal coculture of stem cells retrovirally transfected with antigen-specific T cell receptors (TCRs). Here we show that human umbilical cord blood (UCB)-derived CD34+CD38-/low hematopoietic stem cells can be successfully differentiated into functional, antigen-specific cytotoxic CD8+ T cells without direct stromal coculture or retroviral TCR transfection. Surface-immobilized Notch ligands (DLL1) and stromal cell conditioned medium successfully induced the development of CD1a+CD7+ and CD4+CD8+ early T cells. These cells, upon continued culture with cytomegalovirus (CMV) or influenza-A virus M1 (GIL) epitope-loaded human leukocyte antigen (HLA)-A*0201 tetramers, resulted in the generation of a polyclonal population of CMV-specific or GIL-specific CD8+ T cells, respectively. Upon further activation with antigen-loaded target cells, these antigen-specific, stem cell-derived T cells exhibited cytolytic functionality, specifically CD107a surface mobilization, interferon gamma (IFNg) production, and Granzyme B secretion. Such scalable, in vitro generation of functional, antigen-specific T cells from human stem cells could eventually provide a readily available cell source for adoptive transfer immunotherapies and also allow better understanding of human T cell development.
利用人工Notch信号系统已成功证明小鼠和人类干细胞在体外可分化为早期T细胞。然而，直接从人类干细胞生成成熟的、抗原特异性的功能性T细胞仍然难以实现，除非使用经逆转录病毒转染抗原特异性T细胞受体（TCR）的干细胞进行基质共培养。在这里，我们表明，源自人脐带血（UCB）的CD34+CD38-/低造血干细胞无需直接基质共培养或逆转录病毒TCR转染，即可成功分化为功能性的、抗原特异性的细胞毒性CD8+T细胞。表面固定的Notch配体（DLL1）和基质细胞条件培养基成功诱导了CD1a+CD7+和CD4+CD8+早期T细胞的发育。这些细胞在与负载巨细胞病毒（CMV）或甲型流感病毒M1（GIL）表位的人白细胞抗原（HLA）-A*0201四聚体持续培养后，分别产生了多克隆的CMV特异性或GIL特异性CD8+T细胞群体。在用负载抗原的靶细胞进一步激活后，这些抗原特异性的、源自干细胞的T细胞表现出细胞溶解功能，特别是CD107a表面动员、干扰素γ（IFNγ）产生和颗粒酶B分泌。从人类干细胞如此可扩展地体外生成功能性的、抗原特异性T细胞最终可为过继性转移免疫疗法提供现成的细胞来源，也有助于更好地理解人类T细胞发育。
