DIRECTED STEM CELL HEMATOPOIESIS AND DIFFERENTIATION

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 血细胞形成(造血)的过程已经得到了广泛的研究，但许多基本机制仍然不清楚。Dogma指出，造血分化在本质上通常是随机的。体外干细胞研究表明，干细胞在特定细胞因子的作用下具有一定的定向细胞分化能力，但目前缺乏足够数量的谱系特异性细胞，阻碍了其转化到临床领域。以往对细胞周期相关的造血干细胞(HSC)功能的研究表明，HSC是一个连续体，在整个周期中具有恒定的可逆表型变化。我们发现，细胞周期位置是干细胞对相同刺激的反应结果的一个独立变量。刺激的例子有骨髓移植、祖细胞扩增和分化。我们发现，在施加刺激时，干细胞的细胞周期位置发生了可重复的变化。我们初步研究了这一现象，虽然最初的细胞周期从静止的G0-1状态过渡，但最近观察了最初细胞分裂后的干细胞，发现这种现象继续以可重复的模式与细胞周期位置联系在一起。这些数据表明，早期骨髓干细胞的功能表型随着其遍历细胞周期而发生变化。 我们通过细胞周期探索了造血干细胞分化的功能能力，这是培养中时间的反映，并发现存在向特定谱系分化的点(S)。我们发现，初始细胞分裂前的初始细胞因子诱导信号会影响两周后细胞群体的含量。我们认为，HSC对特定微环境的影响通过循环具有变化的敏感性，这直接影响到HSC的后代。我们用纯化的干/祖细胞[谱系阴性，罗丹明低，Hoescht低细胞]进行了实验，发现在细胞周期的特定位置，高度定向的谱系分化为巨核细胞或粒细胞。这笔赠款的目的是定义临床前小鼠移植模型中与细胞周期相关的定向谱系分化，并研究人类干细胞/祖细胞定向分化的机会。这些研究有望进一步确定造血骨髓干细胞的基本性质，并可能导致有趣的临床前细胞生物处理模型，以选择性地支持各种清髓性治疗方法或祖细胞缺陷状态。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The process of blood cell formation (hematopoiesis) has been extensively studied yet many basic mechanisms continue to be unclear. Dogma states that hematopoietic differentiation is generally stochastic in nature. In vitro stem cell studies indicate that stem cells have some ability toward directed cellular differentiation with use of specific cytokine cocktails, but lack of formation of adequate numbers of lineage specific cells currently prevents translation into the clinical realm. Prior work on cell cycle related hematopoietic stem cell (HSC) function has indicated that HSC are in a continuum, with constant reversible phenotypic variation through cycle. We have found that cell cycle position is an independent variable of the stem cell on the outcome of response to an identical stimulus. Examples of stimuli are bone marrow transplantation, progenitor expansion, and differentiation. We have found reproducible changes in the context of cell cycle position of the stem cell at the time of applying a stimulus. We have primarily studied this though initial cell cycle transit from a quiescent G0-1 state but have more recently looked at these stem cells after initial cell division and have found that this phenomenon continues to be tied to cell cycle position in a reproducible pattern. These data suggest that the functional phenotype of early marrow stem cells shifts as it traverses cell cycle. We have explored the functional ability of HSC differentiation through cell cycle, which is a reflection of time in culture and have found that there are points where differentiation toward specific lineage(s) occurs. We have found that an initial cytokine inductive signal prior to initial cell division influences the content of the cell population two weeks later. We believe that HSC have changing sensitivity to specific microenvironmental influences through cycle and this directly impacts on the HSC's progeny. We have performed experiments using purified stem/progenitors [lineage negative, rhodamine low, Hoescht low cells] and showed highly directed lineage differentiation into megakaryocytes or granulocytes at specific positions in cell cycle. The objectives of this grant are to define cell cycle related directed lineage differentiation in preclinical murine transplant models and to study human stem/progenitor cells for directed differentiation opportunities. These studies promise to further define the basic nature of the hematopoietic marrow stem cells and could lead to interesting preclinical cellular bioprocessing models for selective lineage support of various myeloablative therapy approaches or progenitor deficient states.