Cell Cycle Related Transdifferentation Plasticity

描述(由申请人提供)： 迈赫达德·阿贝迪医学博士是血液肿瘤学博士后研究员，也是彼得·J·奎森伯里博士指导下的K08基金的候选人。申请者希望以干细胞生物学领域的研究为重点从事学术生涯。骨髓来源的细胞具有在适当的组织中或在体外再生许多不同类型细胞的能力。功能干细胞的可塑性与细胞周期的不同阶段有关，这些细胞的跨分化能力还没有得到很好的描述。随着细胞因子驱动的细胞周期传递，黏附蛋白的表达、植入和分化发生可逆的波动，这些波动似乎与细胞周期的特定阶段有关。假设骨髓的跨分化可塑性也会随着细胞周期的不同而波动，这一过程中的关键初始事件与归巢到不同的组织有关。除了为干细胞生物学提供戏剧性的新见解外，该项目还为各种临床组织修复方法提供了独特的潜力。我们计划通过细胞因子诱导细胞周期的所有时间点来研究不同的骨髓部分(全骨髓、LIN-SCA+、罗丹明低Hoescht低、GFP+Hoescht低侧群和克隆性替代骨髓)。我们将确定这些细胞在未经治疗或器官损伤的小鼠中的归巢能力、短期和长期命运以及它们在上述组织中的转分化能力。将使用两种允许细胞追踪的小鼠移植模型：BALB/c雄性到雌性模型和B6 GFP转基因模型。供体细胞将通过Y染色体荧光原位杂交和绿色荧光蛋白荧光染色或抗体染色进行鉴定。将立即进行归巢，用细胞质染料标记细胞。将对冰冻切片或石蜡切片进行双重标记，以确定捐赠者的来源和细胞类型。小鼠也将被重新植入“标记的”骨髓部分，然后在有或没有心脏、骨骼肌或肝脏损伤的情况下进行造血动员，并再次观察细胞的短期和长期命运和跨分化。高速细胞分选、荧光显微镜成像和大事件流式细胞仪分析将评估短期归巢。该项目将探索骨髓转化为非造血组织(心脏、骨骼肌和肝脏)的功能可塑性。

DESCRIPTION (provided by applicant): Mehrdad Abedi MD is a HematologylOncology postdoctorate fellow and a candidate for a K08 grant under the mentorship of Dr. Peter J. Quesenberry. The applicant wishes to pursue an academic career with his research focus in the field of stem cell biology. Marrow derived cells have the capacity to regenerate many different cell types in their appropriate tissues or in vitro. Functional stem cell plasticity connected with different phases of cell cycle and the transdifferentation ability of these cells have not been well charaterized. With cytokine driven cell cycle transit, adhesion protein expression, engraftment and differentiation fluctuates reversibly and these fluctuations appear to be tied to specific phases of cell cycle. The hypothesis is that the transdifferentation plasticity of marrow will also fluctuate with phase of cell cycle and that the critical initial event in this process relates to homing into different tissues. Besides giving dramatic new insights into stem cell biology, this project offers unique potential for various clinical tissue restorative approaches. We plan to study different marrow fractions (whole marrow, lin-sca+, rhodamine low Hoescht low, GFP+ Hoescht low side population, and clonally replaced marrow) through all points in a cytokine induced cell cycle tansit. We will determine the homing ability, short and long-term fate of these cells that go to heart, skeletal muscle and liver in untreated or organ injured mice and their ability to transdifferentiate in the above tissues. Two murine transplant models that allow for cell tracking will be used; the BALB/c male to female model and the B6 GFP transgenic model. Donor cells will be identified using FISH for the Y-chromosome and fluorescence or Ab staining for GFP. Immediate homing will be done marking the cells with a cytoplasmic dye. Double labeling approaches on frozen or paraffin sections will be carried out to identify donor origin and cell type. Mice will also be repopulated with "marked" marrow fractions and then hematopoietic mobilization will be done with or without heart, skeletal muscle or liver injury and again look at short and long-term cell fate and transdifferentation. High-speed cell sorting, fluorescent microscopy imaging and large event FACS analysis will evaluate short term homing. This project will explore the functional plasticity of marrow to transdifferentate into nonhematopoietic tissues (heart, skeletal muscle and in liver).