Hematopoietic Stem Cell Differentiation and Commitment

描述（由申请人提供）：成体干细胞存在于许多组织中，并在整个生命过程中补充这些组织的细胞成分。最近，越来越多的证据表明，至少在某些条件下，成体干细胞的后代可以在其来源组织外产生细胞。在我们的实验室中，我们已经研究了这种所谓的干细胞“可塑性”，在肌肉中的干细胞产生血液的背景下，以及骨髓中的干细胞产生肌肉。肌肉中产生血液的干细胞起源于造血，是上一个资助期的重点。我们最近还表明，造血干细胞（HSC）生成肌肉的能力很可能是由于骨髓细胞直接掺入再生肌肉。在这里，使用肝再生作为模型，我们建议更详细地研究这种非本地细胞掺入的机制，并从长远角度提高其治疗用途的效率。在第一个目标中，我们将明确识别能够参与肝细胞生成的主要造血细胞类型。我们将利用cre-lox谱系标记策略以及细胞移植到遗传缺陷的肝脏模型中来评估血肝转换。我们还将使用多种供体和受体标记物确定在该模型中是否存在非融合肝再生机制的任何证据。在第二个目标中，我们将研究血肝转换是否可以发生在两个不同的肝再生模型的背景下，通过融合依赖性或非依赖性机制。这将揭示这种治疗策略对不同类型的肝脏疾病和再生模式的适用性。最后，在第三个目标中，我们将通过以下方式研究血-肝转化的分子要求：a）在没有骨髓移植的情况下给予细胞时诱导融合，B）使用诱导型病毒融合蛋白提高融合效率，以及c）使用表达标记的组织特异性蛋白的BAC转基因研究融合后发生的核重编程事件。总之，这些研究将提高我们对异型细胞融合的因素以及细胞核重编程以改变细胞身份的步骤的理解。这些见解对于基于干细胞的疗法的治疗应用至关重要。

DESCRIPTION (provided by applicant): Adult stem cells are present in many tissues and replenish the cellular constituents of those tissues throughout life. Recently, evidence has accumulated that at least under some conditions, progeny of adult stem cells can generate cells outside their tissue of origin. In our laboratory, we have examined this so-called stem cell "plasticity" in the context of generation of blood by stem cells in the muscle, and generation of muscle by stem cells from bone marrow. Stem cells in muscle that generate blood are hematopoietic in origin, and were the focus of the previous funding period. We have also recently shown that the ability of hematopoietic stem cells (HSC) to generate muscle is most likely due to the direct incorporation of myeloid cells into regenerating muscle. Here, using liver regeneration as a model, we propose to examine the mechanisms of this non-autochthonous cell incorporation in much more detail, with the long-term view of improving the efficiency for its therapeutic use. In the first Aim, we will definitively identify the major hematopoietic cell types capable of participating in generation of hepatocytes. We will utilize cre-lox lineage marking strategies as well as cell transplantation into a genetically deficient liver model to assess hematohepatic conversion. We will also determine whether there is any evidence for a non-fusion hepatic regeneration mechanism in this model using multiple donor and recipient markers. In the second Aim, we will examine whether hemato-hepatic conversion can occur in the context of two distinct models of liver regeneration through either fusion-dependent or -independent mechanisms. This will reveal the applicability of this therapeutic strategy to different types of liver disease and modes of regeneration. Finally, in the third Aim, we will study the molecular requirements for hemato-hepatic conversion by a) Inducing fusion when cells are given in the absence of bone marrow transplantation, b) Increasing the efficiency of fusion using inducible viral fusion proteins, and c) Studying the nuclear reprogramming events occurring upon fusion using BAC-transgenics expressing tagged tissue-specific proteins. Together, these studies will improve our understanding of the factors that govern heterotypic cell fusion as well as the steps in the reprogramming of nuclei to alter cell identity. These insights will be essential for moving towards therapeutic applications of stem cell-based therapies.