Immunodeficient Mouse Model of Stem Cell Plasticity

描述(由申请人提供)： 最近耐人寻味的观察表明干细胞的可塑性可能存在。 啮齿动物的骨髓细胞被证明对肝脏、骨骼和 心肌。这些非常令人兴奋的发现可能会影响到 一种戏剧性的未来方式，如果适用于人类系统的话。骨髓干细胞 从理论上讲，可以从患者身上获取细胞，并用来修复他的 或者她受损的心脏、肌肉组织或肝脏。为了实现这一点 潜在的，科学界必须确定表型(S) 表现出可塑性的小鼠细胞的人类对应物 确定人类骨髓中的单个全能干细胞是否能够 分化为组织和血液的能力，必须优化分离和 全能细胞的移植。我们假设有真实的 存在于成人骨髓和小鼠骨髓中的全能细胞 新生儿脐带血。我们假设共同的干细胞存在 对于血液和肝脏，以及在人类中，它们的表型将是可塑性的 非粘附性CD34-/LIN-，在小鼠中，它们的表型将是塑料非 贴壁SP或c-Kit+Thy-1.1(Lo)LIN-SCA-1+。我们预测人类和小鼠 能够修复肌肉的细胞实际上是可塑性贴壁间充质干细胞 细胞(MSC)，不会产生血细胞。我们将用我们的小说 免疫缺陷的裸/NOD/SCID小鼠品系，寿命为两年， 作为明确表型的标记干细胞的接受者，从 小鼠骨髓、人骨髓和人脐带血。逆转录病毒 慢病毒载体标记的MSC和HSC组分将来自每个来源 在体内评估它们形成血细胞和肌肉的能力，如下 肝脏损伤后的肌肉损伤，或血细胞和肝脏 收件人。单细胞水平的克隆整合分析，具有 测序，将确定后代是否来自相同的 前体，或来自离散的干细胞。我们还将确定是不是 可能调节人的生存、分化和招募 候选小鼠和人类全能干细胞通过以下方式向特定组织分化 肝细胞生长因子/散射超生理学水平的表达 因子(HGF/SF)。肝细胞生长因子是一种化学诱导剂和活性因子， 在受损的肝脏、心脏和骨骼肌中升高。肝细胞生长因子影响 运动和维持成肌细胞、肝卵圆细胞和 造血干细胞。我们假设HGF，在局部分泌 对组织损伤的反应，是招募全能干细胞的主要因素 细胞从循环进入损伤肌肉或肝脏的部位。我们 假设肝细胞生长因子可能维持招募的全能干细胞的活性 细胞，而它们是由其他诱导性的组织特异性因子引导的 局部微环境来分化，以再生受损组织。 肝细胞生长因子对人的募集、存活和分化的影响 小鼠全能干细胞候选者将在体外和在 活着。我们的研究将提供确凿的证据证明干细胞的可塑性 存在，将鉴定小鼠和人类细胞的表型， 除了血细胞外，还会产生肝脏或肌肉组织，并将确定 方法增强这些细胞在受损组织中的募集。

DESCRIPTION (provided by applicant): Recent intriguing observations indicate that stem cell plasticity may exist. Rodent bone marrow cells have been shown to contribute to liver, skeletal and cardiac muscle. These extremely exciting findings could impact health care in a dramatic way in the future, if applicable to the human system. Marrow stem cells could theoretically be harvested from a patient and used to repair his or her damaged heart, muscle tissue, or liver. In order to realize this potential, the scientific community must determine the phenotype(s) of the human counterparts to the murine cells that have displayed plasticity, must determine whether a single totipotent stem cell from human marrow is capable of differentiation into tissue and blood, and must optimize isolation and transplantation of the totipotent cells. We hypothesize that there are true totipotent cells that reside in the adult human and mouse bone marrow and in neonatal umbilical cord blood. We hypothesize that common stem cells exist for blood and liver, and that in the human, their phenotype will be plastic non-adherent CD34-/lin-, and in the mouse their phenotype will be plastic non- adherent SP or c-kit+Thy-1.1(lo)Lin-sca-1+. We predict that human and murine cells that can repair muscle are actually plastic adherent mesenchymal stem cells (MSC) and will not generate blood cells. We will use our novel immunodeficient nude/NOD/SCID mouse strain, which has a lifespan of two years, as the recipient for marked stem cells of defined phenotype, isolated from murine bone marrow, human marrow, and human umbilical cord blood. Retroviral and lentiviral vector marked MSC and HSC fractions from each source will be assessed in vivo for their capacity to form blood cells and muscle, following muscle injury, or blood cells and liver, following liver injury to the recipient. Clonal integration analysis at the single cell level, with sequencing, will determine whether the progeny were derived from the same precursor, or from discrete stem cells. We will also determine whether it is possible to modulate the survival, differentiation, and recruitment of candidate murine and human totipotent stem cells to specific tissues by expression of supraphysiological levels of Hepatocyte Growth Factor/Scatter Factor (HGF/SF). HGF is a chemoattractant and viability factor that is elevated in injured liver, cardiac, and skeletal muscle. HGF affects the motility and maintains the viability of myoblasts, hepatic oval cells, and hematopoietic stem cells. We hypothesize that HGF, secreted locally in response to tissue injury, is a major factor in recruiting totipotent stem cells from the circulation into the site of muscle or liver injury. We hypothesize that HGF may maintain the viability of recruited, totipotent stem cells, while they are directed by other inductive, tissue-specific factors in the local microenvironment to differentiate, to regenerate the damage tissue. The impact of HGF on the recruitment, survival and differentiation of human and murine totipotent stem cell candidates will be tested in vitro and in vivo. Our studies will provide definitive proof that stem cell plasticity exists, will identify the phenotypes of the murine and human cells that can generate liver or muscle tissue in addition to blood cells, and will determine methods to enhance the recruitment of those cells to damaged tissue.