Stroma as a target to prevent stress-induced cellular senescence of hematopoietic stem cells in vitro and in vivo

基质作为预防体外和体内应激诱导的造血干细胞细胞衰老的靶标

• 批准号: 318433017
• 负责人: Professor Dr. Robert A.J. Oostendorp
• 金额: --
• 依托单位: Klinik und Poliklinik für Innere Medizin III: Hämatologie und Onkologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2016
• 资助国家: 德国
• 起止时间: 2015-12-31 至 2019-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/318433017?language=en
• 关键词: Stroma; target; prevent; stress; induced

Adult stem cells are characterized by their ability to self-renew and regenerate tissues. Under different in vitro and in vivo conditions of stress, hematopoietic stem cells (HSCs) their ability to self-renew is poorly maintained, which ultimately leads to HSC exhaustion. Our previous work shows that selected stromal cells, or their conditioned medium preserve functional HSCs in culture. Gene expression studies using our co-cultures of stromal cells with early hematopoietic cells: Lineage- Sca-1+ Kit+ cells (LSKs) showed a highly dynamic reciprocal interaction of stromal cells and LSK cells. For instance, LSK cells up-regulate the stromal production of connective tissue growth factor (Ctgf), which, in turn, is required to maintain HSCs with long-term repopulating ability in cultures. The underlying mechanism was that HSC-enriched CD34- CD48- CD150+ LSK (CD34- SLAM) cells cultured on Ctgf-knockdown stroma increases the fraction of these cells in G0, which show delayed time to first cell division and a DNA damage response-associated senescence. This study unveils important feedback mechanisms by which hematopoietic cells promote the ability of niche cells to maintain HSC activity by antagonizing HSC senescence. Here, we are interested in translating these studies in vivo, and, more importantly, use the previous results to define strategies, which prevent DNA damage-response-associated HSC senescence under stress conditions in vivo. For this purpose, we will focus on stress-inducible Ctgf to dissect the relevance of different niche populations in preventing HSC senescence using genetic reporter and conditional knockout models. Also, since CTGF is a known regulator of stromal cell proliferation and differentiation in vitro, we will assess the role of stress-induced genes, such as Ctgf, in stromal cell function and regeneration in vivo. In the second part of the project, we will put our previous studies in a broader perspective and translate newly proposed and already existing gene expression analyses to aid in the formulation of stress-induced signaling pathways in niche cells and critical nodes therein. Screening will be performed using single CD34- SLAM cultures in conditioned media of genetically altered stromal cells, and, in a later step, in the presence of small molecule inhibitors. These analyses will be used to define specific niche targets to antagonize HSC senescence. The results of this project may have broader implications for prevention of exhaustion of normal HSC under different conditions of chronic stress, such as observed in in vitro culture. Interestingly, the defined targets may also be of relevance for combating HSC exhaustion in vivo as a result of stress due to wounding, infections, leukemia, or cytostatic therapy, as well as during diabetes or ageing.

成人干细胞的特征是具有自我更新和组织再生的能力。在不同的体外和体内应激条件下，造血干细胞（HSCs）自身的自我更新能力维持不佳，最终导致HSCs衰竭。我们以前的工作表明，选择的基质细胞，或它们的条件培养基在培养中保存功能性HSC。使用我们的基质细胞与早期造血细胞的共培养物的基因表达研究：谱系- Sca-1+ Kit+细胞（LSK）显示基质细胞和LSK细胞的高度动态的相互作用。例如，LSK细胞上调结缔组织生长因子（Ctgf）的基质产生，这反过来又是维持HSC在培养物中具有长期再增殖能力所必需的。潜在的机制是在Ctgf敲低基质上培养的HSC富集的CD 34-CD 48-CD 150 + LSK（CD 34- SLAM）细胞增加了这些细胞在G 0中的分数，其显示出延迟的第一次细胞分裂时间和DNA损伤反应相关的衰老。这项研究揭示了重要的反馈机制，造血细胞促进能力的小生境细胞维持HSC的活性，通过拮抗HSC衰老。在这里，我们有兴趣在体内翻译这些研究，更重要的是，使用以前的结果来定义策略，防止DNA损伤反应相关的HSC衰老在体内应激条件下。为了这个目的，我们将专注于应力诱导的Ctgf解剖不同的生态位人口的相关性，在防止HSC衰老的基因报告和条件敲除模型。此外，由于CTGF是一种已知的调节基质细胞增殖和分化的体外，我们将评估的作用，应力诱导的基因，如Ctgf，在基质细胞的功能和再生在体内。在该项目的第二部分，我们将把我们以前的研究在一个更广泛的角度和翻译新提出的和已经存在的基因表达分析，以帮助制定压力诱导的信号通路在小生境细胞和关键节点。将在遗传改变的基质细胞的条件培养基中使用单一CD 34- SLAM培养物进行筛选，并且在随后的步骤中，在存在小分子抑制剂的情况下进行筛选。这些分析将用于定义拮抗HSC衰老的特定小生境靶标。该项目的结果可能具有更广泛的意义，在不同条件下的慢性应激，如在体外培养中观察到的正常HSC的衰竭的预防。有趣的是，所定义的靶点也可能与对抗由于创伤、感染、白血病或细胞生长抑制疗法以及糖尿病或衰老期间的应激而导致的体内HSC耗竭有关。