MECHANISMS OF SYNERGISTIC REGULATION OF STEM/PROGENITORS

细胞因子在体外起作用以刺激、增强和/或抑制骨髓祖细胞（MPC）的增殖。 在体外观察到的一些细胞因子的作用已经在动物模型中重现，并且一些已经在人类临床试验中显示出功效。 可能的是，在响应于细胞因子的组合，特别是有效的共刺激细胞因子如钢因子的MPC增殖的刺激方面，在体外注意到的协同作用，以及通过趋化因子对这种协同作用的抑制与体内造血有关。 我们相信，对这些生长促进和抑制细胞因子触发的细胞内机制的评估将增强我们对正常MPC调节的理解，并最终将用于设计血液病患者的合理治疗。我们认为，多生长因子诱导的MPC增殖的协同刺激和这种抑制由趋化因子是，至少部分，细胞周期现象。 我们的假设是，细胞周期调节因子，如细胞周期蛋白依赖性激酶抑制剂，p21 cip 1/waf 1，某些细胞周期蛋白依赖性激酶，细胞周期蛋白，和其他细胞内信号是关键的反应介导的增殖协同作用。 我们的生化结果和评价MPC增殖的小鼠功能性缺失的一些这些建议的细胞内介质支持这一假设。 我们的长期目标是确定参与这些效应的关键细胞内分子，并利用这些信息为临床带来益处。 为了实现这些目标，我们提出了以下两个具体目标：1。研究多种生长因子参与的细胞内机制（GM-CSF和钢因子）诱导的MPC增殖的协同刺激，通过：a）评估细胞周期蛋白依赖性激酶抑制剂/调节剂p21 cip 1/waf 1的潜在作用，以及其与细胞周期蛋白依赖性激酶和细胞周期蛋白在这些作用中的关系，和B）阐明转录因子BCL-6和Stat 4的潜在作用，蛋白磷酸酶SHP-1和SHIP以及MAP激酶，以及它们在这些作用中彼此之间以及与p21 cip 1/waf 1之间可能的相互关系。 2.通过评估骨髓抑制性CC的比较效应，评估参与抑制多种生长因子诱导的MPC增殖的协同刺激的细胞内机制（MIP-1 α，MCP-1），CXC（IL-8，IP-10）和C（趋化因子）趋化因子和它们通过以下途径作用的受体：a）细胞周期和控制细胞周期进程的调节分子，和B）与MPC增殖刺激相关的细胞内信号传导分子。

Cytokines act in vitro to stimulate, enhance, and/or suppress proliferation of myeloid progenitor cells (MPC). Actions of some cytokines noted in vitro have been reproduced in animal models, and some have shown efficacy in human clinical trials. It is probable that synergism noted in vitro in terms of stimulation of MPC proliferation in response to combinations of cytokines, especially a potent co-stimulating cytokine such as steel factor, and suppression of this synergism by chemokines are of relevance to hematopoiesis in vivo. It is our belief that an evaluation of intracellular mechanisms triggered by these growth promoting and suppressing cytokines will enhance our understanding of normal MPC regulation, and will ultimately be of use in designing rational treatments for patients with hematological disorders. We believe that multi-growth factor induced synergistic stimulation of MPC proliferation and this inhibition by chemokines is, at least in part, a cell cycle phenomenon. Our hypothesis is that cell cycle regulators such as cyclin-dependent kinase inhibitor, p21cip1/waf1, certain cyclin-dependent kinases, and cyclins, and other intracellular signals are key to responses mediating proliferative synergy. Our biochemical results and evaluation of MPC proliferation from mice functionally deleted in some of these proposed intracellular mediators support this hypothesis. Our long-term goal is to define key intracellular molecules involved in these effects and to utilize this information for clinical benefit. Towards these goals we propose the following two Specific Aims: 1. Investigate intracellular mechanisms involved in multiple growth factor (GM-CSF and steel factor)-induced synergistic stimulation of the proliferation of MPC by: a) assessing a potential role for the cyclin-dependent kinase inhibitor/modulator p21cip1/waf1, and its relationship with cyclin-dependent kinases and cyclins in these effects and b) elucidating a potential role for the transcription factors BCL-6 and Stat4, the protein phosphatases SHP-1 and SHIP, and MAP kinase, and their possible interrelationship(s) with each other and with p21cip1/waf1 in these effects. 2. Evaluate intracellular mechanisms involved in suppression of multiple growth factor-induced synergistic stimulation of the proliferation of MPC by assessing comparative effects of myelosuppressive CC (MIP-1alpha, MCP-1), CXC (IL-8, IP-10), and C (lymphotactin) chemokines and the receptors they act on through: a) the cell cycle and regulatory molecules which control cell cycle progression and b) Intracellular signaling molecules linked to stimulation of MPC proliferation.