MOLECULAR BASIS OF MICROENVIRONMENTAL CELL CYCLE CONTROL

微环境细胞周期控制的分子基础

• 批准号: 8169374
• 负责人: James P Freyer
• 金额: $ 1.67万
• 依托单位: LOS ALAMOS NAT SECTY-LOS ALAMOS NAT LAB
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2011-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169374
• 关键词: Antibodies; Blood flow; Bromodeoxyuridine; Cell Cycle; Cell Cycle Arrest; Cell Cycle Regulation; Cell Proliferation; Cells; Cellular Spheroids; Computer Retrieval of Information on Scientific Projects Database; Cyclin-Dependent Kinases; Cyclins; DNA analysis; Data; Experimental Neoplasms; Funding; Grant; In Vitro; Institution; Label; Link; Location; Measurement; Modeling; Molecular; Molecular Analysis; Nutrient; Penetration; Ploidies; Radiation; Research; Research Personnel; Resources; Signal Transduction; Source; System; Techniques; Tumor Biology; United States National Institutes of Health; base; in vivo; neoplastic cell; tumor; uptake

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. One of the major unsolved mysteries in tumor biology is the mechanism by which tumor cells in vivo exit from the cell cycle in a reversible fashion. An implicit assumption through decades of research is that limitations of nutrient supply, either by limited penetration into the cell mass or restricted blood flow to local regions, create a stressful microenvironment which induces cells to arrest their cell cycle transit. Due to well-known limitations of experimental tumors for such mechanistic studies, we are using the multicellular tumor spheroid model for the majority of this project. Spheroids are ideally suited for such studies, both because of their symmetrical arrangement of microenvironmental and cellular proliferation gradients, and because of our unique ability to experimentally exploit this symmetry. Specifically, we can isolate intact, viable cells from known locations within the spheroid microenvironment for detailed study of the molecular changes associated with cell cycle arrest. In order to provide a link between this in vitro system and the in vivo situation we will determine whether our proposed mechanism is operative in actual tumors. We are pursuing four Specific Aims: 1) to determine the molecular basis for cell cycle arrest in multicellular spheroids; 2) to determine if the same molecular mechanisms are operative in tumors in vivo; 3) to identify the microenvironmental signal(s) which induce cell cycle arrest in spheroids; and 4) to determine the interaction between radiation- and microenvironmentally-induced cell cycle arrest. Flow analysis is used both for routine DNA content analysis, and also for determining the uptake of bromodeoxyuridine by means of a dual-label DNA analysis technique. These cell cycle data are critical for comparison with our molecular analysis. We are also pursuing the measurement of cyclin and cyclin-dependent kinase expression by flow using fluorescently-tagged antibodies.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目及 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 中心，不一定是研究者的机构。 肿瘤生物学中未解的主要谜团之一是体内肿瘤细胞以可逆方式退出细胞周期的机制。 数十年研究的一个隐含假设是，营养供应的限制，无论是通过限制渗透到细胞团还是限制血液流向局部区域，都会产生一个有压力的微环境，诱导细胞阻止其细胞周期转变。 由于此类机制研究的实验肿瘤存在众所周知的局限性，我们在该项目的大部分内容中使用多细胞肿瘤球体模型。 球体非常适合此类研究，既因为它们的微环境和细胞增殖梯度的对称排列，也因为我们通过实验利用这种对称性的独特能力。 具体来说，我们可以从球体微环境中的已知位置分离完整的、活的细胞，以详细研究与细胞周期停滞相关的分子变化。 为了提供该体外系统与体内情况之间的联系，我们将确定我们提出的机制是否在实际肿瘤中有效。 我们正在追求四个具体目标：1）确定多细胞球体中细胞周期停滞的分子基础； 2) 确定相同的分子机制是否在体内肿瘤中发挥作用； 3) 识别诱导球体细胞周期停滞的微环境信号； 4) 确定辐射和微环境诱导的细胞周期停滞之间的相互作用。 流式分析既可用于常规 DNA 含量分析，也可用于通过双标记 DNA 分析技术测定溴脱氧尿苷的摄取。 这些细胞周期数据对于与我们的分子分析进行比较至关重要。 我们还致力于使用荧光标记抗体通过流式测量细胞周期蛋白和细胞周期蛋白依赖性激酶表达。