Determination of myeloid cell subpopulations during intestinal cell invasion

肠细胞侵袭过程中骨髓细胞亚群的测定

• 批准号: 8716846
• 负责人: Zev B Einhorn
• 金额: $ 5.15万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-09 至 2017-05-08
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8716846
• 关键词: Adenocarcinoma; Adenomatous Polyps; Animals; Architecture; Basement membrane; Benign; Biological Assay; Biological Models; Biological Process; Cancer Cell Growth; Cancer Etiology; Cancer Prognosis; Candidate Disease Gene; Cell physiology; Cells; Cessation of life; Colorectal Cancer; Complex; Connective Tissue; Development; Disease; Disseminated Malignant Neoplasm; Dyes; Epithelial; Epithelial Cells; Epithelium; Gene Expression; Genes; Genetic; Goals; Homeostasis; Human; Immune; Immune system; In Situ Hybridization; In Vitro; Infection; Inflammatory; Injury; Intestines; Ischemia; Label; Laboratories; Larva; Lead; Link; Lymphatic System; Malignant Neoplasms; Membrane; Methods; Modeling; Mutation; Myelogenous; Myeloid Cell Activation; Myeloid Cells; Neoplasm Metastasis; Normal tissue morphology; Oncogenic; Organ; Pathogenesis; Pathway interactions; Phenotype; Play; Population; Process; Reporter; Reporter Genes; Role; Signal Pathway; Signal Transduction; Site; Smooth Muscle; Smooth Muscle Myosins; Staging; Stromal Cells; Stromal Invasion; Supporting Cell; System; Testing; Time; Tissues; Transgenic Organisms; Validation; Work; Wound Healing; Zebrafish; cancer cell; cell motility; cell type; clinically significant; density; in vivo; in vivo Model; insight; loss of function; macrophage; molecular phenotype; mutant; neutrophil; novel; novel marker; prognostic; promoter; public health relevance; regenerative; repaired; research study; response; tissue regeneration; tumor; tumor progression

DESCRIPTION (provided by applicant): Stromal cells are increasingly recognized as playing an essential role in tissue regenerative and reparative processes. In the intestine, a rich population of connective tissue and immune cells supports the function and architecture of the continuously renewing epithelium. Disruption of epithelial homeostasis, such as occurs with tissue injury or in cancer, leads to the recruitment and activation of myeloid cells of the innate immune system. These play diverse roles in tissue defense and repair and in certain instances contribute to disease pathogenesis. This is particularly true in colorectal cancer. Recent work has shown that the transition of benign adenomatous polyps to adenocarcinoma is accompanied by an increase in the number of infiltrating immune cells. Furthermore, colorectal cancer survival has been linked to both macrophage and neutrophil density within the tumors. How immune cells regulate cancer progression is poorly understood, but there is good evidence that these cells can contribute to cancer cell growth, invasion and metastasis. The goal of this proposal is to study immune cell function in an in vivo, zebrafish cell invasion model that shares important features with currently accepted models of human cancer cell invasion that were developed using in vitro systems. This proposal consists of two specific aims. The goal of the first aim is to define the innate immune cell population in the intestine of wild type zebrafish larvae and myh11-meltdown mutants that develop epithelial cell invasion in response to unregulated smooth muscle myosin function. Immune cells will be localized in vivo using transgenic lines that express fluorescent reporter genes under the control of established myeloid cell promoters. Additionally, the use of fluorescent reporters and specific labeling dyes will be used to assess the activation pathways upregulated in polarized immune cells. Time-lapse recordings will be used to study interactions between the immune cells and the invasive epithelial cells. Genetic loss-of-function studies will be used to assess what role if any these cells play in the cell invasion phenotype. Comparable studies will be performed in a transgenic line that drives formation of epithelial cell invadopodia but that does not support stromal invasion of the epithelial cells, thus allowing immune cells analyses at specific stages of the invasive process. The goal of the second aim will be to perform transcriptional profiling of innate immune cells in the two zebrafish models, and to correlate changes in gene expression with myeloid cell subtypes, particularly those linked to cancer progression as well as functionally assay the importance of differentially regulated genes to the invasion process. Collectively, these studies will enhance our understanding of how immune cells regulate tissue architecture in the intestine. Furthermore, they may lead to the identification of markers of novel innate immune cell subtypes that have prognostic significance in colorectal cancer.

描述（由申请人提供）：基质细胞在组织再生和修复过程中发挥着重要作用。在肠道中，丰富的结缔组织和免疫细胞支持着不断更新的上皮的功能和结构。上皮稳态的破坏，如发生在组织损伤或癌症中，导致先天免疫系统的骨髓细胞的募集和激活。它们在组织防御和修复中起着不同的作用，在某些情况下有助于疾病的发病机制。在结直肠癌中尤其如此。最近的研究表明，良性腺瘤性息肉向腺癌的转变伴随着浸润性免疫细胞数量的增加。此外，结直肠癌的存活与肿瘤内巨噬细胞和中性粒细胞密度有关。免疫细胞如何调节癌症进展尚不清楚，但有充分的证据表明，这些细胞可以促进癌细胞的生长、侵袭和转移。本提案的目标是研究斑马鱼体内细胞入侵模型中的免疫细胞功能，该模型与目前使用体外系统开发的人类癌细胞入侵模型具有相同的重要特征。这项建议包括两个具体目标。第一个目标是确定野生型斑马鱼幼虫和myh11-熔解突变体肠道内的先天免疫细胞群，myh11-熔解突变体在平滑肌肌球蛋白功能不调节的情况下发生上皮细胞入侵。免疫细胞将被定位在体内使用转基因系表达荧光报告基因在既定的髓细胞启动子的控制下。此外，荧光报告和特异性标记染料的使用将用于评估极化免疫细胞中上调的激活途径。延时录像将用于研究免疫细胞和侵袭性上皮细胞之间的相互作用。基因功能丧失研究将用于评估这些细胞在细胞侵袭表型中发挥的作用。类似的研究将在转基因细胞系中进行，该细胞系驱动上皮细胞侵过性形成，但不支持上皮细胞的基质侵袭，因此允许在侵袭过程的特定阶段进行免疫细胞分析。第二个目标将是在两种斑马鱼模型中执行先天免疫细胞的转录谱分析，并将基因表达变化与骨髓细胞亚型，特别是与癌症进展相关的基因表达变化联系起来，以及功能分析差异调节基因对入侵过程的重要性。总的来说，这些研究将增强我们对免疫细胞如何调节肠道组织结构的理解。此外，它们可能导致新标记的识别