Vascular Biology in Cancer

1.IL-32在炎症性疾病中的作用IL-32是一种促炎细胞因子，在许多人类炎症性疾病中升高。有趣的是，IL-32在人类体内存在，但在啮齿动物中不存在。本项目的目的是了解IL-32在血管炎症中的作用和机制，以便建立更好的小鼠模型来模拟人类疾病。不幸的是，我们2012年的实地考察委员会建议我们停止这方面的研究。为了顺应这一点，我们正在努力完成我们已经开始的研究。这不仅对避免浪费我们在这个项目上使用的资源很重要，而且对参与这个项目的博士后研究员发表研究结果也很重要。目前，我们正在将调查结果总结为两份手稿。与CCR的Howard Young博士和临床中心的Alevizos Ilias博士合作，我们发现干燥综合征患者的IL-32水平升高，并且IL-32通过调节B细胞的迁移和功能促进自身免疫表型。淋巴网络对组织的动态平衡至关重要。淋巴的定向流动部分由淋巴阀控制。我们的发现表明，IL-32通过VEGFR3/NFATc1/FOXC2信号调节淋巴瓣的形成。2.Vav和β-catenin在血管形成和血管内稳态中的相互作用。根据我们现场访问委员会的建议，我们在过去的一年里重新定位了我们对这一领域研究的重点。血管形成是组织生长、修复和再生所必需的。如何区分正常发育中的生理性血管生成和疾病条件下的病理性血管生成是一个非常重要的问题。这对治疗干预具有重要意义。我们报道了一种神经性连环蛋白--β-连环蛋白在血管内皮细胞中也有表达，在小鼠中，只要缺失一个等位基因，就足以损害血管内皮细胞的运动和体外的血管组装以及体内的病理性血管生成，从而抑制肿瘤的生长。相反，Delta-catenin的一个或两个等位基因的缺失对激素诱导的子宫生理性血管生成没有影响。因为只有病理性血管生成对降低的β-连环蛋白水平敏感，这可能为抗血管生成治疗提供一个很好的靶点。进一步的分析表明，Delta-catenin通过与鸟嘌呤核苷酸交换因子Vav1相互作用来调节RhoGTP酶的活性。Vav1在造血细胞中特异表达，调节细胞分化。由于造血细胞和内皮细胞具有共同的祖细胞，Vav1也与血管内皮细胞有关。然而，它在血管系统中的功能还完全不清楚。为了系统和深入地研究Vav1在血管生物学中的作用和机制，我们在本物理年度建立了条件Vav1缺失和条件Vav1转基因小鼠品系。已知Vav1存在于造血细胞中。我们发现Vav1在血管内皮细胞中表达。然而，目前尚不清楚Vav1是否也在其他细胞系中表达，因为有报道称Vav1在几个上皮性癌细胞中表达。我们开发的Vav1小鼠系将使我们能够系统地分析Vav1在发育和疾病条件下的表达，因为报告基因是在内源性Vav1启动子下引入小鼠的。它们将使我们能够以特定细胞类型的方式研究Vav1的功能和机制。在细胞/器官培养和基因工程小鼠的结合中，我们已经证明：1)Vav1通过其针对RhoGTPase的全球环境基金功能来调节内皮细胞的运动和血管生成；2)Vav1通过稳定细胞表面的VE-钙粘连蛋白连接来调节血管的通透性；3)Vav1通过Sirt1调节eNOS的激活，从而调节血管功能和稳态；以及4)Vav1通过调节骨髓来源的血管前体细胞中CXCR4的表达和MMP9的激活来调节内皮祖细胞的产生和动员(血管生成)。有趣的是，我们发现间充质干细胞(MSC)中高水平表达Vav1。与软骨细胞和骨细胞相比，Vav1在MSC向脂肪细胞分化中的作用相反。总之，这些发现说明了Vav1和Delta catenin相互作用在血管形成和完整性中的重要和复杂的功能。目前，我们正在将这些新颖的发现总结成多个手稿，以便出版。合作为了确保研究的成功进展，我们与NIH内部和校外社区的调查人员密切合作。我们一直在与美国国立卫生研究院的Howard Young博士和Alevizos Ilias博士以及约翰霍普金斯大学的Michelle Petri博士合作进行自身免疫研究；与约翰霍普金斯大学的Dan E.Berkowitz博士合作研究Vav1/enos信号和血管功能；与芝加哥大学的Hans Schreiber博士和MD Anderson癌症中心的Tomasz Zal博士合作，通过T细胞介导的间质靶向实时体内成像消除癌症变异；与Vanderbilt大学的Vladimir R.Babaev博士合作研究巨噬细胞ikk-Akt信号在动脉粥样硬化发展中的作用；以及希望城国家医学中心的艾米丽·王博士研究癌细胞衍生的miR-105通过破坏抗转移的天然屏障促进肿瘤侵袭。

1. IL-32 in inflammatory diseases. IL-32 is a pro-inflammatory cytokine that is elevated in a number of inflammatory human diseases. Interestingly, IL-32 is present in humans, but absent in rodents. The purpose of this project was to understand the role and mechanism of IL-32 in vascular inflammation in order to generate better mouse models for modeling human diseases. Unfortunately, our 2012 site visit committee suggested us to discontinue this line of research. To comply with this, we are trying to finish studies we have already started. This is important not only to avoid wasting resources we have used for this project, but also important for post-doc fellows who had worked on this project to publish the findings. Currently, we are in the process to summarize the findings into two manuscripts. In collaboration with Dr. Howard Young at the CCR and Dr. Alevizos Ilias in the Clinical Center, we found that IL-32 is elevated in patient with Sjogren's syndrome, and IL-32 promotes autoimmune phenotype via regulation of B cell migration and function. Lymphatic networks are essential for tissue homeostasis. Directional lymphatic flow is partially controlled by lymphatic valves. Our findings suggest a novel role of IL-32 in regulation of lymphatic valve formation via VEGFR3/NFATc1/FoxC2 signaling. 2. Interaction of Vav and delta-catenin in vascular formation and vascular homeostasis. Based on our site visit committee's recommendation, we reposition our focus on this line of research in the past physical year. Vascular formation is essential for tissue growth, repair and regeneration. What distinguishes physiological angiogenesis during normal development from pathological angiogenesis in disease conditions is a very important question. It has significant implications for therapeutic interventions. We reported that delta-catenin, a neuronal catenin, is also expressed in vascular endothelial cells, and deletion of only one allele of delta-catenin in mice is sufficient to impair endothelial cell motility and vascular assembly in vitro and pathological angiogenesis in vivo, thereby inhibiting tumor growth. In contrast, deletion of one or both allele of delta-catenin had no effects on hormone-induced physiological angiogenesis in the uterus. Because only pathological angiogenesis is sensitive to decreased levels of delta-catenin, this may provide a good target for anti-angiogenic therapy. Further analysis suggests that delta-catenin regulates RhoGTPase activity via interacting with Vav1, a guanine nucleotide exchange factor (GEF). Vav1 is specifically expressed in hematopoietic cells and regulates cell differentiation. Since hematopoietic cells and endothelial cells share a common progenitor, Vav1 was also implicated in vascular endothelium. However, its function in the vasculature is totally unknown. To systemically and thoroughly study the role and mechanism of Vav1 in vascular biology, we have generated conditional Vav1 null and conditional Vav1 transgenic mouse lines during this physical year. Vav1 is known to be present in hematopoietic cells. We showed expression of Vav1 in vascular endothelial cells. However, it is currently unknown if Vav1 is also expressed in other cell lineages considering reports of Vav1 expression in several epithelial cancer cells. The Vav1 mouse lines we developed will allow us to systemically analyze Vav1 expression during development and disease conditions since a reporter gene was introduced in the mice under the endogenous Vav1 promoter. They will allow us to investigate Vav1 function and mechanisms in a cell type specific manner. In combination of cell/organ cultures and genetically engineered mice, we have demonstrated that 1) Vav1 regulates endothelial cell motility and angiogenesis via its GEF function for RhoGTPase; 2) Vav1 regulates vascular permeability through stabilization VE-Cadherin junctions on cell surface; 3) Vav1 regulates eNOS activation via Sirt1 thereby regulating vascular function and homeostasis; and 4) Vav1 regulates endothelial progenitor cell production and mobilization (vasculogenesis) via regulating CXCR4 expression and MMP9 activation in bone marrow derived vascular progenitor cells. Interestingly, 5) we found high levels of Vav1 expression in mesenchymal stem cells (MSC). Vav1 plays opposite roles in MSC differentiation toward adipocyte in comparison to chondrocyte and osteocyte. Together, these findings illustrate important and complex functions of Vav1 and delta catenin interaction in vascular formation and integrity. Currently, we are in the process to summarize these novel findings into multiple manuscripts for publication. Collaborations To ensure successful progress in research, we have worked closely with investigators within the NIH and extramural community. We have been working with Drs. Howard Young and Alevizos Ilias at the NIH, and Dr. Michelle Petri at the Johns Hopkins University on autoimmune studies; with Dr. Dan E. Berkowitz at the Johns Hopkins University on Vav1/eNOS signaling and vascular functions; with Drs. Hans Schreiber at the University of Chicago and Tomasz Zal at the MD Anderson Cancer Center on real-time in vivo imaging of cancer variant elimination through T cell-mediated stromal targeting; with Dr. Vladimir R. Babaev at the Vanderbilt University on macrophage IKK-Akt signaling in atherosclerosis development; and Dr. Emily Wang at the City of Hope National Medical Center on cancer cell-derived miR-105 in promoting tumor invasion through destroying the natural barriers against metastasis.