Vascular Biology in Cancer

1. 过继T细胞转移后实体瘤破坏的纵向共聚焦显微镜成像。一种基于荧光的高分辨率成像方法被开发出来，用于纵向可视化T细胞介导的肿瘤破坏过程中展开的细胞事件。T细胞、癌细胞和间质细胞的动态相互作用——在体内都有颜色编码——在植入小鼠背部的窗户后形成的实体瘤中进行了分析。在过继性T细胞治疗之前、期间和之后，可以在精确的同一肿瘤区域内重复跟踪事件，从而首次实现对长期事件的纵向体内评估，这是手术暴露肿瘤的终末成像无法实现的分析。研究了T细胞浸润、基质相互作用和血管破坏及其功能后果，包括消除癌细胞和癌细胞变异。微小的血管周围T细胞浸润引起肿瘤肿块内血管破坏，最终导致肉眼可见的中央肿瘤坏死。T细胞与肿瘤抗原交叉呈递基质细胞的长时间接触与高干扰素γ细胞因子释放和抗原阴性癌细胞的旁观者消除相关。这里描述的这种高分辨率、纵向、体内成像方法将有助于进一步更好地理解T细胞和其他抗癌疗法根除肿瘤的机制。这项研究发表在2013年的《肿瘤免疫学》杂志上。2. 肿瘤分泌的miR-105破坏血管内皮屏障促进转移。在这项研究中，我们着手鉴定通过适应生态位细胞参与癌症转移的癌症分泌的mirna。我们的研究结果表明，miR-105通过靶向细胞紧密连接，在早期转移前生态位形成过程中破坏宿主血管内皮屏障中发挥重要作用。在乳腺癌患者中，循环中miR-105水平的升高可以在转移前阶段检测到，并与转移的发生相关。Anti-miR-105治疗抑制转移并消除肿瘤源性miR-105对生态位适应的全身作用。因此，这些观察结果强烈提示miR-105作为预测或早期诊断血源性标志物以及乳腺癌转移的治疗靶点的临床应用。该研究作为专题文章发表在2014年的Cancer Cell杂志上，并在Sci上发表评论。2 .信号、自然医学、自然遗传学、自然癌症、自然药物发现、SciBX和癌症研究。Vav1在保护心脏免受缺血损伤的血管稳态中的关键作用。心肌灌注是心脏稳态的重要组成部分。不能诱导足够的灌注是心肌功能障碍和心力衰竭的主要原因。这项研究揭示了Vav1在缺血应激下维持心脏稳态的新功能。Vav1是一种已知在造血细胞中表达的GEF蛋白。由于血液和内皮细胞具有共同的祖细胞，我们发现Vav1在心脏冠状动脉内皮中表达，但在心肌细胞中不表达。值得注意的是，小鼠Vav1基因缺失会导致病理性心脏肥厚的发展，并随着年龄的增长导致左心室功能受损，这与冠状动脉血管密度的显著降低有关。这一发现与GEF在调节内皮细胞运动和血管生成中的功能是一致的。尽管幼年时Vav1缺失小鼠和WT小鼠在左室功能和血管密度上没有差异，但当异丙肾上腺素诱导心肌肥厚时，Vav1缺失小鼠表现出脆弱的心脏表型。此外，Vav1缺陷小鼠在冠状动脉结扎后表现出猝死倾向，这与血管渗漏和内皮细胞凋亡增加有关。分子分析表明，Vav1通过Rac1调控VE-cadherin粘附连接的形成，并通过p38/HIF1a/ vegf信号通路调控内皮细胞存活。重要的是，Vav1在缺血性心脏病患者和缺血性小鼠模型的心脏组织中升高。总的来说，这些发现有力地支持了Vav1在缺血应激下冠状动脉血管生物学和心脏稳态中的保护作用，这是通过其对血管生成、血管完整性和内皮细胞存活的功能介导的。4. C/ ebp - α和- δ在髓源性抑制细胞（MDSCs）的扩增中发挥相反的作用，C/EBP-d激活内皮细胞中的自分泌VEGF信号。MDSCs和血管内皮细胞是构成肿瘤微环境的两个重要组成部分。靶向这些细胞提供了阻止肿瘤生长的潜力。MDSCs在癌症患者和荷瘤小鼠体内大量扩增。它们渗透到肿瘤中，通过抑制免疫和促进血管生成来调节肿瘤微环境。然而，是什么调控了MDSCs在肿瘤条件下的扩增尚不清楚。在一系列研究中，我们发现C/EBP-a和-d在MDSCs的扩增中发挥相反的作用。肿瘤条件抑制了C/EBPa的表达，同时增加了C/EBP-d在MDSCs中的表达。髓细胞中C/EBPa的条件性缺失对正常发育的MDSCs数量没有影响。有趣的是，它在肿瘤条件下显著增强MDSC的增殖和扩张，提示在肿瘤诱导的MDSC扩张中具有特定作用。此外，MDSCs中C/EBPa的缺失通过诱导VEGF、MMP9和NO来增强细胞的血管生成功能，并通过增加精氨酸酶1和iNOS的产生来增强免疫抑制活性。因此，在C/EBPa髓系条件缺失小鼠中生长的肿瘤表现出更多的MDSC浸润，增加的血管化和加速的肿瘤生长。另一方面，C/EBP-d在肿瘤来源的MDSCs中升高。在小鼠中，该基因的基因缺失会显著损害MDSC在肿瘤进展中的扩增，但对正常发育中MDSC的产生没有影响。此外，C/EBP-d也在血管内皮细胞和淋巴内皮细胞中表达。它直接调节细胞运动，以及血管和淋巴内皮网络的形成。值得注意的是，C/EBP-d的缺失特异性抑制血管内皮细胞中VEGFR2和淋巴内皮细胞中VEGFR3的表达。此外，C/EBP-d分别调节血管内皮细胞和淋巴内皮细胞中内源性VEGFA和VEGFC的表达，以及细胞生存所必需的自分泌VEGF信号。因此，缺乏C/EBP-d的小鼠在缺血条件下表现出内皮细胞凋亡增加、组织损伤和器官功能受损。在分子水平上，缺氧通过HIF-1a诱导C/EBP-d表达，C/EBP-d与HIF-1a启动子结合，激活HIF-1a转录，形成内源性VEGF生成、自分泌VEGF信号传导和血管存活的正反馈循环。总的来说，这些研究确定了C/EBP-a和C/EBP-d在内皮细胞中MDSC扩增和VEGF/VEGFR表达中的关键作用。考虑到内皮细胞中MDSCs和VEGF信号在肿瘤进展中的重要性，以及缺血应激下血管存活和组织稳态中VEGF自分泌信号的重要性，这些结果在肿瘤生物学和血管生物学中具有更广泛的意义。

1. Longitudinal confocal microscopy imaging of solid tumor destruction following adoptive T cell transfer. A fluorescence-based, high-resolution imaging approach was developed to visualize longitudinally the cellular events unfolding during T cell-mediated tumor destruction. The dynamic interplay of T cells, cancer cells, and stromal cells-all color-coded in vivo-was analyzed in established, solid tumors that had developed behind windows implanted on the backs of mice. Events could be followed repeatedly within precisely the same tumor region-before, during and after adoptive T cell therapy-thereby enabling for the first time a longitudinal in vivo evaluation of protracted events, an analysis not possible with terminal imaging of surgically exposed tumors. T cell infiltration, stromal interactions, and vessel destruction, as well as the functional consequences thereof, including the elimination of cancer cells and cancer cell variants were studied. Minimal perivascular T cell infiltrates initiated vascular destruction inside the tumor mass eventually leading to macroscopic central tumor necrosis. Prolonged engagement of T cells with tumor antigen-cross presenting stromal cells correlated with high IFNgamma cytokine release and bystander elimination of antigen-negative cancer cells. This high-resolution, longitudinal, in vivo imaging approach described here will help to further a better mechanistic understanding of tumor eradication by T cells and other anti-cancer therapies. This work was published in Oncoimmunology 2013. 2. Cancer-secreted miR-105 destroys vascular endothelial barriers to promote metastasis. In this study, we set out to identify cancer-secreted miRNAs that participate in cancer metastasis by adapting the niche cells. Our results demonstrate an important role of miR-105 in destroying the vascular endothelial barriers in the host during early premetastatic niche formation by targeting the cellular tight junctions. In breast cancer patients, increased levels of miR-105 in the circulation can be detected at the premetastatic stage and correlate with the occurrence of metastasis. Anti-miR-105 treatment suppresses metastasis and abolishes the systemic effect of tumor-derived miR-105 on niche adaptation. Therefore, these observations strongly suggest clinical applications of miR-105 as a predictive or early diagnostic blood-borne marker as well as a therapeutic target for breast cancer metastasis. This work was published as a featured article in Cancer Cell 2014, received commentary in Sci. Signal, Nature Medicine, Nature Review Genetics, Nature Review Cancer, Nature Review Drug Discovery, SciBX, and Cancer Research. 3. Critical roles of Vav1 in vascular homeostasis that protects the heart from ischemia-induced injury. Myocardial perfusion is a key component of cardiac homeostasis. Failure to induce sufficient perfusion represents a major cause of myocardial dysfunction and heart failure. This study reveals a novel function of Vav1 in maintaining cardiac homeostasis under ischemic stress. Vav1 is a GEF protein known to be expressed in hematopoietic cells. Since blood and endothelial cells share a common progenitor, we found expression of Vav1 in coronary endothelium in the heart, but not in myocardiocytes. Notably, genetic deletion of Vav1 in mice leads to the development of pathological cardiac hypertrophy with impaired left ventricular (LV) function with aging, which is associated with a significant reduction of coronary vascular density. This finding is consistent with its GEF function in regulating endothelial cell motility and angiogenesis. Although there is no difference in LV function or vascular density between the Vav1 null and WT mice at a young age, the Vav1 null mice show a fragile cardiac phenotype when cardiac hypertrophy is induced by isoproterenol administration. Moreover, Vav1 deficient mice exhibit a tendency to sudden death after coronary ligation, which is associated with leaky vasculature and increased endothelial apoptosis. Molecular analysis reveals that Vav1 regulates VE-cadherin adherens junctions formation through Rac1 and endothelial survival via the p38/HIF1a/VEGF-signaling pathway. Importantly, Vav1 is elevated in heart tissues from patients with ischemic heart conditions and ischemic mouse models. Collectively, these findings strongly support a protective role of Vav1 in coronary vascular biology and cardiac homeostasis under ischemic stress, and this is mediated through its functions on angiogenesis, vascular integrity and endothelial survival. 4. C/EBP-alpha and -delta play opposite roles in the expansion of myeloid derived suppressor cells (MDSCs), and C/EBP-d activates autocrine VEGF signaling in endothelial cells. MDSCs and vascular endothelial cells are two important components that constitute the tumor microenvironment. Targeting these cells offers the potential to halt tumor growth. MDSCs are greatly expanded in cancer patients and tumor-bearing mice. They infiltrate into tumors and modulate the tumor microenvironment through immune suppression and promotion of angiogenesis. However, what regulates the expansion of MDSCs in tumor conditions is less clear. In a series of studies, we found that C/EBP-a and -d play opposite roles in the expansion of MDSCs. Tumor conditions inhibit the expression of C/EBPa and concomitantly increases expression of C/EBP-d in MDSCs. Conditional deletion of C/EBPa in myeloid cells has no effect on the number of MDSCs in normal development. Interestingly, it results in significantly enhanced MDSC proliferation and expansion in tumor conditions, suggesting a specific role in tumor-induced MDSC expansion. In addition, deletion of C/EBPa in MDSCs enhances the angiogenic function of the cells via an induction of VEGF, MMP9 and NO, as well as immune suppression activity through increased arginase 1 and iNOS production. Accordingly, tumors growing in C/EBPa myeloid conditional null mice display greater MDSC infiltration, increased vascularization and accelerated tumor growth. On the other hand, C/EBP-d is elevated in tumor derived MDSCs. Genetic deletion of the gene in mice significantly impairs MDSC expansion in response to tumor progression, but it has no effect on MDSC production in normal development. Moreover, C/EBP-d is also expressed in vascular and lymphatic endothelial cells. It directly regulates cell motility, and vascular and lymphatic endothelial network formation. Notably, loss of C/EBP-d specifically inhibits the expression of VEGFR2 in vascular and VEGFR3 in lymphatic endothelial cells. In addition, C/EBP-d regulates the expression of endogenous VEGFA and VEGFC in vascular and lymphatic endothelial cells, respectively, and autocrine VEGF signaling essential for cell survival. Accordingly, mice without C/EBP-d exhibit increased endothelial apoptosis, tissue damage, and compromised organ function under ischemic conditions. At the molecular level, hypoxia induces C/EBP-d expression via HIF-1a, and C/EBP-d binds to the HIF-1a promoter and activates HIF-1a transcription, thereby forming a positive feedback loop to propagate endogenous VEGF production, autocrine VEGF signaling and vascular survival. Collectively, these studies identify critical roles of C/EBP-a and C/EBP-d in MDSC expansion and VEGF/VEGFR expression in endothelial cells. Considering the importance of MDSCs and VEGF signaling in endothelium in tumor progression as well as VEGF autocrine signaling in vascular survival and tissue homeostasis under ischemic stress, these results have broader implications in tumor biology and vascular biology.