It is now widely recognized that a strong correlation exists between cancer and aberrant hemostasis. Patients with various types of cancers, including pancreatic, colorectal, and gastric cancer, often develop thrombosis, a phenomenon commonly referred to as Trousseau syndrome. Reciprocally, components from the coagulation cascade also influence cancer progression. The primary initiator of coagulation, the transmembrane receptor tissue factor (TF), has gained considerable attention as a determinant of tumor progression. On complex formation with its ligand, coagulation factor VIIa, TF influences protease-activated receptor-dependent tumor cell behavior, and regulates integrin function, which facilitate tumor angiogenesis both in vitro and in mouse models. Furthermore, evidence exists that an alternatively spliced isoform of TF also affects tumor growth and tumor angiogenesis. In patient material, TF expression and TF cytoplasmic domain phosphorylation correlate with disease outcome in many, but not in all, cancer subtypes, suggesting that TFdependent signal transduction events are a potential target for therapeutic intervention in selected types of cancer. In this review, we summarize our current understanding of the role of TF in tumor growth and metastasis, and speculate on anticancer therapy by targeting TF. (Blood. 2012; 119(4):924-932) IntroductionAfter Trousseau's description of thrombophlebitis as a complication of pancreatic cancer in the 19th century, the notion that increased expression of tissue factor (TF) underlies the relation between coagulation and cancer has become generally accepted. Full-length TF (flTF) is a 47-kDa membrane-bound glycoprotein that is present on subendothelial cells. 1 In the classic concept of coagulation, it is thought that endothelial disruption leads to exposure of flTF to the bloodstream. Exposed flTF can bind to its natural ligand factor VII (FVII), which then becomes activated FVII (FVIIa). The thus formed flTF/FVIIa complex converts factor X (FX) to factor Xa (FXa), and FXa in turn activates prothrombin leading to formation of thrombin (factor IIa). Thrombin subsequently activates platelets and converts fibrinogen into fibrin, 2 essential components of a stable hemostatic plug. 1 The primary function of subendothelial flTF is to serve as a hemostatic envelope surrounding the vasculature. However, under certain conditions, the expression of flTF is induced in monocytes and endothelial cells. flTF is also often expressed on cancer cells and the tumor vasculature, 2 and flTF-bearing microparticles can become shed by these cells. 3 These flTF-bearing microparticles are important contributors to the thrombotic phenotype in cancer patients. 3 The flTF/FVIIa complex also influences pathways that do not lead to blood coagulation, but rather activate cell-bound proteaseactivated receptors (PARs) that are of importance during the inflammatory and angiogenic response to injury. 4 Furthermore, a soluble variant of flTF, known as alternatively spliced TF (asTF), stimulates angiogen...

The initiator of coagulation, full-length tissue factor (flTF), in complex with factor VIIa, influences angiogenesis through PAR-2. Recently, an alternatively spliced variant of TF (asTF) was discovered, in which part of the TF extracellular domain, the transmembrane, and cytoplasmic domains are replaced by a unique C terminus. Subcutaneous tumors produced by asTF-secreting cells revealed increased angiogenesis, but it remained unclear if and how angiogenesis is regulated by asTF. Here, we show that asTF enhances angiogenesis in matrigel plugs in mice, whereas a soluble form of flTF only modestly enhances angiogenesis. asTF dose-dependently upregulates angiogenesis ex vivo independent of either PAR-2 or VIIa. Rather, asTF was found to ligate integrins, resulting in downstream signaling. asTF-␣V␤3 integrin interaction induces endothelial cell migration, whereas asTFdependent formation of capillaries in vitro is dependent on ␣6␤1 integrin. Finally, asTF-dependent aortic sprouting is sensitive to ␤1 and ␤3 integrin blockade and a TF-antibody that disrupts asTFintegrin interaction. We conclude that asTF, unlike flTF, does not affect angiogenesis via PAR-dependent pathways but relies on integrin ligation. These findings indicate that asTF may serve as a target to prevent pathological angiogenesis.cancer ͉ coagulation ͉ endothelial cells ͉ integrins

Full-length tissue factor (flTF), the coagulation initiator, is overexpressed in breast cancer (BrCa), but associations between flTF expression and clinical outcome remain controversial. It is currently not known whether the soluble alternatively spliced TF form (asTF) is expressed in BrCa or impacts BrCa progression. We are unique in reporting that asTF, but not flTF, strongly associates with both tumor size and grade, and induces BrCa cell proliferation by binding to β1 integrins. asTF promotes oncogenic gene expression, anchorage-independent growth, and strongly up-regulates tumor expansion in a luminal BrCa model. In basal BrCa cells that constitutively express both TF isoforms, asTF blockade reduces tumor growth and proliferation in vivo. We propose that asTF plays a major role in BrCa progression acting as an autocrine factor that promotes tumor progression. Targeting asTF may comprise a previously unexplored therapeutic strategy in BrCa that stems tumor growth, yet does not impair normal hemostasis.regulated pre-mRNA processing | outside-in signaling I n breast cancer (BrCa), proteins that modulate splicing events such as ASF/SF2 and SR(Serine/Arginine-rich)p55, are frequently up-regulated and contribute to cell transformation (1, 2). BrCa cells exhibit specific alternative splicing signatures that were proposed as potential prognostic factors in BrCa (3). Alternative splicing of proteins, such as spleen tyrosine kinase (Syk), p53, phosphatase and tensin homolog (PTEN), chemokine (C-X-C motif) receptor 3 (CXCR3), and ras-related C3 botulinum toxin substrate 1 (Rac1) impacts BrCa cell behavior and therefore, disease progression (4-8).Full-length tissue factor (flTF) is the initiator of blood coagulation (9). Following vascular damage, flTF binds its ligand FVII(a), which triggers clot formation. Aside from subendothelial tissues, flTF is also abundant on cancer cells (9) and fuels tumor progression by modulating integrin α3β1 function, cell migration (10), and FVIIa-dependent protease activated receptor (PAR)2 activation, but flTF-β1 integrin complexation enhances PAR2 activation (11). flTF-dependent PAR2 activation results in the production of VEGF, CXCL1, and IL-8, thus promoting the angiogenic switch and consequently, tumor growth in vivo (10, 11).Alternative splicing of TF pre-mRNA results in the deletion of exon 5 and thus a frameshift in exon 6, yielding a transmembrane domain-lacking isoform that can be secreted (12). Human and murine alternatively spliced TF (asTF) contain novel C termini with poor homology to one another or any other protein, and the murine asTF C terminus is longer than that of human asTF (12,13). High expression of asTF in tumor cell lines suggests a role in tumor progression (10,14). Subcutaneous growth of pancreatic cancer cells overexpressing asTF results in larger and more vascularized tumors (15). We recently discovered that asTF induces angiogenesis, independent of PAR2 activation, by acting as an integrin ligand (16). Thus, flTF and asTF facilitate cellular signaling via ...

Ischemia/reperfusion injury (IRI) is a central phenomenon in kidney transplantation and AKI. Integrity of the renal peritubular capillary network is an important limiting factor in the recovery from IRI. facilitates vascular regeneration by functioning as an angiomiR and by modulating mobilization of hematopoietic stem/progenitor cells. We hypothesized that overexpression of miR-126 in the hematopoietic compartment could protect the kidney against IRI via preservation of microvascular integrity. Here, we demonstrate that hematopoietic overexpression of miR-126 increases neovascularization of subcutaneously implanted Matrigel plugs in mice. After renal IRI, mice overexpressing miR-126 displayed a marked decrease in urea levels, weight loss, fibrotic markers, and injury markers (such as kidney injury molecule-1 and neutrophil gelatinase-associated lipocalin). This protective effect was associated with a higher density of the peritubular capillary network in the corticomedullary junction and increased numbers of bone marrow-derived endothelial cells. Hematopoietic overexpression of miR-126 increased the number of circulating Lin 2 /Sca-1 + /cKit + hematopoietic stem and progenitor cells. Additionally, miR-126 overexpression attenuated expression of the chemokine receptor CXCR4 on Lin 2 /Sca-1 + /cKit + cells in the bone marrow and increased renal expression of its ligand stromal cell-derived factor 1, thus favoring mobilization of Lin 2 /Sca-1 + /cKit + cells toward the kidney. Taken together, these results suggest overexpression of miR-126 in the hematopoietic compartment is associated with stromal cell-derived factor 1/CXCR4-dependent vasculogenic progenitor cell mobilization and promotes vascular integrity and supports recovery of the kidney after IRI.

Summary Background TF is highly expressed in cancerous and atherosclerotic lesions. Monocyte recruitment is a hallmark of disease progression in these pathological states. Objective To examine the role of integrin signaling in TF-dependent recruitment of monocytes by endothelial cells. Methods The expression of flTF and asTF in cervical cancer and atherosclerotic lesions was examined. Biologic effects of the exposure of primary microvascular endothelial cells (MVEC) to truncated flTF ectodomain (LZ-TF) and recombinant asTF were assessed. Results flTF and asTF exhibited nearly identical expression patterns in cancer lesions and lipid-rich plaques. Tumor lesions as well as stromal CD68+ monocytes/macrophages expressed both TF forms. Primary MVEC rapidly adhered to asTF and LZ-TF, and this was completely blocked by anti-β1 integrin antibody. asTF- and LZ-TF-treatment of MVEC promoted adhesion of peripheral blood mononuclear cells (PBMC) under orbital shear conditions and under laminar flow; asTF-elicited adhesion was more pronounced than that elicited by LZ-TF. Expression profiling and western blotting revealed a broad activation of cell adhesion molecules (CAMs) in MVEC following asTF treatment including E-selectin, ICAM-1, and VCAM-1. In transwell assays, asTF potentiated PMBC migration through MVEC monolayers by ~3 fold under MCP-1 gradient. Conclusions TF splice variants ligate β1 integrins on MVEC, which induces the expression of CAMs in MVEC and leads to monocyte adhesion and transendothelial migration. asTF appears more potent than flTF in eliciting these effects. Our findings underscore the pathophysiologic significance of non-proteolytic, integrin-mediated signaling by the two naturally occurring TF variants in cancer and atherosclerosis.

Belemnitellid cephalopods from the Maastrichtian stratotype area (southeast Netherlands) are shown to be comparatively well preserved. Although partial diagenetic alteration has been observed, micromilling techniques have permitted the extraction of pristine belemnite calcite, suitable for the reconstruction of strontium (Sr), oxygen (O) and carbon (C) isotope variation of Maastrichtian seawater. A distinct Sr isotope pattern in the Maastricht record can be matched stratigraphically with records from Hemmoor (northern Germany), El Kef (Tunisia) and ODP site 690 (Maud Rise, Antarctica), leading to a new chemostratigraphical age model for the Maastrichtian stratotype section. Our data improve currently applied strontium isotope stratigraphical reference curves by revealing an Sr isotope inflection pattern near the lower/upper Maastrichtian boundary that is a potentially diagnostic feature for intra-Maastrichtian stratigraphical correlation between distant sections. Belemnites further show significant stratigraphical oxygen isotope variation through the Maastrichtian. We interpret this variation to have resulted from palaeoceanographic reorganisations in the Atlantic Ocean during this time interval.

See also Swystun LL, Mukherjee S, Liaw PC. Breast cancer chemotherapy induces the release of cell‐free DNA, a novel procoagulant stimulus. This issue, pp 2313–21.

This study was performed to determine whether murine alternatively spliced tissue factor (masTF) acts analogously to human alternatively spliced tissue factor (hasTF) in promoting neovascularization via integrin ligation. Immunohistochemical evaluation of a spontaneous murine pancreatic ductal adenocarcinoma model revealed increased levels of masTF and murine full-length tissue factor (mflTF) in tumor lesions compared with benign pancreas; furthermore, masTF colocalized with mflTF in spontaneous aortic plaques of Ldlr -/-mice, indicating that masTF is likely involved in atherogenesis and tumorigenesis. Recombinant masTF was used to perform in vitro and ex vivo studies examining its integrin-mediated biologic activity. Murine endothelial cells (ECs) rapidly adhered to masTF in a β3-dependent fashion. Using adult and embryonic murine ECs, masTF potentiated cell migration in transwell assays. Scratch assays were performed using murine and primary human ECs; the effects of masTF and hasTF were comparable in murine ECs, but in human ECs, the effects of hasTF were more pronounced. In aortic sprouting assays, the potency of masTF-triggered vessel growth was undistinguishable from that observed with hasTF. The proangiogenic effects of masTF were found to be Ccl2-mediated, yet independent of vascular endothelial growth factor. In murine ECs, masTF and hasTF upregulated genes involved in inflammatory responses; murine and human ECs stimulated with masTF and hasTF exhibited increased interaction with murine monocytic cells under orbital shear. We propose that masTF is a functional homolog of hasTF, exerting some of its key effects via β3 integrins. Our findings have implications for the development of murine models to examine the interplay between blood coagulation, atherosclerosis and cancer.