Key Points
-
Modalities of cell–cell communication of relevance to angiogenesis, inflammation and fibrosis include paracrine signalling, mechanosignalling, direct signal transduction via gap junctions and tunnelling nanotubes, and communication via the release and uptake of exosomes. Intercellular communication might be targeted by altering gap junctions and nanotubes, and exosomes are potential drug delivery vehicles as well as diagnostic markers.
-
Leukocytes interact with microvascular cells to affect angiogenesis. These interactions are potentially druggable processes for the regulation of angiogenesis.
-
Angiogenesis can occur through three different mechanisms — sprouting, intussusception and looping — which should be taken into account when designing new pharmacological treatments to reduce or increase angiongesis.
-
Different classes of leukocytes are potential targets to reduce uncontrolled fibrosis. Inhibited recruitment of inflammatory leukocytes together with on-site reprogramming of macrophages could promote fibrosis resolution.
-
In response to different signals from the environment, such as interferons, Toll-like receptor ligands or interleukins, macrophages undergo classical (M1) or alternative activation, which results in a continuum of diverse phenotypes depending on activation states. The identification of the underlying regulation of macrophage plasticity and polarized activation provides targets for macrophage-centred therapies.
-
Delineation of the endothelial heterogeneity is important to the development of strategies for the targeted delivery of drugs to organ-specific vascular beds to limit adverse effects.
Abstract
Regulation of vascular permeability, recruitment of leukocytes from blood to tissue and angiogenesis are all processes that occur at the level of the microvasculature during both physiological and pathological conditions. The interplay between microvascular cells and leukocytes during inflammation, together with the emerging roles of leukocytes in the modulation of the angiogenic process, make leukocyte–vascular interactions prime targets for therapeutics to potentially treat a wide range of diseases, including pathological and dysfunctional vessel growth, chronic inflammation and fibrosis. In this Review, we discuss how the different cell types that are present in and around microvessels interact, cooperate and instruct each other, and in this context we highlight drug targets as well as emerging druggable processes that can be exploited to restore tissue homeostasis.
Similar content being viewed by others
Main
The discovery that the growth of solid tumours beyond a few millimetres in diameter is dependent on angiogenesis1 has inspired the development of research fields related to blood vessel formation since the 1970s. Tumours were subsequently thought of as analogous to wounds that do not heal2, and angiogenesis and components of the immune system have been shown to have crucial roles in tumour growth, wound healing and tissue restoration. Angiogenesis is clearly regulated by many different cell types that are present in the local vascular microenvironment, including specialized subpopulations of immune cells3,4,5,6. During inflammation, endothelial cells have a crucial role in recruiting circulating leukocytes to the tissue7, and mural cells are also involved in the guidance of leukocytes to sites of inflammation8. The local signals that regulate angiogenesis and inflammation are transmitted between cells by paracrine factors, secreted hormones and exosomes, homophilic and heterophilic cell–cell interactions, gap junctions and the recently described tunnelling nanotubes (TnTs)9. Importantly, the involvement of specific cell types in these processes can now be determined by high-resolution intravital microscopy. Cell migration, cell interactions and the presence and involvement of specific signalling molecules can thus be directly visualized in experimental in vivo models of development and disease, which, together with single cell gene profiling, will help to accelerate the identification of new drug targets10,11. In this Review, we discuss some of the interactions between the different cells that regulate angiogenesis and inflammation. Several new drug targets related to the communication between leukocytes and vascular cells — including interleukins, chemotactic cytokines (chemokines), and molecules within the Notch, WNT and transforming growth factor-β (TGFβ) signalling pathways — could be exploited for the treatment of diseases that involve angiogenesis or inflammation.
Microvascular cells, leukocytes and angiogenesis
The microvasculature is the part of the vascular system that holds the smallest vessels: the terminal arterioles, the capillary network and the postcapillary venules (Fig. 1). Meticulous control of blood flow and vascular permeability are central to tissue and organ performance and are regulated by the microvasculature. Most of the gas, solute and hormone exchange between blood and tissue occurs in the microvasculature, which also has the highest regenerative capacity and is consequently the most active vascular compartment during angiogenesis. Leukocyte extravasation during inflammation also occurs predominantly in postcapillary venules.
Mural cells and perivascular cells that can influence the functions and growth of the microvasculature are shown. The microvasculature is the most active vascular compartment during angiogenesis. Recent findings suggest that many cells have the capacity to affect angiogenesis. Leukocytes, including neutrophils, macrophages and mast cells, are potent regulators of angiogenesis and can release growth factors that modulate angiogenesis. In addition, it was recently suggested that macrophages mediate the fusion of angiogenic sprouts. Pericytes have been shown to mediate blood vessel maturation and leukocyte trafficking. Furthermore, smooth muscle cells, pericytes and mesenchymal stem cells (MSCs) can give rise to fibroblasts and myofibroblasts that are important regulators of angiogenesis, wound healing and fibrosis. Nerves are often located close to vessels and can modulate vascular tone as well as vessel formation.
Endothelial cells. Endothelial cells form the inner lining of all blood vessels and are in contact with each other through intercellular tight junctions and adherens junctions. Endothelial junctions control various key aspects of vascular homeostasis. For example, the regulated opening and closing of endothelial junctions occurs during leukocyte extravasation to inflamed tissue and also controls vascular permeability and affects communication between endothelial cells during angiogenesis12,13. Increased vascular permeability is manifested during many malignant and nonmalignant diseases. Oedema is a cardinal sign of inflammation and is the consequence of locally released inflammatory agents (such as bradykinin, platelet-activating factor and histamine) that affect adjacent blood vessels to increase junctional leakage.
The pathology of solid tumours as well as chronic inflammatory conditions, age-related macular degeneration, psoriasis, hereditary haemorrhagic telangiectasia and the formation of atherosclerotic plaques all involve the formation of microvascular networks with abnormal structure and function. In these pathologies, excessive expression of angiogenic factors such as vascular endothelial growth factor A (VEGFA) or of cytokines (such as bradykinin) and histamine destabilize vascular–endothelial cell cadherin (VE-cadherin)-mediated endothelial cell–cell interactions13. The resulting pathologically increased vascular permeability leads to disease progression with local oedema, elevated interstitial pressure and impaired blood flow. Means to normalize the functions of the microvasculature can both curb the development of disease and enhance the delivery of blood-borne pharmaceuticals to the target tissue, owing to reduced vascular leakage with concomitantly reduced interstitial pressure and increased blood flow14,15.
Endothelial cell heterogeneity. Endothelial cells are equipped with sensors that measure the level of shear stress generated by blood flow16,17. Shear stress induces signals that lead to the adaptation of individual endothelial cells and the development of situation-dependent functional properties. Endothelial cells in high-flow arterioles and arteries are elongated and narrow, whereas endothelial cells in low-flow veins and venules have cobblestone-like morphologies. Notably, the endothelial cells of arteries, capillaries and veins have differential gene expression and molecular profiles18 before the onset of circulatory flow19, indicating the importance of local microenvironmental factors to endothelial cell identity. Some key regulators of arterial–venous specification have been identified, including sonic hedgehog. Sonic hedgehog induces the expression of VEGFA20, which, via vascular endothelial growth factor receptor 2 (VEGFR2) and the phospholipase Cγ (PLCγ)–RAF–extracellular signal-regulated kinase (ERK) cascade, increases Notch and ephrin B2 expression19,21. These molecules, together with reduced ephrin type-B receptor 4, enhance the arterial identity.
Delineation of the endothelial heterogeneity during health and disease is important to the development of strategies for the targeted delivery of pharmaceutical compounds to organ-specific vascular beds, as this could limit adverse effects. For example, leukocytes use different adhesion molecules to interact with the vascular beds of different organs, which could be utilized to direct drugs to a specific sites22. In a model of glomerulonephritis, expression of the adhesion molecule E-selectin by activated glomerular endothelium was used to direct E-selectin-targeting immunoliposomes containing dexamethasone predominantly to the glomerular endothelium23. As a result, pro-inflammatory gene expression was strongly reduced in the inflamed glomeruli and adverse effects normally associated with dexamethasone were absent23,24.
The transcription factor SOX17 was recently shown to be selectively expressed in arteries and suggested to act upstream of Notch, but downstream of WNT25. Canonical WNT signalling, which acts through β-catenin, is activated by the binding of WNTs to Frizzled receptor complexes, but also by cell–cell interactions between cadherins. Abnormally high levels of WNT signalling have been observed both in cancer26 and during wound healing and lead to the formation of abnormal and dysfunctional vessels. Drugs that target WNTs or β-catenin could inhibit the formation of pathological blood vessels. WNTs are also involved in the communication between endothelium and leukocytes: WNTs produced by the endothelium induce matrix metalloproteinase (MMP) expression in T lymphocytes that carry Frizzled receptors, which accelerates extravasation27. Both neutrophils and macrophages carry Frizzled receptors and respond to WNT5a28,29. Thus, it is important to keep in mind that targeting WNT and β-catenin may not only affect communication within the endothelial cell population, but will also probably affect the communication between the endothelium and pro-angiogenic leukocytes.
Mural cells. The traditional mural cells are vascular smooth muscle cells and pericytes. Mural cells surround and stabilize the endothelial cell tube and also regulate angiogenesis, vascular function and leukocyte recruitment. The identities, phenotypes and numbers of perivascular cells differ between organs and vessel types and depend on microenvironmental conditions30,31,32,33,34. Smooth muscle cells form several layers around the endothelial cells in the larger arteries and veins, whereas pericytes partially cover the endothelial cell tube of arterioles, capillaries and postcapillary venules. Mural cells may contract in response to stimuli to influence organ perfusion and downstream blood flow. Pericytes share a basement membrane with endothelial cells, interact directly with the endothelium via cell surface receptors and secrete signalling molecules to support vessel maturation and function30,31,35,36.
Perivascular leukocytes. Many types of leukocytes are found in perivascular positions. Tissue-resident mast cells are often adjacent to blood vessels in skin and mucosa. In response to inflammatory stimuli, the mast cells quickly release permeability-increasing agents such as histamine and VEGFA, which have potent paracrine effects on endothelial cells37. Resident macrophages, located perivascularly, sometimes have a pericyte-like appearance. In addition to functioning as sentinel cells crucial for host defence38, certain macrophages are pro-angiogenic and modulate vascular branching morphogenesis during embryonic development39.
Multiple macrophage subclasses are being characterized, and they differ in surface receptors, cytokine production and phenotype. These macrophage subclasses are believed to result from either different precursor cells or from microenvironmental cues that lead to the activation of distinct macrophage signalling pathways, including transcription factors, post-transcriptional regulators and epigenetic regulators40. Classically activated macrophages (designated M1) are activated by interferon-γ (IFNγ), express high levels of pro-inflammatory cytokines (interleukin-1β (IL-1β), IL-6, IL-12, IL-23 and tumour necrosis factor (TNF)) as well as reactive oxygen species (ROS) and nitric oxide (NO); inducible nitric oxide synthase (iNOS) is an M1 hallmark. M1 macrophages are important inducers and effectors of the T helper 1 (TH1) lymphocyte response40, and M1 macrophages clearly contribute to the development of inflammatory diseases.
Other roles have been described for alternatively activated macrophages, such as M2 macrophages. These macrophages are associated with tissue remodelling, wound healing, angiogenesis and tumour progression, and they are less efficient than M1 macrophages at killing bacteria41,42,43,44. Alternatively activated macrophages are induced by IL-4 and IL-13 and produce anti-inflammatory cytokines (IL-10 and TGFβ)40. The signals and mechanisms that induce organ-resident maintenance macrophages to become cells that either actively fuel or inhibit the inflammatory process are currently being delineated and are discussed below. It was recently demonstrated that resident macrophages are transcriptionally diverse, with very limited overlap between different populations, suggesting that there are probably additional classes of macrophages whose precise function remains to be defined45.
Microvascular communication
The behaviour and function of the different cell types in the microvascular compartment are regulated by intercellular communication either through direct cell–cell contact or by the transfer of secreted molecules. There are four basic modalities for cell–cell communication at this site: paracrine signalling; mechanosignalling, which is mediated by direct interactions between surface molecules on adjacent cells; direct signal transduction via gap junctions (connexins) and TnTs; and communication via the release and uptake of cell-derived vesicles such as exosomes. The relative functional importance of these modalities in different situations remains to be fully established.
Paracrine signalling is the most well-studied mode of communication between neighbouring cells. Classic examples of paracrine signalling in the microvasculature include mast cells releasing permeability-inducing factors, which then affect endothelial cells during inflammation37, and the release of platelet-derived growth factor (PDGF) from endothelial cells to recruit pericytes that express PDGF receptor-β (PDGFRβ)46. Paracrine signalling is also very relevant to stem cell therapies, including those for tissue regeneration and angiogenesis, that are now being evaluated in an increasing number of clinical trials47,48. Interestingly, a substantial part of the beneficial effects of stem cell therapies may be due to their release of paracrine factors. Undifferentiated stem cells secrete a range of factors that instruct nearby cells in the microvascular compartment to participate in angiogenesis. In addition, stem cells also secrete factors that are immunosuppressive, anti-apoptotic or that may stimulate proliferation49. Characterizing the secretome of stem cells and circulating progenitor and/or myeloid populations could identify alternatives to stem cell therapy, as a locally administered cocktail of stem-cell-secreted factors could be used to promote angiogenesis, thereby circumventing some of the problems with systemic injections of stem cells, such as nonspecific stem cell homing and insufficient engraftment in the tissues of interest50,51.
The recruitment of circulating leukocytes is directed by mechanosignalling — between adhesion molecules on the endothelium and integrins on leukocytes52. Integrin inside-out signalling occurs following chemokine receptor ligation on leukocytes and results in conformational changes in the extracellular integrin domains, which then induces clustering of integrins and increases the avidity of integrins for their ligands. Following the binding of integrin to its counter-receptor on the endothelium, integrin outside-in signalling occurs, resulting in cytoskeletal rearrangements, strong cellular adhesion, cell spreading and leukocyte migration. Distinct adhesion molecules are involved in the different steps of the recruitment cascade, ultimately resulting in leukocyte extravasation; this process is further discussed below. Notably, distinct leukocyte populations and vascular beds express different combinations of adhesion molecules that can enable tissue-specific therapeutic targeting23,24.
Connexins are membrane proteins that form gap junctions and enable direct signal transduction between adjacent cells by allowing rapid transfer of ions, second messengers and metabolites. Gap junction channels that contain connexin 43 (CX43, also known as gap junction α-1) and CX45 (also known as gap junction γ-1) and are between endothelial cells and mesenchymal cells are important for mesenchymal cells to obtain a mural cell fate during blood vessel formation53,54. CX43 channels are also involved in communication between leukocytes and epithelial cells in both the lung and intestine, resulting in immune suppression in the lung55 and oral tolerance in the intestine56. In addition, in vitro studies have suggested that connexin-dependent communication occurs between leukocytes and endothelial cells57,58. Interestingly, the synthetic peptide ACT1, which mimics the carboxy terminus of CX43, stabilized gap junctions and decreased infiltration of inflammatory neutrophils into wounds59. Application of Granexin gel containing ACT1 also improved healing of diabetic foot ulcers in clinical trials (Clinical Trials Registry India identifier: CTRI/2011/09/001984)60. Targeting connexins to modulate the communication between epithelial or endothelial cells and leukocytes could be an exciting approach to improve wound healing60,61.
Cell–cell signalling also occurs via cell-derived microvesicles, also called exosomes, that shuttle proteins, lipids, microRNAs and mRNAs between cells62,63. Exosomes are released from most cell types, including leukocytes and endothelial cells, in response to various stimuli. Platelets also constitute a rich source of exosomes owing to their abundance in blood (≈ 1011 platelets per l). Platelets are crucial for haemostasis and thrombosis but are also involved in the regulation of both inflammation and vascular homeostasis64,65. Depending on the stimulus, platelets can release exosomes containing specific molecules; for example, exosomes containing pro-angiogenic VEGFA can be released in response to ADP or exosomes containing endostatin can be released in response to thromboxane A2 (Ref. 66). In the United States, many clinical trials investigating exosomes have been initiated, and in one of these, exosomes will be tested as an alternative vehicle to liposomes for drug delivery (ClinicalTrials.gov identifier: NCT01294072).
As drug delivery vehicles, exosomes could, in principle, be injected into the target tissue; however, in most studies conducted thus far the exosomes have been administered systemically. An exciting possibility is that exosomes could be designed to express surface molecules that target them to specific vascular beds67. For systemic delivery, an important question to address is whether exosomes generally home to the activated endothelium of inflamed tissues. Interestingly, exosomes from rat pancreatic adenocarcinoma cells (the ASML cell line) bind to and are taken up by all leukocyte subpopulations in vivo to support leukocyte effector functions, and it was suggested that exosomes could function as adjuvants in immunotherapy68. Somewhat contradictory to these findings, exosome removal has also been suggested as a therapeutic adjuvant in cancer69. Clearly, depending on the exosome content, removing or adding exosomes will have different effects. Ongoing clinical trials are also testing whether circulating exosomes can be used as prognostic biomarkers, for example in gastric cancer (the EXO–PPP Study; NCT01779583).
Recent findings link exocytosis mediated by exocyst complex component 7 (EXO70) with both epithelial–mesenchymal transition (EMT)70 and the dynamic cell-shape changes required for cell migration and morphogenesis71, suggesting that selectively expressed exocyst components, or isoforms thereof, could be targeted to modulate EMT, cell migration and fibrosis. The exocyst complex has been suggested to be involved in the formation of exosomes72 and TnTs9. TnTs are F-actin-containing structures up to 200 nm in diameter that can be several cell diameters long (over 70 μm in length)73, which, together with gap junctions, are the only described way for cells to directly exchange intracellular components. TnTs seem to allow transport of larger cargo than gap junctions do. Mitochondrial RHO GTPase 1 (MIRO1)-expressing mesenchymal stem cells (as well as smooth muscle cells and fibroblasts) have very recently been shown to assist in the repair of damaged epithelial cells by donating mitochondria through TnTs, suggesting that overexpression of MIRO1 to induce TnT formation could be exploited in stem cell therapy74. The actin-bundling protein fascin has been identified as one of the protein components of TnTs73, and there is interest in developing fascin-targeting drugs, such as migrastatin analogues, that could impede tumour growth by blocking cell–cell communication between cancer cells and stromal cells (mesenchymal stem cells (MSCs) or endothelial cells)75, as such interactions may increase chemoresistance in cancer cells76. TnTs are also utilized to transfer signalling proteins such as HRAS from B cells to T cells77, and dying endothelial cells can be rescued by TnT-mediated transfer of substances from adjacent endothelial progenitor cells78 or mesenchymal cells79. The roles of TnTs in leukocyte–endothelial exchange have yet to be explored.
Angiogenesis
Several antibody-based drugs that inhibit angiogenesis have reached the market for treating cancer or abnormal blood vessel formation in wet age-related macular degeneration (AMD)80. Despite its benefits in some forms of cancer and in AMD, anti-angiogenic treatments have also been associated with severe adverse effects, and this has led to an increased interest in therapies aimed at normalizing pathological vessels rather than eliminating them81. Therapeutic means to induce the formation of functional blood vessels are also helpful for the treatment of local ischaemia during peripheral ischaemic disease, stroke or engraftment following transplantation. A better understanding of the different angiogenic mechanisms, as well as the roles of microvascular cell–leukocyte communication in regulating angiogenesis, is required to develop new therapeutic agents with higher specificity and efficacy.
Mechanisms of angiogenesis. The main paradigm guiding the field of anti-angiogenic therapies has been that angiogenesis occurs predominantly via sprouting angiogenesis together with some degree of intussusceptive angiogenesis (vessel splitting)82,83 (Fig. 2). Notably, our understanding of sprouting angiogenesis is based mainly on data obtained from mouse and zebrafish models of developmental angiogenesis as well as on data from advanced 3D in vitro models84,85. During developmental angiogenesis, reproducible patterning of the vasculature occurs in response to spatiotemporally regulated guidance cues (such as morphogen gradients) that induce expression of target genes to regulate the formation of angiogenic sprouts headed by a tip cell84. Multiple extracellular concentration gradients of guidance cues help the vascular cells to know their position and guide the migrating tip cells86. Comparatively less is known about the mechanisms of angiogenesis in adult tissues during wound healing and inflammation. In contrast to the organized angiogenic process during development, a wound may appear in any location at any time. Acute and generic repair mechanisms are therefore crucial for vessel formation in association with wound healing.
Early embryonic angiogenesis is thought to mainly occur through sprouting angiogenesis (top right). In this process, new vessel sprouts headed by tip cells respond to highly regulated guidance cues often present in the form of concentration gradients of chemotactic and pro-angiogenic factors. The conditions for angiogenesis during wound healing are very different from those during development. During wound healing, biomechanical forces elicited by myofibroblasts during the wound closure process induce looping and splitting angiogenesis, processes of vessel neoformation that are faster that sprouting (bottom left). Modulation and maturation of the primitive vascular network occurs through a combination of mechanisms, including sprout fusion, vessel splitting and pruning, as the tissue expands during development (bottom right). All mechanisms of vessel formation probably occur simultaneously, albeit to different extents in different situations. Whereas sprouting dominates early embryonic angiogenesis, looping and splitting may have more prominent roles during initial wound healing. Notably, the end result — irrespective of the degree and types of angiogenic mechanisms at play — would be the same, such that the final vessel density is ultimately regulated by the functional demands set by oxygen tension.
A rapid angiogenic response, termed looping angiogenesis, which is compatible with the special requirements of wound healing, was recently proposed87,88,89 (Fig. 2). During wound closure, looping angiogenesis occurs when contractile myofibroblasts pull adjacent intact tissue — including fully perfused vessels — into the wounded area89, leading to vessel elongation and the formation of extended vascular loops. Interestingly, tumours have previously been described as wounds that do not heal2, and data obtained by time-lapse intravital multiphoton microscopy indicate that physical forces in rapidly growing tumours expand the vasculature in a process that could be considered as looping angiogenesis, which occurs together with sprouting and splitting angiogenesis (see Supplementary information S1 (video) in Ref. 90). The biomechanical hypothesis that explains the driving forces for looping angiogenesis is probably of general relevance to the development of both anti- and pro-angiogenic therapies, although the mechanism was first described in relation to wound healing87. The formation of contractile myofibroblasts that generate the forces that lead to vessel elongation and looping is induced by TGFβ and can be inhibited by TGFβ-specific antibodies such as fresolimumab and 1D11 (Ref. 91), small-molecule receptor serine/threonine kinase inhibitors such as LY2157299 (Ref. 92) and blocking peptides such as P144 (a 14-mer peptide derived from the TGFβ1 proteoglycan receptor betaglycan (also known as TGFR3))93.
Modulation of extracellular matrix production will also affect all modes of angiogenesis. Inhibition of collagen production can be achieved using small interfering RNA (siRNA)-mediated approaches, whereas inhibitors against cathepsin K, MMPs and other enzymes with collagen-degrading activity can increase levels of extracellular collagen. The precise contributions of different leukocyte populations to looping angiogenesis remains to be established, but the abilities of leukocytes to affect the extracellular matrix and to produce TGFβ suggest that they could have a role94,95.
Modulation of angiogenesis by leukocytes. In addition to accumulating in inflamed tissue, leukocytes are recruited in large numbers to sites of hypoxia and sterile injury. Tissue-resident macrophages and recruited neutrophils and eosinophils clearly contribute to angiogenesis as they alter the microenvironment by secreting factors such as VEGFA and MMPs that promote angiogenesis43,96. Interestingly, a direct association between perivascular macrophages and angiogenic blood vessels has also been reported during developmental angiogenesis in the mouse hindbrain and in zebrafish, in which the macrophages were found to directly partake in the fusion of endothelial sprouts39.
Monocyte recruitment to damaged and hypoxic tissue involves chemokine receptors (including CXC-chemokine receptor 4 (CXCR4) and CC-chemokine receptor 2 (CCR2)) as well as sphingosine 1 phosphate receptor 3 (S1PR3), all of which are upregulated during ischaemia97,98,99. CXC chemokine ligand 12 (CXCL12), the ligand of CXCR4 (Ref. 100), is upregulated around larger vessels in a VEGFA-dependent manner101, whereas sphingosine 1-phosphate, produced by platelets and endothelial cells, is blood-borne and crucial for vessel maturation31. At the affected site, the monocytes and/or macrophages acquire enhanced pro-angiogenic properties following 'on-site' education by microenvironmental cues102. Circulating pro-angiogenic neutrophils are also recruited to hypoxic tissue, where they are crucial for the initiation of angiogenesis because they deliver the pro-angiogenic enzyme MMP9 (Refs 4,43,103,104). MMP9 may promote angiogenesis both by degrading the extracellular matrix and by increasing the release of matrix-bound VEGFA. Thus, uncovering the underlying mechanisms regulating the recruitment and on-site education of leukocytes will probably provide new druggable targets to influence angiogenesis.
Inhibiting angiogenesis. Anti-angiogenic therapy to combat cancer progression has been discussed since the discovery, in the 1970s, that inhibition of vascular growth can slow down the growth of solid tumours1. In 1991, Kerbel suggested that the genetically stable endothelial cells in tumours could be targeted to halt tumour growth105. To date, the best results for anti-angiogenic (that is, VEGF-specific) treatments are observed when treating macular degeneration, including proliferative diabetic retinopathy106. However, using a VEGF-specific treatment (Table 1), such as the humanized antibody bevacizumab (Avastin; Genentech/Roche), to combat cancer through inhibition of tumour angiogenesis has yielded modest results relative to expectations. The addition of VEGF-specific therapy to radiotherapy and systemic chemotherapy has, in some studies, been beneficial107,108,109,110, in agreement with the vascular normalization hypothesis81. This hypothesis states that normalization of pathological and disorganized tumour vessels leads to decreased vascular leakage and normalization of interstitial pressure, and at least transiently improves drug uptake81,111. However, many adverse effects associated with anti-angiogenic drugs have been reported, such as bleeding, proteinurea, hypertension, reduced wound healing and increased risk for thromboembolic events112,113.
It seems that the complexity of the angiogenic process was underestimated, at least initially, and a number of compensatory mechanisms come into play when VEGF is inhibited. The modes of resistance to VEGFA pathway inhibitors have been discussed in detail in an expert review114, and proposed escape mechanisms typically include a compensatory increase of the production of other pro-angiogenic compounds. It seems plausible that in order for anti-angiogenic therapy to be more efficient in the context of inhibiting tumour growth, multiple molecules expressed by endothelial cells or cells of the microvascular compartment (including angiomodulatory leukocytes) should be simultaneously targeted by combination therapies. Examples include combinations of antibodies targeting VEGFR2 or VEGFA together with CXCR4 antagonists, which inhibit angiogenesis and reduce tumour growth in experimental mouse models115,116. Furthermore, a combination therapy using antibodies against both VEGFA and the VEGFR2 co-receptor neuropilin 1 (NRP1) reduced vessel formation and tumour growth in xenograft models, and blockade of NRP1 also resulted in reduced pericyte coverage117. Pericytes are crucial for blood vessel formation and function and are therefore prospective targets for anti-angiogenic therapies. As pericytes depend on PDGF signalling, many small multi-targeting kinase inhibitors of PDGF signalling have been investigated for their effects on pericytes and angiogenesis (Box 1; Table 2).
Metabolic targets to inhibit angiogenesis. Several recent papers suggest that targeting endothelial cell metabolism could limit angiogenesis, as the rate of glycolysis can regulate vessel branching118,119,120. Recently, it was shown that 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3), an activator of glycolysis, promotes vascular growth and that the PFKFB3 inhibitor 3-PO inhibits endothelial cell proliferation in mouse and zebrafish models121. Interestingly, 3-PO counteracts hyperbranching induced by inhibition of Notch and augments the effects of VEGF-specific treatment. The effects were indeed partial and transient, but 3-PO could be a new type of anti-angiogenic compound, and the related molecule ACT-PFK-158 is now in a Phase I safety study of patients with cancer (NCT02044861).
Recently, the oxygen sensor prolyl hydroxylase 2 (PHD2), which promotes degradation of hypoxia-inducible factor 1α (HIF1α), has received a lot of attention. Haplodeficiency of the gene encoding PHD2 (PHD2, also known as EGLN1) resulted in normalization of tumour vasculature, which increased tumour blood perfusion, decreased tumour interstitial pressure (caused by decreased vessel leakage) and inhibited metastasis122,123. In a model of muscle ischaemia, mice haplodeficient for Phd2 were less susceptible to ischaemia-induced tissue necrosis124. Interestingly, increased numbers of alternatively activated macrophages secreting pro-arteriogenic factors were found in the ischaemic muscle of Phd2+/− mice, and either chronic or acute deletion of Phd2 in macrophages skewed them to a pro-angiogenic phenotype124. Furthermore, in situ education of pro-arteriogenic macrophages after vascular occlusion has been shown to rely on angiopoietin 1 (ANG1)–ANG1 receptor (TIE2, also known as TEK) signalling, and ANG1 was shown to downregulate PHD2 expression via activation of TIE2 in macrophages125. Thus, ANG1, which is stored in platelets and mural cells and released during tissue damage, can assist in macrophage reprogramming. Clearly, the homeostatic roles of the HIF pathway and factors regulating HIFs, such as PHD2, are of great interest as therapeutic targets, and screens to identify inhibitors and activators are ongoing126.
Targeting Notch signalling to inhibit angiogenesis. Notch signalling is central to many aspects of vascular development, including differentiation of endothelial and mural cells as well as regulation of sprouting. Tip cells that are positive for the Notch ligand delta-like protein 4 (DLL4) hold adjacent Notch 1-positive stalk cells in check by lateral inhibition127,128. Interestingly, inhibition of Notch signalling has recently been proposed to increase infiltration of mononuclear cells into tissue, which is followed by intussusceptive angiogenesis129, and Notch signalling in macrophages is required for efficient interactions with DLL4-positive tip cells to mediate sprout fusion130.
Whereas DLL4 in endothelial tip cells inhibits sprouting from adjacent stalk cells, DLL4 stimulates VEGFA production in pro-angiogenic macrophages. Pharmacological inhibition or activation of DLL4 may have very different net effects on inflammatory neovascularization and tumour vascularization, for example, depending on the balance of Notch signalling in endothelial cells and macrophages, both of which express Notch 1 and Notch 4 (Ref. 131). Furthermore, blockade of the DLL4–Notch interaction leads to increased homing of leukocytes expressing CD45 (also known as receptor-type tyrosine-protein phosphatase C) and αM integrin (also known as CD11b) to ischaemic tissue, and IL-8 release by activated monocytes is strongly inhibited by endothelial cells through a DLL4–Notch-mediated mechanism132. Thus, it is conceivable that if the number of pro-angiogenic macrophages and other DLL4-responsive leukocytes is large, the effect of DLL4 on leukocytes will partially balance the inhibitory effect of DLL4 on endothelial cell activation. Notably, the regulation of the functional phenotypes of macrophages has also been reported to be partially downstream of Notch receptors133,134, and it is thus currently difficult to predict the effects of Notch pathway inhibitors on angiogenic events that are modified by the immune system.
Targeting macrophages to inhibit angiogenesis. Myeloid-derived immune cells, predominantly macrophages, supply tumours with VEGFA, which increases the number of tumour vessels of an immature and leaky phenotype135. Specific deletion of VEGFA in myeloid-derived cells resulted in large tumours with normalized vasculature, which were sensitive to chemotherapy135. Numerous clinical trials are evaluating anti-macrophage therapies, including antibodies and small-molecule inhibitors such as colony stimulating factor 1 (CSF1)-specific therapies, which reduce macrophage differentiation and survival, or CC-chemokine ligand 2 (CCL2)-specific therapies, which inhibit monocyte recruitment in patients with cancer136,137. However, a recent study revealed a hitherto unknown complexity of chemokine biology, as the withdrawal of CCL2-specific therapy in mice with breast cancer tumours accelerated lung metastasis and death138. The reason for this unexpected twist might be a feedback mechanism that results in increased CCL2 production upon the release of CCL2 inhibition, which then recruits monocytes to tumours. In addition, CCL2-specific therapy blocks monocyte recruitment from bone marrow, resulting in an increased pool of CCR2-positive monocytes that, following cessation of CCL2-specific treatment, are released into the blood and recruited to metastatic tumours where they contribute to angiogenesis by releasing VEGFA.
Efforts are currently being made to develop treatments that shift tumour-associated and pro-angiogenic alternatively activated macrophages to an antitumour M1 subtype40. The opposite shift (from M1 to alternatively activated macrophages) would be useful in situations, such as peripheral ischaemia, in which increased angiogenesis would be beneficial. Several promising candidates for these opposing scenarios are listed in Table 3, some of which are under investigation and some of which are already on the market. As with all immunomodulating therapies, systemic alterations of the immune system must be considered with great caution.
Macrophages have anti-angiogenic functions during retinal development that depend on non-canonical WNT signalling and the concomitant production of soluble VEGF receptor 1 (sVEGFR1, also known as sFLT1), which competitively inhibits VEGF signalling139. This was recently demonstrated to be important during early phases of wound healing140. Abnormally high levels of WNT signalling are observed during both wound healing and cancer, and the drugs that (at least in part) target WNT signalling and are currently in clinical trials for the treatment of malignancies and/or vascular conditions include the small molecular weight porcupine inhibitor LGK974 (for WNT-dependent malignancies; NCT01351103), as well as resveratrol (for coronary artery disease; NCT02137421), sulindac and curcumin (for acute kidney injury or abdominal aortic aneurysm; NCT01225094).
Inflammation
A regulated inflammatory response is vital for life after birth, and dysregulated inflammation is involved in the progression of many diseases. Similar cues drive inflammation irrespective of whether the underlying trigger is a bacterial infection or sterile tissue damage. Blunt and nonspecific anti-inflammatory therapies are associated with serious adverse effects, such as increased risk for opportunistic infections and cancer141. The fine-tuning mechanism that regulates the amplitude and prolongation of the local inflammatory process should be deciphered, to prevent chronic inflammation while maintaining a sufficient host defence.
Mechanisms of inflammation. The cellular 'whistleblowers' that initiate inflammation following damage or bacterial detection are primarily the resident leukocyte populations (such as macrophages) that are found in most organs. Classic tasks of these sentinel cells are to maintain organ and system homeostasis by cleaning up after dying cells and defeating potential threats. The macrophages have unique means to monitor their environment, and in response to certain alterations and challenges they release multiple paracrine factors to instruct cells in the microenvironment. In response to macrophage-produced cytokines and damage- or pathogen-associated molecular pattern molecules (DAMPs or PAMPs) released by injured cells or the invaders, the postcapillary venular endothelium upregulates adhesion molecules involved in the leukocyte recruitment cascade. This cascade involves mechanosignalling between leukocytes in the circulation and endothelial cells, and it ultimately results in extravasation of leukocytes, of which neutrophils are the first to arrive to the affected site (Fig. 3) (reviewed in Refs 7, 142,143,144). Some leukocyte adhesion molecules — namely the β2 integrins: αLβ2 integrin (also known as lymphocyte function-associated antigen 1 (LFA1)) and αMβ2 integrin (also known as macrophage-1 antigen (MAC1)) — switch from low-affinity to high-affinity states following the binding of chemokines to G protein coupled receptors on leukocytes. As a consequence, the neutrophils become activated and start to adhere to144,145 and crawl on the vessel wall along chemokine gradients146,147. The intravascular crawling continues until the neutrophil reaches one of the optimal sites for transmigration, which are most often located at endothelial junctions147,148 and coincide with areas of low expression of laminin 10, collagen IV and nidogen 2 in the basement membrane between pericytes149,150.
Leukocyte rolling, adhesion, crawling and transmigration involve distinct adhesion molecules on leukocytes and endothelium and result in leukocytes being recruited from the circulation to inflamed tissues. Some key molecules that mediate leukocyte–endothelial cell interactions in each step of the recruitment cascade are listed. Drugs and compounds that bind to these targets could prevent interactions between leukocytes and the endothelium to impede or block leukocyte extravasation during inflammation. ESAM, endothelial cell-selective adhesion molecule; ESL1, E-selectin ligand 1; ICAM1, intercellular adhesion molecule 1; JAMA, junctional adhesion molecule A; MAdCAM1, mucosal addressin cell adhesion molecule 1; PECAM1, platelet endothelial cell adhesion molecule; PSGL1, P-selectin glycoprotein ligand 1; PTX3, pentraxin-related protein; VCAM1, vascular cell adhesion protein 1; VE-cadherin, vascular–endothelial cell cadherin; VEGFA, vascular endothelial growth factor A.
Neutrophil extravasation requires bidirectional crosstalk between the neutrophil and endothelial cells and may involve the formation of endothelial protrusions that engulf the transmigrating neutrophils in dome-like structures151,152,153. This process involves ligation of numerous endothelial adhesion molecules (intracellular adhesion protein 1 (ICAM1), ICAM2 and vascular cell adhesion protein 1 (VCAM1)) and junctional adhesion molecules (platelet endothelial cell adhesion molecule (PECAM1), CD99 and junctional adhesion molecules (JAMs)) and ultimately results in the regulated opening of endothelial junctions, which probably limits vascular leakage143. Data suggest that the endothelium functions as a barrier regulating transendothelial migration, which has spurred new interest in the role of endothelial cells in leukocyte transmigration153,154,155. This gating could be regulated by targeting the junctional and cytoskeletal rearrangement of the endothelium, which would limit detrimental inflammation by reducing leukocyte transmigration and/or vascular permeability154,155. In addition, intravascular neutrophils that adhere to the endothelium have the potential to modify the endothelial barrier. A recent study demonstrated that neutrophils release TNF in close proximity to endothelial junctions in response to chemoattractants156 and locally increase microvascular permeability. This permeability increase was abolished in mice null for the TNF receptor (Tnfr−/− mice), even though leukocyte transmigration was not affected156. These observations highlight the bidirectional communication between leukocytes and endothelial cells during inflammation, an area in which further studies are warranted.
Pericytes support neutrophil transmigration, and outside the endothelial lining, abluminal crawling ultimately directs the neutrophils to gaps between adjacent pericytes157. These gaps become enlarged in response to pro-inflammatory chemokines, which also induce contractions and conformational changes in pericytes. Furthermore, extravasated neutrophils and monocytes chemotactically move towards and interact with arterial pericytes in a model of sterile skin inflammation8, a process that is dependent on the inflammation-induced pericyte presentation of macrophage migration inhibitory factor (MMIF, also known as MIF). These interactions result in increased expression of pro-migratory receptors as well as prosurvival molecules on the leukocytes, thereby increasing the efficiency with which they navigate to the site of injury. Furthermore, neutrophil transmigration was recently demonstrated to occur at close proximity to perivascular macrophages in a skin infection model using Staphylococcus aureus158. The virulence factor α-haemolysin (produced by the bacteria) lysed the perivascular macrophages, thus reducing neutrophil extravasation. These data demonstrate that the different cells of the microvascular compartment collaborate during leukocyte recruitment to inflamed tissue. Together, these cells — as well as their communication — comprise possible targets for anti-inflammatory therapies.
General mechanisms to inhibit inflammation. Anti-inflammatory drugs on the market span from general immune-suppressing therapies to highly specific monoclonal antibodies targeting distinct adhesion molecules or downstream intracellular signalling proteins. The major problems with the general anti-inflammatory therapies are the adverse effects associated with global immunosuppression, which render the patient more prone to infections and also increase the risk of acquiring other pathological conditions associated with impaired function of the immune system, such as cancer. Clinically available approaches to reduce inflammation include inhibiting the cell-specific effects triggered by inflammatory molecules, such as nonsteroidal anti-inflammatory drugs (NSAIDs) that inhibit cyclooxygenase (COX), neutralizing antibodies that target chemokines and cytokines and inhibitory antibodies that target adhesion molecules159. In recent years, the treatment of autoimmune diseases such as colitis and arthritis has been revolutionized by the introduction of TNF-specific therapies159. Other means to reduce the induction of inflammation that are currently under clinical evaluation include the inhibition of pattern recognition receptors, such as toll-like receptor 2 (TLR2), which is targeted by the monoclonal antibody OPN-305 (NCT01794663); and inhibition of the master regulator nuclear factor κB (NF-κB) by small molecule inhibitors of upstream activators such as the IκB kinases (IKKs), JUN N-terminal kinases (JNKs), Janus kinases (JAKs)160 or mitogen-activated protein kinases (MAPKs)159. However, as long as the targeted pathways are also crucial for the efficient host response that is required for restoration of homeostasis, the general immune-suppressing effects will result in unpleasant adverse effects and thus reinforce the need to identify site-specific drug targets.
Targeting adhesion molecules. The adhesion molecules involved in leukocyte–endothelial cell interactions during leukocyte recruitment are obvious drug targets that have been explored over the past decade, and several ongoing clinical trials are currently examining this strategy161 (Table 4). However, the broad cellular expression of many of the targeted adhesion molecules prevents specific inhibition and hence these strategies are associated with severe adverse effects. For instance, the life threatening condition progressive multifocal leukoencephalopathy (PML) was observed during inhibition of either α4 or αL integrin, which was due to reactivation of John Cunningham (JC) virus in the immunosuppressed patient161,162,163,164. However, certain endothelial adhesion molecules are expressed in an organ-specific manner, and strategies to block these adhesion molecules could provide localized solutions with fewer adverse effects. For example, in recent clinical trials (the Gemini studies)165,166, treatment with humanized monoclonal antibodies targeting α4β7 integrin showed promising results in the management of both Crohn disease and ulcerative colitis. The α4β7 ligand mucosal addressin cell adhesion molecule 1 (MAdCAM1) is specifically expressed in blood vessels of the intestinal tract167,168,169. As a consequence, α4β7 inhibition results in local effects in the intestine and does not alter the phenotype of cerebrospinal fluid T lymphocytes that are believed to underlie the development of PML in response to treatment with the α4 integrin-specific antibody natalizumab170,171,172. No case of PML has yet been reported during α4β7 inhibition, further stressing the importance of developing drugs that only target selected organs.
Targeting chemokines and their receptors. Circulating leukocytes are directed to inflammatory sites by the release of chemokines that bind to extracellular or cell-surface associated glycosaminoglycans (GAGs), mainly through electrostatic interactions. The chemokines are released by cells in the affected tissue, including endothelial cells, which also present the chemokines to leukocytes to recruit them from the blood stream. Targeting the activity of these chemokines and/or their cognate G protein-coupled chemokine receptors on leukocytes are attractive approaches to limit leukocyte influx. Several ways to bind and inactivate chemokines in the circulation or in tissue are currently being evaluated experimentally and include the cytokine receptor domain fused to the crystallizable fragment (Fc) region of immunogluobulin G (IgG)173, nanobodies174 and GAG-mimetics175,176. Several therapies targeting chemokines or their receptors have been tested in clinical trials, including the small molecule CCX282-B, which targets CCR9 and has been investigated for the treatment of Crohn disease; eldelumab, a CXCL10-specific antibody that has been investigated for the treatment of ulcerative colitis; and MLN1202, a CCR2-specific antibody that has been investigated for the treatment of cardiovascular disease177. Small molecular receptor antagonists that have made it to the market include the CCR5 antagonist maraviroc (Pfizer; for HIV infection178) and a CXCR4 antagonist AMD3100, which is a stem cell mobilizer179.
Another possible approach to limit chemokine activity could be to increase inhibitory post-translational modifications. In this context, the enzyme protein-arginine deiminase (PAD) changes peptide-bound arginine into citrulline and leads to citrullination of chemokines such as CXCL8 (IL-8), CXCL10 and TNF, which reduces their activities180,181. Interestingly, neutrophils — as well as certain bacterial pathogens such as Porphyromonas gingivalis that also express PAD — might thus be able to inactivate chemokines to impede leukocyte infiltration, and in the case of bacteria this could allow them to escape immune recognition181,182. Whether citrullination can be induced pharmacologically remains to be established.
Accelerating resolution of inflammation. An attractive but relatively unexplored approach to limit inflammation is to accelerate the inherent mechanisms underlying resolution of inflammation. In the best-case scenario, the inflammatory process is self-limiting and tissue homeostasis is completely restored. Inflammation is often resolved by expanding the population of alternatively activated macrophages, which are crucial for the resolution phase183. The mechanisms underlying the re-education of macrophages and their subsequent differentiation into distinct phenotypes are now considered potential targets for a wide range of therapies owing to the divergent effector functions of these cells (reviewed in Ref. 40) and numerous therapies are being evaluated or are already in the clinic (Table 3). Efferocytosis, the removal of dying cells by phagocytosis, is a key event during resolution of inflammation that is executed by resident macrophages; during this process these cells acquire an alternatively activated phenotype184. Simultaneously, the macrophages switch their production of arachidonic acid-derived eicosanoids from prostaglandins and leukotrienes to anti-inflammatory lipoxins (leukotriene A4 and leukotriene B4)185. Lipoxins, together with the omega-3-derived resolvins, protectins and maresins, were recently confirmed to be expressed in human tissues186 and are specialized pro-resolving mediators that inhibit leukocyte recruitment and enhance macrophage efferocytosis (reviewed in Ref. 187).
In animal models, addition of resolvin E1 suppresses the recruitment of leukocytes and the expression of inflammatory mediators (such as TNF and IL-1β188) during sterile inflammation, reduces herpes simplex virus-induced ocular inflammatory lesions189 and accelerates healing of diabetic wounds190. In a double-blind clinical trial of infants with eczema, topical application of 15(R/S)-methyl-lipoxin 4 relieved symptoms and improved quality of life191. Additional clinical applications are now being tested: mimetics of resolvin E1 are being examined for ocular indications (NCT01639846 and NCT01675570; also see the Auven Therapeutics website) and protectin D1 is being examined for neurodegenerative disorders (see the Anida Pharma website). The existence of these specialized pro-resolving mediators in humans as well as their ability to enhance resolution of inflammation without concomitant immunosuppression are indeed promising, even though their specific anti-inflammatory mechanisms of action and their functional importance in human health need to be better understood. Altering the microenvironmental cues provided by cells in microvessels and their close surroundings to re-educate resident and infiltrating leukocytes is an attractive approach to limit inflammation. A recently developed zebrafish model of sterile injury might be helpful for screening for new compounds that limit chronic inflammation and stimulate resolution192.
Combatting the development of fibrosis
Vasculitis, the inflammation of blood vessels, may lead to vessel damage, tissue destruction (necrosis) and eventually fibrosis. The diagnosis of many microvascular pathologies is based on symptoms and relatively little is known regarding the precise underlying mechanisms and molecular aetiologies. The pathogenesis of vasculitis is considered to be predominantly autoimmune, but infection by bacteria or viruses may also spark its progression. Fibrosis is generally the result of prolonged injury and deregulated wound healing (reviewed in Ref. 193).
Mechanisms of fibrosis. Fibrosis is a protective mechanism that follows inflammation and tissue injury whereby large quantities of extracellular matrix (mainly type I collagen) are secreted by cells that invade the wounded area in order to reconstitute and strengthen the damaged tissue and accelerate the healing process (Fig. 4). Dysregulated fibrosis that results from repeated tissue damage leads to continued activation of matrix-secreting fibroblasts and myofibroblasts, which eventually alters tissue function and can ultimately lead to organ dysfunction. Leukocytes have key roles as modulators of the fibrotic process and are thus targets for reducing uncontrolled fibrosis. Continued wounding or irritant-elicited responses can lead to overactive leukocytes that damage the affected tissue, whereas a fine-tuned and self-regulated leukocyte response that is gradually turned off promotes tissue repair.
Repeated tissue damage can lead to high levels of matrix deposition following myofibroblast activation and the continued secretion of pro-inflammatory factors in the wounded tissue. Recruitment of M1 macrophages, mast cells and neutrophils, together with on-site formation of myofibroblasts, supports fibrosis that will eventually lead to tissue dysfunction. Different classes of leukocytes, as well as myofibroblasts, are potential targets to reduce uncontrolled fibrosis. Elimination of the inflammatory triggers together with on-site reprogramming of macrophages and myofibroblasts as well as inhibition of recruitment of inflammatory leukocytes can be used to promote fibrosis resolution. Processes that can be targeted therapeutically are indicated. DAMPs, damage-associated molecular pattern molecules; EMT, epithelial–mesenchymal transition; EndMT, endothelial–mesenchymal transition; MSC, mesenchymal stem cell; PDGF, platelet-derived growth factor; rM, resolution-phase macrophage; TGFβ, transforming growth factor-β; VEGF, vascular endothelial growth factor.
The fibrotic process is activated by the destruction of cell membranes, which releases cell contents and DAMPs. The DAMPs activate mesenchymal cells and mural cells located in the proximity of the damaged site, followed by the recruitment of monocytes and macrophages via chemokine-induced chemotaxis (for example through activation of the CCL2–CCR2 signalling axis). The recruited macrophages phagocytose dead cells and secrete additional chemokines to recruit other types of leukocytes, including neutrophils.
Inhibition of fibrosis. EMT occurs during fibrosis, and TGFβ and related signalling proteins are major mediators of both EMT and fibrosis in many organs194. Activated macrophages release TGFβ and other pro-inflammatory mediators, which leads to the generation of fibroblasts from several cell sources, as well as to transdifferentiation of fibroblasts into myofibroblasts195,196. Interestingly, αV integrins on myofibroblasts are involved in the activation of extracellular TGFβ, and pharmacological blockade of αV-containing integrins using the small-molecule inhibitor CWHM 12 can reduce fibrosis in both the lung and liver197. Furthermore, the EMT transcriptional programme is anti-apoptotic and generally promotes cell migration, such that cells undergoing EMT often move away from their original position over time. Thus, reducing EMT-dependent formation and migration of myofibroblasts (Fig. 4) could be of interest to target fibrosis and also to target angiogenesis198. The anti-fibrotic properties of the tyrosine-kinase inhibitor nintedanib (Ofev; Boehringer Ingelheim; approved by for treatment of idiopathic pulmonary fibrosis) were recently described199. Nintedanib was shown to have anti-angiogenic effects and to inhibit tumour growth in vivo, and it did not promote EMT, despite inducing hypoxia.
Attempts are ongoing to identify targets within pro-fibrotic signalling cascades such as the TGFβ pathway197,200. Of interest, signalling via TLR4 can downregulate BMP and activin membrane-bound inhibitor homolog (BAMBI), the competitive and non-signalling pseudoreceptor for TGFβ201. Thus, it is possible that inhibition of TLRs or activation of BAMBI could be novel ways to modulate or counteract TGFβ-dependent fibrosis. Accordingly, the TLR4-specific monoclonal antibody NI-0101 has entered clinical trials (NCT01808469) for chronic inflammation and fibrosis. Additionally, targeting of 5′-AMP-activated protein kinase (AMPK) by metformin leads to increased BAMBI expression, which could explain some of the reported anti-fibrotic effects of this small organic compound, which is normally used to manage type 2 diabetes202.
Furthermore, release of PDGF subunit B (PDGFB) from platelets and activated macrophages in the wounded area will also attract and activate myofibroblasts, and indeed a PDGFB-specific monoclonal antibody reduces the development of liver fibrosis in experimental mouse models203.
Accumulating evidence supports the notion that pathological angiogenesis contributes to fibrosis. For example, blockade of VEGFR2 attenuates the steatosis and inflammation that can precede fibrosis in mice204. Interestingly, fibrotic scars, mainly composed of type I collagen, can be degraded by MMPs that are regulated by endogenous tissue inhibitors of metalloproteinases (TIMPs). Notably, neutrophils are the only cell type that release MMP9 without concomitant release of its specific inhibitor TIMP1, and this unique property enables them to deliver highly active MMP9 to sites of hypoxia205. Targeting MMPs is generally of great interest to control cell invasiveness, matrix remodelling and pathological angiogenesis. An interesting concept was recently presented in which injectable and bioresponsive hydrogels were used to locally deliver TIMPs: in response to MMP activity, MMP-degradable hydrogels released TIMPs locally 'on-demand' to limit unwanted effects of the inhibitor in other tissues206. Monocytes may also mediate fibrosis-associated angiogenesis: inhibiting infiltration of monocytes expressing increased levels of VEGF and MMP9 using the novel inhibitor mNOX-E36 that targets CCL2 was shown to prevent fibrosis-associated angiogenesis6.
An additional macrophage subtype — resolution-phase macrophages (rMs) — that has increased expression of the M1 markers iNOS and COX2 is also present during resolution of inflammation207. Differentiation into this subtype is controlled by cAMP levels; M1 macrophages shift to rMs when cAMP levels are high and the opposite occurs when cAMP levels are low. Furthermore, in a model of liver fibrosis, regression of fibrosis was dependent on restorative post-phagocytic macrophages that express high levels of MMPs, growth factors and phagocytosis-related genes208. The restorative macrophage phenotype and concomitant reduction in scar formation could be induced by liposome administration, which promoted macrophage phagocytic behaviour208. Inducing the rM phenotype could thus be a potential therapeutic option. These results further demonstrate the wide range of effector functions exerted by different types of macrophages.
Conclusions
As of now, information about the various cell types in and around microvessels that influence angiogenesis, inflammation and the development of fibrosis have been compiled, but the integrative effects of these different cells remain to be fully established, and comprehensive molecular and cellular interaction maps have not yet been built. This 'knowledge integration' is now taking place at an increasingly rapid pace, facilitated by direct in vivo observations of how cells of the microvasculature move and interact during angiogenesis and inflammation. A full appreciation of how these cells interact will not only tell us more about how angiogenesis and inflammation are regulated, but should also unveil many new potential methods to influence the microenvironment in order to steer cellular functions towards the restoration of homeostasis. New indications will also probably be found for existing drugs as a result of a more detailed molecular understanding of the communication between leukocytes and vascular cells.
References
Folkman, J. Tumor angiogenesis: therapeutic implications. N. Engl. J. Med. 285, 1182–1186 (1971).
Dvorak, H. F. Tumors: wounds that do not heal. Similarities between tumor stroma generation and wound healing. N. Engl. J. Med. 315, 1650–1659 (1986).
Casazza, A. et al. Impeding macrophage entry into hypoxic tumor areas by Sema3A/Nrp1 signaling blockade inhibits angiogenesis and restores antitumor immunity. Cancer Cell 24, 695–709 (2013).
Christoffersson, G. et al. VEGF-A recruits a proangiogenic MMP-9-delivering neutrophil subset that induces angiogenesis in transplanted hypoxic tissue. Blood 120, 4653–4662 (2012).
Chung, A. S. et al. An interleukin-17-mediated paracrine network promotes tumor resistance to anti-angiogenic therapy. Nat. Med. 19, 1114–1123 (2013).
Ehling, J. et al. CCL2-dependent infiltrating macrophages promote angiogenesis in progressive liver fibrosis. Gut 63, 1960–1971 (2014).
Kolaczkowska, E. & Kubes, P. Neutrophil recruitment and function in health and inflammation. Nat. Rev. Immunol. 13, 159–175 (2013).
Stark, K. et al. Capillary and arteriolar pericytes attract innate leukocytes exiting through venules and 'instruct' them with pattern-recognition and motility programs. Nat. Immunol. 14, 41–51 (2013).
Hase, K. et al. M-Sec promotes membrane nanotube formation by interacting with Ral and the exocyst complex. Nat. Cell Biol. 11, 1427–1432 (2009).
Kristensen, V. N. et al. Principles and methods of integrative genomic analyses in cancer. Nat. Rev. Cancer 14, 299–313 (2014).
Islam, S. et al. Quantitative single-cell RNA-seq with unique molecular identifiers. Nat. Meth. 11, 163–166 (2014).
Bentley, K. et al. The role of differential VE-cadherin dynamics in cell rearrangement during angiogenesis. Nat. Cell Biol. 16, 309–321 (2014).
Dejana, E., Tournier-Lasserve, E. & Weinstein, B. M. The control of vascular integrity by endothelial cell junctions: molecular basis and pathological implications. Dev. Cell 16, 209–221 (2009).
Chauhan, V. P., Stylianopoulos, T., Boucher, Y. & Jain, R. K. Delivery of molecular and nanoscale medicine to tumors: transport barriers and strategies. Annu. Rev. Chem. Biomol. Eng. 2, 281–298 (2011).
Heldin, C. H., Rubin, K., Pietras, K. & Ostman, A. High interstitial fluid pressure — an obstacle in cancer therapy. Nat. Rev. Cancer 4, 806–813 (2004).
Conway, D. E. et al. Fluid shear stress on endothelial cells modulates mechanical tension across VE-cadherin and PECAM-1. Curr. Biol. 23, 1024–1030 (2013).
Tzima, E. et al. A mechanosensory complex that mediates the endothelial cell response to fluid shear stress. Nature 437, 426–431 (2005).
Langenkamp, E. & Molema, G. Microvascular endothelial cell heterogeneity: general concepts and pharmacological consequences for anti-angiogenic therapy of cancer. Cell Tissue Res. 335, 205–222 (2009).
Swift, M. R. & Weinstein, B. M. Arterial-venous specification during development. Circ. Res. 104, 576–588 (2009).
Lawson, N. D., Vogel, A. M. & Weinstein, B. M. Sonic hedgehog and vascular endothelial growth factor act upstream of the Notch pathway during arterial endothelial differentiation. Dev. Cell 3, 127–136 (2002).
Wang, H. U., Chen, Z. F. & Anderson, D. J. Molecular distinction and angiogenic interaction between embryonic arteries and veins revealed by ephrin-B2 and its receptor Eph-B4. Cell 93, 741–753 (1998).
D'Onofrio, N. et al. Vascular-homing peptides for targeted drug delivery and molecular imaging: meeting the clinical challenges. Biochim. Biophys. Acta 1846, 1–12 (2014).
Ageirsdottir, S. A. et al. Site-specific inhibition of glomerulonephritis progression by targeted delivery of dexamethasone to glomerular endothelium. Mol. Pharmacol. 72, 121–131 (2007). This study demonstrates means to direct drug delivery to specific organs and thereby limit systemic adverse effects by decorating exosomes with relevant adhesion molecules.
Ageirsdottir, S. A. et al. Inhibition of proinflammatory genes in anti-GB; glomerulonephritis by targeted dexamethasone-loaded AbEsel liposomes. Am. J. Renal Physiol. 294, F554–F561 (2008).
Corada, M. et al. Sox17 is indispensable for acquisition and maintenance of arterial identity. Nat. Commun. 4, 2609 (2013).
Fancy, S. P. et al. Parallel states of pathological Wnt signaling in neonatal brain injury and colon cancer. Nat. Neurosci. 17, 506–512 (2014).
Wu, B., Crampton, S. P. & Hughes, C. C. Wnt signaling induces matrix metalloproteinase expression and regulates T cell migration. Immunity 26, 227–239 (2007).
Jung, Y. S. et al. Wnt5a stimulates chemotactic migration and chemokine production in human neutrophils. Exp. Mol. Med. 45, e27 (2013).
Maiti, G., Naskar, D. & Sen, M. The Wingless homolog Wnt5a stimulates phagocytosis but not bacterial killing. Proc. Natl Acad. Sci. USA 109, 16600–16605 (2012).
Armulik, A. et al. Pericytes regulate the blood-brain barrier. Nature 468, 557–561 (2010).
Gaengel, K. et al. The sphingosine-1-phosphate receptor S1PR1 restricts sprouting angiogenesis by regulating the interplay between VE-cadherin and VEGFR2. Dev. Cell 23, 587–599 (2012). This study shows for the first time that sphingosine 1-phosphate (produced by, for example, platelets) is required for vessel maturation via the inhibition of VEGFA signalling and stabilization of VE-cadherin-based junctions.
Krueger, M. & Bechmann, I. CNS pericytes: concepts, misconceptions, and a way out. Glia 58, 1–10 (2010).
Sims, D. E. The pericyte — a review. Tissue Cell 18, 153–174 (1986).
Sims, D. E. Recent advances in pericyte biology — implications for health and disease. Can. J. Cardiol. 7, 431–443 (1991).
Falcón, B. L. et al. Contrasting actions of selective inhibitors of angiopoietin-1 and angiopoietin-2 on the normalization of tumor blood vessels. Am. J. Pathol. 175, 2159–2170 (2009).
Gerhardt, H., Wolburg, H. & Redies, C. N-cadherin mediates pericytic-endothelial interaction during brain angiogenesis in the chicken. Dev. Dyn. 218, 472–479 (2000).
Wernersson, S. & Pejler, G. Mast cell secretory granules: armed for battle. Nat. Rev. Immunol. 14, 478–494 (2014).
Pollard, A. J., Perrett, K. P. & Beverley, P. C. Maintaining protection against invasive bacteria with protein-polysaccharide conjugate vaccines. Nat. Rev. Immunol. 9, 213–220 (2009).
Fantin, A. et al. Tissue macrophages act as cellular chaperones for vascular anastomosis downstream of VEGF-mediated endothelial tip cell induction. Blood 116, 829–840 (2010). This study demonstrates for the first time the role of tissue resident macrophages in vessel fusion in the developing brain.
Sica, A. & Mantovani, A. Macrophage plasticity and polarization: in vivo veritas. J. Clin. Invest. 122, 787–795 (2012).
Mantovani, A. Molecular pathways linking inflammation and cancer. Curr. Mol. Med. 10, 369–373 (2010).
Mantovani, A. & Sica, A. Macrophages, innate immunity and cancer: balance, tolerance, and diversity. Curr. Opin. Immunol. 22, 231–237 (2010).
Nozawa, H., Chiu, C. & Hanahan, D. Infiltrating neutrophils mediate the initial angiogenic switch in a mouse model of multistage carcinogenesis. Proc. Natl Acad. Sci. USA 103, 12493–12498 (2006).
Wynn, T. A., Chawla, A. & Pollard, J. W. Macrophage biology and health and disease. Nature 496, 445–455 (2013).
Gautier, E. L. et al. Gene-expression profiles and transcriptional regulatory pathways that underlie the identity and diversity of mouse tissue macrophages. Nat. Immunol. 13, 1118–1128 (2012).
Armulik, A., Genové, G. & Betsholtz, C. Pericytes: development, physiological, and pathological perspectives, problems, and promises. Dev. Cell 21, 193–215 (2011).
Wei, X. et al. Mesenchymal stem cells: a new trend for cell therapy. Acta Pharmacol. Sin. 34, 747–754 (2013).
Leeper, N. J., Hunter, A. L. & Cooke, J. P. Stem cell therapy for vascular regeneration. Circulation 122, 517–526 (2010).
Schinköthe, T., Bloch, W. & Schmidt, A. In vitro secreting profile of human mesenchymal stem cells. Stem Cells Dev. 17, 199–206 (2008).
Hsiao, S. T. et al. Comparative analysis of paracrine factor expression in human adult mesenchymal stem cells derived from bone marrow, adipose tissue, and dermal tissue. Stem Cells Dev. 21, 2189–2203 (2012).
Rustad, K. C. & Gurtner, G. C. Mesenchymal stem cells home to sites of injury and inflammation. Adv. Wound Care 1, 147–152 (2012).
Abram, C. L. & Lowell, C. A. The ins and outs of leukocyte integrin signaling. Annu. Rev. Immunol. 27, 339–362 (2009).
Fang, J. S., Dai, C., Kurjiaka, D. T., Burt, J. M. & Hirschi, K. K. Connexin45 regulates endothelial-induced mesenchymal cell differentiation toward a mural cell phenotype. Arterioscler. Thromb. Vasc. Biol. 33, 362–368 (2013).
Hirschi, K. K., Burt, J. M., Hirschi, K. D. & Dai, C. Gap junction communication mediates transforming growth factor-β activation and endothelial-induced mural cell differentiation. Circ. Res. 93, 429–437 (2003). This study shows that CX43 gap junction communication between endothelial cells and mesenchymal progenitors is necessary for the differentiation of progenitors into mural cells.
Westphalen, K. et al. Sessile alveolar macrophages communicate with alveolar epithelium to modulate immunity. Nature 506, 503–506 (2014).
Mazzini, E., Massimiliano, L., Penna, G. & Rescigno, M. Oral tolerance can be established via gap junction transfer of fed antigens from CX3CR1+ macrophages to CD103+ dendritic cells. Immunity 40, 248–261 (2014).
Oviedo-Orta, E., Errington, R. J. & Evans, W. H. Gap junction intercellular communication during lymphocyte transendothelial migration. Cell. Biol. Int. 26, 253–263 (2002).
Zahler, S. et al. Gap-junctional coupling between neutrophils and endothelial cells: a novel modulator of transendothelial migration. J. Leuk. Biol. 73, 118–126 (2003).
Ghatnekar, G. S. et al. Connexin43 carboxyl-terminal peptides reduce scar progenitor and promote regenerative healing following skin wounding. Regen. Med. 4, 205–223 (2009).
Grek, C. S., Rhett, J. M. & Ghatnekar, G. S. Cardiac to cancer: connecting connexins to clinical opportunity. FEBS Lett. 588, 1349–1364 (2014).
Ghatnekar, G. S., Grek, C. L., Armstrong, D. G., Desai, S. C. & Gourdie, R. G. The effect of a connexin43-based peptide on the healing of chronic venous leg ulcers: a multicenter, randomized trial. J. Invest. Dermatol. 135, 289–298 (2015).
van Balkom, B. W. et al. Endothelial cells require miR-214 to secrete exosomes that suppress senescence and induce angiogenesis in human and mouse endothelial cells. Blood 121, 3997–4006 (2013).
Valadi, H. et al. Exosome-mediated transfer of mRNAs and microRNAs is a novel mechanism of genetic exchange between cells. Nat. Cell Biol. 9, 654–659 (2007).
Patzelt, J. & Langer, H. F. Platelets in angiogenesis. Curr. Vasc. Pharmacol. 10, 570–577 (2012).
Sreeramkumar, V. et al. Neutrophils scan for activated platelets to initiate inflammation. Science 346, 1234–1238 (2014).
Battinelli, E. M., Markens, B. A. & Italiano, J. E. Jr. Release of angiogenesis regulatory proteins from platelet alpha granules: modulation of physiologic and pathologic angiogenesis. Blood 118, 1359–1369 (2011).
Alvarez-Erviti, L. et al. Delivery of siRNA to the mouse brain by systemic injection of targeted exosomes. Nat. Biotech. 29, 341–345 (2011). This study presents evidence that exosomes from self-derived dendritic cells, engineered to express the neuron-specific rabies viral glycoprotein (RVG) peptide, can selectively deliver functional siRNA to distinct cell populations in the brain.
Zech, D., Sanyukta, R., Büchler, M. W. & Xöller, M. Tumor-crosstalk and leukocyte activation: an ambivalent crosstalk. Cell Comm. Signal. 10, 37 (2012).
Marleau, A. M., Chen, C. S., Joyce, J. A. & Tullis, R. H. Exosome removal as a therapeutic adjuvant in cancer. J. Transl. Med. 10, 134 (2012).
Lu, H. et al. Exo70 isoform switching upon epithelial–mesenchymal transition mediates cancer cell invasion. Dev. Cell 27, 560–573 (2013).
Zhao, Y. et al. Exo70 generates membrane curvature for morphogenesis and cell migration. Dev. Cell 26, 266–278 (2013).
Chacon-Heszele, M. F. et al. The exocyst and regulatory GTPases in urinary exosomes. Physiol. Rep. 2, e12116 (2014).
Lou, E. et al. Tunneling nanotubes provide a unique conduit for intercellular transfer of cellular contents in human malignant pleural mesothelioma. PLoS ONE 7, e33093 (2012).
Ahmad, T. et al. Miro1 regulates intercellular mitochondrial transport and enhances mesenchymal stem cell rescue efficacy. EMBO J. 33, 994–1010 (2014).
Chen, L., Yang, S., Jakoncic, J., Zhang, J. J. & Huang, X. Y. Migrastatin analogues target fascin to block tumour metastasis. Nature 464, 1062–1066 (2010).
Pasquier, J. et al. Preferential transfer of mitochondria from endothelial to cancer cells through tunneling nanotubes modulates chemoresistance. J. Transl. Med. 11, 94 (2013).
Rainy, N. et al. H-Ras transfers from B to T cells via tunneling nanotubes. Cell Death Dis. 4, e726 (2013).
Yasuda, K. et al. Tunneling nanotubes mediate rescue of prematurely senescent endothelial cells by endothelial progenitors: exchange of lysosomal pool. Aging 3, 597–608 (2011).
Liu, K. et al. Mesenchymal stem cells rescue injured endothelial cells in an in vitro ischemia–reperfusion model via tunneling nanotube like structure-mediated mitochondrial transfer. Microvasc. Res. 92, 10–18 (2014).
Claesson-Welsh, L. & Welsh, M. VEGFA and tumour angiogenesis. J. Intern. Med. 273, 114–127 (2013).
Jain, R. K. Normalizing tumor vasculature with anti-angiogenic therapy: a new paradigm for combination therapy. Nat. Med. 7, 987–989 (2001).
Adams, R. H. & Alitalo, K. Molecular regulation of angiogenesis and lymphangiogenesis. Nat. Rev. Mol. Cell. Biol. 8, 464–478 (2007).
Styp-Rekowska, B., Hlushchuk, R., Pries, A. R. & Djonov, V. Intussusceptive angiogenesis: pillars against the blood flow. Acta Physiol. (Oxf.) 202, 213–223 (2011).
Adams, R. H. & Eichmann, A. Axon guidance molecules in vascular patterning. Cold Spring Harb. Perspect. Biol. 2, a001875 (2010).
Jakobsson, L., Kreuger, J. & Claesson-Welsh, L. Building blood vessels-stem cell models in vascular biology. J. Cell Biol. 177, 751–755 (2007).
Lander, A. D. How cells know where they are. Science 339, 923–927 (2013).
Benest, A. V. & Augustin, H. G. Tension in the vasculature. Nat. Med. 15, 608–610 (2009).
Boerckel, J. D., Uhrig, B. A., Willett, N. J., Huebsch, N. & Guldberg, R. E. Mechanical regulation of vascular growth and tissue regeneration in vivo. Proc. Natl Acad. Sci. USA 108, E674–E680 (2011).
Kilarski, W. W., Samolov, B., Petersson, L., Kvanta, A. & Gerwins, P. Biomechanical regulation of blood vessel growth during tissue vascularization. Nat. Med. 15, 657–664 (2009). This study demonstrates that during wound healing activated myofibroblasts generate mechanical forces that rapidly pull and elongate vessels from intact vascular beds into the wounded area. The vascular network is thereafter modulated by sprouting, splitting and regression.
Vakoc, B. J. et al. Three-dimensional microscopy of the tumor microenvironment in vivo using optical frequency domain imaging. Nat. Med. 15, 1219–1223 (2009).
Tabe, Y. et al. TGF-β-neutralizing antibody 1D11 enhances cytarabine-induced apoptosis in AML cells in the bone marrow microenvironment. PLoS ONE 8, e62785 (2011).
Dituri, F. et al. Differential inhibition of the TGF-β signaling pathway in HCC cells using the small molecule inhibitor LY2157299 and the D10 monoclonal antibody against TGF-β receptor type II. PLoS ONE 8, e67109 (2013).
Santiago, B. et al. Topical application of a peptide inhibitor of transforming growth factor-β1 ameliorates bleomycin-induced skin fibrosis. J. Invest. Dermatol. 125, 450–455 (2005).
Khalil, N., Bereznay, O., Sporn, M. & Greenberg, A. H. Macrophage production of transforming growth factor β and fibroblast collagen synthesis in chronic pulmonary inflammation. J. Exp. Med. 170, 727–737 (1989).
Nacu, N. et al. Macrophages produce TGF-β-induced (β-ig-h3) following ingestion of apoptotic cells and regulate MMP14 levels and collagen turnover in fibroblasts. J. Immunol. 180, 5036–5044 (2008).
Taichman, N. S., Young, S., Cruchley, A. T., Taylor, P. & Paleoplog, E. Human neutrophils secrete vascular endothelial growth factor. J. Leuk. Biol. 62, 397–400 (1997).
Auffray, C. et al. Monitoring of blood vessels and tissue by a population of monocytes with patrolling behavior. Science 317, 666–670 (2007).
Awojoodu, A. O. et al. Sphingosine 1-phosphate receptor 3 regulates recruitment of anti-inflammatory monocytes to microvessels during implant arteriogenesis. Proc. Natl Acad. Sci. USA 110, 13785–13790 (2013).
Carlin, L. M. et al. Nr4a1-dependent Ly6C(low) monocytes monitor endothelial cells and orchestrate their disposal. Cell 153, 362–375 (2013).
Ganju, R. K. et al. The α-chemokine, stromal cell-derived factor-1α, binds to the transmembrane G-protein-coupled CXCR-4 receptor and activates multiple signal transduction pathways. J. Biol. Chem. 273, 23169–23175 (1998).
Grunewald, M. et al. VEGF-induced adult neovascuarisation, recruitment, retention and role of accessory cells. Cell 124, 175–189 (2006).
Avraham-Davidi, I. et al. On-site education of VEGF-recruited monocytes improves their performance as angiogenesis and arteriogenic accessory cells. J. Exp. Med. 210, 2611–2625 (2013). This study demonstrates that local VEGF 're-educates' monocytes to attain a pro-angiogenic phenotype.
Christoffersson, G. et al. Clinical and experimental pancreatic islet transplantation to striated muscle: establishment of a vascular system similar to that in native islets. Diabetes 59, 2569–2578 (2010).
Massena, S. et al. Identification and characterization of VEGF-A-responsive neutrophils expressing CD49d, VEGFR1 and CXCR4 in mice and humans. Blood 126, 2016–2026 (2015).
Kerbel, R. S. Inhibition of tumor angiogenesis as a strategy to circumvent acquired resistance to anti-cancer therapeutic agents. Bioessays 13, 31–36 (1991).
Lim, L. S., Mitchell, P., Seddon, J. M., Holz, F. G. & Wong, T. Y. Age-related macular degeneration. Lancet 379, 1728–1738 (2012).
Reck, M. et al. Phase III trial of cisplatin plus gemcitabine with either placebo or bevacizumab as first-line therapy for nonsquamous non-small-cell lung cancer: AVAiL. J. Clin. Oncol. 27, 1227–1234 (2009).
Saltz, L. B. et al. Bevacizumab in combination with oxaliplatin-based chemotherapy as first-line therapy in metastatic colorectal cancer: a randomized Phase III study. J. Clin. Oncol. 26, 2013–2019 (2008).
Sandler, A. et al. Paclitaxel–carboplatin alone or with bevacizumab for non-small-cell lung cancer. N. Engl. J. Med. 355, 2542–2550 (2006).
Willett, C. G. et al. Direct evidence that the VEGF-specific antibody bevacizumab has antivascular effects in human renal cancer. Nat. Med. 10, 145–147 (2004).
Batchelor, T. T. et al. Improved tumor oxygenation and survival in glioblastoma patients who show increased blood perfusion after cediranib and chemoradiation. Proc. Natl Acad. Sci. USA 47, 19059–19064 (2013).
Armstrong, T. S., Wen, P. Y., Gilbert, M. R. & Schiff, D. Management of treatment-associated toxicites of anti-angiogenic therapy in patients with brain tumors. Neuro. Oncol. 14, 1203–1214 (2012).
Wu, J. M. & Staton, C. A. Anti-angiogenic drug discovery: lessons from the past and thoughts for the future. Expert Opin. Drug Discov. 7, 723–743 (2012).
Bergers, G. & Hanahan, D. Modes of resistance to anti-angiogenic therapy. Nat. Rev. Cancer 8, 592–603 (2008).
Barone, A. et al. Combined VEGF and CXCR4 antagonism targets the GBM stem cell population and synergistically improves survival in an intracranial mouse model of glioblastoma. Oncotarget 5, 9811–9822 (2014). In a mouse model of glioblastoma, this study shows that angiogenesis, tumour growth and metastasis are reduced by treatment with the CXCR4 inhibitor CTCE-9908 in combination with docetaxel or a VEGF-specific antibody.
Hassan, S. et al. CXCR4 peptide antagonist inhibits primary breast tumor growth, metastasis and enhances the efficacy of anti-VEGF treatment or docetaxel in a transgenic mouse model. Int. J. Cancer 129, 225–232 (2011).
Pan, Q. et al. Blocking neuropilin-1 function has an additive effect with anti-VEGF to inhibit tumor growth. Cancer Cell 11, 53–67 (2007).
Ruan, G.-X. & Kazlauskas, A. Lactate engages receptor tyrosine kinases Axl, Tie2, and vascular endothelial growth factor receptor 2 to activate phosphoinositide 3-kinase/Akt and promote angiogenesis. J. Biol. Chem. 288, 21161–21172 (2013).
De Bock, K. et al. Role of PFKFB3-driven glycolysis in vessel sprouting. Cell 154, 651–663 (2013). This study demonstrates that loss of the glycolytic activator PFKFB3 in endothelial cells results in impaired angiogenesis. PFKFB3 is required for proliferation but also regulates filopodia formation and chemotaxis.
Leopold, J. A. et al. Glucose-6-phosphate dehydrogenase modulates vascular endothelial growth factor-mediated angiogenesis. J. Biol. Chem. 278, 32100–32106 (2003).
Schoors, S. et al. Partial and transient reduction of glycolysis by PFKFB3 blockade reduces pathological angiogenesis. Cell. Metab. 19, 37–48 (2014).
Leite de Oliveira, R. et al. Gene-targeting of Phd2 improves tumor response to chemotherapy and prevents side-toxicity. Cancer Cell 22, 263–277 (2012).
Mazzone, M. et al. Heterozygous deficiency of PHD2 restores tumor oxygenation and inhibits metastasis via endothelial normalization. Cell 136, 839–851 (2009).
Takeda, Y. et al. Macrophage skewing by Phd2 haplodeficiency prevents ischaemia by inducing arteriogenesis. Nature 479, 122–126 (2011).
Hamm, A. et al. PHD2 regulates arteriogenic macrophages through TIE2 signalling. EMBO Mol. Med. 5, 843–857 (2013).
Flagg, S. C., Martin, C. B., Taabazuing, C. Y., Holmes, B. E. & Knapp, M. J. Screening chelating inhibitors of HIF–prolyl hydroxylase domain 2 (PHD2) and factor inhibiting HIF (FIH). J. Inorg. Biochem. 113, 25–30 (2012).
Hellström, M. et al. Dll4 signalling through Notch1 regulates formation of tip cells during angiogenesis. Nature 445, 776–780 (2007).
Phng, L. K. et al. Nrarp coordinates endothelial Notch and Wnt signaling to control vessel density in angiogenesis. Dev. Cell 16, 70–82 (2009).
Dimova, I. et al. Inhibition of Notch signaling induces extensive intussusceptive neo-angiogenesis by recruitment of mononuclear cells. Angiogenesis 16, 921–937 (2013).
Outtz, H. H., Tattersall, I. W., Kofler, N. M., Steinbach, N. & Kitajewski, J. Notch1 controls macrophage recruitment and Notch signaling is activated at sites of endothelial cell anastomosis during retinal angiogenesis in mice. Blood 118, 3436–3439 (2011).
Camelo, S. et al. Delta-like 4 inhibits choroidal neovascularization despite opposing effects on vascular endothelium and macrophages. Angiogenesis 15, 609–622 (2012).
Haj Zen, A. A. et al. Inhibition of delta-like-4-mediated signaling impairs reparative angiogenesis after ischemia. Circ. Res. 107, 283–293 (2010).
Wang, Y. C. et al. Notch signaling determines the M1 versus M2 polarization of macrophages in antitumor immune responses. Cancer Res. 70, 4840–4849 (2010).
Xu, H. et al. Notch–RBP-J signaling regulates the transcription factor IRF8 to promote inflammatory macrophage polarization. Nat. Immunol. 13, 642–650 (2012).
Stockmann, C. et al. Deletion of vascular endothelial growth factor in myeloid cells accelerates tumorigenesis. Nature 456, 814–819 (2008).
DeNardo, D. G. et al. Leukocyte complexity predicts breast cancer survival and functionally regulates response to chemotherapy. Cancer Discov. 1, 54–67 (2011).
Ries, C. H. et al. Targeting tumor-associated macrophages with anti-CSF-1R antibody reveals a strategy for cancer therapy. Cancer Cell 25, 846–859 (2014).
Bonapace, L. et al. Cessation of CCL2 inhibition accelerates breast cancer metastasis by promoting angiogenesis. Nature 515, 130–133 (2014).
Stefater, J. A. 3rd et al. Regulation of angiogenesis by a non-canonical Wnt–Flt1 pathway in myeloid cells. Nature 474, 511–515 (2011).
Stefater, J. A. 3rd et al. Macrophage Wnt–Calcineurin–Flt1 signaling regulates mouse wound angiogenesis and repair. Blood 121, 2574–2578 (2013).
Ross, K. For organ transplant recipients, cancer threatens long-term survival. J. Nat. Cancer Inst. 99, 421–422 (2007).
Ley, K., Laudanna, C., Cybulsky, M. I. & Nourshargh, S. Getting to the site of inflammation: the leukocyte adhesion cascade updated. Nat. Rev. Immunol. 7, 678–689 (2007).
Nourshargh, S., Hordijk, P. L. & Sixt, M. Breaching multiple barriers: leukocyte motility through venular walls and the interstitium. Nat. Rev. Mol. Cell. Biol. 11, 366–378 (2010).
Phillipson, M. & Kubes, P. The neutrophil in vascular inflammation. Nat. Med. 17, 1381–1390 (2011).
Luo, B. H., Carman, C. V. & Springer, T. A. Structural basis of integrin regulation and signaling. Annu. Rev. Immunol. 25, 619–647 (2007).
Massena, S. et al. A chemotactic gradient sequestered on endothelial heparan sulfate induces directional intraluminal crawling of neutrophils. Blood 116, 1924–1931 (2010). This study demonstrates that an intravascular chemokine gradient on endothelial heparan sulfate directs crawling neutrophils towards the site of chemokine origin and accelerates their transmigration.
Phillipson, M. et al. Intraluminal crawling of neutrophils to emigration sites: a molecularly distinct process from adhesion in the recruitment cascade. J. Exp. Med. 203, 2569–2575 (2006).
Schenkel, A. R., Mamdouh, Z. & Muller, W. A. Locomotion of monocytes on endothelium is a critical step during extravasation. Nat. Immunol. 5, 393–400 (2004).
Voisin, M. B., Pröbstl, D. & Nourshargh, S. Venular basement membranes ubiquitously express matrix protein low-expression regions: characterization in multiple tissues and remodeling during inflammation. Am. J. Pathol. 176, 482–495 (2010).
Wang, S. et al. Venular basement membranes contain specific matrix protein low expression regions that act as exit points for emigrating neutrophils. J. Exp. Med. 203, 1519–1532 (2006).
Carman, C. V. & Springer, T. A. A transmigratory cup in leukocyte diapedesis both through individual vascular endothelial cells and between them. J. Cell Biol. 167, 377–388 (2004).
Petri, B. et al. Endothelial LSP1 is involved in endothelial dome formation, minimizing vascular permeability changes during neutrophil transmigration in vivo. Blood 117, 942–952 (2011).
Phillipson, M., Kaur, J., Colarusso, P., Ballantyne, C. M. & Kubes, P. Endothelial domes encapsulate adherent neutrophils and minimize increases in vascular permeability in paracellular and transcellular emigration. PLoS ONE 3, e1649 (2008).
Liu, L. et al. LSP1 is an endothelial gatekeeper of leukocyte transendothelial migration. J. Exp. Med. 201, 409–418 (2005).
Broermann, A. et al. Dissociation of VE-PTP from VE-cadherin is required for leukocyte extravasation and for VEGF-induced vascular permeability in vivo. J. Exp. Med. 208, 2393–2401 (2011).
Finsterbusch, M., Voisin, M. B., Beyrau, M., Williams, T. J. & Nourshargh, S. Neutrophils recruited by chemoattractants in vivo induce microvascular plasma protein leakage through secretion of TNF. J. Exp. Med. 211, 1307–1314 (2014).
Proebstl, D. et al. Pericytes support neutrophil subendothelial cell crawling and breaching of venular walls in vivo. J. Exp. Med. 209, 1219–1234 (2012). This study demonstrates that pericytes support neutrophil migration to inflamed tissue by providing a surface for neutrophil crawling and by enlarging the inter-pericyte spaces by contraction.
Abtin, A. et al. Perivascular macrophages mediate neutrophil recruitment during bacterial skin infection. Nat. Immunol. 15, 45–53 (2014). This study shows that perivascular macrophages guide neutrophil extravasation and comprise targets during bacterial immune evasion.
Tabas, I. & Glass, C. K. Anti-inflammatory therapy in chronic disease: challenges and opportunities. Science 339, 166–172 (2013).
Fox, D. A. Kinase inhibition — a new approach to the treatment of rheumatoid arthritis. N. Engl. J. Med. 367, 565–567 (2012).
Hoepner, R., Faissner, S., Salmen, A., Gold, R. & Chan, A. Efficacy and side effects of natalizumab therapy in patients with multiple sclerosis. J. Cent. Nerv. Syst. Dis. 6, 41–49 (2014).
Kothary, N. et al. Progressive multifocal leukoencephalopathy associated with efalizumab use in psoriasis patients. J. Am. Acad. Dermatol. 65, 546–551 (2011).
Schwab, N. et al. Fatal PML associated with efalizumab therapy: insights into integrin αLβ2 in JC virus control. Neurology 78, 458–467 (2012).
Van Assche, G. et al. Progressive multifocal leukoencephalopathy after natalizumab therapy for Crohn's disease. N. Engl. J. Med. 353, 362–368 (2005).
Feagan, B. G. et al. Vedolizumab as induction and maintenance therapy for ulcerative colitis. N. Eng. J. Med. 369, 699–710 (2013). In this clinical study, the beneficial effect of integrin-specific antibody therapy targeting α4β7 in patients with ulcerative colitis is demonstrated.
Sandborn, W. J. et al. Vedolizumab as induction and maintenance therapy for Crohn's disease. N. Eng. J. Med. 369, 711–721 (2013).
Briskin, M. et al. Human mucosal addressin cell adhesion molecule-1 is preferentially expressed in intestinal tract and associated lymphoid tissue. Am. J. Pathol. 151, 97–110 (1997).
Danese, S. & Panés, J. Development of drugs to target interactions between leukocytes and endothelial cells and treatment algorithms for inflammatory bowel diseases. Gastroenterology 147, 981–989 (2014).
Fedyk, E. R. et al. Exclusive antagonism of the α4β7 integrin by vedolizumab confirms the gut-selectivity of this pathway in primates. Inflamm. Bowel. Dis. 18, 2107–2119 (2012).
Haanstra, K. G. et al. Antagonizing the α4β1 integrin, but not α4β7, inhibits leukocytic infiltration of the central nervous system in rhesus monkey experimental autoimmune encephalomyelitis. J. Immunol. 190, 1961–1973 (2013).
Milch, C. et al. Vedolizumab, a monoclonal antibody to the gut homing α4β7 integrin, does not affect cerebrospinal fluid T-lymphocyte immunophenotype. J. Neuroimmunol. 264, 123–126 (2013).
Watanabe, M. et al. 370 AJM300, an oral α4 integrin antagonist, for active ulcerative colitis: a multicenter, randomized, double-blind, placebo-controlled Phase 2A study. Gastroenterology 146, S-82 (2014).
Economides, A. N. et al. Cytokine traps: multi-component, high-affinity blockers of cytokine action. Nat. Med. 9, 47–52 (2003).
Blanchetot, C. et al. Neutralizing nanobodies targeting diverse chemokines effectively inhibit chemokine function. J. Biol. Chem. 288, 25173–25182 (2013).
Berghmans, N. et al. Rescue from acute neuroinflammation by pharmacological chemokine-mediated deviation of leukocytes. J. Neuroinfl. 9, 243 (2012).
Li, S. et al. Interference with glycosaminoglycan-chemokine interactions with a probe to alter leukocyte recruitment and inflammation in vivo. PLoS ONE 9, e104107 (2014).
Klarenbeek, A. et al. Targeting chemokines and chemokine receptors with antibodies. Drug Discov. Today Tech. 9, 237–244 (2012).
Perry, C. M. Maraviroc: a review of its use in the management of CCR5-tropic HIV-1 infection. Drugs 70, 1189–1213 (2010).
De Clercq, E. The AMD3100 story: the path to the discovery of a stem cell mobilizer (Mozobil). Biochem. Pharmacol. 77, 1655–1664 (2009).
Moelants, E. A. et al. Citrullination of TNF-α by peptidylarginine deiminases reduces its capacity to stimulate the production of inflammatory chemokines. Cytokine 61, 161–167 (2013).
Proost, P. et al. Citrullination of CXCL8 by peptidylarginine deiminase alters receptor usage, prevents proteolysis, and dampens tissue inflammation. J. Exp. Med. 205, 2085–2097 (2008). This study demonstrates that inflammation can be regulated by post-translational modifications of chemokines to greatly affect chemokine activity and stability.
Moelants, E. A. et al. Citrullination and proteolytic processing of chemokines by Porphyromonas gingivalis. Infect. Immun. 82, 2511–2519 (2014).
Mantovani, A., Biswas, S. K., Galdiero, M. R., Sica, A. & Locati, M. Macrophage plasticity and polarization in tissue repair and remodelling. J. Pathol. 229, 176–185 (2013).
Korns, D., Frasch, S. C., Fernandez-Boyanapalli, R., Henson, P. M. & Bratton, D. L. Modulation of macrophage efferocytosis in inflammation. Front. Immunol. 2, 57 (2011).
Markworth, J. F. et al. Human inflammatory and resolving lipid mediator responses to resistance exercise and ibuprofen treatment. Am. J. Physiol. Regul. Integr. Comp. Physiol. 305, R1281–R1296 (2013).
Colas, R. A., Shinohara, M., Dalli, J., Chiang, N. & Serhan, C. N. Identification and signature profiles for pro-resolving and inflammatory lipid mediators in human tissue. Am. J. Physiol. Cell Physiol. 307, C39–C54 (2014). This study identifies the mediators involved in resolution of acute inflammation, including lipoxins, resolvins, protectins and maresins in human plasma and lymphoid organs.
Serhan, C. N. Pro-resolving lipid mediators are leads for resolution physiology. Nature 510, 92–101 (2014).
Jin, Y. et al. Anti-angiogenesis effect of the novel anti-inflammatory and pro-resolving lipid mediators. Invest. Ophthalmol. Vis. Sci. 50, 4743–4752 (2009).
Rajasagi, N. K. et al. Controlling herpes simplex virus-induced ocular inflammatory lesions with the lipid-derived mediator resolvin E1. J. Immunol. 186, 1735–1746 (2011).
Tang, Y. et al. Proresolution therapy for the treatment of delayed healing of diabetic wounds. Diabetes 62, 618–627 (2013).
Wu, S. H., Chen, X. Q., Liu, B., Wu, H. J. & Dong, L. Efficacy and safety of 15(R/S)-methyl-lipoxin A4 in topical treatment of infantile eczema. Br. J. Dermatol. 168, 172–178 (2013).
Robertson, A. L. et al. A zebrafish compound screen reveals modulation of neutrophil reverse migration as an anti-inflammatory mechanism. Sci. Transl. Med. 6, 225ra29 (2014).
Pellicoro, A., Ramachandran, P., Iredale, J. P. & Fallowfield, J. A. Liver fibrosis and repair: immune regulation of wound heling in solid organs. Nat. Rev. Immunol. 14, 181–194 (2014).
Moustakas, A. & Heldin, P. TGFβ and matrix regulated epithelial to mesenchymal transition. Biochim. Biophys. Acta 1840, 2621–2634 (2014).
Brenner, D. A. et al. Origin of myofibroblasts in liver fibrosis. Fibrogen. Tissue Repair 5, S17 (2012).
Hinz, B. et al. The myofibroblast: one function, multiple origins. Am. J. Pathol. 170, 1807–1816 (2007).
Henderson, N. C. et al. Targeting of αV integrin identifies a core molecular pathway that regulates fibrosis in several organs. Nat. Med. 19, 1617–1624 (2013). Depletion of αV integrin in mouse hepatic stellate cells was shown to protect against fibrosis in several organs. The authors suggest that pharmacological targeting of αV integrin may be of broad clinical utility.
Thiery, J. P., Acloque, H., Huang, R. Y. J. & Nieto, M. A. Epithelial-mesenchymal transitions in development and disease. Cell 139, 871–890 (2009).
Cenik, B. K., Ostapoff, K. T., Gerber, D. E. & Brekken, R. A. BIBF 1120 (nintedanib), a triple angiokinase inhibitor, induces hypoxia but not EMT and blocks progression of preclinical models of lung and pancreatic cancer. Mol. Cancer Ther. 12, 992–1001 (2013).
Mehal, W. Z., Iredale, J. & Friedman, S. L. Scraping fibrosis: expressway to the core of fibrosis. Nat. Med. 17, 552–553 (2011).
Seki, E. et al. TLR4 enhances TGF-β signaling and hepatic fibrosis. Nat. Med. 13, 1324–1332 (2007).
Subramaniam, N. et al. Metformin-mediated BAMBI expression in hepatic stellate cells induces prosurvival Wnt/β catenin signaling. Cancer Prev. Res. 5, 553–561 (2012).
Ogawa, S. et al. Anti-PDGF-B monoclonal antibody reduces liver fibrosis development. Hepatol. Res. 40, 1128–1141 (2010).
Coulon, S. et al. Role of vascular endothelial growth factor in the pathophysiology of nonalcoholic steatohepatitis in two rodent models. Hepatology 57, 1793–19805 (2013).
Ardi, V. C., Kupriyanova, T. A., Deryugina, E. I. & Quigley, J. P. Human neutrophils uniquely release TIMP-free MMP-9 to provide a potent catalytic stimulator of angiogenesis. Proc. Natl Acad. Sci. USA 104, 20262–20267 (2007).
Purcell, B. P. et al. Injectable and bioresponsive hydrogels for on-demand matrix metalloproteinase inhibition. Nat. Mater. 13, 653–661 (2014).
Bystrom, J. et al. Resolution-phase macrophages possess a unique inflammatory phenotype that is controlled by cAMP. Blood 112, 4117–4127 (2008).
Ramachandran, P. et al. Differential Ly-6C expression identifies the recruited macrophage phenotype, which orchestrates the regression of murine liver fibrosis. Proc. Natl Acad. Sci. USA 109, E3186–E3195 (2012). This paper describes the first identification and characterization of the 'restorative macrophage', which can promote tissue remodelling and resolution of fibrosis.
Hellström, M. et al. Lack of pericytes leads to endothelial hyperplasia and abnormal vascular morphogenesis. J. Cell. Biol. 153, 543–553 (2001).
Lindblom, P. et al. Endothelial PDGF-B retention is required for proper investment of pericytes in the microvessel wall. Genes Dev. 17, 1835–1840 (2003).
Lindahl, P., Johansson, B. R., Leveen, P. & Betsholtz, C. Pericyte loss and microaneurysm formation in PDGF-B-deficient mice. Science 277, 242–245 (1997).
Kelly-Goss, M. R., Sweat, R. S., Stapor, P. C., Peirce, S. M. & Murfee, W. L. Targeting pericytes for angiogenic therapies. Microcirculation 21, 345–357 (2014).
Ayres-Sander, C. E. et al. Transendothelial migration enables subsequent transmigration of neutrophils through underlying pericytes. PLoS ONE 8, e60025 (2013).
Ren, S. et al. LRP-6 is a coreceptor for multiple fibrogenic signaling pathways in pericytes and myofibroblasts that are inhibited by DKK-1. Proc. Natl Acad. Sci. USA 110, 1440–1445 (2013).
Humphreys, B. D. et al. Fate tracing reveals the pericyte and not epithelial origin of myofibroblasts in kidney fibrosis. Am. J. Pathol. 176, 85–97 (2010).
Lee, W. Y. et al. An intravascular immune response to Borrelia burgdorferi involves Kupffer cells and iNKT cells. Nat. Immunol. 11, 295–302 (2010).
Chintalgattu, V. et al. Coronary microvascular pericytes are the cellular target of sunitinib malate-induced cardiotoxicity. Sci. Transl. Med. 5, 187ra69 (2013).
Ehnman, M. et al. Distinct effects of ligand-induced PDGFRα and PDGFRβ signaling in the human rhabdomyosarcoma tumor cell and stroma cell compartments. Cancer Res. 73, 2139–2149 (2013).
Ruan, J. et al. Imatinin disrupts lymphoma angiogenesis by targeting vascular pericytes. Blood 121, 5192–5202 (2013).
Reck, M. et al. Docetaxel plus nintedanib versus docetaxel plus placebo in patients with previously treated non-small-cell lung cancer (LUME-Lung 1): a phase 3, double-blind, randomised controlled trial. Lancet Oncol. 15, 143–155 (2014).
Purow, B. Notch inhibition as a promising new approach to cancer therapy. Adv. Exp. Med. Biol. 727, 305–319 (2012).
Sun, L. et al. Discovery of 5-[5-fluoro-2-oxo-1,2- dihydroindol-(3Z)-ylidenemethyl]-2,4- dimethyl- 1H-pyrrole-3-carboxylic acid (2-diethylaminoethyl)amide, a novel tyrosine kinase inhibitor targeting vascular endothelial and platelet-derived growth factor receptor tyrosine kinase. J. Med. Chem. 46, 1116–1119 (2003).
Roberts, W. G. et al. Antiangiogenic and antitumor activity of a selective PDGFR tyrosine kinase inhibitor, CP-673,451. Cancer Res. 65, 957–966 (2005).
Wilhelm, S. M. et al. BAY 43–9006 exhibits broad spectrum oral antitumor activity and targets the RAF/MEK/ERK pathway and receptor tyrosine kinases involved in tumor progression and angiogenesis. Cancer Res. 64, 7099–7109 (2004).
Laird, A. D. et al. SU6668 is a potent antiangiogenic and antitumor agent that induces regression of established tumors. Cancer Res. 60, 4152–4160 (2000).
Pan, B. S. et al. MK-2461, a novel multitargeted kinase inhibitor, preferentially inhibits the activated c-Met receptor. Cancer Res. 70, 1524–1533 (2010).
Albert, D. H. et al. Preclinical activity of ABT-869, a multitargeted receptor tyrosine kinase inhibitor. Mol. Can. Therap. 5, 995–1006 (2006).
Hu-Lowe, D. D. et al. Nonclinical antiangiogenesis and antitumor activities of axitinib (AG-013736), an oral, potent, and selective inhibitor of vascular endothelial growth factor receptor tyrosine kinases 1, 2, 3. Clin. Cancer Res. 14, 7272–7283 (2008).
Buchdunger, E. et al. Selective inhibition of the platelet-derived growth factor signal transduction pathway by a protein-tyrosine kinase inhibitor of the 2-phenylaminopyrimidine class. Proc. Natl Acad. Sci. USA 92, 2558–2562 (1995).
Heinrich, M. C. et al. Inhibition of c-KIT receptor tyrosine kinase activity by STI 571, a selective tyrosine kinase inhibitor. Blood 96, 925–932 (2000).
Heinrich, M. C. et al. Crenolanib inhibits the drug-resistant PDGFRA D842V mutation associated with imatinib-resistant gastrointestinal stromal tumors. Clin. Cancer Res. 18, 4375–4384 (2012).
Trudel, S. et al. CHIR-258, a novel, multitargeted tyrosine kinase inhibitor for the potential treatment of t(4;14) multiple myeloma. Blood 105, 2941–2948 (2005).
Hilberg, F. et al. BIBF 1120: triple angiokinase inhibitor with sustained receptor blockade and good antitumor efficacy. Cancer Res. 68, 4774–4782 (2008).
Pyonteck, S. M. et al. CSF-1R inhibition alters macrophage polarization and blocks glioma progression. Nat. Med. 19, 1264–1272 (2013).
Qian, B. Z. et al. CCL2 recruits inflammatory monocytes to facilitate breast-tumour metastasis. Nature 475, 222–225 (2011).
Allavena, P. et al. Intraperitoneal recombinant gamma-interferon in patients with recurrent ascitic ovarian carcinoma: modulation of cytotoxicity and cytokine production in tumor-associated effectors and of major histocompatibility antigen expression on tumor cells. Cancer Res. 50, 7318–7323 (1990).
Krieg, A. M. Therapeutic potential of Toll-like receptor 9 activation. Nat. Rev. Drug Discov. 5, 471–484 (2006).
Sun, Z., Yao, Z., Liu, S., Tang, H. & Yan, X. An oligonucleotide decoy for Stat3 activates the immune response of macrophages to breast cancer. Immunobiology 211, 199–209 (2006).
Hemmi, H. et al. Small anti-viral compounds activate the immune cells via the TLR7 MyD88-dependent signaling pathway. Nat. Immunol. 3, 196–200 (2002).
Rolny, C. et al. HRG inhibits tumor growth and metastasis by inducing macrophage polarization and vessel normalization through downregulation of PlGF. Cancer Cell 19, 31–44 (2011).
Freemerman, A. J. et al. Metabolic reprogramming of macrophages: glucose transporter 1 (GLUT1)-mediated glucose metabolism drives a proinflammatory phenotype. J. Biol. Chem. 289, 7884–7896 (2014).
Beatty, G. L. et al. CD40 agonists alter tumor stroma and show efficacy against pancreatic carcinoma in mice and humans. Science 331, 1612–1616 (2011).
Odegaard, J. I. et al. Macrophage-specific PPARγ controls alternative activation and improves insulin resistance. Nature 447, 1116–1120 (2007).
Lutgens, E. et al. Deficient CD40–TRAF6 signaling in leukocytes prevents atherosclerosis by skewing the immune response toward an antiinflammatory profile. J. Exp. Med. 207, 391–404 (2010).
Kurowska-Stolarska, M. et al. IL-33 amplifies the polarization of alternatively activated macrophages that contribute to airway inflammation. J. Immunol. 183, 6469–6499 (2009).
Germano, G. et al. Antitumor and anti-inflammatory effects of trabectedin on human myxoid liposarcoma cells. Cancer Res. 70, 2235–2244 (2010).
Acknowledgements
The authors would like to thank the members of their laboratories and many of their colleagues, especially L. Claesson-Welsh, A. Moustakas and M. Welsh for sharing their insights and for stimulating discussions. The authors apologize to all colleagues whose relevant work has not been cited owing to space limitations. The authors are supported by the Swedish Medical Research Council, the Swedish Cancer Society, the Royal Swedish Academy of Sciences, the Swedish Diabetes Foundation, the Swedish Foundation for Strategic Research, the Novo Nordisk Foundation, the Ragnar Söderberg foundation, the Knut and Alice Wallenberg Foundation, the Ruth and Nils-Erik Stenbäcks Foundation, the Foundations for Proteoglycan Research, the Marie Sklodowska-Curie Innovative Training Network InCeM (Integrated Component Cycling in Epithelial Cell Motility) and Uppsala University.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Competing interests
The authors declare no competing financial interests.
Related links
Glossary
- Leukocytes
-
White blood cells of the immune system. Different classes of leukocytes may fight infections or modulate processes such as angiogenesis, inflammation and fibrosis.
- Mural cells
-
The vascular smooth muscle cells and pericytes that surround and support endothelial cells.
- Tunnelling nanotubes
-
Very thin cytoplasmic extensions containing actin that connect various cell types to allow direct exchange of intracellular components.
- Intravital microscopy
-
A microscopy technique that enables direct observations of biological processes in vivo.
- Extravasation
-
Emigration of immune cells from the vasculature into the surrounding tissue.
- Immunoliposomes
-
Antibody-conjugated spheres of phospholipids that can be used to deliver drugs to tissues.
- Alternatively activated macrophages
-
These macrophages are less efficient at killing bacteria and are activated by different cytokines than are the classic M1 inflammatory macrophages. Alternatively activated macrophages are involved in tissue remodelling, wound healing, angiogenesis and tumour progression.
- Inside-out signalling
-
Leukocyte integrin inside-out signalling occurs following chemokine receptor ligation and activates the ligand-binding function of the integrins.
- Outside-in signalling
-
Integrin outside-in signalling occurs following the binding of leukocyte integrins to their counter-receptors on the endothelium and results in cytoskeletal rearrangements, strengthening of adhesion, cell spreading and migration.
- Epithelial–mesenchymal transition
-
A process in which normally stationary epithelial cells lose cell–cell adhesions and cell polarity, thus becoming migratory.
Rights and permissions
About this article
Cite this article
Kreuger, J., Phillipson, M. Targeting vascular and leukocyte communication in angiogenesis, inflammation and fibrosis. Nat Rev Drug Discov 15, 125–142 (2016). https://doi.org/10.1038/nrd.2015.2
Published:
Issue Date:
DOI: https://doi.org/10.1038/nrd.2015.2
This article is cited by
-
Design of therapeutic biomaterials to control inflammation
Nature Reviews Materials (2022)
-
Prediction of the Postoperative Fat Volume Retention Rate After Augmentation Mammoplasty with Autologous Fat Grafting: From the Perspective of Preoperative Inflammatory Level
Aesthetic Plastic Surgery (2022)
-
Sex differences in stroke outcome correspond to rapid and severe changes in gut permeability in adult Sprague-Dawley rats
Biology of Sex Differences (2021)
-
Three-dimensional vascular microenvironment landscape in human glioblastoma
Acta Neuropathologica Communications (2021)
-
The secreted Ly6/uPAR-related protein-1 suppresses neutrophil binding, chemotaxis, and transmigration through human umbilical vein endothelial cells
Scientific Reports (2019)
