Quantification and cell-to-cell variation of vascular endothelial growth factor receptors

The vascular endothelial growth factor receptors (VEGFR) play a significant role in angiogenesis, the formation of new blood vessels from existing vasculature. Systems biology offers promising approaches to better understand angiogenesis by computational modeling the key molecular interactions in this process. Such modeling requires quantitative knowledge of cell surface density of pro-angiogenic receptors versus anti-angiogenic receptors, their regulation, and their cell-to-cell variability. Using quantitative fluorescence, we systematically characterized the endothelial surface density of VEGFRs and neuropilin-1 (NRP1). We also determined the role of VEGF in regulating the surface density of these receptors. Applying cell-by-cell analysis revealed heterogeneity in receptor surface density and VEGF tuning of this heterogeneity. Altogether, we determine inherent differences in the surface expression levels of these receptors and the role of VEGF in regulating the balance of anti-angiogenic or modulatory (VEGFR1) and pro-angiogenic (VEGFR2) receptors.     

GPR126 Protein Regulates Developmental and Pathological Angiogenesis through Modulation of VEGFR2 Receptor Signaling

Angiogenesis, the formation of new blood vessels from pre-existing ones, is essential for development, wound healing, and tumor progression. The VEGF pathway plays irreplaceable roles during angiogenesis, but how other signals cross-talk with and modulate VEGF cascades is not clearly elucidated. Here, we identified that Gpr126, an endothelial cell-enriched gene, plays an important role in angiogenesis by regulating endothelial cell proliferation, migration, and tube formation. Knockdown of Gpr126 in the mouse retina resulted in the inhibition of hypoxia-induced angiogenesis. Interference of Gpr126 expression in zebrafish embryos led to defects in intersegmental vessel formation. Finally, we identified that GPR126 regulated the expression of VEGFR2 by targeting STAT5 and GATA2 through the cAMP-PKA-cAMP-response element-binding protein signaling pathway during angiogenesis. Our findings illustrate that GPR126 modulates both physiological and pathological angiogenesis through VEGF signaling, providing a potential target for the treatment of angiogenesis-related diseases.     

Targeting neuropilin-1 to inhibit VEGF signaling in cancer: Comparison of therapeutic approaches

Angiogenesis (neovascularization) plays a crucial role in a variety of physiological and pathological conditions including cancer, cardiovascular disease, and wound healing. Vascular endothelial growth factor (VEGF) is a critical regulator of angiogenesis. Multiple VEGF receptors are expressed on endothelial cells, including signaling receptor tyrosine kinases (VEGFR1 and VEGFR2) and the nonsignaling co-receptor Neuropilin-1. Neuropilin-1 binds only the isoform of VEGF responsible for pathological angiogenesis (VEGF₁₆₅), and is thus a potential target for inhibiting VEGF signaling. Using the first molecularly detailed computational model of VEGF and its receptors, we have shown previously that the VEGFR–Neuropilin interactions explain the observed differential effects of VEGF isoforms on VEGF signaling in vitro, and demonstrated potent VEGF inhibition by an antibody to Neuropilin-1 that does not block ligand binding but blocks subsequent receptor coupling. In the present study, we extend that computational model to simulation of in vivo VEGF transport and binding, and predict the in vivo efficacy of several Neuropilin-targeted therapies in inhibiting VEGF signaling: (a) blocking Neuropilin-1 expression; (b) blocking VEGF binding to Neuropilin-1; (c) blocking Neuropilin–VEGFR coupling. The model predicts that blockade of Neuropilin–VEGFR coupling is significantly more effective than other approaches in decreasing VEGF–VEGFR2 signaling. In addition, tumor types with different receptor expression levels respond differently to each of these treatments. In designing human therapeutics, the mechanism of attacking the target plays a significant role in the outcome: of the strategies tested here, drugs with similar properties to the Neuropilin-1 antibody are predicted to be most effective. The tumor type and the microenvironment of the target tissue are also significant in determining therapeutic efficacy of each of the treatments studied.     

Signal transduction by VEGF receptors in regulation of angiogenesis and lymphangiogenesis

The VEGF/VPF (vascular endothelial growth factor/vascular permeability factor) ligands and receptors are crucial regulators of vasculogenesis, angiogenesis, lymphangiogenesis and vascular permeability in vertebrates. VEGF-A, the prototype VEGF ligand, binds and activates two tyrosine kinase receptors: VEGFR1 (Flt-1) and VEGFR2 (KDR/Flk-1). VEGFR1, which occurs in transmembrane and soluble forms, negatively regulates vasculogenesis and angiogenesis during early embryogenesis, but it also acts as a positive regulator of angiogenesis and inflammatory responses, playing a role in several human diseases such as rheumatoid arthritis and cancer. The soluble VEGFR1 is overexpressed in placenta in preeclampsia patients. VEGFR2 has critical functions in physiological and pathological angiogenesis through distinct signal transduction pathways regulating proliferation and migration of endothelial cells. VEGFR3, a receptor for the lymphatic growth factors VEGF-C and VEGF-D, but not for VEGF-A, regulates vascular and lymphatic endothelial cell function during embryogenesis. Loss-of-function variants of VEGFR3 have been identified in lymphedema. Formation of tumor lymphatics may be stimulated by tumor-produced VEGF-C, allowing increased spread of tumor metastases through the lymphatics. Mapping the signaling system of these important receptors may provide the knowledge necessary to suppress specific signaling pathways in major human diseases.     

Localized cysteine sulfenic acid formation by vascular endothelial growth factor: role in endothelial cell migration and angiogenesis

Abstract

Reactive oxygen species (ROS) are important mediators for VEGF receptor 2 (VEGFR2) signalling involved in angiogenesis. The initial product of Cys oxidation, cysteine sulfenic acid (Cys-OH), is a key intermediate in redox signal transduction; however, its role in VEGF signalling is unknown. We have previously demonstrated IQGAP1 as a VEGFR2 binding scaffold protein involved in ROS-dependent EC migration and post-ischemic angiogenesis. Using a biotin-labelled Cys-OH trapping reagent, we show that VEGF increases protein-Cys-OH formation at the lamellipodial leading edge where it co-localizes with NADPH oxidase and IQGAP1 in migrating ECs, which is prevented by IQGAP1 siRNA or trapping of Cys-OH with dimedone. VEGF increases IQGAP1-Cys-OH formation, which is prevented by N-acetyl cysteine or dimedone, which inhibits VEGF-induced EC migration and capillary network formation. In vivo, hindlimb ischemia in mice increases Cys-OH formation in small vessels and IQGAP1 in ischemic tissues. In summary, VEGF stimulates localized formation of Cys-OH-IQGAP1 at the leading edge, thereby promoting directional EC migration, which may contribute to post-natal angiogenesis in vivo. Thus, targeting Cys-oxidized proteins at specific compartments may be the potential therapeutic strategy for various angiogenesis-dependent diseases.     

Novel role of p66Shc in ROS-dependent VEGF signaling and angiogenesis in endothelial cells

p66Shc, a longevity adaptor protein, is demonstrated as a key regulator of reactive oxygen species (ROS) metabolism involved in aging and cardiovascular diseases. Vascular endothelial growth factor (VEGF) stimulates endothelial cell (EC) migration and proliferation primarily through the VEGF receptor-2 (VEGFR2). We have shown that ROS derived from Rac1-dependent NADPH oxidase are involved in VEGFR2 autophosphorylation and angiogenic-related responses in ECs. However, a role of p66Shc in VEGF signaling and physiological responses in ECs is unknown. Here we show that VEGF promotes p66Shc phosphorylation at Ser36 through the JNK/ERK or PKC pathway as well as Rac1 binding to a nonphosphorylated form of p66Shc in ECs. Depletion of endogenous p66Shc with short interfering RNA inhibits VEGF-induced Rac1 activity and ROS production. Fractionation of caveolin-enriched lipid raft demonstrates that p66Shc plays a critical role in VEGFR2 phosphorylation in caveolae/lipid rafts as well as downstream p38MAP kinase activation. This in turn stimulates VEGF-induced EC migration, proliferation, and capillary-like tube formation. These studies uncover a novel role of p66Shc as a positive regulator for ROS-dependent VEGFR2 signaling linked to angiogenesis in ECs and suggest p66Shc as a potential therapeutic target for various angiogenesis-dependent diseases.     

Bisphosphonates induce the osteogenic gene expression in co‐cultured human endothelial and mesenchymal stem cells

Bisphosphonates (BPs) are known to affect bone homeostasis and also to have anti-angiogenic properties. Because of the intimate relationship between angiogenesis and osteogenesis, this study analysed the effects of Alendronate (AL) and Zoledronate (ZL) in the expression of endothelial and osteogenic genes on interacting endothelial and mesenchymal stem cells, an issue that was not previously addressed. Alendronate and ZL, 10⁻¹²–10⁻⁶ M, were evaluated in a direct co-culture system of human dermal microvascular endothelial cells (HDMEC) and human bone marrow mesenchymal stem cells (HMSC), over a period of 14 days. Experiments with the respective monocultures were run in parallel. Alendronate and ZL caused an initial dose-dependent stimulation in the cell proliferation in the monocultures and co-cultures, and did not interfere with their cellular organization. In HDMEC monocultures, the expression of the endothelial genes CD31, VE-cadherin and VEGFR2 was down-regulated by AL and ZL. In HMSC monocultures, the BPs inhibited VEGF expression, but up-regulated the expression of the osteogenic genes alkaline phosphatase (ALP), bone morphogenic protein-2 (BMP-2) and osteocalcin (OC) and, to a greater extent, osteoprotegerin (OPG), a negative regulator of the osteoclastic differentiation, and increased ALP activity. In co-cultured HDMEC/HMSC, AL and ZL decreased the expression of endothelial genes but elicited an earlier and sustained overexpression of ALP, BMP-2, OC and OPG, compared with the monocultured cells; they also induced ALP activity. This study showed for the first time that AL and ZL greatly induced the osteogenic gene expression on interacting endothelial and mesenchymal stem cells.     

Expression of VEGF Receptors on Endothelial Cells in Mouse Skeletal Muscle

VEGFR surface localization plays a critical role in converting extracellular VEGF signaling towards angiogenic outcomes, and the quantitative characterization of these parameters is critical for advancing computational models; however the levels of these receptors on blood vessels is currently unknown. Therefore our aim is to quantitatively determine the VEGFR localization on endothelial cells from mouse hindlimb skeletal muscles. We contextualize this VEGFR quantification through comparison to VEGFR-levels on cells in vitro. Using quantitative fluorescence we measure and compare the levels of VEGFR1 and VEGFR2 on endothelial cells isolated from C57BL/6 and BALB/c gastrocnemius and tibialis anterior hindlimb muscles. Fluorescence measurements are calibrated using beads with known numbers of phycoerythrin molecules. The data show a 2-fold higher VEGFR1 surface localization relative to VEGFR2 with 2,000–3,700 VEGFR1/endothelial cell and 1,300–2,000 VEGFR2/endothelial cell. We determine that endothelial cells from the highly glycolytic muscle, tibialis anterior, contain 30% higher number of VEGFR1 surface receptors than gastrocnemius; BALB/c mice display ∼17% higher number of VEGFR1 than C57BL/6. When we compare these results to mouse fibroblasts in vitro, we observe high levels of VEGFR1 (35,800/cell) and very low levels of VEGFR2 (700/cell), while in human endothelial cells in vitro, we observe that the balance of VEGFRs is inverted, with higher levels VEGFR2 (5,800/cell) and lower levels of VEGFR1 (1,800/cell). Our studies also reveal significant cell-to-cell heterogeneity in receptor expression, and the quantification of these dissimilarities ex vivo for the first time provides insight into the balance of anti-angiogenic or modulatory (VEGFR1) and pro-angiogenic (VEGFR2) signaling.     

Toxicity and anti-angiogenicity evaluation of Pak1 inhibitor IPA-3 using zebrafish embryo model

p21-activated kinase 1 (Pak1)—a key node protein kinase regulating various cellular process including angiogenesis—has been recognised to be a therapeutic target for multitude of diseases, and hence, various small molecule inhibitors targeting its activity have been tested. However, the direct toxic and anti-angiogenic effects of these pharmacologic agents have not been examined. In this study, we evaluate the translational efficacy of Pak1 inhibitor IPA-3 using zebrafish toxicity model system to stratify its anti-angiogenic potential and off-target effects to streamline the compound for further therapeutic usage. The morphometric analysis has shown explicit delay in hatching, tail bending, pericardial sac oedema and abnormal angiogenesis. We provide novel evidence that Pak1 inhibitor could act as anti-angiogenic agents by impeding the development of sub-intestinal vessel (SIV) and intersegmental vessels (ISVs) by suppressing the expression of vascular endothelial growth factor (VEGF), VEGF receptor 2 (VEGFR2), neurophilin 1 (NRP1) and its downstream genes matrix metalloproteinase (MMP)-2 and MMP-9. Knockdown studies using 2-O-methylated oligoribonucleotides targeting Pak1 also revealed similar phenotypes with inhibition of angiogenesis accompanied with deregulation of major angiogenic factor and cardiac-specific genes. Taken together, our findings indicate that Pak1 signalling facilitates enhanced angiogenesis and also advocated the design and use of small molecule inhibitors of Pak1 as potent anti-angiogenic agents and suggest their utility in combinatorial therapeutic approaches targeting anomalous angiogenesis.     

Activation of alternative pathways of angiogenesis and involvement of stem cells following anti-angiogenesis treatment in glioma

Malignant gliomas are hypervascular tumors that are highly resistant to all the currently available multimodal treatments. Therefore, anti-angiogenic therapies targeting VEGF or VEGF receptors (VEGFRs) were designed and thought to be an effective tool for controlling the growth of malignant gliomas. However, recent results of early clinical trials using humanized monoclonal antibodies against VEGF (Bevacizumab), as well as small-molecule tyrosine kinase inhibitors that target different VEGF receptors (VEGFRs) (Vatalanib, Vandetanib, Sunitinib, Sorafenib, etc) alone or in combination with other therapeutic agents demonstrated differing outcomes, with the majority of reports indicating that glioma developed resistance to the employed anti-angiogenic treatments. It has been noted that continued anti-angiogenic therapy targeting only the VEGF-VEGFR system might affect pro-angiogenic factors other than VEGF, such as basic fibroblast growth factor (bFGF), stromal derived factor 1 (SDF-1) and Tie-2. These factors may in turn stimulate angiogenesis by mobilizing bone marrow derived precursor cells, such as endothelial progenitor cells (EPCs), which are known to promote angiogenesis and vasculogenesis. In this short review, the current antiangiogenic treatments, possible mechanisms of activation of alternative pathways of angiogenesis, and possible involvement of bone marrow derived progenitor cells in the failure of anti-angiogenic treatments are discussed.     

Differential roles of vascular endothelial growth factor receptor-1 and receptor-2 in angiogenesis

Vascular endothelial growth factor (VEGF)-A, a major regulator for angiogenesis, binds and activates two tyrosine kinase receptors, VEGFR1 (Flt-1) and VEGFR2 (KDR/Flk-1). These receptors regulate physiological as well as pathological angiogenesis. VEGFR2 has strong tyrosine kinase activity, and transduces the major signals for angiogenesis. However, unlike other representative tyrosine kinase receptors which use the Ras pathway, VEGFR2 mostly uses the Phospholipase-Cgamma-Protein kinase-C pathway to activate MAP-kinase and DNA synthesis. VEGFR2 is a direct signal transducer for pathological angiogenesis including cancer and diabetic retinopathy, thus, VEGFR2 itself and the signaling appear to be critical targets for the suppression of these diseases. VEGFR1 plays dual role, a negative role in angiogenesis in the embryo most likely by trapping VEGF-A, and a positive role in adulthood in a tyrosine kinase-dependent manner. VEGFR1 is expressed not only in endothelial cells but also in macrophage-lineage cells, and promotes tumor growth, metastasis, and inflammation. Furthermore, a soluble form of VEGFR1 was found to be present at abnormally high levels in the serum of preeclampsia patients, and induces proteinurea and renal dysfunction. Therefore, VEGFR1 is also an important target in the treatment of human diseases. Recently, the VEGFR2-specific ligand VEGF-E (Orf-VEGF) was extensively characterized. Interestingly, the activation of VEGFR2 via VEGF-E in vivo results in a strong angiogenic response in mice with minor side effects such as inflammation compared with VEGF-A, suggesting VEGF-E to be a novel material for pro-angiogenic therapy.     

A novel angiogenesis inhibitor impairs lovo cell survival via targeting against human VEGFR and its signaling pathway of phosphorylation

Colorectal cancer represents the fourth commonest malignancy, and constitutes a major cause of significant morbidity and mortality among other diseases. However, the chemical therapy is still under development. Angiogenesis plays an important role in colon cancer development. We developed HMQ18–22 (a novel analog of taspine) with the aim to target angiogenesis. We found that HMQ18–22 significantly reduced angiogenesis of chicken chorioallantoic membrane (CAM) and mouse colon tissue, and inhibited cell migration and tube formation as well. Then, we verified the interaction between HMQ18–22 and VEGFR2 by AlphaScreen P-VEGFR assay, screened the targets on angiogenesis by VEGF Phospho Antibody Array, validated the target by western blot and RNAi in lovo cells. We found HMQ18–22 could decrease phosphorylation of VEGFR2(Tyr¹²¹⁴), VEGFR1(Tyr¹³³³), Akt(Tyr³²⁶), protein kinase Cα (PKCα) (Tyr⁶⁵⁷) and phospholipase-Cγ-1 (PLCγ-1) (Tyr⁷⁷¹). Most importantly, HMQ18–22 inhibited proliferation of lovo cell and tumor growth in a human colon tumor xenografted model of athymic mice. Compared with normal lovo cells proliferation, the inhibition on proliferation of knockdown cells (VEGFR2, VEGFR1, Akt, PKCα and PLCγ-1) by HMQ18–22 decreased. These results suggested that HMQ18–22 is a novel angiogenesis inhibitor and can be a useful therapeutic candidate for colon cancer intervention.     

VEGFR1 (Flt1) Regulates Rab4 Recycling to Control Fibronectin Polymerization and Endothelial Vessel Branching

The cell's main receptor for VEGF, VEGFR2 (Kdr) is one of the most important positive regulators of new blood vessel growth and its downstream signalling is well characterized. By contrast, VEGFR1 (Flt1) and the mechanisms by which this VEGF receptor promotes branching morphogenesis in angiogenesis remain relatively unclear. Here we report that engagement of VEGFR1 activates a Rab4A-dependent pathway that transports αvβ3 integrin from early endosomes to the plasma membrane, and that this is required for VEGF-driven fibronectin polymerization in endothelial cells. Furthermore, VEGFR1 acts to promote endothelial tubule branching in an organotypic model of angiogenesis via a mechanism that requires Rab4A and αvβ3 integrin. We conclude that a recycling pathway regulated by Rab4A is a critical effector of VEGFR1 during branching morphogenesis of the vasculature.     

Molecular MRI assessment of vascular endothelial growth factor receptor‐2 in rat C6 gliomas

Angiogenesis is essential to tumour progression and a precise evaluation of angiogenesis is important for tumour early diagnosis and treatment. The quantitative and dynamic in vivo assessment of tumour angiogenesis can be achieved by molecular magnetic resonance imaging (mMRI). Vascular endothelial growth factor (VEGF) and VEGF receptors (VEGFRs) are the main regulatory systems in angiogenesis and have been used as hot targets for radionuclide‐based molecular imaging. However, little research has been accomplished in targeting VEGF/VEGFRs by mMRI. In our study, we aimed to assess the expression of VEGFR2 in C6 gliomas by using a specific molecular probe with mMRI. The differential uptake of the probe conjugated to anti‐VEGFR2 monoclonal antibody, shown by varied increases in T₁ signal intensity during a 2 hr period, demonstrated the heterogeneous expression of VEGFR2 in different tumour regions. Microscopic fluorescence imaging, obtained for the biotin group in the probe with streptavidin‐Cy3, along with staining for cellular VEGFR2 levels, laminin and CD45, confirmed the differential distribution of the probe which targeted VEGFR2 on endothelial cells. The angiogenesis process was also assessed using magnetic resonance angiography, which quantified tumour blood volume and provided a macroscopic view and a dynamic change of the correlation between tumour vasculature and VEGFR2 expression. Together these results suggest mMRI can be very useful in assessing and characterizing the expression of specific angiogenic markers in vivo and help evaluate angiogenesis associated with tumour progression.     

Localized cysteine sulfenic acid formation by vascular endothelial growth factor: role in endothelial cell migration and angiogenesis

Reactive oxygen species (ROS) are important mediators for VEGF receptor 2 (VEGFR2) signalling involved in angiogenesis. The initial product of Cys oxidation, cysteine sulfenic acid (Cys-OH), is a key intermediate in redox signal transduction; however, its role in VEGF signalling is unknown. We have previously demonstrated IQGAP1 as a VEGFR2 binding scaffold protein involved in ROS-dependent EC migration and post-ischemic angiogenesis. Using a biotin-labelled Cys-OH trapping reagent, we show that VEGF increases protein-Cys-OH formation at the lamellipodial leading edge where it co-localizes with NADPH oxidase and IQGAP1 in migrating ECs, which is prevented by IQGAP1 siRNA or trapping of Cys-OH with dimedone. VEGF increases IQGAP1-Cys-OH formation, which is prevented by N-acetyl cysteine or dimedone, which inhibits VEGF-induced EC migration and capillary network formation. In vivo, hindlimb ischemia in mice increases Cys-OH formation in small vessels and IQGAP1 in ischemic tissues. In summary, VEGF stimulates localized formation of Cys-OH-IQGAP1 at the leading edge, thereby promoting directional EC migration, which may contribute to post-natal angiogenesis in vivo. Thus, targeting Cys-oxidized proteins at specific compartments may be the potential therapeutic strategy for various angiogenesis-dependent diseases.     

Phage-derived fully human antibody scFv fragment directed against human vascular endothelial growth factor receptor 2 blocked its interaction with VEGF

Vascular endothelial growth factor receptor 2 (VEGFR‐2) plays a critical role in tumor angiogenesis. None therapeutic antibodies targeting VEGFR‐2 are available in clinical use. Herein, we describe the screening of a new single‐chain antibody fragment (scFv) targeting extracellular domain 3 of human VEGFR‐2 (kinase insert domain‐containing receptor [KDR]3) from Griffin phage display scFv library. A comprehensive sequence analysis was performed to assign the framework and complementary‐determining regions. The scFv exerted particular binding sites to KDR3 on molecular docking, and the binding affinity was further convinced by binding analysis both in quantitative ELISA and real‐time kinetic determination by biosensors (K_D = 40 nM). Finally, the scFv was revealed to inhibit VEGF‐stimulated proliferation of human umbilical vein endothelial cells (HUVECs; IC₅₀ = 5 nM) and to inhibit HUVEC migration significantly at 17 nM. Taken together, our results indicate that we have successfully isolated a scFv which differentially recognizes KDR3 and has potential clinical applications in the treatment of angiogenesis related diseases. © 2012 American Institute of Chemical Engineers Biotechnol. Prog., 28: 981–989, 2012     

Evaluation of inflammatory and angiogenic factors in patients with non-alcoholic fatty liver disease

The liver is a major target of injury in obese patients. Non-alcoholic fatty liver disease (NAFLD) is present in 60-90% of obese Americans and can range from simple steatosis to the more severe non-alcoholic steatohepatitis (NASH). The onset of a chronic inflammatory reaction marks the progression from simple steatosis to NASH and the expansion of adipose tissue is strongly associated with angiogenesis. Therefore, we determined the serum concentration of inflammatory [tumor necrosis factor alpha (TNFα) and interleukin 6 (IL6)] and angiogenic [vascular endothelial growth factor A (VEGF)] cytokines and soluble VEGF receptors 1 and 2 (sVEGFR1, sVEGFR2) in the serum of an obese population with simple steatosis and NASH compared to healthy controls. Moreover, we determined the TNFα, IL6, VEGF, VEGFR1 and VEGFR2 gene expression in the liver of these simple steatosis and NASH patients. The population consisted of 30 obese patients, which were diagnosed with simple steatosis and 32 patients with NASH and compared to 30 age-and-sex matched healthy controls. Mean serum TNFα levels were elevated in the serum of simple steatosis and NASH patients compared to healthy controls, reaching significance in NASH patients. IL6 was significantly increased in simple steatosis and NASH patients compared to the healthy controls. VEGF levels were significantly elevated in patients with simple steatosis and borderline significantly elevated in NASH patients compared to the serum levels of healthy control subjects. The concentration of sVEGFR1 was significantly increased in serum of simple steatosis and NASH patients compared to controls. sVEGFR2 concentration was not significantly different in the three groups. TNFα mRNA expression was higher in NASH patients compared to simple steatosis patients. Hepatic gene expression of VEGF, VEGFR1 and VEGFR2 were slightly decreased in NASH patients compared to simple steatosis patients. These data indicate the involvement of inflammatory (TNFα and IL6), angiogenic (VEGF) cytokines and sVEGFR1 in the pathophysiology of NAFLD.     

Receptor Tyrosine Kinase-Mediated Angiogenesis

The endothelial cell is the essential cell type forming the inner layer of the vasculature. Two families of receptor tyrosine kinases (RTKs) are almost completely endothelial cell specific: the vascular endothelial growth factor (VEGF) receptors (VEGFR1-3) and the Tie receptors (Tie1 and Tie2). Both are key players governing the generation of blood and lymphatic vessels during embryonic development. Because the growth of new blood and lymphatic vessels (or the lack thereof) is a central element in many diseases, the VEGF and the Tie receptors provide attractive therapeutic targets in various diseases. Indeed, several drugs directed to these RTK signaling pathways are already on the market, whereas many are in clinical trials. Here we review the VEGFR and Tie families, their involvement in developmental and pathological angiogenesis, and the different possibilities for targeting them to either block or enhance angiogenesis and lymphangiogenesis.