%0 Journal Article %J Duodecim Lääketieteellinen Aikakauskirja %D 2020 %T Lymphatics and the eye. [Finnish] %A Gucciardo, Erika %A Lehti, Timo A. %A Korhonen, Ani %A Salvén, Petri %A Lehti, Kaisa %A Jeltsch, Michael %A Loukovaara, Sirpa %B Duodecim Lääketieteellinen Aikakauskirja %V 136 %P 1777-1788 %8 2020/02/10/ %G eng %U https://www.duodecimlehti.fi/lehti/2020/16/duo15739 %N 16 %9 review %! Duodecim %& 1777 %R 10.5281/zenodo.4005517 %0 Journal Article %J Lymphologie in Forschung und Praxis %D 2019 %T The Proteolytic Activation of Vascular Endothelial Growth Factor-C %A Lackner, Marcel %A Schmotz, Constanze %A Jeltsch, Michael %K Lymphangiogenesis %K proteinases %K proteolysis %K VEGF-C %X The enzymatic cleavage of the protein backbone (proteolysis) is integral to many biological processes, e.g. for the break­down of proteins in the digestive system. Specific proteolytic cleavages are also used to turn on or off the activity of proteins. For example, the lymphangiogenic vascular endothelial growth factor-­C (VEGF-­C) is synthesized as a precursor molecule that must be converted to a mature form by the enzymatic removal of C-­ and N-­terminal propeptides before it can bind and activate its receptors. The constitutive C-­terminal cleavage is mediated by proprotein convertases such as furin. The subsequent ac­tivating cleavage can be mediated by at least four different proteases: by plasmin, ADAMTS3, prostate­-specific antigen (PSA) and cathepsin D. Processing by different proteases results in distinct forms of "ma­ture" VEGF­-C, that differ in their affinity and their receptor activation potential. This processing is tightly regulated by the CCBE1 protein. CCBE1 regulates the acti­vating cleavage of VEGF­C by ADAMTS3 and PSA, but not by plasmin. During embryonic development of the lymphatic system, VEGF-­C is activated primarily by the ADAMTS3 protease. In contrast, it is believed that plasmin is responsible for wound healing lymphangiogenesis and PSA for tumor-­associated pathological lym­phangiogenesis. Cathepsin D has also been implicated in tumor lymphangiogenesis. In addition, cathepsin D in saliva might activate latent VEGF­C upon wound licking, thereby accelerating wound healing. The molecular details of proteolytic activation of VEGF­-C are only recently extensively explored, and we likely do not know yet all activating proteases. It appears that the activity of VEGF­-C is regulated for different specific functions by different proteinases. Although VEGF­-C clearly plays a pivotal role for tumor progression and metastasis in experimental animal studies, the rele­vance of most correlative studies on the role of VEGF­-C in human cancers is quite limited until now, also due to the lack of methods to differentiate between inactive and active forms. %B Lymphologie in Forschung und Praxis %V 23 %P 88 - 98 %8 2019/12/18/ %G eng %U https://doi.org/10.5281/zenodo.3629263 %N 2 %9 Review %! LymphForsch %( Die proteolytische Aktivierung des Vaskulären Endothelzellwachstumsfaktors-C %R 10.5281/zenodo.3629263 %0 Journal Article %J Blood %D 2011 %T Structural determinants of vascular endothelial growth factor-D receptor binding and specificity %A Leppänen, Veli-Matti %A Jeltsch, Michael %A Anisimov, Andrey %A Tvorogov, Denis %A Aho, Kukka %A Kalkkinen, Nisse %A Toivanen, Pyry %A Ylä-Herttuala, Seppo %A Ballmer-Hofer, Kurt %A Alitalo, Kari %X Vascular endothelial growth factors (VEGFs) and their tyrosine kinase receptors (VEGFR-1-3) are central mediators of angiogenesis and lymphangiogenesis. VEGFR-3 ligands VEGF-C and VEGF-D are produced as precursor proteins with long N- and C-terminal propeptides and show enhanced VEGFR-2 and VEGFR-3 binding on proteolytic removal of the propeptides. Two different proteolytic cleavage sites have been reported in the VEGF-D N-terminus. We report here the crystal structure of the human VEGF-D Cys117Ala mutant at 2.9 Å resolution. Comparison of the VEGF-D and VEGF-C structures shows similar extended N-terminal helices, conserved overall folds, and VEGFR-2 interacting residues. Consistent with this, the affinity and the thermodynamic parameters for VEGFR-2 binding are very similar. In comparison with VEGF-C structures, however, the VEGF-D N-terminal helix was extended by 2 more turns because of a better resolution. Both receptor binding and functional assays of N-terminally truncated VEGF-D polypeptides indicated that the residues between the reported proteolytic cleavage sites are important for VEGF-D binding and activation of VEGFR-3, but not of VEGFR-2. Thus, we define here a VEGFR-2-specific form of VEGF-D that is angiogenic but not lymphangiogenic. These results provide important new insights into VEGF-D structure and function. %B Blood %V 117 %P 1507 - 15 %8 2011/Feb/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/21148085 %N 5 %! Blood %0 Journal Article %J Journal of Heart and Lung Transplantation %D 2011 %T VEGF-C/VEGFR-3 Signaling Regulates Inflammatory Response in Development of Obliterative Airway Disease %A Krebs, R %A Tikkanen, JM %A Ropponen, JO %A Jeltsch, M %A Jokinen, JJ %A Ylä-Herttuala, S %A Koskinen, PK %A Nykänen, AI %A Lemström, KB %B Journal of Heart and Lung Transplantation %V 30 %P S118 - S118 %8 2011// %G eng %0 Journal Article %J Genes Dev %D 2010 %T Claudin-like protein 24 interacts with the VEGFR-2 and VEGFR-3 pathways and regulates lymphatic vessel development %A Saharinen, Pipsa %A Helotera, Hanna %A Miettinen, Juho %A Norrmen, Camilla %A D'Amico, Gabriela %A Jeltsch, Michael %A Langenberg, Tobias %A Vandevelde, Wouter %A Ny, Annelii %A Dewerchin, Mieke %A Carmeliet, Peter %A Alitalo, Kari %X The Claudin-like protein of 24 kDa (CLP24) is a hypoxia-regulated transmembrane protein of unknown function. We show here that clp24 knockdown in Danio rerio and Xenopus laevis results in defective lymphatic development. Targeted disruption of Clp24 in mice led to enlarged lymphatic vessels having an abnormal smooth muscle cell coating. We also show that the Clp24(-/-) phenotype was further aggravated in the Vegfr2(+/LacZ) or Vegfr3(+/LacZ) backgrounds and that CLP24 interacts with vascular endothelial growth factor receptor-2 (VEGFR-2) and VEGFR-3 and attenuates the transcription factor CREB phosphorylation via these receptors. Our results indicate that CLP24 is a novel regulator of VEGFR-2 and VEGFR-3 signaling pathways and of normal lymphatic vessel structure. %B Genes Dev %V 24 %P 875 - 80 %8 2010/May/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/20439428 %N 9 %! Genes & Development %0 Journal Article %J Cancer Cell %D 2010 %T Effective suppression of vascular network formation by combination of antibodies blocking VEGFR ligand binding and receptor dimerization %A Tvorogov, Denis %A Anisimov, Andrey %A Zheng, Wei %A Leppänen, Veli-Matti %A Tammela, Tuomas %A Laurinavicius, Simonas %A Holnthoner, Wolfgang %A Heloterä, Hanna %A Holopainen, Tanja %A Jeltsch, Michael %A Kalkkinen, Nisse %A Lankinen, Hilkka %A Ojala, Päivi M %A Alitalo, Kari %X Antibodies that block vascular endothelial growth factor (VEGF) have become an integral part of antiangiogenic tumor therapy, and antibodies targeting other VEGFs and receptors (VEGFRs) are in clinical trials. Typically receptor-blocking antibodies are targeted to the VEGFR ligand-binding site. Here we describe a monoclonal antibody that inhibits VEGFR-3 homodimer and VEGFR-3/VEGFR-2 heterodimer formation, signal transduction, as well as ligand-induced migration and sprouting of microvascular endothelial cells. Importantly, we show that combined use of antibodies blocking ligand binding and receptor dimerization improves VEGFR inhibition and results in stronger inhibition of endothelial sprouting and vascular network formation in vivo. These results suggest that receptor dimerization inhibitors could be used to enhance antiangiogenic activity of antibodies blocking ligand binding in tumor therapy. %B Cancer Cell %V 18 %P 630 - 40 %8 2010/Dec/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/21130043 %N 6 %! Cancer Cell %0 Journal Article %J Proceedings of the National Academy of Sciences of the United States of America %D 2010 %T Structural determinants of growth factor binding and specificity by VEGF receptor 2 %A Leppänen, Veli-Matti %A Prota, Andrea E %A Jeltsch, Michael %A Anisimov, Andrey %A Kalkkinen, Nisse %A Strandin, Tomas %A Lankinen, Hilkka %A Goldman, Adrian %A Ballmer-Hofer, Kurt %A Alitalo, Kari %X Vascular endothelial growth factors (VEGFs) regulate blood and lymph vessel formation through activation of three receptor tyrosine kinases, VEGFR-1, -2, and -3. The extracellular domain of VEGF receptors consists of seven immunoglobulin homology domains, which, upon ligand binding, promote receptor dimerization. Dimerization initiates transmembrane signaling, which activates the intracellular tyrosine kinase domain of the receptor. VEGF-C stimulates lymphangiogenesis and contributes to pathological angiogenesis via VEGFR-3. However, proteolytically processed VEGF-C also stimulates VEGFR-2, the predominant transducer of signals required for physiological and pathological angiogenesis. Here we present the crystal structure of VEGF-C bound to the VEGFR-2 high-affinity-binding site, which consists of immunoglobulin homology domains D2 and D3. This structure reveals a symmetrical 22 complex, in which left-handed twisted receptor domains wrap around the 2-fold axis of VEGF-C. In the VEGFs, receptor specificity is determined by an N-terminal alpha helix and three peptide loops. Our structure shows that two of these loops in VEGF-C bind to VEGFR-2 subdomains D2 and D3, while one interacts primarily with D3. Additionally, the N-terminal helix of VEGF-C interacts with D2, and the groove separating the two VEGF-C monomers binds to the D2/D3 linker. VEGF-C, unlike VEGF-A, does not bind VEGFR-1. We therefore created VEGFR-1/VEGFR-2 chimeric proteins to further study receptor specificity. This biochemical analysis, together with our structural data, defined VEGFR-2 residues critical for the binding of VEGF-A and VEGF-C. Our results provide significant insights into the structural features that determine the high affinity and specificity of VEGF/VEGFR interactions. %B Proceedings of the National Academy of Sciences of the United States of America %V 107 %P 2425 - 30 %8 02/2010 %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/20145116 %N 6 %0 Journal Article %J PLoS ONE %D 2010 %T Suppressive effects of vascular endothelial growth factor-B on tumor growth in a mouse model of pancreatic neuroendocrine tumorigenesis %A Albrecht, Imke %A Kopfstein, Lucie %A Strittmatter, Karin %A Schomber, Tibor %A Falkevall, Annelie %A Hagberg, Carolina E %A Lorentz, Pascal %A Jeltsch, Michael %A Alitalo, Kari %A Eriksson, Ulf %A Christofori, Gerhard %A Pietras, Kristian %X BACKGROUND: The family of vascular endothelial growth factors (VEGF) contains key regulators of blood and lymph vessel development, including VEGF-A, -B, -C, -D, and placental growth factor. The role of VEGF-B during physiological or pathological angiogenesis has not yet been conclusively delineated. Herein, we investigate the function of VEGF-B by the generation of mouse models of cancer with transgenic expression of VEGF-B or homozygous deletion of Vegfb. METHODOLOGY/PRINCIPAL FINDINGS: Ectopic expression of VEGF-B in the insulin-producing β-cells of the pancreas did not alter the abundance or architecture of the islets of Langerhans. The vasculature from transgenic mice exhibited a dilated morphology, but was of similar density as that of wildtype mice. Unexpectedly, we found that transgenic expression of VEGF-B in the RIP1-Tag2 mouse model of pancreatic neuroendocrine tumorigenesis retarded tumor growth. Conversely, RIP1-Tag2 mice deficient for Vegfb presented with larger tumors. No differences in vascular density, perfusion or immune cell infiltration upon altered Vegfb gene dosage were noted. However, VEGF-B acted to increase blood vessel diameter both in normal pancreatic islets and in RIP1-Tag2 tumors. CONCLUSIONS/SIGNIFICANCE: Taken together, our results illustrate the differences in biological function between members of the VEGF family, and highlight the necessity of in-depth functional studies of VEGF-B to fully understand the effects of VEGFR-1 inhibitors currently used in the clinic. %B PLoS ONE %V 5 %P e14109 %8 2010// %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/21124841 %N 11 %! PLoS ONE %0 Journal Article %J Circulation %D 2010 %T Vascular endothelial growth factor-B acts as a coronary growth factor in transgenic rats without inducing angiogenesis, vascular leak, or inflammation %A Bry, Maija %A Kivelä, Riikka %A Holopainen, Tanja %A Anisimov, Andrey %A Tammela, Tuomas %A Soronen, Jarkko %A Silvola, Johanna %A Saraste, Antti %A Jeltsch, Michael %A Korpisalo, Petra %A Carmeliet, Peter %A Lemström, Karl B %A Shibuya, Masabumi %A Ylä-Herttuala, Seppo %A Alhonen, Leena %A Mervaala, Eero %A Andersson, Leif C %A Knuuti, Juhani %A Alitalo, Kari %X BACKGROUND: Vascular endothelial growth factor-B (VEGF-B) binds to VEGF receptor-1 and neuropilin-1 and is abundantly expressed in the heart, skeletal muscle, and brown fat. The biological function of VEGF-B is incompletely understood. METHODS AND RESULTS: Unlike placenta growth factor, which binds to the same receptors, adeno-associated viral delivery of VEGF-B to mouse skeletal or heart muscle induced very little angiogenesis, vascular permeability, or inflammation. As previously reported for the VEGF-B(167) isoform, transgenic mice and rats expressing both isoforms of VEGF-B in the myocardium developed cardiac hypertrophy yet maintained systolic function. Deletion of the VEGF receptor-1 tyrosine kinase domain or the arterial endothelial Bmx tyrosine kinase inhibited hypertrophy, whereas loss of VEGF-B interaction with neuropilin-1 had no effect. Surprisingly, in rats, the heart-specific VEGF-B transgene induced impressive growth of the epicardial coronary vessels and their branches, with large arteries also seen deep inside the subendocardial myocardium. However, VEGF-B, unlike other VEGF family members, did not induce significant capillary angiogenesis, increased permeability, or inflammatory cell recruitment. CONCLUSIONS: VEGF-B appears to be a coronary growth factor in rats but not in mice. The signals for the VEGF-B-induced cardiac hypertrophy are mediated at least in part via the endothelium. Because cardiomyocyte damage in myocardial ischemia begins in the subendocardial myocardium, the VEGF-B-induced increased arterial supply to this area could have therapeutic potential in ischemic heart disease. %B Circulation %V 122 %P 1725 - 33 %8 2010/Oct/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/20937974 %N 17 %! Circulation %0 Journal Article %J Circ Res %D 2009 %T Activated forms of VEGF-C and VEGF-D provide improved vascular function in skeletal muscle %A Anisimov, Andrey %A Alitalo, Annamari %A Korpisalo, Petra %A Soronen, Jarkko %A Kaijalainen, Seppo %A Leppänen, Veli-Matti %A Jeltsch, Michael %A Ylä-Herttuala, Seppo %A Alitalo, Kari %X The therapeutic potential of vascular endothelial growth factor (VEGF)-C and VEGF-D in skeletal muscle has been of considerable interest as these factors have both angiogenic and lymphangiogenic activities. Previous studies have mainly used adenoviral gene delivery for short-term expression of VEGF-C and VEGF-D in pig, rabbit, and mouse skeletal muscles. Here we have used the activated mature forms of VEGF-C and VEGF-D expressed via recombinant adeno-associated virus (rAAV), which provides stable, long-lasting transgene expression in various tissues including skeletal muscle. Mouse tibialis anterior muscle was transduced with rAAV encoding human or mouse VEGF-C or VEGF-D. Two weeks later, immunohistochemical analysis showed increased numbers of both blood and lymph vessels, and Doppler ultrasound analysis indicated increased blood vessel perfusion. The lymphatic vessels further increased at the 4-week time point were functional, as shown by FITC-lectin uptake and transport. Furthermore, receptor activation and arteriogenic activity were increased by an alanine substitution mutant of human VEGF-C (C137A) having an increased dimer stability and by a chimeric CAC growth factor that contained the VEGF receptor-binding domain flanked by VEGF-C propeptides, but only the latter promoted significantly more blood vessel perfusion when compared to the other growth factors studied. We conclude that long-term expression of VEGF-C and VEGF-D in skeletal muscle results in the generation of new functional blood and lymphatic vessels. The therapeutic value of intramuscular lymph vessels in draining tissue edema and lymphedema can now be evaluated using this model system. %B Circ Res %V 104 %P 1302 - 12 %8 2009/Jun/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/19443835 %N 11 %! Circulation Research %0 Journal Article %J Circ Res %D 2008 %T Overexpression of vascular endothelial growth factor-B in mouse heart alters cardiac lipid metabolism and induces myocardial hypertrophy %A Karpanen, Terhi %A Bry, Maija %A Ollila, Hanna M %A Seppänen-Laakso, Tuulikki %A Liimatta, Erkki %A Leskinen, Hanna %A Kivelä, Riikka %A Helkamaa, Teemu %A Merentie, Mari %A Jeltsch, Michael %A Paavonen, Karri %A Andersson, Leif C %A Mervaala, Eero %A Hassinen, Ilmo E %A Ylä-Herttuala, Seppo %A Oresic, Matej %A Alitalo, Kari %X Vascular endothelial growth factor (VEGF)-B is poorly angiogenic but prominently expressed in metabolically highly active tissues, including the heart. We produced mice expressing a cardiac-specific VEGF-B transgene via the alpha-myosin heavy chain promoter. Surprisingly, the hearts of the VEGF-B transgenic mice showed concentric cardiac hypertrophy without significant changes in heart function. The cardiac hypertrophy was attributable to an increased size of the cardiomyocytes. Blood capillary size was increased, whereas the number of blood vessels per cell nucleus remained unchanged. Despite the cardiac hypertrophy, the transgenic mice had lower heart rate and blood pressure than their littermates, and they responded similarly to angiotensin II-induced hypertension, confirming that the hypertrophy does not compromise heart function. Interestingly, the isolated transgenic hearts had less cardiomyocyte damage after ischemia. Significantly increased ceramide and decreased triglyceride levels were found in the transgenic hearts. This was associated with structural changes and eventual lysis of mitochondria, resulting in accumulation of intracellular vacuoles in cardiomyocytes and increased death of the transgenic mice, apparently because of mitochondrial lipotoxicity in the heart. These results suggest that VEGF-B regulates lipid metabolism, an unexpected function for an angiogenic growth factor. %B Circ Res %V 103 %P 1018 - 26 %8 2008/Oct/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/18757827 %N 9 %! Circulation Research %0 Journal Article %J Arterioscler Thromb Vasc Biol %D 2008 %T Reevaluation of the role of VEGF-B suggests a restricted role in the revascularization of the ischemic myocardium %A Li, Xuri %A Tjwa, Marc %A Van Hove, Inge %A Enholm, Berndt %A Neven, Elke %A Paavonen, Karri %A Jeltsch, Michael %A Juan, Toni Diez %A Sievers, Richard E %A Chorianopoulos, Emmanuel %A Wada, Hiromichi %A Vanwildemeersch, Maarten %A Noel, Agnes %A Foidart, Jean-Michel %A Springer, Matthew L %A von Degenfeld, Georges %A Dewerchin, Mieke %A Blau, Helen M %A Alitalo, Kari %A Eriksson, Ulf %A Carmeliet, Peter %A Moons, Lieve %X OBJECTIVE: The endogenous role of the VEGF family member vascular endothelial growth factor-B (VEGF-B) in pathological angiogenesis remains unclear. METHODS AND RESULTS: We studied the role of VEGF-B in various models of pathological angiogenesis using mice lacking VEGF-B (VEGF-B(-/-)) or overexpressing VEGF-B(167). After occlusion of the left coronary artery, VEGF-B deficiency impaired vessel growth in the ischemic myocardium whereas, in wild-type mice, VEGF-B(167) overexpression enhanced revascularization of the infarct and ischemic border zone. By contrast, VEGF-B deficiency did not affect vessel growth in the wounded skin, hypoxic lung, ischemic retina, or ischemic limb. Moreover, VEGF-B(167) overexpression failed to enhance vascular growth in the skin or ischemic limb. CONCLUSIONS: VEGF-B appears to have a relatively restricted angiogenic activity in the ischemic heart. These insights might offer novel therapeutic opportunities. %B Arterioscler Thromb Vasc Biol %V 28 %P 1614 - 20 %8 2008/Sep/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/18511699 %N 9 %! Arteriosclerosis, Thrombosis, and Vascular Biology %0 Journal Article %J Circ Res %D 2007 %T Distinct architecture of lymphatic vessels induced by chimeric vascular endothelial growth factor-C/vascular endothelial growth factor heparin-binding domain fusion proteins %A Tammela, Tuomas %A He, Yulong %A Lyytikkä, Johannes %A Jeltsch, Michael %A Markkanen, Johanna %A Pajusola, Katri %A Ylä-Herttuala, Seppo %A Alitalo, Kari %X Vascular endothelial growth factor (VEGF)-C and VEGF-D are composed of the receptor-binding VEGF homology domain and a carboxy-terminal silk homology domain that requires proteolytic cleavage for growth factor activation. Here, we explored whether the C-terminal heparin-binding domain of the VEGF(165) or VEGF(189) isoform also containing neuropilin-binding sequences could substitute for the silk homology domain of VEGF-C. Such VEGF-C/VEGF-heparin-binding domain chimeras were produced and shown to activate VEGF-C receptors, and, when expressed in tissues via adenovirus or adeno-associated virus vectors, stimulated lymphangiogenesis in vivo. However, both chimeras induced a distinctly different pattern of lymphatic vessels when compared with VEGF-C. Whereas VEGF-C-induced vessels were initially a dense network of small diameter vessels, the lymphatic vessels induced by the chimeric growth factors tended to form directly along tissue borders, along basement membranes that are rich in heparan sulfate. For example, in skeletal muscle, the chimeras induced formation of lumenized lymphatic vessels more efficiently than wild-type VEGF-C. We conclude that the matrix-binding domain of VEGF can target VEGF-C activity to heparin-rich basement membrane structures. These properties may prove useful for tissue engineering and attempts to regenerate lymphatic vessels in lymphedema patients. %B Circ Res %V 100 %P 1468 - 75 %8 2007/May/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/17478733 %N 10 %! Circulation Research %0 Journal Article %J Circ Res %D 2007 %T Enhanced capillary formation stimulated by a chimeric vascular endothelial growth factor/vascular endothelial growth factor-C silk domain fusion protein %A Keskitalo, Salla %A Tammela, Tuomas %A Lyytikka, Johannes %A Karpanen, Terhi %A Jeltsch, Michael %A Markkanen, Johanna %A Yla-Herttuala, Seppo %A Alitalo, Kari %X Vascular endothelial growth factor (VEGF)-C and VEGF-D require proteolytic cleavage of the carboxy terminal silk-homology domain for activation. To study the functions of the VEGF-C propeptides, we engineered a chimeric growth factor protein, VEGF-CAC, composed of the amino- and carboxy-terminal propeptides of VEGF-C fused to the receptor-activating core domain of VEGF. Like VEGF-C, VEGF-CAC underwent proteolytic cleavage, and like VEGF, it bound to and activated VEGF receptor-1 and VEGF receptor-2, but not the VEGF-C receptor VEGF receptor-3. VEGF-CAC also bound to neuropilins in a heparin-dependent manner. Strikingly, when VEGF-CAC was expressed via an adenovirus vector in the ear skin of immunodeficient mice, it proved to be a more potent inducer of capillary angiogenesis than VEGF. The VEGF-CAC-induced vessels differed greatly from those induced by VEGF, as they formed a very dense and fine network of pericyte and basement membrane-covered capillaries that were functional, as shown by lectin perfusion experiments. VEGF-CAC could prove useful in proangiogenic therapies in patients experiencing tissue ischemia. %B Circ Res %V 100 %P 1460 - 7 %8 2007/May/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/17478734 %N 10 %! Circulation Research %0 Journal Article %J J Biol Chem %D 2006 %T Vascular endothelial growth factor (VEGF)/VEGF-C mosaic molecules reveal specificity determinants and feature novel receptor binding patterns %A Jeltsch, Michael %A Karpanen, Terhi %A Strandin, Tomas %A Aho, Kukka %A Lankinen, Hilkka %A Alitalo, Kari %X Vascular endothelial growth factors (VEGFs) and their receptors play key roles in angiogenesis and lymphangiogenesis. VEGF activates VEGF receptor-1 (VEGFR-1) and VEGFR-2, whereas VEGF-C activates VEGFR-2 and VEGFR-3. We have created a library of VEGF/VEGF-C mosaic molecules that contains factors with novel receptor binding profiles, notably proteins binding to all three VEGF receptors ("super-VEGFs"). The analyzed super-VEGFs show both angiogenic and lymphangiogenic effects in vivo, although weaker than the parental molecules. The composition of the VEGFR-3 binding molecules and scanning mutagenesis revealed determinants of receptor binding and specificity. VEGFR-2 and VEGFR-3 showed striking differences in their requirements for VEGF-C binding; extracellular domain 2 of VEGFR-2 was sufficient, whereas in VEGFR-3, both domains 1 and 2 were necessary. %B J Biol Chem %V 281 %P 12187 - 95 %8 2006/Apr/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/16505489 %N 17 %! The Journal of Biological Chemistry %0 Journal Article %J J Clin Invest %D 2005 %T Pathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation %A Baluk, Peter %A Tammela, Tuomas %A Ator, Erin %A Lyubynska, Natalya %A Achen, Marc G %A Hicklin, Daniel J %A Jeltsch, Michael %A Petrova, Tatiana V %A Pytowski, Bronislaw %A Stacker, Steven A %A Ylä-Herttuala, Seppo %A Jackson, David G %A Alitalo, Kari %A McDonald, Donald M %X Edema occurs in asthma and other inflammatory diseases when the rate of plasma leakage from blood vessels exceeds the drainage through lymphatic vessels and other routes. It is unclear to what extent lymphatic vessels grow to compensate for increased leakage during inflammation and what drives the lymphangiogenesis that does occur. We addressed these issues in mouse models of (a) chronic respiratory tract infection with Mycoplasma pulmonis and (b) adenoviral transduction of airway epithelium with VEGF family growth factors. Blood vessel remodeling and lymphangiogenesis were both robust in infected airways. Inhibition of VEGFR-3 signaling completely prevented the growth of lymphatic vessels but not blood vessels. Lack of lymphatic growth exaggerated mucosal edema and reduced the hypertrophy of draining lymph nodes. Airway dendritic cells, macrophages, neutrophils, and epithelial cells expressed the VEGFR-3 ligands VEGF-C or VEGF-D. Adenoviral delivery of either VEGF-C or VEGF-D evoked lymphangiogenesis without angiogenesis, whereas adenoviral VEGF had the opposite effect. After antibiotic treatment of the infection, inflammation and remodeling of blood vessels quickly subsided, but lymphatic vessels persisted. Together, these findings suggest that when lymphangiogenesis is impaired, airway inflammation may lead to bronchial lymphedema and exaggerated airflow obstruction. Correction of defective lymphangiogenesis may benefit the treatment of asthma and other inflammatory airway diseases. %B J Clin Invest %V 115 %P 247 - 57 %8 2005/Feb/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/15668734 %N 2 %! The Journal of Clinical Investigation %0 Journal Article %J FASEB J %D 2003 %T Intrinsic versus microenvironmental regulation of lymphatic endothelial cell phenotype and function %A Veikkola, Tanja %A Lohela, Marja %A Ikenberg, Kristian %A Mäkinen, Taija %A Korff, Thomas %A Saaristo, Anne %A Petrova, Tatania %A Jeltsch, Michael %A Augustin, Hellmut G %A Alitalo, Kari %X Vascular endothelial cells are characterized by a high degree of functional and phenotypic plasticity, which is controlled both by their pericellular microenvironment and their intracellular gene expression programs. To gain further insight into the mechanisms regulating the endothelial cell phenotype, we have compared the responses of lymphatic endothelial cells (LECs) and blood vascular endothelial cells (BECs) to vascular endothelial growth factors (VEGFs). VEGFR-3-specific signals are sufficient for LEC but not BEC proliferation, as shown by the ability of the specific ligand VEGF-C156S to stimulate cell cycle entry only in LECs. On the other hand, we found that VEGFR-3 stimulation did not induce LEC cell shape changes typical of VEGFR-2-stimulated LECs, indicating receptor-specific differences in the cytoskeletal responses. Genes induced via VEGFR-2 also differed between BECs and LECs: angiopoietin-2 (Ang-2) was induced via VEGFR-2 in BECs and LECs, but the smooth muscle cell (SMC) chemoattractant BMP-2 was induced only in BECs. Both BECs and LECs were able to promote SMC chemotaxis, but contact with SMCs led to down-regulation of VEGFR-3 expression in BECs in a 3-dimensional coculture system. This was consistent with the finding that VEGFR-3 is down-regulated in vivo at sites of endothelial cell-pericyte/smooth muscle cell contacts. Collectively, these data show intrinsic cell-specific differences of BEC and LEC responses to VEGFs and identify a pericellular regulatory mechanism for VEGFR-3 down-regulation in endothelial cells. %B FASEB J %V 17 %P 2006 - 13 %8 2003/Nov/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/14597670 %N 14 %! The FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology %0 Journal Article %J J Cell Biol %D 2003 %T VEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia %A Gerhardt, Holger %A Golding, Matthew %A Fruttiger, Marcus %A Ruhrberg, Christiana %A Lundkvist, Andrea %A Abramsson, Alexandra %A Jeltsch, Michael %A Mitchell, Christopher %A Alitalo, Kari %A Shima, David %A Betsholtz, Christer %X Vascular endothelial growth factor (VEGF-A) is a major regulator of blood vessel formation and function. It controls several processes in endothelial cells, such as proliferation, survival, and migration, but it is not known how these are coordinately regulated to result in more complex morphogenetic events, such as tubular sprouting, fusion, and network formation. We show here that VEGF-A controls angiogenic sprouting in the early postnatal retina by guiding filopodial extension from specialized endothelial cells situated at the tips of the vascular sprouts. The tip cells respond to VEGF-A only by guided migration; the proliferative response to VEGF-A occurs in the sprout stalks. These two cellular responses are both mediated by agonistic activity of VEGF-A on VEGF receptor 2. Whereas tip cell migration depends on a gradient of VEGF-A, proliferation is regulated by its concentration. Thus, vessel patterning during retinal angiogenesis depends on the balance between two different qualities of the extracellular VEGF-A distribution, which regulate distinct cellular responses in defined populations of endothelial cells. %B J Cell Biol %V 161 %P 1163 - 77 %8 2003/Jun/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/12810700 %N 6 %! The Journal of Cell Biology %0 Journal Article %J Circulation %D 2000 %T Intravascular adenovirus-mediated VEGF-C gene transfer reduces neointima formation in balloon-denuded rabbit aorta %A Hiltunen, M O %A Laitinen, M %A Turunen, M P %A Jeltsch, M %A Hartikainen, J %A Rissanen, T T %A Laukkanen, J %A Niemi, M %A Kossila, M %A Häkkinen, T P %A Kivelä, A %A Enholm, B %A Mansukoski, H %A Turunen, A M %A Alitalo, K %A Ylä-Herttuala, S %X BACKGROUND: Gene transfer to the vessel wall may provide new possibilities for the treatment of vascular disorders, such as postangioplasty restenosis. In this study, we analyzed the effects of adenovirus-mediated vascular endothelial growth factor (VEGF)-C gene transfer on neointima formation after endothelial denudation in rabbits. For comparison, a second group was treated with VEGF-A adenovirus and a third group with lacZ adenovirus. Clinical-grade adenoviruses were used for the study. METHODS AND RESULTS: Aortas of cholesterol-fed New Zealand White rabbits were balloon-denuded, and gene transfer was performed 3 days later. Animals were euthanized 2 and 4 weeks after the gene transfer, and intima/media ratio (I/M), histology, and cell proliferation were analyzed. Two weeks after the gene transfer, I/M in the lacZ-transfected control group was 0. 57+/-0.04. VEGF-C gene transfer reduced I/M to 0.38+/-0.02 (P:<0.05 versus lacZ group). I/M in VEGF-A-treated animals was 0.49+/-0.17 (P:=NS). The tendency that both VEGF groups had smaller I/M persisted at the 4-week time point, when the lacZ group had an I/M of 0.73+/-0.16, the VEGF-C group 0.44+/-0.14, and the VEGF-A group 0. 63+/-0.21 (P:=NS). Expression of VEGF receptors 1, 2, and 3 was detected in the vessel wall by immunocytochemistry and in situ hybridization. As an additional control, the effect of adenovirus on cell proliferation was analyzed by performing gene transfer to intact aorta without endothelial denudation. No differences were seen in smooth muscle cell proliferation or I/M between lacZ adenovirus and 0.9% saline-treated animals. CONCLUSIONS: Adenovirus-mediated VEGF-C gene transfer may be useful for the treatment of postangioplasty restenosis and vessel wall thickening after vascular manipulations. %B Circulation %V 102 %P 2262 - 8 %8 2000/Oct/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/11056103 %N 18 %! Circulation %0 Journal Article %J Science (80- ) %D 1997 %T Hyperplasia of lymphatic vessels in VEGF-C transgenic mice %A Jeltsch, M %A Kaipainen, A %A Joukov, V %A Meng, X %A Lakso, M %A Rauvala, H %A Swartz, M %A Fukumura, D %A Jain, R K %A Alitalo, K %X No growth factors specific for the lymphatic vascular system have yet been described. Vascular endothelial growth factor (VEGF) regulates vascular permeability and angiogenesis, but does not promote lymphangiogenesis. Overexpression of VEGF-C, a ligand of the VEGF receptors VEGFR-3 and VEGFR-2, in the skin of transgenic mice resulted in lymphatic, but not vascular, endothelial proliferation and vessel enlargement. Thus, VEGF-C induces selective hyperplasia of the lymphatic vasculature, which is involved in the draining of interstitial fluid and in immune function, inflammation, and tumor metastasis. VEGF-C may play a role in disorders involving the lymphatic system and may be of potential use in therapeutic lymphangiogenesis. %B Science (80- ) %V 276 %P 1423 - 5 %8 1997/May/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/9162011 %N 5317 %! Science %0 Journal Article %J Development %D 1996 %T VEGF-C receptor binding and pattern of expression with VEGFR-3 suggests a role in lymphatic vascular development %A Kukk, E %A Lymboussaki, A %A Taira, S %A Kaipainen, A %A Jeltsch, M %A Joukov, V %A Alitalo, K %X The vascular endothelial growth factor family has recently been expanded by the isolation of two new VEGF-related factors, VEGF-B and VEGF-C. The physiological functions of these factors are largely unknown. Here we report the cloning and characterization of mouse VEGF-C, which is produced as a disulfide-linked dimer of 415 amino acid residue polypeptides, sharing an 85% identity with the human VEGF-C amino acid sequence. The recombinant mouse VEGF-C protein was secreted from transfected cells as VEGFR-3 (Flt4) binding polypeptides of 30-32x10(3) Mr and 22-23x10(3) Mr which preferentially stimulated the autophosphorylation of VEGFR-3 in comparison with VEGFR-2 (KDR). In in situ hybridization, mouse VEGF-C mRNA expression was detected in mesenchymal cells of postimplantation mouse embryos, particularly in the regions where the lymphatic vessels undergo sprouting from embryonic veins, such as the perimetanephric, axillary and jugular regions. In addition, the developing mesenterium, which is rich in lymphatic vessels, showed strong VEGF-C expression. VEGF-C was also highly expressed in adult mouse lung, heart and kidney, where VEGFR-3 was also prominent. The pattern of expression of VEGF-C in relation to its major receptor VEGFR-3 during the sprouting of the lymphatic endothelium in embryos suggests a paracrine mode of action and that one of the functions of VEGF-C may be in the regulation of angiogenesis of the lymphatic vasculature. %B Development %V 122 %P 3829 - 37 %8 1996/Dec/ %G eng %U http://view.ncbi.nlm.nih.gov/pubmed/9012504 %N 12 %! Development (Cambridge, England)