@article {694,
	title = {Lymphatics and the eye. [Finnish]},
	journal = {Duodecim L{\"a}{\"a}ketieteellinen Aikakauskirja},
	volume = {136},
	year = {2020},
	month = {2020/02/10/},
	pages = {1777-1788},
	type = {review},
	chapter = {1777},
	issn = {2242-3281},
	doi = {10.5281/zenodo.4005517},
	url = {https://www.duodecimlehti.fi/lehti/2020/16/duo15739},
	author = {Gucciardo, Erika and Lehti, Timo A. and Korhonen, Ani and Salv{\'e}n, Petri and Lehti, Kaisa and Jeltsch, Michael and Loukovaara, Sirpa}
}
@article {536,
	title = {Functional Importance of a Proteoglycan Co-Receptor in Pathologic Lymphangiogenesis},
	journal = {Circulation Research},
	volume = {119},
	year = {2016},
	month = {05/2016},
	pages = {210-221},
	abstract = {Rationale: Lymphatic vessel growth is mediated by major pro-lymphangiogenic factors such as VEGF-C and -D, among other endothelial effectors. Heparan sulfate is a linear polysaccharide expressed on proteoglycan core proteins on cell-membranes and matrix, playing roles in angiogenesis, although little is known regarding any function(s) in lymphatic remodeling in vivo.
Objective: To explore the genetic basis and mechanisms whereby heparan sulfate proteoglycans mediate pathologic lymphatic remodeling.
Methods and Results: Lymphatic endothelial deficiency in the major heparan sulfate biosynthetic enzyme N-deacetylase/N-sulfotransferase-1 (Ndst1; involved in glycan-chain sulfation) was associated with reduced lymphangiogenesis in pathologic models, including spontaneous neoplasia. Mouse mutants demonstrated tumor-associated lymphatic vessels with apoptotic nuclei. Mutant lymphatic endothelia demonstrated impaired mitogen (Erk) and survival (Akt) pathway signaling as well as reduced VEGF-C mediated protection from starvation-induced apoptosis. Lymphatic endothelial specific Ndst1 deficiency (in Ndst1f/fProx1+/CreERT2 mice) was sufficient to inhibit VEGF-C dependent lymphangiogenesis. Lymphatic heparan sulfate deficiency reduced phosphorylation of the major lymphatic growth receptor VEGFR-3 in response to multiple VEGF-C species. Syndecan-4 was the dominantly expressed heparan sulfate proteoglycan in mouse lymphatic endothelia, and pathologic lymphangiogenesis was impaired in Sdc4(-/-) mice. On the lymphatic cell surface, VEGF-C induced robust association between syndecan-4 and VEGFR-3 which was sensitive to glycan disruption. Moreover, VEGFR-3 mitogen and survival signaling was reduced in the setting of Ndst1 or Sdc4 deficiency.
Conclusions: These findings demonstrate the genetic importance of heparan sulfate and the major lymphatic proteoglycan syndecan-4 in pathologic lymphatic remodeling. This may introduce novel future strategies to alter pathologic lymphatic-vascular remodeling.},
	keywords = {endothelial cell growth, glycosaminoglycan, lymphatic capillary, Proteoglycan, vascular endothelial growth factor receptor},
	doi = {10.1161/CIRCRESAHA.116.308504},
	url = {http://circres.ahajournals.org/content/early/2016/05/25/CIRCRESAHA.116.308504},
	author = {Johns, Scott C. and Yin, Xin and Jeltsch, Michael and Bishop, Joseph R. and Schuksz, Manuela and Ghazal, Roland El and Wilcox-Adelman, Sarah A. and Alitalo, Kari and Fuster, Mark M.}
}
@article {442,
	title = {Structural and mechanistic insights into VEGF receptor 3 ligand binding and activation},
	journal = {Proceedings of the National Academy of Sciences of the United States of America},
	volume = {110},
	year = {2013},
	month = {08/2013},
	pages = {12960 - 12965},
	abstract = {Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are key drivers of blood and lymph vessel formation in development, but also in several pathological processes. VEGF-C signaling through VEGFR-3 promotes lymphangiogenesis, which is a clinically relevant target for treating lymphatic insufficiency and for blocking tumor angiogenesis and metastasis. The extracellular domain of VEGFRs consists of seven Ig homology domains; domains 1-3 (D1-3) are responsible for ligand binding, and the membrane-proximal domains 4-7 (D4-7) are involved in structural rearrangements essential for receptor dimerization and activation. Here we analyzed the crystal structures of VEGF-C in complex with VEGFR-3 domains D1-2 and of the VEGFR-3 D4-5 homodimer. The structures revealed a conserved ligand-binding interface in D2 and a unique mechanism for VEGFR dimerization and activation, with homotypic interactions in D5. Mutation of the conserved residues mediating the D5 interaction (Thr446 and Lys516) and the D7 interaction (Arg737) compromised VEGF-C induced VEGFR-3 activation. A thermodynamic analysis of VEGFR-3 deletion mutants showed that D3, D4-5, and D6-7 all contribute to ligand binding. A structural model of the VEGF-C/VEGFR-3 D1-7 complex derived from small-angle X-ray scattering data is consistent with the homotypic interactions in D5 and D7. Taken together, our data show that ligand-dependent homotypic interactions in D5 and D7 are essential for VEGFR activation, opening promising possibilities for the design of VEGFR-specific drugs.},
	keywords = {Amino Acid Sequence, Binding Sites, Binding, Competitive, Crystallography, X-Ray, Electrophoresis, Polyacrylamide Gel, Humans, Ligands, Microscopy, Electron, Models, Molecular, Molecular Sequence Data, Multiprotein Complexes, Mutation, Protein Binding, Protein Multimerization, Protein Structure, Tertiary, Scattering, Small Angle, Sequence Homology, Amino Acid, Thermodynamics, Vascular Endothelial Growth Factor C, Vascular Endothelial Growth Factor Receptor-3, X-Ray Diffraction},
	isbn = {1091-6490},
	doi = {10.1073/pnas.1301415110},
	url = {http://www.pnas.org/content/110/32/12960.long},
	author = {Lepp{\"a}nen, Veli-Matti and Tvorogov, Denis and Kisko, Kaisa and Prota, Andrea E and Jeltsch, Michael and Anisimov, Andrey and Markovic-Mueller, Sandra and Stuttfeld, Edward and Goldie, Kenneth N and Ballmer-Hofer, Kurt and Alitalo, Kari}
}
@article {425,
	title = {A truncation allele in vascular endothelial growth factor c reveals distinct modes of signaling during lymphatic and vascular development.},
	journal = {Development},
	volume = {140},
	year = {2013},
	month = {2013 Apr},
	pages = {1497-506},
	abstract = {<p>Vascular endothelial growth factor C (Vegfc) is a secreted protein that guides lymphatic development in vertebrate embryos. However, its role during developmental angiogenesis is not well characterized. Here, we identify a mutation in zebrafish vegfc that severely affects lymphatic development and leads to angiogenesis defects on sensitized genetic backgrounds. The um18 mutation prematurely truncated Vegfc, blocking its secretion and paracrine activity but not its ability to activate its receptor Flt4. When expressed in endothelial cells, vegfc(um18) could not rescue lymphatic defects in mutant embryos, but induced ectopic blood vessel branching. Furthermore, vegfc-deficient endothelial cells did not efficiently contribute to tip cell positions in developing sprouts. Computational modeling together with assessment of endothelial cell dynamics by time-lapse analysis suggested that an autocrine Vegfc/Flt4 loop plays an important role in migratory persistence and filopodia stability during sprouting. Our results suggest that Vegfc acts in two distinct modes during development: as a paracrine factor secreted from arteries to guide closely associated lymphatic vasculature and as an autocrine factor to drive migratory persistence during angiogenesis.</p>},
	keywords = {Alleles, Animals, Animals, Genetically Modified, Autocrine Communication, Blood Vessels, Cell Movement, Codon, Nonsense, Embryo, Nonmammalian, Female, Lymphatic System, Mice, Mice, Knockout, Neovascularization, Physiologic, Paracrine Communication, Protein Isoforms, Signal Transduction, Vascular Endothelial Growth Factor C, Zebrafish, Zebrafish Proteins},
	issn = {1477-9129},
	doi = {10.1242/dev.084152},
	author = {Villefranc, Jacques A and Nicoli, Stefania and Bentley, Katie and Jeltsch, Michael and Zarkada, Georgia and Moore, John C and Gerhardt, Holger and Alitalo, Kari and Lawson, Nathan D}
}
@article {424,
	title = {Vascular endothelial growth factor-angiopoietin chimera with improved properties for therapeutic angiogenesis},
	journal = {Circulation},
	volume = {127},
	year = {2013},
	month = {01/2013},
	pages = {424-434},
	chapter = {424},
	abstract = {BACKGROUND:

There is an unmet need for proangiogenic therapeutic molecules for the treatment of tissue ischemia in cardiovascular diseases. However, major inducers of angiogenesis such as vascular endothelial growth factor (VEGF/VEGF-A) have side effects that limit their therapeutic utility in vivo, especially at high concentrations. Angiopoietin-1 has been considered to be a blood vessel stabilization factor that can inhibit the intrinsic property of VEGF to promote vessel leakiness. In this study, we have designed and tested the angiogenic properties of chimeric molecules consisting of receptor-binding parts of VEGF and angiopoietin-1. We aimed at combining the activities of both factors into 1 molecule for easy delivery and expression in target tissues.
METHODS AND RESULTS:

The VEGF-angiopoietin-1 (VA1) chimeric protein bound to both VEGF receptor-2 and Tie2 and induced the activation of both receptors. Detailed analysis of VA1 versus VEGF revealed differences in the kinetics of VEGF receptor-2 activation and endocytosis, downstream kinase activation, and VE-cadherin internalization. The delivery of a VA1 transgene into mouse skeletal muscle led to increased blood flow and enhanced angiogenesis. VA1 was also very efficient in rescuing ischemic limb perfusion. However, VA1 induced less plasma protein leakage and myeloid inflammatory cell recruitment than VEGF. Furthermore, angioma-like structures associated with VEGF expression were not observed with VA1.
CONCLUSIONS:

The VEGF-angiopoietin-1 chimera is a potent angiogenic factor that triggers a novel mode of VEGF receptor-2 activation, promoting less vessel leakiness, less tissue inflammation, and better perfusion in ischemic muscle than VEGF. These properties of VA1 make it an attractive therapeutic tool.},
	author = {Andrey Anisimov and Denis Tvorogov and Annamari Alitalo and Veli-Matti Lepp{\"a}nen and Y An and EC Han and F Orsenigo and EI Ga{\'a}l and Tanja Holopainen and YJ Koh and Tuomas Tammela and P Korpisalo and Salla Keskitalo and Michael Jeltsch and Seppo Yl{\"a}-Herttuala and Elisabetta Dejana and GY Koh and C Choi and Pipsa Saharinen and Kari Alitalo}
}
@article {40,
	title = {Structural determinants of growth factor binding and specificity by VEGF receptor 2},
	journal = {Proceedings of the National Academy of Sciences of the United States of America},
	volume = {107},
	year = {2010},
	month = {02/2010},
	pages = {2425 - 30},
	abstract = {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.},
	url = {http://view.ncbi.nlm.nih.gov/pubmed/20145116},
	author = {Lepp{\"a}nen, Veli-Matti and Prota, Andrea E and Jeltsch, Michael and Anisimov, Andrey and Kalkkinen, Nisse and Strandin, Tomas and Lankinen, Hilkka and Goldman, Adrian and Ballmer-Hofer, Kurt and Alitalo, Kari}
}
@article {26,
	title = {VEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia},
	journal = {J Cell Biol},
	volume = {161},
	year = {2003},
	month = {2003/Jun/},
	pages = {1163 - 77},
	abstract = {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.},
	url = {http://view.ncbi.nlm.nih.gov/pubmed/12810700},
	author = {Gerhardt, Holger and Golding, Matthew and Fruttiger, Marcus and Ruhrberg, Christiana and Lundkvist, Andrea and Abramsson, Alexandra and Jeltsch, Michael and Mitchell, Christopher and Alitalo, Kari and Shima, David and Betsholtz, Christer}
}
@article {17,
	title = {Vascular endothelial growth factor B (VEGF-B) binds to VEGF receptor-1 and regulates plasminogen activator activity in endothelial cells},
	journal = {Proc Natl Acad Sci U S A},
	volume = {95},
	year = {1998},
	month = {1998/Sep/},
	pages = {11709 - 14},
	abstract = {The vascular endothelial growth factor (VEGF) family has recently expanded by the identification and cloning of three additional members, namely VEGF-B, VEGF-C, and VEGF-D. In this study we demonstrate that VEGF-B binds selectively to VEGF receptor-1/Flt-1. This binding can be blocked by excess VEGF, indicating that the interaction sites on the receptor are at least partially overlapping. Mutating the putative VEGF receptor-1/Flt-1 binding determinants Asp63, Asp64, and Glu67 to alanine residues in VEGF-B reduced the affinity to VEGF receptor-1 but did not abolish binding. Mutational analysis of conserved cysteines contributing to VEGF-B dimer formation suggest a structural conservation with VEGF and platelet-derived growth factor. Proteolytic processing of the 60-kDa VEGF-B186 dimer results in a 34-kDa dimer containing the receptor-binding epitopes. The binding of VEGF-B to its receptor on endothelial cells leads to increased expression and activity of urokinase type plasminogen activator and plasminogen activator inhibitor 1, suggesting a role for VEGF-B in the regulation of extracellular matrix degradation, cell adhesion, and migration.},
	url = {http://view.ncbi.nlm.nih.gov/pubmed/9751730},
	author = {Olofsson, B and Korpelainen, E and Pepper, M S and Mandriota, S J and Aase, K and Kumar, V and Gunji, Y and Jeltsch, M M and Shibuya, M and Alitalo, K and Eriksson, U}
}
