00823nas a2200253 4500008004100000022001400041245012000055210007100175260001600246300001000262490000700272100002100279700001900300700001800319700001800337700002800355700001700383700002100400700002100421700002200442700002100464700002200485856006200507 2020 eng d a2324-926900aInvestigation on the role of biallelic variants in VEGF‐C found in a patient affected by Milroy‐like lymphedema0 aInvestigation on the role of biallelic variants in VEGF‐C found cFeb-06-2022 ae13890 v001 aMukenge, Sylvain1 aJha, Sawan, K.1 aCatena, Marco1 aManara, Elena1 aLeppänen, Veli‐Matti1 aLenti, Elisa1 aNegrini, Daniela1 aBertelli, Matteo1 aBrendolan, Andrea1 aJeltsch, Michael1 aAldrighetti, Luca uhttps://onlinelibrary.wiley.com/doi/abs/10.1002/mgg3.138900566nas a2200205 4500008004100000022001400041245003800055210003500093260001600128300001400144490000800158100002100166700002000187700001800207700001900225700001700244700002100261700002200282856005600304 2020 eng d a2242-328100aLymphatics and the eye. [Finnish]0 aLymphatics and the eye Finnish c2020/02/10/ a1777-17880 v1361 aGucciardo, Erika1 aLehti, Timo, A.1 aKorhonen, Ani1 aSalvén, Petri1 aLehti, Kaisa1 aJeltsch, Michael1 aLoukovaara, Sirpa uhttps://www.duodecimlehti.fi/lehti/2020/16/duo1573900911nas a2200217 4500008004100000245006800041210006700109260001600176300003100192520016400223100001900387700001500406700001700421700002600438700002100464700002300485700002100508700001900529700001800548856012700566 2020 eng d00aVEGF-C Protects the Integrity of Bone Marrow Perivascular Niche0 aVEGFC Protects the Integrity of Bone Marrow Perivascular Niche c2020/07/31/ aaccepted - for publication3 aKey Points. Vegfc deletion in endothelial or LepR+ cells compromises the bone marrow perivascular niche and hematopoietic stem cell maintenance.Exogenous admin1 aFang, Shentong1 aChen, Shuo1 aNurmi, Harri1 aLeppänen, Veli-Matti1 aJeltsch, Michael1 aScadden, David, T.1 aSilberstein, Lev1 aMikkola, Hanna1 aAlitalo, Kari uhttps://ashpublications.org/blood/article/doi/10.1182/blood.2020005699/463465/VEGF-C-Protects-the-Integrity-of-Bone-Marrow01971nas a2200337 4500008004100000020001400041245005600055210005300111260001600164300001100180490000600191520106000197653001901257653001601276653003401292653001301326653002201339653001001361653001101371653001101382100002201393700002201415700002001437700002001457700002701477700002101504700002401525700001801549700002101567856004501588 2019 eng d a2050-084X00aKLK3/PSA and cathepsin D activate VEGF-C and VEGF-D0 aKLK3PSA and cathepsin D activate VEGFC and VEGFD c2019/05/17/ ae444780 v83 aVascular endothelial growth factor-C (VEGF-C) acts primarily on endothelial cells, but also on non-vascular targets, e.g. in the CNS and immune system. Here we describe a novel, unique VEGF-C form in the human reproductive system produced via cleavage by kallikrein-related peptidase 3 (KLK3), aka prostate-specific antigen (PSA). KLK3 activated VEGF-C specifically and efficiently through cleavage at a novel N-terminal site. We detected VEGF-C in seminal plasma, and sperm liquefaction occurred concurrently with VEGF-C activation, which was enhanced by collagen and calcium binding EGF domains 1 (CCBE1). After plasmin and ADAMTS3, KLK3 is the third protease shown to activate VEGF-C. Since differently activated VEGF-Cs are characterized by successively shorter N-terminal helices, we created an even shorter hypothetical form, which showed preferential binding to VEGFR-3. Using mass spectrometric analysis of the isolated VEGF-C-cleaving activity from human saliva, we identified cathepsin D as a protease that can activate VEGF-C as well as VEGF-D.10acancer biology10aCathepsin D10akallikrein-related peptidases10aKLK3/PSA10aLymphangiogenesis10amouse10aVEGF-C10aVEGF-D1 aJha, Sawan, Kumar1 aRauniyar, Khushbu1 aChronowska, Ewa1 aMattonet, Kenny1 aMaina, Eunice, Wairimu1 aKoistinen, Hannu1 aStenman, Ulf-Håkan1 aAlitalo, Kari1 aJeltsch, Michael uhttps://elifesciences.org/articles/4447802718nas a2200205 4500008004100000245007100041210006600112260001600178300001200194490000700206520212700213653002202340653001602362653001602378653001102394100002002405700002302425700002102448856004302469 2019 eng d00aThe Proteolytic Activation of Vascular Endothelial Growth Factor-C0 aProteolytic Activation of Vascular Endothelial Growth FactorC c2019/12/18/ a88 - 980 v233 aThe 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.10aLymphangiogenesis10aproteinases10aproteolysis10aVEGF-C1 aLackner, Marcel1 aSchmotz, Constanze1 aJeltsch, Michael uhttps://doi.org/10.5281/zenodo.362926302383nas a2200277 4500008004100000022001400041245009400055210006900149260001200218490000600230520155100236653001201787653001001799653002801809653001901837653002201856653001501878653002701893653002301920653001901943653001101962100002201973700001801995700002102013856007102034 2018 eng d a2296-418500aBiology of Vascular Endothelial Growth Factor C in the Morphogenesis of Lymphatic Vessels0 aBiology of Vascular Endothelial Growth Factor C in the Morphogen c02/20180 v63 aBecause virtually all tissues contain blood vessels, the importance of hemevascularization has been long recognized in regenerative medicine and tissue engineering. However, the lymphatic vasculature has only recently become a subject of interest. Central to the task of growing a lymphatic network are lymphatic endothelial cells (LECs), which constitute the innermost layer of all lymphatic vessels. The central molecule that directs proliferation and migration of LECs during embryogenesis is Vascular Endothelial Growth Factor-C (VEGF-C). VEGF-C is, therefore, an important ingredient for LEC culture and attempts to (re)generate lymphatic vessels and networks. During its biosynthesis, VEGF-C undergoes a stepwise proteolytic processing, during which its properties and affinities for its interaction partners change. Many of these fundamental aspects of VEGF-C biosynthesis have only recently been uncovered. So far, most - if not all - applications of VEGF-C do not discriminate between different forms of VEGF-C. However, for lymphatic regeneration and engineering purposes, it appears mandatory to understand these differences, since they relate e.g. to such important aspects as biodistribution and receptor activation potential. In this review, we discuss the molecular biology of VEGF-C as it relates to the growth of LECs and lymphatic vessels. However, the properties of VEGF-C are similarly relevant for the cardiovascular system, since both old and recent data show that VEGF-C can have a profound effect on the blood vasculature.10aADAMTS310aCCBE110agrowth factor signaling10agrowth factors10aLymphatic Vessels10alymphedema10aproteolytic processing10aTissue Engineering10aVEGF receptors10aVEGF-C1 aRauniyar, Khushbu1 aJha, Sawan, K1 aJeltsch, Michael uhttps://www.frontiersin.org/articles/10.3389/fbioe.2018.00007/full02194nas a2200169 4500008004100000020001300041245007300054210006900127260001600196300001200212490000800224520168300232100001201915700001601927700001501943856006601958 2018 eng d a0940960200aKey molecules in lymphatic development, function, and identification0 aKey molecules in lymphatic development function and identificati c2018/09/01/ a25 - 340 v2193 aWhile both blood and lymphatic vessels transport fluids and thus share many similarities, they also show functional and structural differences, which can be used to differentiate them. Specific visualization of lymphatic vessels has historically been and still is a pivot point in lymphatic research. Many of the proteins that are investigated by molecular biologists in lymphatic research have been defined as marker molecules, i.e. to visualize and distinguish lymphatic endothelial cells (LECs) from other cell types, most notably from blood vascular endothelial cells (BECs) and cells of the hematopoietic lineage.Among the factors that drive the developmental differentiation of lymphatic structures from venous endothelium, Prospero homeobox protein 1 (PROX1) is the master transcriptional regulator. PROX1 maintains lymphatic identity also in the adult organism and thus is a universal LEC marker. Vascular endothelial growth factor receptor-3 (VEGFR-3) is the major tyrosine kinase receptor that drives LEC proliferation and migration. The major activator for VEGFR-3 is vascular endothelial growth factor-C (VEGF-C). However, before VEGF-C can signal, it needs to be proteolytically activated by an extracellular protein complex comprised of Collagen and calcium binding EGF domains 1 (CCBE1) protein and the protease A disintegrin and metallopeptidase with thrombospondin type 1 motif 3 (ADAMTS3).This minireview attempts to give an overview of these and a few other central proteins that scientific inquiry has linked specifically to the lymphatic vasculature. It is limited in scope to a brief description of their main functions, properties and developmental roles.1 aJha, SK1 aRauniyar, K1 aJeltsch, M uhttp://linkinghub.elsevier.com/retrieve/pii/S094096021830071200711nas a2200205 4500008004100000245014500041210006900186260001600255300000900271490000600280100001200286700002200298700002200320700002600342700002300368700002000391700001800411700002100429856005500450 2017 eng d00aEfficient activation of the lymphangiogenic growth factor VEGF-C requires the C-terminal domain of VEGF-C and the N-terminal domain of CCBE10 aEfficient activation of the lymphangiogenic growth factor VEGFC c2017/07/07/ a49160 v71 aJha, SK1 aRauniyar, Khushbu1 aKärpänen, Terhi1 aLeppänen, Veli-Matti1 aBrouillard, Pascal1 aVikkula, Miikka1 aAlitalo, Kari1 aJeltsch, Michael uhttps://www.nature.com/articles/s41598-017-04982-100395nas a2200097 4500008004100000245007600041210006900117260004400186100002100230856004600251 2017 eng d00aWhat you should know about VEGF-C when working with lymphatics [German]0 aWhat you should know about VEGFC when working with lymphatics Ge aBad Soden (Frankfurt), Germanyc06/20171 aJeltsch, Michael uhttps://jeltsch.org/Abstrakt-BadSoden201700739nas a2200241 4500008004100000245009400041210006900135260001600204300000900220490000900229653002100238653001200259653001900271653002200290653001200312653002000324100003200344700002000376700002200396700002100418700002300439856003500462 2016 eng d00aFactors regulating the substrate specificity of cytosolic phospholipase A2-alpha in vitro0 aFactors regulating the substrate specificity of cytosolic phosph c2016/07/01/ a15970 v186110aArachidonic acid10aBilayer10aCatalytic site10aMass spectrometry10aMicelle10aPhospholipase A1 aBatchu, Krishna, Chaithanya1 aHänninen, Satu1 aJha, Sawan, Kumar1 aJeltsch, Michael1 aSomerharju, Pentti uhttps://jeltsch.org/Batchu201602911nas a2200109 4500008004100000245008700041210006900128260003000197520249000227100002102717856006302738 2016 eng d00aFrom Molecular Genetics and Biology to Effective Treatments of Lymphatic Disorders0 aFrom Molecular Genetics and Biology to Effective Treatments of L aMulhouse, Francec05/20163 aIn 1971, Judah Folkman proposed the concept of anti-angiogenic tumor therapy. 12 years later, Harold Dvorak isolated the responsible growth factor VEGF. Nine years later, Napoleone Ferrara reported the generation of neutralizing monoclonal antibodies against VEGF. Another five years later, Phase I trials started with the humanized version of one of the monoclonals: bevacizumab. Since 2004, when it received FDA approval, it has been marketed under the brand name Avastin. The translation of basic biomedical research into tangible benefits for patients appears sometimes agonizingly slow. The public has been promised much by hyped scientific breakthroughs [4]. Scientific journals and scientists have played along in over-hyping scientific breakthroughs in the hope of impact factors and citations in order to secure and justify funding and fame. Not surprisingly, practitioners ask when the discoveries from basic research will finally improve the standard of care for their patients. Lymphatic research is no exception. Practitioners are largely still limited to symptomatic treatment and there seems to be still an invisible, but perceptible divide between those who do the molecular biology research and those who treat patients. The Avastin story is a plea for basic research: it might be complicated and it might take time, but it eventually does pay off. How is the lymphatic research community doing concerning the translation of research results into treatment options? Examples of lymphatic research in or shortly before the clinical trial stage include: - Growth factor enhanced lymph node transplantation to treat secondary lymphedema - Utilizing the Schlemm channel's lymphatic character in glaucoma treatment - Anti-angiogenic tumor treatment with anti-lymphangiogenic agents Treatment of primary lymphedema with VEGF-C has been proposed. However, our understanding of the physiological process of lymph vessel development is far from complete, despite significant recent progress in our understanding of developmental lymphangiogenesis and first attempts at tissue-engineering lymphatic vessels. If the results from high throughput cancer profilings are predictive of lymphatic conditions, then many patients will feature very individual, multifactorial disease profiles. Even more challenging than the identification of such causes will be the development of treatment regimens that rapidly can be tailored to such individual needs.1 aJeltsch, Michael uhttp://www.eurolymphology.org/JOURNAL/VOL28-N74-2016/#p=1402952nas a2200289 4500008004100000245008800041210006900129260001200198300001200210490000800222520202000230653002802250653002202278653002402300653001702324653004802341100002102389700001302410700002102423700002302444700002102467700002302488700003002511700001802541700002102559856008202580 2016 eng d00aFunctional Importance of a Proteoglycan Co-Receptor in Pathologic Lymphangiogenesis0 aFunctional Importance of a Proteoglycan CoReceptor in Pathologic c05/2016 a210-2210 v1193 aRationale: 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.10aendothelial cell growth10aglycosaminoglycan10alymphatic capillary10aProteoglycan10avascular endothelial growth factor receptor1 aJohns, Scott, C.1 aYin, Xin1 aJeltsch, Michael1 aBishop, Joseph, R.1 aSchuksz, Manuela1 aGhazal, Roland, El1 aWilcox-Adelman, Sarah, A.1 aAlitalo, Kari1 aFuster, Mark, M. uhttp://circres.ahajournals.org/content/early/2016/05/25/CIRCRESAHA.116.30850400962nas a2200181 4500008004100000245007000041210006900111260000900180300000900189490000700198520040300205100002400608700002100632700002300653700001600676700002500692856006300717 2016 eng d00aLymphatic Vessels in Regenerative Medicine and Tissue Engineering0 aLymphatic Vessels in Regenerative Medicine and Tissue Engineerin c2016 a1-130 v223 aOnce a DOI is available for this article, the final publication will be available from Mary Ann Liebert, Inc., publishers at http://dx.doi.org/10.1089/TEN.TEB.2016.0034. The postprint manuscript is available from here and for the next 30 days also from the publisher via this bit.ly shortcut: http://bit.ly/1VKjjMk. 1 aSchaupper, Mira, V.1 aJeltsch, Michael1 aRohringer, Sabrina1 aRedl, Heinz1 aHolnthoner, Wolfgang uhttp://online.liebertpub.com/doi/10.1089/ten.TEB.2016.003403585nas a2200589 4500008004100000022001400041245010200055210006900157260001500226300001400241490000800255520182400263653001202087653001802099653002902117653001302146653003102159653002202190653002802212653004602240653002902286653002702315653001302342653002002355653001702375653001102392653003302403653002202436653001502458653000902473653002102482653001302503653001402516653002002530653004302550653002402593653001702617653003002634653004102664653001402705653002302719100001702742700002602759700002002785700002102805700002602826700001802852700001802870700002702888700002002915856006002935 2015 eng d a1524-457100aFunctional Dissection of the CCBE1 Protein: A Crucial Requirement for the Collagen Repeat Domain.0 aFunctional Dissection of the CCBE1 Protein A Crucial Requirement c2015 May 8 a1660-16690 v1163 a
RATIONALE: Collagen- and calcium-binding EGF domain-containing protein 1 (CCBE1) is essential for lymphangiogenesis in vertebrates and has been associated with Hennekam syndrome. Recently, CCBE1 has emerged as a crucial regulator of vascular endothelial growth factor-C (VEGFC) signaling.
OBJECTIVE: CCBE1 is a secreted protein characterized by 2 EGF domains and 2 collagen repeats. The functional role of the different CCBE1 protein domains is completely unknown. Here, we analyzed the functional role of the different CCBE1 domains in vivo and in vitro.
METHODS AND RESULTS: We analyzed the functionality of several CCBE1 deletion mutants by generating knock-in mice expressing these mutants, by analyzing their ability to enhance Vegfc signaling in vivo in zebrafish, and by testing their ability to induce VEGFC processing in vitro. We found that deleting the collagen domains of CCBE1 has a much stronger effect on CCBE1 activity than deleting the EGF domains. First, although CCBE1ΔCollagen mice fully phenocopy CCBE1 knock-out mice, CCBE1ΔEGF knock-in embryos still form rudimentary lymphatics. Second, Ccbe1ΔEGF, but not Ccbe1ΔCollagen, could partially substitute for Ccbe1 to enhance Vegfc signaling in zebrafish. Third, CCBE1ΔEGF, similarly to CCBE1, but not CCBE1ΔCollagen could activate VEGFC processing in vitro. Furthermore, a Hennekam syndrome mutation within the collagen domain has a stronger effect than a Hennekam syndrome mutation within the EGF domain.
CONCLUSIONS: We propose that the collagen domains of CCBE1 are crucial for the activation of VEGFC in vitro and in vivo. The EGF domains of CCBE1 are dispensable for regulation of VEGFC processing in vitro, however, they are necessary for full lymphangiogenic activity of CCBE1 in vivo.
10aAnimals10aBinding Sites10aCalcium-Binding Proteins10aCollagen10aCraniofacial Abnormalities10aEndothelial Cells10aEpidermal Growth Factor10aGene Expression Regulation, Developmental10aGene Knock-In Techniques10aGenital Diseases, Male10aGenotype10aGestational Age10aHEK293 Cells10aHumans10aLymphangiectasis, Intestinal10aLymphatic Vessels10alymphedema10aMice10aMice, Transgenic10aMutation10aPhenotype10aProtein Binding10aProtein Interaction Domains and Motifs10aSignal Transduction10aTransfection10aTumor Suppressor Proteins10aVascular Endothelial Growth Factor C10aZebrafish10aZebrafish Proteins1 aRoukens, Guy1 aPeterson-Maduro, Josi1 aPadberg, Yvonne1 aJeltsch, Michael1 aLeppänen, Veli-Matti1 aBos, Frank, L1 aAlitalo, Kari1 aSchulte-Merker, Stefan1 aSchulte, Dörte uhttp://circres.ahajournals.org/content/116/10/1660.long02227nas a2200193 4500008004100000020002200041245004600063210004100109250000600150260002900156300001200185520168900197100002001886700001901906700002101925700002201946700002701968856003801995 2015 eng d a978-3-934371-53-800aThe genetic causes of primary lymphedema.0 agenetic causes of primary lymphedema a6 aColognebViavital Verlag a210-2293 aEnglish: Primary lymphedema can be treated, but not cured. In addition, their diagnosis is due to heterogeneous phenotypes often ambiguous. However, these problems can be tackled by identifying the edema-causing genetic lesions to yield unambiguous diagnoses and by developing specific treatments that address the underlying, molecular cause. New developments in molecular biology are providing the necessary tools for these tasks and in the recent years the genetic causes of many forms of primary lymphedema have been identified, notably by exome sequencing. For a significant proportion of lymphatic disorders multifactorial genetic causes are suspected. This chapter provides an overview of the current knowledge on the genetic origin, the categorization as well as the molecular and biochemical causes of primary lymphedema. German: Primäre Lymphödeme sind behandelbar, aber nicht heilbar. Zudem ist die Diagnostik aufgrund heterogener Phänotypen oft nicht eindeutig. Um diese Probleme anzugehen, müssen die das Ödem verursachenden genetischen Ursachen gefunden, diagnostiziert und gezielt behandelt werden. Die hierzu notwendigen Techniken liefern die neuen Entwicklungen in der Molekularbiologie. Insbesondere durch die Technik der Exom-Sequenzierung wurden in den letzten Jahren die genetischen Ursachen vieler primärer Lymphödeme identifiziert. Für einen weiteren großen Anteil dieser Erkrankungen werden multifaktorielle genetische Dispositionen vermutet. Dieses Kapitel gibt einen Überblick über den derzeitigen Kenntnisstand der genetischen Ursachen, der Kategorisierung sowie der molekularbiologischen und biochemischen Grundlagen primärer Lymphödeme.1 aMattonet, Kenny1 aWilting, Jörg1 aJeltsch, Michael1 aWeissleder, Horst1 aSchuchhardt, Christian uhttps://jeltsch.org/Mattonet2015a01275nas a2200157 4500008004100000022001400041245006700055210006300122260001200185300001000197490000700207520079300214100002001007700002101027856006901048 2015 eng d a1433-525500aHeterogeneity of the origin of the lymphatic system. [German].0 aHeterogeneity of the origin of the lymphatic system German c12/2015 a84-880 v193 aThe question “How does the lymphatic system develop?” may be a simple one, but it is fundamental to our understanding of lymphatic malformations in children and the regeneration of lymphatics in adults. The question is by no means new and was already explored in the early 20 century. This resulted in a long-lasting controversy, which until recently had been far from being settled. The interest in the lymphatic system has greatly increased in recent years due to its implications in a variety of diseases. Several studies published this year address the heterogeneity of lymphatic endothelial cell development and unite previous controversially discussed data in a coherent model. These remarkable results, as well as the studies that paved their way, are discussed in this review.1 aMattonet, Kenny1 aJeltsch, Michael uhttp://www.dglymph.de/fileadmin/global/pdfs/LymphForsch_2-15.pdf02348nas a2200277 4500008004100000245010600041210006900147260001200216300001400228490000700242520151100249100001901760700001901779700002201798700001601820700002401836700001701860700001401877700001701891700001601908700002101924700001601945700002101961700002201982856006602004 2015 eng d00aIschemia-Reperfusion Injury Enhances Lymphatic Endothelial VEGFR3 and Rejection in Cardiac Allografts0 aIschemiaReperfusion Injury Enhances Lymphatic Endothelial VEGFR3 c12/2015 a1160-11720 v163 aOrgan damage and innate immunity during heart transplantation may evoke adaptive immunity with serious consequences. Because lymphatic vessels bridge innate and adaptive immunity, they are critical in immune surveillance; however, their role in ischemia–reperfusion injury (IRI) in allotransplantation remains unknown. We investigated whether the lymphangiogenic VEGF-C/VEGFR3 pathway during cardiac allograft IRI regulates organ damage and subsequent interplay between innate and adaptive immunity. We found that cardiac allograft IRI, within hours, increased graft VEGF-C expression and lymphatic vessel activation in the form of increased lymphatic VEGFR3 and adhesion protein expression. Pharmacological VEGF-C/VEGFR3 stimulation resulted in early lymphatic activation and later increase in allograft inflammation. In contrast, pharmacological VEGF-C/VEGFR3 inhibition during cardiac allograft IRI decreased early lymphatic vessel activation with subsequent dampening of acute and chronic rejection. Genetic deletion of VEGFR3 specifically in the lymphatics of the transplanted heart recapitulated the survival effect achieved by pharmacological VEGF-C/VEGFR3 inhibition. Our results suggest that tissue damage rapidly changes lymphatic vessel phenotype, which, in turn, may shape the interplay of innate and adaptive immunity. Importantly, VEGF-C/VEGFR3 inhibition during solid organ transplant IRI could be used as lymphatic-targeted immunomodulatory therapy to prevent acute and chronic rejection.1 aDashkevich, A.1 aRaissadati, A.1 aSyrjälä, S., O.1 aZarkada, G.1 aKeränen, M., A. I.1 aTuuminen, R.1 aKrebs, R.1 aAnisimov, A.1 aJeltsch, M.1 aLeppänen, V.-M.1 aAlitalo, K.1 aNykänen, A., I.1 aLemström, K., B. uhttp://onlinelibrary.wiley.com/doi/10.1111/ajt.13564/abstract03405nas a2200109 4500008004100000245004400041210004400085260006500129520301700194100002103211856006303232 2015 eng d00aLymphangiogenesis in Health and Disease0 aLymphangiogenesis in Health and Disease aLausanne, SwitzerlandbEuropean Group of Lymphologyc06/20153 aDespite the intensive research on the lymphangiogenic VEGF-C/VEGFR-3 signaling pathway in the last two decades, new and unexpected findings do not cease to be made. Diseases that involve the lymphatic system have helped to uncover mechanisms of its normal functioning and development. A recent example of new basic knowledge that resulted from the investigation of a human disease is Hennekam lymphangiectasia lymphedema syndrome (OMIM 235510). It is an autosomal recessive condition, which can co-segregate with mutations in the collagen- and calcium-binding EGF domains 1 (CCBEJ) or the protocadherin Fat 4 (FAT4) gene. Both CCBEI and the lymphangiogenic vascular endothelial growth factor C (VEGF-C) are necessary for the early lymphatic development, namely for the budding and migration of endothelial cells from the cardinal vein (CV) and for the formation of the early lymphatic structures. These processes fail in embryos deficient of either Ccbel or Vegfc. In Vegfc-deficient embryos pro-spective lymphatic endothelial cells fail to sprout from the CV, whereas in Ccbel-deficient embryos, the sprouting is abnormal and does not result in the formation of discrete lymphatic structures. The similar phenotypes of Ccbe- and Vegfc-deficient embryos result from the interaction of CCBEI with the VEGF-C growth factor signaling pathway, which is critical in embryonic and adult lymphangiogenesis. VEGF-C is synthesized as an inactive proprotein and needs to be processed by at least two distinct proteases to become fully active. The presence of CCBEI promotes VEGF-C by two independent mechanisms. The C-terminal domain of CCBEI boosts VEGF-C function via increased ADAMTS3-mediated proteolytic activation of VEGF-C, while the N-terminal domain of CCBEI concentrates pro-VEGF-C on endothelial cell-surfaces, where it can be activated in situ by cell-surface associated proteases. Both mechanisms lead to increased VEGFR-3 signaling and increased lymphangiogenesis. These results show that CCBEI is integral to lymphangiogenesis by increasing the levels of active VEGF-C at the endothelial cell surface. Because some forms of lymphedema appear to be treatable by increasing the amount of VEGFR-3 signaling, the first clinical trials designed around a pro-lymphangiogenic concept use VEGF-C. The goal in these trials is to enhance the integration of lymph nodes into the lymphatic vasculature after autologous transfer to treat postmastectomy lymphedema. On the other hand, VEGF-C induced lymphangiogenesis enhances tumor metastasis and VEGF-C-induced tumor angiogenesis in several mouse models. Blocking VEGF-C might be for these reasons an attractive adjuvant treatment to supplement current cancer treatment regimens that include anti angiogenic drugs. The right balance between pro- and anti-Iymphangiogenic stimuli might therefore differ between individuals and CCBEI is an attractive drug target to adjust pro- and anti-Iymphangiogenic stimuli by tuning the rate of VEGF-C activation. 1 aJeltsch, Michael uhttp://www.eurolymphology.org/JOURNAL/VOL26-N72-2015/#p=1002234nas a2200157 4500008004100000245010800041210007000149260001500219520165500234100003201889700001801921700002101939700002001960700002301980856007302003 2015 eng d00aSubstrate efflux propensity is the key determinant of iPLA-β-mediated glycerophospholipid hydrolysised0 aSubstrate efflux propensity is the key determinant of iPLAβmedia c2015/02/233 aA-type phospholipases (PLAs) are key players in glycerophospholipid (GPL)homeostasis and in mammalian cells, Ca2+-independent PLA-beta (iPLAβ) in particular has been implicated in this essential process.However, the regulation of this enzyme,which is necessary to avoid futile competition between synthesis and degradation, is not understood. Recently, we provided evidence that the efflux of the substrate molecules from the bilayer is the rate-limiting step in the hydrolysis of GPLs by some secretory nonhomeostatic) PLAs. To study if this is the case with iPLAβ as well a mass-spectrometric assay was employed to determine the rate of hydrolysis of multiple saturated and unsaturated GPL species in parallel using micelles or vesicle bilayers as the macrosubstrate. With micelles, the hydrolysis decreased with increasing acyl chain length independent of unsaturation and modest discrimination between acyl positional isomers was observed, presumably due to the differences in the structure of the sn1 and sn2 acyl binding sites of the protein. In striking contrast, no significant discrimination between positional isomers was observed with bilayers, and the rate of hydrolysis decreased with the acyl chain length logarithmically and far more than with micelles. These data provide compelling evidence that efflux of the substrate molecule from the bilayer, which also decreases monotonously with acyl chain length, is the rate-determining step in iPLAβ- mediated hydrolysis of GPLs in membranes. This finding is intriguing as it may help to understand how homeostatic PLAs are regulated and how degradation and biosynthesis are coordinated.1 aBatchu, Krishna, Chaithanya1 aHokynar, Kati1 aJeltsch, Michael1 aMattonet, Kenny1 aSomerharju, Pentti uhttp://www.jbc.org/content/early/2015/02/23/jbc.M115.642835.abstract00835nas a2200301 4500008004100000022002200041245002800063210002400091260003800115300001200153653000800165653001700173653001000190653002100200653002100221653002300242653000900265653000900274653002500283100002100308700002100329700002100350700002600371700001800397700002300415700001800438856007700456 2015 eng d a978-3-319-11888-800aThe TIE Receptor Family0 aTIE Receptor Family bSpringer International Publishing a743-77510aANG10aAngiopoietin10aANGPT10aEndothelial cell10aLymphatic vessel10aNeovascularization10aTIE110aTIE210aVascular dysfunction1 aSaharinen, Pipsa1 aJeltsch, Michael1 aSantoyo, MayteM.1 aLeppänen, Veli-Matti1 aAlitalo, Kari1 aWheeler, Deric, L.1 aYarden, Yosef uhttps://link.springer.com/content/pdf/10.1007%2F978-3-319-11888-8_16.pdf02518nas a2200337 4500008004100000245007600041210006900117260001200186490000800198520151000206653001201716653001701728653001001745653001601755653003301771653002201804653002201826653001601848653001101864100002101875700002201896700002001918700002101938700002601959700002201985700002002007700002102027700002202048700001802070856009202088 2014 eng d00aCCBE1 enhances lymphangiogenesis via ADAMTS3-mediated VEGF-C activation0 aCCBE1 enhances lymphangiogenesis via ADAMTS3mediated VEGFC activ c05/20140 v1293 aBackground—Hennekam lymphangiectasia-lymphedema syndrome (OMIM 235510) is a rare autosomal recessive disease, which is associated with mutations in the collagen- and calcium-binding EGF domains 1 (CCBE1) gene. Because of the striking phenotypic similarity of embryos lacking either the Ccbe1 gene or the lymphangiogenic growth factor Vegfc gene, we searched for CCBE1 interactions with the VEGF-C growth factor signaling pathway, which is critical in embryonic and adult lymphangiogenesis. Methods and Results—By analyzing VEGF-C produced by CCBE1-transfected cells, we found that while CCBE1 itself does not process VEGF-C, it promotes proteolytic cleavage of the otherwise poorly active 29/31-kDa form of VEGF-C by the A disintegrin and metalloprotease with thrombospondin motifs-3 (ADAMTS3) protease, resulting in the mature 21/23-kDa form of VEGF-C, which induces increased VEGF-C receptor signaling. Adeno-associated viral vector (AAV) mediated transduction of CCBE1 into mouse skeletal muscle enhanced lymphangiogenesis and angiogenesis induced by AAV-VEGF-C. Conclusions—These results identify ADAMTS3 as a VEGF-C activating protease and reveal a novel type of regulation of a vascular growth factor by a protein that enhances its proteolytic cleavage and activation. The results suggest CCBE1 is a potential therapeutic tool for the modulation of lymphangiogenesis and angiogenesis in a variety of diseases that involve the lymphatic system, such as lymphedema or lymphatic metastasis.10aADAMTS310aangiogenesis10aCCBE110aendothelium10agrowth factors and cytokines10aHennekam Syndrome10ametalloproteinase10avasculature10aVEGF-C1 aJeltsch, Michael1 aJha, Sawan, Kumar1 aTvorogov, Denis1 aAnisimov, Andrey1 aLeppänen, Veli-Matti1 aHolopainen, Tanja1 aKivelä, Riikka1 aOrtega, Sagrario1 aKärpänen, Terhi1 aAlitalo, Kari uhttp://circ.ahajournals.org/content/early/2014/02/19/CIRCULATIONAHA.113.002779.abstract00713nas a2200181 4500008004100000245007900041210006900120260007600189520012600265653001200391653001000403653002200413653001500435653001100450653001200461100001500473856004300488 2014 eng d00aFrom molecular biology to a causal treatment of lymphatic system disorders0 aFrom molecular biology to a causal treatment of lymphatic system aHalle (Saale), GermanybDeutsche Gesellschaft für Lymphologiec10/20143 aOriginal Conference Abstracts (in German)10aADAMTS310aCCBE110aLymphangiogenesis10alymphedema10aVEGF-C10aVEGFR-31 aJeltsch, M uhttps://jeltsch.org/abstract-Halle201401874nas a2200289 4500008004100000022001400041245011000055210006900165260000900234300000900243490000600252520103100258653000901289653002901298653002401327653001101351653001201362100002101374700002601395700002001421700002101441700002101462700002201483700002301505700001801528856003801546 2013 eng d a1937-914500aThe basis for the distinct biological activities of vascular endothelial growth factor receptor-1 ligands0 abasis for the distinct biological activities of vascular endothe c2013 ara520 v63 aVascular endothelial growth factors (VEGFs) regulate blood and lymphatic vessel development through VEGF receptors (VEGFRs). The VEGFR immunoglobulin homology domain 2 (D2) is critical for ligand binding, and D3 provides additional interaction sites. VEGF-B and placenta growth factor (PlGF) bind to VEGFR-1 with high affinity, but only PlGF is angiogenic in most tissues. We show that VEGF-B, unlike other VEGFs, did not require D3 interactions for high-affinity binding. VEGF-B with a PlGF-derived L1 loop (B-L1(P)) stimulated VEGFR-1 activity, whereas PlGF with a VEGF-B-derived L1 loop (P-L1(B)) did not. Unlike P-L1(B) and VEGF-B, B-L1(P) and PlGF were also angiogenic in mouse skeletal muscle. Furthermore, B-L1(P) also bound to VEGFR-2 and activated downstream signaling. These results establish a role for L1-mediated D3 interactions in VEGFR activation in endothelial cells and indicate that VEGF-B is a high-affinity VEGFR-1 ligand that, unlike PlGF, cannot efficiently induce signaling downstream of VEGFR-1.
10aPlGF10areceptor tyrosine kinase10aSignal Transduction10aVEGF-B10aVEGFR-11 aAnisimov, Andrey1 aLeppänen, Veli-Matti1 aTvorogov, Denis1 aZarkada, Georgia1 aJeltsch, Michael1 aHolopainen, Tanja1 aKaijalainen, Seppo1 aAlitalo, Kari uhttps://jeltsch.org/Anisimov2013b01999nas a2200217 4500008004100000245016300041210006900204260001200273300001300285490000700298520124700305653001901552653002201571653001501593653002701608653001101635653001101646100001801657700002101675856008501696 2013 eng d00aDie lymphangiogenic growth factors VEGF-C and VEGF-D. Part 2: The role of VEGF-C and VEGF-D in diseases of the lymphatic system. [bilingual: English, German].0 aDie lymphangiogenic growth factors VEGFC and VEGFD Part 2 The ro c11/2013 a96 - 1040 v173 aVEGF-C and VEGF-D are the two central signaling molecules that stimulate the develop- ment and growth of the lymphatic system. Both belong to the vascular endothelial growth factor (VEGF) protein family, which plays important roles in the growth of blood vessels (angiogenesis) and lymphatic vessels (lymphangiogenesis). In mammals, the VEGF family comprises five members: VEGF-A, PlGF, VEGF-B, VEGF-C and VEGF-D. The family was named after VEGF-A, the first member to be discovered. VEGF-C and VEGF-D form a subgroup within this family in terms of function and structure. Their distinctive biosynthesis differentiates them from the other VEGFs: they are produced as inactive precursors and need to be activated by proteolytic removal of their long N- and C-terminal propeptides. Unlike the other VEGFs, VEGF-C and VEGF-D are direct stimulators of lymphatic vessel growth. They exert their lymphangiogenic function via VEGF receptor 3, which is expressed in the adult organism almost exclusively on lymphatic endothelial cells. In this review, we provide an overview of the VEGF protein family and their receptors. We focus on the lymphangiogenic VEGF-C and VEGF-D, discussing their biosynthesis and their role in embryonic lymphangiogenesis.10agrowth factors10aLymphangiogenesis10alymphedema10alymphogenic metastasis10aVEGF-C10aVEGF-D1 aKrebs, Rainer1 aJeltsch, Michael uhttp://jeltsch.org/sites/jeltsch.org/files/JeltschMichael_Lymphforsch2013_96.pdf01909nas a2200193 4500008004100000245014000041210006900181260001200250300001200262490000700274520124700281653001901528653002201547653001101569653001101580100001801591700002101609856008501630 2013 eng d00aThe lymphangiogenic growth factors VEGF-C and VEGF-D. Part 1: Basic principles and embryonic development. [bilingual: English, German].0 alymphangiogenic growth factors VEGFC and VEGFD Part 1 Basic prin c05/2013 a30 - 370 v173 aVEGF-C and VEGF-D are the two central signaling molecules that stimulate the develop- ment and growth of the lymphatic system. Both belong to the vascular endothelial growth factor (VEGF) protein family, which plays important roles in the growth of blood vessels (angiogenesis) and lymphatic vessels (lymphangiogenesis). In mammals, the VEGF family comprises five members: VEGF-A, PlGF, VEGF-B, VEGF-C and VEGF-D. The family was named after VEGF-A, the first member to be discovered. VEGF-C and VEGF-D form a subgroup within this family in terms of function and structure. Their distinctive biosynthesis differentiates them from the other VEGFs: they are produced as inactive precursors and need to be activated by proteolytic removal of their long N- and C-terminal propeptides. Unlike the other VEGFs, VEGF-C and VEGF-D are direct stimulators of lymphatic vessel growth. They exert their lymphangiogenic function via VEGF receptor 3, which is expressed in the adult organism almost exclusively on lymphatic endothelial cells. In this review, we provide an overview of the VEGF protein family and their receptors. We focus on the lymphangiogenic VEGF-C and VEGF-D, discussing their biosynthesis and their role in embryonic lymphangiogenesis.10agrowth factors10aLymphangiogenesis10aVEGF-C10aVEGF-D1 aKrebs, Rainer1 aJeltsch, Michael uhttp://jeltsch.org/sites/jeltsch.org/files/JeltschMichael_Lymphforsch2013_30.pdf01449nas a2200169 4500008004100000020001600041245005100057210005000108260000900158490000600167520096300173100002101136700002601157700002101183700001801204856005701222 2013 eng d a, 1943-026400aReceptor Tyrosine Kinase-Mediated Angiogenesis0 aReceptor Tyrosine KinaseMediated Angiogenesis c20130 v53 aThe 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.1 aJeltsch, Michael1 aLeppänen, Veli-Matti1 aSaharinen, Pipsa1 aAlitalo, Kari uhttp://cshperspectives.cshlp.org/content/5/9/a00918303178nas a2200517 4500008004100000020001400041245009100055210006900146260001200215300001800227490000800245520156900253653002401822653001801846653002501864653002701889653004001916653001101956653001201967653002501979653002202004653002802026653002702054653001302081653002002094653002802114653003202142653002802174653003402202653001902236653004102255653005002296653002202346100002602368700002002394700001702414700002102431700002102452700002102473700002902494700002202523700002302545700002402568700001802592856005002610 2013 eng d a1091-649000aStructural and mechanistic insights into VEGF receptor 3 ligand binding and activation0 aStructural and mechanistic insights into VEGF receptor 3 ligand c08/2013 a12960 - 129650 v1103 aVascular 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.10aAmino Acid Sequence10aBinding Sites10aBinding, Competitive10aCrystallography, X-Ray10aElectrophoresis, Polyacrylamide Gel10aHumans10aLigands10aMicroscopy, Electron10aModels, Molecular10aMolecular Sequence Data10aMultiprotein Complexes10aMutation10aProtein Binding10aProtein Multimerization10aProtein Structure, Tertiary10aScattering, Small Angle10aSequence Homology, Amino Acid10aThermodynamics10aVascular Endothelial Growth Factor C10aVascular Endothelial Growth Factor Receptor-310aX-Ray Diffraction1 aLeppänen, Veli-Matti1 aTvorogov, Denis1 aKisko, Kaisa1 aProta, Andrea, E1 aJeltsch, Michael1 aAnisimov, Andrey1 aMarkovic-Mueller, Sandra1 aStuttfeld, Edward1 aGoldie, Kenneth, N1 aBallmer-Hofer, Kurt1 aAlitalo, Kari uhttp://www.pnas.org/content/110/32/12960.long02689nas a2200469 4500008004100000022001400041245014300055210006900198260001300267300001300280490000800293520127600301653001201577653001201589653003401601653002801635653001801663653001801681653002001699653002501719653001101744653002101755653000901776653001901785653003601804653002801840653002101868653002401889653004101913653001401954653002301968100002701991700002102018700001902039700002102058700002102079700001902100700002102119700001802140700002202158856003902180 2013 eng d a1477-912900aA truncation allele in vascular endothelial growth factor c reveals distinct modes of signaling during lymphatic and vascular development.0 atruncation allele in vascular endothelial growth factor c reveal c2013 Apr a1497-5060 v1403 aVascular 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.
10aAlleles10aAnimals10aAnimals, Genetically Modified10aAutocrine Communication10aBlood Vessels10aCell Movement10aCodon, Nonsense10aEmbryo, Nonmammalian10aFemale10aLymphatic System10aMice10aMice, Knockout10aNeovascularization, Physiologic10aParacrine Communication10aProtein Isoforms10aSignal Transduction10aVascular Endothelial Growth Factor C10aZebrafish10aZebrafish Proteins1 aVillefranc, Jacques, A1 aNicoli, Stefania1 aBentley, Katie1 aJeltsch, Michael1 aZarkada, Georgia1 aMoore, John, C1 aGerhardt, Holger1 aAlitalo, Kari1 aLawson, Nathan, D uhttps://jeltsch.org/Villefranc201302825nas a2200361 4500008004100000245011400041210006900155260001200224300001200236490000800248520180400256100002102060700002002081700002202101700002602123700001002149700001202159700001602171700001402187700002202201700001202223700002002235700001702255700002102272700002102293700002602314700002302340700001202363700001202375700002102387700001802408856003702426 2013 eng d00aVascular endothelial growth factor-angiopoietin chimera with improved properties for therapeutic angiogenesis0 aVascular endothelial growth factorangiopoietin chimera with impr c01/2013 a424-4340 v1273 aBACKGROUND: 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.1 aAnisimov, Andrey1 aTvorogov, Denis1 aAlitalo, Annamari1 aLeppänen, Veli-Matti1 aAn, Y1 aHan, EC1 aOrsenigo, F1 aGaál, EI1 aHolopainen, Tanja1 aKoh, YJ1 aTammela, Tuomas1 aKorpisalo, P1 aKeskitalo, Salla1 aJeltsch, Michael1 aYlä-Herttuala, Seppo1 aDejana, Elisabetta1 aKoh, GY1 aChoi, C1 aSaharinen, Pipsa1 aAlitalo, Kari uhttps://jeltsch.org/Anisimov201303199nas a2200529 4500008004100000022001400041245013000055210006900185260001300254300001200267490000800279520162800287653002201915653001201937653002901949653001501978653001701993653002002010653003702030653002002067653002102087653001502108653002002123653002102143653002002164653001702184653002202201653001602223653001602239653000902255653002402264653001502288653001202303653003202315653001802347653004102365653005002406100001802456700002302474700002302497700002102520700002202541700002602563700002302589700002302612856003402635 2012 eng d a1525-219100aCritical role of VEGF-C/VEGFR-3 signaling in innate and adaptive immune responses in experimental obliterative bronchiolitis.0 aCritical role of VEGFCVEGFR3 signaling in innate and adaptive im c2012 Nov a1607-200 v1813 aChronic inflammation, a hallmark of obliterative bronchiolitis, is known to induce lymphangiogenesis. We therefore studied the role of lymphangiogenic vascular endothelial growth factor C (VEGF-C), its receptor VEGFR-3, and lymphangiogenesis during development of experimental obliterative bronchiolitis [ie, obliterative airway disease (OAD)] in rat tracheal allografts. The functional importance of VEGF-C was investigated by adenovirus-mediated overexpression of VEGF-C (AdVEGF-C), and by inhibition of VEGF-C activity with VEGFR-3-Ig (AdVEGFR-3-Ig). Analyses included histology, immunohistochemistry, and real-time RT-PCR 10 and 30 days after transplantation. In the course of OAD development, lymphangiogenesis was induced in the airway wall during the alloimmune response, which was reversed by cyclosporine A in a dose-dependent fashion. VEGF-C overexpression in tracheal allografts induced epithelial activation, neutrophil chemotaxis, and a shift toward a Th17 adaptive immune response, followed by enhanced lymphangiogenesis and the development of OAD. In contrast, inhibition of VEGF-C activity with VEGFR-3-Ig inhibited lymphangiogenesis and angiogenesis and reduced infiltration of CD4(+) T cells and the development of OAD. Lymphangiogenesis was linked to T-cell responses during the development of OAD, and VEGF-C/VEGFR-3 signaling modulated innate and adaptive immune responses in the development of OAD in rat tracheal allografts. Our results thus suggest VEGFR-3-signaling as a novel strategy to regulate T-cell responses in the development of obliterative bronchiolitis after lung transplantation.
10aAdaptive Immunity10aAnimals10aBronchiolitis Obliterans10aChemotaxis10aCyclosporine10aDendritic Cells10aDose-Response Relationship, Drug10aDown-Regulation10aEpithelial Cells10aEpithelium10aGraft Rejection10aImmunity, Innate10aImmunoglobulins10aInflammation10aLymphangiogenesis10aMacrophages10aNeutrophils10aRats10aSignal Transduction10aTh17 Cells10aTrachea10aTransplantation, Homologous10aUp-Regulation10aVascular Endothelial Growth Factor C10aVascular Endothelial Growth Factor Receptor-31 aKrebs, Rainer1 aTikkanen, Jussi, M1 aRopponen, Jussi, O1 aJeltsch, Michael1 aJokinen, Janne, J1 aYlä-Herttuala, Seppo1 aNykänen, Antti, I1 aLemström, Karl, B uhttps://jeltsch.org/Krebs201202103nas a2200241 4500008004100000245010100041210006900142260001400211300001400225490000800239520135400247100002601601700002101627700002101648700002001669700001501689700002101704700001901725700002601744700002401770700001801794856004901812 2011 eng d00aStructural determinants of vascular endothelial growth factor-D receptor binding and specificity0 aStructural determinants of vascular endothelial growth factorD r c2011/Feb/ a1507 - 150 v1173 aVascular 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.1 aLeppänen, Veli-Matti1 aJeltsch, Michael1 aAnisimov, Andrey1 aTvorogov, Denis1 aAho, Kukka1 aKalkkinen, Nisse1 aToivanen, Pyry1 aYlä-Herttuala, Seppo1 aBallmer-Hofer, Kurt1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/2114808500655nas a2200217 4500008004100000245010700041210006900148260001100217300001600228490000700244100001300251700001700264700001700281700001500298700001600313700002200329700001700351700001700368700001800385856003400403 2011 eng d00aVEGF-C/VEGFR-3 Signaling Regulates Inflammatory Response in Development of Obliterative Airway Disease0 aVEGFCVEGFR3 Signaling Regulates Inflammatory Response in Develop c2011// aS118 - S1180 v301 aKrebs, R1 aTikkanen, JM1 aRopponen, JO1 aJeltsch, M1 aJokinen, JJ1 aYlä-Herttuala, S1 aKoskinen, PK1 aNykänen, AI1 aLemström, KB uhttps://jeltsch.org/Krebs201101603nas a2200265 4500008004100000245011900041210006900160260001400229300001300243490000700256520077800263100002101041700002001062700002001082700002101102700002201123700002101145700002301166700002301189700001601212700002101228700002101249700001801270856004901288 2010 eng d00aClaudin-like protein 24 interacts with the VEGFR-2 and VEGFR-3 pathways and regulates lymphatic vessel development0 aClaudinlike protein 24 interacts with the VEGFR2 and VEGFR3 path c2010/May/ a875 - 800 v243 aThe 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.1 aSaharinen, Pipsa1 aHelotera, Hanna1 aMiettinen, Juho1 aNorrmen, Camilla1 aD'Amico, Gabriela1 aJeltsch, Michael1 aLangenberg, Tobias1 aVandevelde, Wouter1 aNy, Annelii1 aDewerchin, Mieke1 aCarmeliet, Peter1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/2043942801838nas a2200289 4500008004100000245014100041210006900182260001400251300001300265490000700278520091500285100002001200700002101220700001501241700002601256700002001282700002701302700002501329700002101354700002201375700002101397700002101418700002101439700002101460700001801481856004901499 2010 eng d00aEffective suppression of vascular network formation by combination of antibodies blocking VEGFR ligand binding and receptor dimerization0 aEffective suppression of vascular network formation by combinati c2010/Dec/ a630 - 400 v183 aAntibodies 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.1 aTvorogov, Denis1 aAnisimov, Andrey1 aZheng, Wei1 aLeppänen, Veli-Matti1 aTammela, Tuomas1 aLaurinavicius, Simonas1 aHolnthoner, Wolfgang1 aHeloterä, Hanna1 aHolopainen, Tanja1 aJeltsch, Michael1 aKalkkinen, Nisse1 aLankinen, Hilkka1 aOjala, Päivi, M1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/2113004302522nas a2200241 4500008004100000245008800041210006900129260001200198300001400210490000800224520178600232100002602018700002102044700002102065700002102086700002102107700002002128700002102148700002002169700002402189700001802213856004902231 2010 eng d00aStructural determinants of growth factor binding and specificity by VEGF receptor 20 aStructural determinants of growth factor binding and specificity c02/2010 a2425 - 300 v1073 aVascular 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.1 aLeppänen, Veli-Matti1 aProta, Andrea, E1 aJeltsch, Michael1 aAnisimov, Andrey1 aKalkkinen, Nisse1 aStrandin, Tomas1 aLankinen, Hilkka1 aGoldman, Adrian1 aBallmer-Hofer, Kurt1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/2014511602377nas a2200265 4500008004100000245014000041210006900181260001100250300001100261490000600272520152800278100001901806700002101825700002401846700002001870700002301890700002501913700002001938700002101958700001801979700001801997700002502015700002202040856004902062 2010 eng d00aSuppressive effects of vascular endothelial growth factor-B on tumor growth in a mouse model of pancreatic neuroendocrine tumorigenesis0 aSuppressive effects of vascular endothelial growth factorB on tu c2010// ae141090 v53 aBACKGROUND: 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.1 aAlbrecht, Imke1 aKopfstein, Lucie1 aStrittmatter, Karin1 aSchomber, Tibor1 aFalkevall, Annelie1 aHagberg, Carolina, E1 aLorentz, Pascal1 aJeltsch, Michael1 aAlitalo, Kari1 aEriksson, Ulf1 aChristofori, Gerhard1 aPietras, Kristian uhttp://view.ncbi.nlm.nih.gov/pubmed/2112484102729nas a2200349 4500008004100000245015500041210006900196260001400265300001400279490000800293520163900301100001501940700002001955700002201975700002101997700002002018700002002038700002102058700001902079700002102098700002102119700002102140700002302161700002202184700002602206700001902232700001902251700002302270700001902293700001802312856004902330 2010 eng d00aVascular endothelial growth factor-B acts as a coronary growth factor in transgenic rats without inducing angiogenesis, vascular leak, or inflammation0 aVascular endothelial growth factorB acts as a coronary growth fa c2010/Oct/ a1725 - 330 v1223 aBACKGROUND: 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.1 aBry, Maija1 aKivelä, Riikka1 aHolopainen, Tanja1 aAnisimov, Andrey1 aTammela, Tuomas1 aSoronen, Jarkko1 aSilvola, Johanna1 aSaraste, Antti1 aJeltsch, Michael1 aKorpisalo, Petra1 aCarmeliet, Peter1 aLemström, Karl, B1 aShibuya, Masabumi1 aYlä-Herttuala, Seppo1 aAlhonen, Leena1 aMervaala, Eero1 aAndersson, Leif, C1 aKnuuti, Juhani1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/2093797402406nas a2200229 4500008004100000245009500041210006900136260001400205300001400219490000800233520168800241100002101929700002201950700002101972700002001993700002302013700002602036700002102062700002602083700001802109856004902127 2009 eng d00aActivated forms of VEGF-C and VEGF-D provide improved vascular function in skeletal muscle0 aActivated forms of VEGFC and VEGFD provide improved vascular fun c2009/Jun/ a1302 - 120 v1043 aThe 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.1 aAnisimov, Andrey1 aAlitalo, Annamari1 aKorpisalo, Petra1 aSoronen, Jarkko1 aKaijalainen, Seppo1 aLeppänen, Veli-Matti1 aJeltsch, Michael1 aYlä-Herttuala, Seppo1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1944383502437nas a2200325 4500008004100000245014100041210006900182260001400251300001400265490000800279520142200287100002001709700001501729700002101744700003101765700002001796700002001816700002001836700002001856700001901876700002101895700002001916700002301936700001901959700002201978700002602000700001802026700001802044856004902062 2008 eng d00aOverexpression of vascular endothelial growth factor-B in mouse heart alters cardiac lipid metabolism and induces myocardial hypertrophy0 aOverexpression of vascular endothelial growth factorB in mouse h c2008/Oct/ a1018 - 260 v1033 aVascular 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.1 aKarpanen, Terhi1 aBry, Maija1 aOllila, Hanna, M1 aSeppänen-Laakso, Tuulikki1 aLiimatta, Erkki1 aLeskinen, Hanna1 aKivelä, Riikka1 aHelkamaa, Teemu1 aMerentie, Mari1 aJeltsch, Michael1 aPaavonen, Karri1 aAndersson, Leif, C1 aMervaala, Eero1 aHassinen, Ilmo, E1 aYlä-Herttuala, Seppo1 aOresic, Matej1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1875782702098nas a2200385 4500008004100000245011800041210006900159260001400228300001400242490000700256520095200263100001301215700001501228700001901243700001901262700001601281700002001297700002101317700002001338700002401358700002901382700002001411700002901431700001601460700002501476700002501501700002301526700002101549700001901570700001801589700001801607700002101625700001701646856004901663 2008 eng d00aReevaluation of the role of VEGF-B suggests a restricted role in the revascularization of the ischemic myocardium0 aReevaluation of the role of VEGFB suggests a restricted role in c2008/Sep/ a1614 - 200 v283 aOBJECTIVE: 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.1 aLi, Xuri1 aTjwa, Marc1 aVan Hove, Inge1 aEnholm, Berndt1 aNeven, Elke1 aPaavonen, Karri1 aJeltsch, Michael1 aJuan, Toni Diez1 aSievers, Richard, E1 aChorianopoulos, Emmanuel1 aWada, Hiromichi1 aVanwildemeersch, Maarten1 aNoel, Agnes1 aFoidart, Jean-Michel1 aSpringer, Matthew, L1 aDegenfeld, Georges1 aDewerchin, Mieke1 aBlau, Helen, M1 aAlitalo, Kari1 aEriksson, Ulf1 aCarmeliet, Peter1 aMoons, Lieve uhttp://view.ncbi.nlm.nih.gov/pubmed/1851169902705nas a2200229 4500008004100000245014300041210006900184260001400253300001400267490000700281520193200288100002502220700002202245700002102267700002102288700002102309700002602330700002202356700003002378700001802408856004902426 2008 eng d00aThe tyrosine kinase inhibitor cediranib blocks ligand-induced vascular endothelial growth factor receptor-3 activity and lymphangiogenesis0 atyrosine kinase inhibitor cediranib blocks ligandinduced vascula c2008/Jun/ a4754 - 620 v683 aSolid tumors express a range of factors required to sustain their growth and promote their dissemination. Among these are vascular endothelial growth factor-A (VEGF-A), the key angiogenic stimulant, and VEGF-C, a primary mediator of lymphangiogenesis. Small molecule tyrosine kinase inhibitors offer the potential to inhibit more than one kinase and impede tumor growth by multiple mechanisms. However, their potency toward individual targets can vary. Cediranib (RECENTIN; AZD2171) is an inhibitor of VEGF signaling that has been shown in experimental models to prevent VEGF-A-induced angiogenesis and primary tumor growth, yet the effects of cediranib on VEGF receptor (VEGFR)-3-mediated endothelial cell function and lymphangiogenesis are unknown. To better understand the activity of cediranib against VEGFR-3 and its associated signaling events compared with its activity against VEGFR-2, we used the receptor-specific ligands VEGF-E and VEGF-C156S. In human endothelial cells, cediranib inhibited VEGF-E-induced phosphorylation of VEGFR-2 and VEGF-C156S-induced phosphorylation of VEGFR-3 at concentrations of =1nmol/L and inhibited activation of downstream signaling molecules. Additionally, cediranib blocked VEGF-C156S-induced and VEGF-E-induced proliferation, survival, and migration of lymphatic and blood vascular endothelial cells. In vivo, cediranib (6 mg/kg/d) prevented angiogenesis and lymphangiogenesis induced by VEGF-E-expressing and VEGF-C156S-expressing adenoviruses, respectively. Cediranib (6 mg/kg/day) also blocked angiogenesis and lymphangiogenesis induced by adenoviruses expressing VEGF-A or VEGF-C and compromised the blood and lymphatic vasculatures of VEGF-C-expressing tumors. Cediranib may, therefore, be an effective means of preventing tumor progression, not only by inhibiting VEGFR-2 activity and angiogenesis, but also by concomitantly inhibiting VEGFR-3 activity and lymphangiogenesis.1 aHeckman, Caroline, A1 aHolopainen, Tanja1 aWirzenius, Maria1 aKeskitalo, Salla1 aJeltsch, Michael1 aYlä-Herttuala, Seppo1 aWedge, Stephen, R1 aJürgensmeier, Juliane, M1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1855952202160nas a2200217 4500008004100000245017800041210006900219260001400288300001400302490000800316520140200324100002001726700001501746700002401761700002101785700002301806700002001829700002601849700001801875856004901893 2007 eng d00aDistinct architecture of lymphatic vessels induced by chimeric vascular endothelial growth factor-C/vascular endothelial growth factor heparin-binding domain fusion proteins0 aDistinct architecture of lymphatic vessels induced by chimeric v c2007/May/ a1468 - 750 v1003 aVascular 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.1 aTammela, Tuomas1 aHe, Yulong1 aLyytikkä, Johannes1 aJeltsch, Michael1 aMarkkanen, Johanna1 aPajusola, Katri1 aYlä-Herttuala, Seppo1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1747873301875nas a2200217 4500008004100000245015700041210006900198260001400267300001300281490000800294520113500302100002101437700002001458700002301478700002001501700002101521700002301542700002501565700001801590856004901608 2007 eng d00aEnhanced capillary formation stimulated by a chimeric vascular endothelial growth factor/vascular endothelial growth factor-C silk domain fusion protein0 aEnhanced capillary formation stimulated by a chimeric vascular e c2007/May/ a1460 - 70 v1003 aVascular 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.1 aKeskitalo, Salla1 aTammela, Tuomas1 aLyytikka, Johannes1 aKarpanen, Terhi1 aJeltsch, Michael1 aMarkkanen, Johanna1 aYla-Herttuala, Seppo1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1747873402828nas a2200181 4500008004100000245013000041210006900171260007100240520211900311100002602430700002202456700002102478700002102499700002102520700002302541700003002564856005202594 2007 eng d00aInhibiton of VEGF-C-induced VEGFR-3 activity and lymphatic endothelial cell function by the tyrosine kinase inhibitor AZD21710 aInhibiton of VEGFCinduced VEGFR3 activity and lymphatic endothel aLos Angeles, CAbAmerican Association for Cancer Researchc2007///3 aSolid tumors express a range of growth factors required to sustain their growth and promote their dissemination. Among these factors is vascular endothelial growth factor-A (VEGF-A), the key angiogenic stimulant, and VEGF-C, a primary mediator of lymphangiogenesis. Small molecule tyrosine kinase inhibitors can prevent VEGF signaling activity by targeting the VEGF receptors and are an effective approach to impede tumor progression. The indole-ether quinazoline AZD2171 is a highly potent ATP-competitive inhibitor of VEGFR-2 (KDR) kinase, with additional activity against VEGFR-1 (Flt-1) and -3 (Flt-4), that has been shown in experimental models to prevent VEGF-A-induced angiogenesis and primary tumor growth (Wedge et al. Cancer Res 2005;65:4389-4400). For these studies we wished to further assess the ability of AZD2171 to inhibit VEGFR-3 and its associated functions. Upon binding its ligands VEGF-C or -D, VEGFR-3 becomes activated with the resulting signaling cascade eventually translated into increased proliferation, survival and migration of lymphatic and blood vascular endothelial cells. At concentrations of ≤1 nM AZD2171 inhibited VEGFR-3 phosphorylation in porcine aortic endothelial cells selectively expressing the human receptor, and in human dermal microvascular endothelial cells (HDMVECs). In HDMVECs, AZD2171 prevented phosphorylation of signaling molecules downstream of VEGFR-2 and -3, ERK1/2, Akt and CREB, induced by the VEGFR-2 and -3-specific ligands VEGF-E and -C156S, respectively. Additionally, AZD2171 blocked VEGF-E- and -C156S-induced proliferation of both lymphatic and blood vascular endothelial cells at similar concentrations, and prevented ligand-induced endothelial cell cord formation in a Matrigel assay. The effects of AZD2171 on VEGF-C-induced lymphangiogenesis are currently being assessed in vivo. These studies, together with previous results, not only demonstrate that AZD2171 may be an effective means of preventing tumor progression by inhibition of VEGFR-2 activity and angiogenesis, but may also prevent further tumor spread by inhibiting VEGFR-3 activity1 aHeckman, Caroline, A.1 aHolopainen, Tanja1 aWirzenius, Maria1 aKeskitalo, Salla1 aJeltsch, Michael1 aWedge, Stephen, R.1 aJurgensmeier, Juliane, M. uhttp://dx.doi.org/10.1158/0008-5472.CAN-07-580901805nas a2200217 4500008004100000245007400041210006900115260001400184300001400198490000700212520115600219100002201375700002501397700002101422700002101443700001801464700001801482700002001500700001801520856004901538 2006 eng d00aFunctional interaction of VEGF-C and VEGF-D with neuropilin receptors0 aFunctional interaction of VEGFC and VEGFD with neuropilin recept c2006/Jul/ a1462 - 720 v203 aLymphatic vascular development is regulated by vascular endothelial growth factor receptor-3 (VEGFR-3), which is activated by its ligands VEGF-C and VEGF-D. Neuropilin-2 (NP2), known to be involved in neuronal development, has also been implicated to play a role in lymphangiogenesis. We aimed to elucidate the mechanism by which NP2 is involved in lymphatic endothelial cell signaling. By in vitro binding studies we found that both VEGF-C and VEGF-D interact with NP2, VEGF-C in a heparin-independent and VEGF-D in a heparin-dependent manner. We also mapped the domains of VEGF-C and NP2 required for their binding. The functional importance of the interaction of NP2 with the lymphangiogenic growth factors was demonstrated by cointernalization of NP2 along with VEGFR-3 in endocytic vesicles of lymphatic endothelial cells upon stimulation with VEGF-C or VEGF-D. NP2 also interacted with VEGFR-3 in coprecipitation studies. Our results show that NP2 is directly involved in an active signaling complex with the key regulators of lymphangiogenesis and thus suggest a mechanism by which NP2 functions in the development of the lymphatic vasculature.1 aKärpänen, Terhi1 aHeckman, Caroline, A1 aKeskitalo, Salla1 aJeltsch, Michael1 aOllila, Hanna1 aNeufeld, Gera1 aTamagnone, Luca1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1681612101519nas a2200193 4500008004100000245014600041210006900187260001400256300001500270490000800285520086800293100002101161700002001182700002001202700001501222700002101237700001801258856004901276 2006 eng d00aVascular endothelial growth factor (VEGF)/VEGF-C mosaic molecules reveal specificity determinants and feature novel receptor binding patterns0 aVascular endothelial growth factor VEGFVEGFC mosaic molecules re c2006/Apr/ a12187 - 950 v2813 aVascular 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.1 aJeltsch, Michael1 aKarpanen, Terhi1 aStrandin, Tomas1 aAho, Kukka1 aLankinen, Hilkka1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1650548900372nas a2200133 4500008004100000245001900041210001900060250000700079260001000086100002100096700001800117710001600135856008700151 2006 eng d00aVEGF Receptors0 aVEGF Receptors a5. bSigma1 aJeltsch, Michael1 aAlitalo, Kari1 aWatling, K. uhttp://www.sigmaaldrich.com/technical-documents/articles/biology/rbi-handbook.html02389nas a2200289 4500008004100000245009100041210006900132260001400201300001300215490000800228520151500236100001701751700002001768700001501788700002301803700001901826700002301845700002101868700002401889700002401913700002301937700002601960700002201986700001802008700002402026856004902050 2005 eng d00aPathogenesis of persistent lymphatic vessel hyperplasia in chronic airway inflammation0 aPathogenesis of persistent lymphatic vessel hyperplasia in chron c2005/Feb/ a247 - 570 v1153 aEdema 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.1 aBaluk, Peter1 aTammela, Tuomas1 aAtor, Erin1 aLyubynska, Natalya1 aAchen, Marc, G1 aHicklin, Daniel, J1 aJeltsch, Michael1 aPetrova, Tatiana, V1 aPytowski, Bronislaw1 aStacker, Steven, A1 aYlä-Herttuala, Seppo1 aJackson, David, G1 aAlitalo, Kari1 aMcDonald, Donald, M uhttp://view.ncbi.nlm.nih.gov/pubmed/1566873402358nas a2200241 4500008004100000245016500041210006900206260001400275300001400289490000700303520155700310100001501867700001901882700002001901700002101921700002201942700002501964700002001989700001702009700002302026700001802049856004902067 2005 eng d00aVascular endothelial cell growth factor receptor 3-mediated activation of lymphatic endothelium is crucial for tumor cell entry and spread via lymphatic vessels0 aVascular endothelial cell growth factor receptor 3mediated activ c2005/Jun/ a4739 - 460 v653 aLymphangiogenic growth factors vascular endothelial growth factor (VEGF)-C and VEGF-D have been shown to promote lymphatic metastasis by inducing tumor-associated lymphangiogenesis. In this study, we have investigated how tumor cells gain access into lymphatic vessels and at what stage tumor cells initiate metastasis. We show that VEGF-C produced by tumor cells induced extensive lymphatic sprouting towards the tumor cells as well as dilation of the draining lymphatic vessels, suggesting an active role of lymphatic endothelial cells in lymphatic metastasis. A significant increase in lymphatic vessel growth occurred between 2 and 3 weeks after tumor xenotransplantation, and lymph node metastasis occurred at the same stage. These processes were blocked dose-dependently by inhibition of VEGF receptor 3 (VEGFR-3) signaling by systemic delivery of a soluble VEGFR-3-immunoglobulin (Ig) fusion protein via adenoviral or adeno-associated viral vectors. However, VEGFR-3-Ig did not suppress lymph node metastasis when the treatment was started at a later stage after the tumor cells had already spread out, suggesting that tumor cell entry into lymphatic vessels is a key step during tumor dissemination via the lymphatics. Whereas lymphangiogenesis and lymph node metastasis were significantly inhibited by VEGFR-3-Ig, some tumor cells were still detected in the lymph nodes in some of the treated mice. This indicates that complete blockade of lymphatic metastasis may require the targeting of both tumor lymphangiogenesis and tumor cell invasion.1 aHe, Yulong1 aRajantie, Iiro1 aPajusola, Katri1 aJeltsch, Michael1 aHolopainen, Tanja1 aYla-Herttuala, Seppo1 aHarding, Thomas1 aJooss, Karin1 aTakahashi, Takashi1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1593029201674nas a2200265 4500008004100000245011900041210006900160260001400229300001200243490000600255520085100261100002601112700001701138700001801155700001901173700001901192700002401211700002101235700002201256700001901278700002001297700002401317700001801341856004901359 2004 eng d00aVascular endothelial growth factor C is required for sprouting of the first lymphatic vessels from embryonic veins0 aVascular endothelial growth factor C is required for sprouting o c2004/Jan/ a74 - 800 v53 aLymphatic vessels are essential for immune surveillance, tissue fluid homeostasis and fat absorption. Defects in lymphatic vessel formation or function cause lymphedema. Here we show that the vascular endothelial growth factor C (VEGF-C) is required for the initial steps in lymphatic development. In Vegfc-/- mice, endothelial cells commit to the lymphatic lineage but do not sprout to form lymph vessels. Sprouting was rescued by VEGF-C and VEGF-D but not by VEGF, indicating VEGF receptor 3 specificity. The lack of lymphatic vessels resulted in prenatal death due to fluid accumulation in tissues, and Vegfc+/- mice developed cutaneous lymphatic hypoplasia and lymphedema. Our results indicate that VEGF-C is the paracrine factor essential for lymphangiogenesis, and show that both Vegfc alleles are required for normal lymphatic development.1 aKarkkainen, Marika, J1 aHaiko, Paula1 aSainio, Kirsi1 aPartanen, Juha1 aTaipale, Jussi1 aPetrova, Tatiana, V1 aJeltsch, Michael1 aJackson, David, G1 aTalikka, Marja1 aRauvala, Heikki1 aBetsholtz, Christer1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1463464601736nas a2200169 4500008004100000245005000041210005000091260001400141300001200155490000800167520126400175100002101439700002001460700001801480700001901498856004901517 2003 eng d00aGenesis and pathogenesis of lymphatic vessels0 aGenesis and pathogenesis of lymphatic vessels c2003/Oct/ a69 - 840 v3143 aThe lymphatic system is generally regarded as supplementary to the blood vascular system, in that it transports interstitial fluid, macromolecules, and immune cells back into the blood. However, in insects, the open hemolymphatic (or lymphohematic) system ensures the circulation of immune cells and interstitial fluid through the body. The Drosophila homolog of the mammalian vascular endothelial growth factor receptor (VEGFR) gene family is expressed in hemocytes, suggesting a close relationship to the endothelium that develops later in phylogeny. Lymph hearts are typical organs for the propulsion of lymph in lower vertebrates and are still transiently present in birds. The lymphatic endothelial marker VEGFR-3 is transiently expressed in embryonic blood vessels and is crucial for their development. We therefore regard the question of whether the blood vascular system or the lymphatic system is primary or secondary as open. Future molecular comparisons should be performed without any bias based on the current prevalence of the blood vascular system over the lymphatic system. Here, we give an overview of the structure, function, and development of the lymphatics, with special emphasis on the recently discovered lymphangiogenic growth factors.1 aJeltsch, Michael1 aTammela, Tuomas1 aAlitalo, Kari1 aWilting, Jörg uhttp://view.ncbi.nlm.nih.gov/pubmed/1294236202307nas a2200241 4500008004100000245010400041210006900145260001400214300001400228490000700242520156400249100002001813700001801833700002301851700002001874700001801894700001901912700002101931700002101952700002501973700001801998856004902016 2003 eng d00aIntrinsic versus microenvironmental regulation of lymphatic endothelial cell phenotype and function0 aIntrinsic versus microenvironmental regulation of lymphatic endo c2003/Nov/ a2006 - 130 v173 aVascular 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.1 aVeikkola, Tanja1 aLohela, Marja1 aIkenberg, Kristian1 aMäkinen, Taija1 aKorff, Thomas1 aSaaristo, Anne1 aPetrova, Tatania1 aJeltsch, Michael1 aAugustin, Hellmut, G1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1459767001908nas a2200253 4500008004100000245007800041210006900119260001400188300001400202490000800216520113900224100002101363700002101384700002201405700002501427700002201452700002501474700002101499700002601520700001801546700001701564700002401581856004901605 2003 eng d00aVEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia0 aVEGF guides angiogenic sprouting utilizing endothelial tip cell c2003/Jun/ a1163 - 770 v1613 aVascular 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.1 aGerhardt, Holger1 aGolding, Matthew1 aFruttiger, Marcus1 aRuhrberg, Christiana1 aLundkvist, Andrea1 aAbramsson, Alexandra1 aJeltsch, Michael1 aMitchell, Christopher1 aAlitalo, Kari1 aShima, David1 aBetsholtz, Christer uhttp://view.ncbi.nlm.nih.gov/pubmed/1281070002304nas a2200277 4500008004100000245015900041210006900200260001400269300001300283490000700296520139900303100001901702700002001721700001901741700002001760700001901780700002001799700002001819700002101839700002601860700002501886700002201911700002601933700001801959856004901977 2002 eng d00aAdenoviral VEGF-C overexpression induces blood vessel enlargement, tortuosity, and leakiness but no sprouting angiogenesis in the skin or mucous membranes0 aAdenoviral VEGFC overexpression induces blood vessel enlargement c2002/Jul/ a1041 - 90 v163 aVascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are important regulators of blood and lymphatic vessel growth and vascular permeability. The VEGF-C/VEGFR-3 signaling pathway is crucial for lymphangiogenesis, and heterozygous inactivating missense mutations of the VEGFR-3 gene are associated with hereditary lymphedema. However, VEGF-C can have potent effects on blood vessels because its receptor VEGFR-3 is expressed in certain blood vessels and because the fully processed form of VEGF-C also binds to the VEGFR-2 of blood vessels. To characterize the in vivo effects of VEGF-C on blood and lymphatic vessels, we have overexpressed VEGF-C via adenovirus- and adeno-associated virus-mediated transfection in the skin and respiratory tract of athymic nude mice. This resulted in dose-dependent enlargement and tortuosity of veins, which, along with the collecting lymphatic vessels were found to express VEGFR-2. Expression of angiopoietin 1 blocked the increased leakiness of the blood vessels induced by VEGF-C whereas vessel enlargement and lymphangiogenesis were not affected. However, angiogenic sprouting of new blood vessels was not observed in response to AdVEGF-C or AAV-VEGF-C. These results show that virally produced VEGF-C induces blood vessel changes, including vascular leak, but its angiogenic potency is much reduced compared with VEGF in normal skin.1 aSaaristo, Anne1 aVeikkola, Tanja1 aEnholm, Berndt1 aHytönen, Maija1 aArola, Johanna1 aPajusola, Katri1 aTurunen, Païvi1 aJeltsch, Michael1 aKarkkainen, Marika, J1 aKerjaschki, Dontscho1 aBueler, Hansruedi1 aYlä-Herttuala, Seppo1 aAlitalo, Kari uhttp://view.ncbi.nlm.nih.gov/pubmed/1208706502637nas a2200109 4500008004500000245004200045210004100087490001500128520230100143100002102444856006202465 2002 Engldsh 00aVEGFR-3 Ligands and Lymphangiogenesis0 aVEGFR3 Ligands and Lymphangiogenesis0 vPhD Thesis3 aMost of us have seen their own blood at one or the other time. The occasion might have been a small accident or in unfortunate cases a severe blood loss caused by a major injury. We also can feel our heart beating and the resulting pressure wave, the pulse. The existence of the car- diovascular system is obvious to us. Unlike the cardiovascular system, the lymphatic system has, until recently, escaped notable attention not only by the laymen, but also by the scientific community. It is unclear why the lymphatic system originally developed in higher vertebrates. Now, its main function seems to be to collect fluid that has leaked from the blood vessels and to return it into the cardiovascular system. Much of our knowledge about the development and structure of the lymphatic system is of considerable age, and it has been said that there has not been any progress in our understanding since the fine structure of the lymphatics was described with the introduction of the electron microscope. Vascular endothelial growth factor (VEGF) is the principal direct inducer of blood vessel growth, but it does not promote the growth of lymphatic vessels. This study demonstrates for the first time specific lymphangiogenesis as a response to the VEGF homologue VEGF-C. Overexpression of full length VEGF-C under the keratin-14 promoter in the skin of transgenic mice caused a proliferation of the lymphatic endothelium and lymphatic vessel enlargement. In the chorioallantoic membrane assay, the mature form of VEGF-C was also largely specific for lymphatic endothelial cells. A newly discovered close homologue of VEGF-C, VEGF-D was then shown to have the same receptor-binding pattern as VEGF-C. Contrary to the interaction of VEGF with its receptors, VEGF-C interaction with VEGFR-3 has not been analyzed at the molecular level. The structural determinants of VEGFR-3 binding were characterized in relation to VEGF using a non-random family shuffling approach with VEGF and VEGF-C as parent molecules. This approach led to the identification of VEGF/VEGF-C mosaic molecules that showed novel receptor binding profiles and a panel of these molecules was used to delineate the requirements of specific receptors in the induction of angiogenesis versus lymphangiogenesis.1 aJeltsch, Michael uhttp://ethesis.helsinki.fi/julkaisut/mat/bioti/vk/jeltsch01909nas a2200229 4500008004100000245010400041210006900145260001400214300001200228490000700240520124400247100001401491700001601505700001501521700001201536700001601548700001301564700001701577700002101594700001501615856004901630 2001 eng d00aAdenoviral expression of vascular endothelial growth factor-C induces lymphangiogenesis in the skin0 aAdenoviral expression of vascular endothelial growth factorC ind c2001/Mar/ a623 - 90 v883 aThe growth of blood and lymphatic vasculature is mediated in part by secreted polypeptides of the vascular endothelial growth factor (VEGF) family. The prototype VEGF binds VEGF receptor (VEGFR)-1 and VEGFR-2 and is angiogenic, whereas VEGF-C, which binds to VEGFR-2 and VEGFR-3, is either angiogenic or lymphangiogenic in different assays. We used an adenoviral gene transfer approach to compare the effects of these growth factors in adult mice. Recombinant adenoviruses encoding human VEGF-C or VEGF were injected subcutaneously into C57Bl6 mice or into the ears of nude mice. Immunohistochemical analysis showed that VEGF-C upregulated VEGFR-2 and VEGFR-3 expression and VEGF upregulated VEGFR-2 expression at 4 days after injection. After 2 weeks, histochemical and immunohistochemical analysis, including staining for the lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1), the vascular endothelial marker platelet-endothelial cell adhesion molecule-1 (PECAM-1), and the proliferating cell nuclear antigen (PCNA) revealed that VEGF-C induced mainly lymphangiogenesis in contrast to VEGF, which induced only angiogenesis. These results have significant implications in the planning of gene therapy using these growth factors.1 aEnholm, B1 aKarpanen, T1 aJeltsch, M1 aKubo, H1 aStenback, F1 aPrevo, R1 aJackson, D G1 aYla-Herttuala, S1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/1128289701998nas a2200265 4500008004100000245012000041210006900161260001400230300001400244490000700258520123100265100001601496700001501512700001501527700001601542700001501558700001701573700001201590700001601602700001801618700001501636700001701651700001501668856004901683 2001 eng d00aSignalling via vascular endothelial growth factor receptor-3 is sufficient for lymphangiogenesis in transgenic mice0 aSignalling via vascular endothelial growth factor receptor3 is s c2001/Mar/ a1223 - 310 v203 aVascular endothelial growth factor receptor-3 (VEGFR-3) has an essential role in the development of embryonic blood vessels; however, after midgestation its expression becomes restricted mainly to the developing lymphatic vessels. The VEGFR-3 ligand VEGF-C stimulates lymphangiogenesis in transgenic mice and in chick chorioallantoic membrane. As VEGF-C also binds VEGFR-2, which is expressed in lymphatic endothelia, it is not clear which receptors are responsible for the lymphangiogenic effects of VEGF-C. VEGF-D, which binds to the same receptors, has been reported to induce angiogenesis, but its lymphangiogenic potential is not known. In order to define the lymphangiogenic signalling pathway we have created transgenic mice overexpressing a VEGFR-3-specific mutant of VEGF-C (VEGF-C156S) or VEGF-D in epidermal keratinocytes under the keratin 14 promoter. Both transgenes induced the growth of lymphatic vessels in the skin, whereas the blood vessel architecture was not affected. Evidence was also obtained that these growth factors act in a paracrine manner in vivo. These results demonstrate that stimulation of the VEGFR-3 signal transduction pathway is sufficient to induce specifically lymphangiogenesis in vivo.1 aVeikkola, T1 aJussila, L1 aMakinen, T1 aKarpanen, T1 aJeltsch, M1 aPetrova, T V1 aKubo, H1 aThurston, G1 aMcDonald, D M1 aAchen, M G1 aStacker, S A1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/1125088900578nas a2200181 4500008004100000245008300041210006900124260003800193100001600231700001700247700001600264700001700280700001700297700001400314700001600328700001600344856003600360 2001 eng d00aSignalling via VEGFR-3 is sufficient for lymphangiogenesis in transgenic mice.0 aSignalling via VEGFR3 is sufficient for lymphangiogenesis in tra aMiami Beach, Floridac2001/10/29/1 aJussila, L.1 aVeikkola, T.1 aJeltsch, M.1 aThurston, G.1 aMcDonald, D.1 aAchen, M.1 aStacker, S.1 aAlitalo, K. uhttps://jeltsch.org/Jussila200102021nas a2200289 4500008004100000245009500041210006900136260001400205300001300219490000700232520122500239100001901464700001501483700001501498700001601513700001501529700001301544700001501557700001401572700001701586700001701603700001201620700001501632700001901647700001601666856004901682 2001 eng d00aVascular endothelial growth factor-C-mediated lymphangiogenesis promotes tumour metastasis0 aVascular endothelial growth factorCmediated lymphangiogenesis pr c2001/Feb/ a672 - 820 v203 aMetastasis is a frequent and lethal complication of cancer. Vascular endothelial growth factor-C (VEGF-C) is a recently described lymphangiogenic factor. Increased expression of VEGF-C in primary tumours correlates with dissemination of tumour cells to regional lymph nodes. However, a direct role for VEGF-C in tumour lymphangiogenesis and subsequent metastasis has yet to be demonstrated. Here we report the establishment of transgenic mice in which VEGF-C expression, driven by the rat insulin promoter (Rip), is targeted to beta-cells of the endocrine pancreas. In contrast to wild-type mice, which lack peri-insular lymphatics, RipVEGF-C transgenics develop an extensive network of lymphatics around the islets of Langerhans. These mice were crossed with Rip1Tag2 mice, which develop pancreatic beta-cell tumours that are neither lymphangiogenic nor metastatic. Double-transgenic mice formed tumours surrounded by well developed lymphatics, which frequently contained tumour cell masses of beta-cell origin. These mice frequently developed pancreatic lymph node metastases. Our findings demonstrate that VEGF-C-induced lymphangiogenesis mediates tumour cell dissemination and the formation of lymph node metastases.1 aMandriota, S J1 aJussila, L1 aJeltsch, M1 aCompagni, A1 aBaetens, D1 aPrevo, R1 aBanerji, S1 aHuarte, J1 aMontesano, R1 aJackson, D G1 aOrci, L1 aAlitalo, K1 aChristofori, G1 aPepper, M S uhttp://view.ncbi.nlm.nih.gov/pubmed/1117921202694nas a2200313 4500008004100000245011900041210006900160260001400229300001300243490000800256520179900264100001802063700001602081700001702097700001502114700001902129700001802148700001702166700001302183700001502196700001902211700001502230700001402245700001802259700001702277700001502294700002202309856004902331 2000 eng d00aIntravascular adenovirus-mediated VEGF-C gene transfer reduces neointima formation in balloon-denuded rabbit aorta0 aIntravascular adenovirusmediated VEGFC gene transfer reduces neo c2000/Oct/ a2262 - 80 v1023 aBACKGROUND: 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.1 aHiltunen, M O1 aLaitinen, M1 aTurunen, M P1 aJeltsch, M1 aHartikainen, J1 aRissanen, T T1 aLaukkanen, J1 aNiemi, M1 aKossila, M1 aHäkkinen, T P1 aKivelä, A1 aEnholm, B1 aMansukoski, H1 aTurunen, A M1 aAlitalo, K1 aYlä-Herttuala, S uhttp://view.ncbi.nlm.nih.gov/pubmed/1105610300881nas a2200169 4500008004100000245004100041210003900082260001400121300001300135490000700148520044500155100001600600700001500616700001600631700001500647856004900662 1999 eng d00aCurrent biology of VEGF-B and VEGF-C0 aCurrent biology of VEGFB and VEGFC c1999/Dec/ a528 - 350 v103 aEndothelial growth factors and their receptors may provide important therapeutic tools for the treatment of pathological conditions characterised by defective or aberrant angiogenesis. Vascular endothelial growth factor (VEGF) is pivotal for vasculogenesis and for angiogenesis in normal and pathological conditions. VEGF-B and VEGF-C provide this gene family with additional functions, for example, VEGF-C also regulates lymphangiogenesis.1 aOlofsson, B1 aJeltsch, M1 aEriksson, U1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/1060068901922nas a2200253 4500008004100000245014100041210006900182260001400251300001500265490000700280520116200287100001601449700001901465700001601484700001901500700001201519700001301531700001301544700001701557700001501574700001501589700001601604856004801620 1998 eng d00aVascular endothelial growth factor B (VEGF-B) binds to VEGF receptor-1 and regulates plasminogen activator activity in endothelial cells0 aVascular endothelial growth factor B VEGFB binds to VEGF recepto c1998/Sep/ a11709 - 140 v953 aThe 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.1 aOlofsson, B1 aKorpelainen, E1 aPepper, M S1 aMandriota, S J1 aAase, K1 aKumar, V1 aGunji, Y1 aJeltsch, M M1 aShibuya, M1 aAlitalo, K1 aEriksson, U uhttp://view.ncbi.nlm.nih.gov/pubmed/975173001686nas a2200217 4500008004100000245013700041210006900178260001400247300001300261490000700274520102000281100001501301700001501316700001201331700001601343700001401359700001501373700001501388700001701403856004801420 1998 eng d00aVascular endothelial growth factor D (VEGF-D) is a ligand for the tyrosine kinases VEGF receptor 2 (Flk1) and VEGF receptor 3 (Flt4)0 aVascular endothelial growth factor D VEGFD is a ligand for the t c1998/Jan/ a548 - 530 v953 aWe have identified a member of the VEGF family by computer-based homology searching and have designated it VEGF-D. VEGF-D is most closely related to VEGF-C by virtue of the presence of N- and C-terminal extensions that are not found in other VEGF family members. In adult human tissues, VEGF-D mRNA is most abundant in heart, lung, skeletal muscle, colon, and small intestine. Analyses of VEGF-D receptor specificity revealed that VEGF-D is a ligand for both VEGF receptors (VEGFRs) VEGFR-2 (Flk1) and VEGFR-3 (Flt4) and can activate these receptors. However. VEGF-D does not bind to VEGFR-1. Expression of a truncated derivative of VEGF-D demonstrated that the receptor-binding capacities reside in the portion of the molecule that is most closely related in primary structure to other VEGF family members and that corresponds to the mature form of VEGF-C. In addition, VEGF-D is a mitogen for endothelial cells. The structural and functional similarities between VEGF-D and VEGF-C define a subfamily of the VEGFs.1 aAchen, M G1 aJeltsch, M1 aKukk, E1 aMäkinen, T1 aVitali, A1 aWilks, A F1 aAlitalo, K1 aStacker, S A uhttp://view.ncbi.nlm.nih.gov/pubmed/943522902306nas a2200181 4500008004100000245020900041210006900250260001400319300001300333490000800346520164400354100001601998700001902014700001502033700001302048700001502061856004802076 1998 eng d00aVascular endothelial growth factor (VEGF)-C synergizes with basic fibroblast growth factor and VEGF in the induction of angiogenesis in vitro and alters endothelial cell extracellular proteolytic activity0 aVascular endothelial growth factor VEGFC synergizes with basic f c1998/Dec/ a439 - 520 v1773 aVascular endothelial growth factor-C (VEGF-C) is a recently characterized member of the VEGF family of angiogenic polypeptides. We demonstrate here that VEGF-C is angiogenic in vitro when added to bovine aortic or lymphatic endothelial (BAE and BLE) cells but has little or no effect on bovine microvascular endothelial (BME) cells. As reported previously for VEGF, VEGF-C and basic fibroblast growth factor (bFGF) induced a synergistic in vitro angiogenic response in all three cells lines. Unexpectedly, VEGF and VEGF-C also synergized in the in vitro angiogenic response when assessed on BAE cells. Characterization of VEGF receptor (VEGFR) expression revealed that BME, BAE, and BLE cell lines express VEGFR-1 and -2, whereas of the three cell lines assessed, only BAE cells express VEGFR-3. We also demonstrate that VEGF-C increases plasminogen activator (PA) activity in the three bovine endothelial cell lines and that this is accompanied by a concomitant increase in PA inhibitor-1. Addition of alpha2-antiplasmin to BME cells co-treated with bFGF and VEGF-C partially inhibited collagen gel invasion. These results demonstrate, first, that by acting in concert with bFGF or VEGF, VEGF-C has a potent synergistic effect on the induction of angiogenesis in vitro and, second, that like VEGF and bFGF, VEGF-C is capable of altering endothelial cell extracellular proteolytic activity. These observations also highlight the notion of context, i.e., that the activity of an angiogenesis-regulating cytokine depends on the presence and concentration of other cytokines in the pericellular environment of the responding endothelial cell.1 aPepper, M S1 aMandriota, S J1 aJeltsch, M1 aKumar, V1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/980815202100nas a2200145 4500008004500000245004500045210004300090260003700133490002000170520168300190653001101873653001101884100002101895856003801916 1997 Engldsh 00aFunctional Analysis of VEGF-B and VEGF-C0 aFunctional Analysis of VEGFB and VEGFC aHelsinkibUniveristy of Helsinki0 vMaster's Thesis3 aVascular endothelial growth factor (VEGF) is an important regulator of endothelial cell proliferation and migration dur- ing embryonic vasculogenesis and angiogenesis as well as in pathological angiogenesis. The recently cloned new factors structurally homologous to VEGF were designated as VEGF-B/VRF and VEGF-C/VRP. The receptor for VEGF-B is unknown. VEGF-C is the ligand for FLT4, a receptor tyrosine kinase whose expression becomes restricted largely to lymphatic endothelia during development and that is related to VEGF receptors FLT1 and KDR. In this study keratin 14-promoter-directed VEGF-C overexpression in the basal epidermis of transgenic mice was capa- ble of promoting an abundant growth of extensive lymphatic-like vessel structures in the dermis, including large vessel lacunae resembling in their histopathology the human condition known as lymphangioma. Thus, VEGF-C appears to induce selective angiogenesis of the lymphatic vessels in vivo. In contrast, preliminary data on mice, which overexpress VEGF-B under the same promoter, does not yet allow us draw any conclusions about its possible biological function. Recombinant biologically active human VEGF-C was produced using the baculovirus system. Unpurified and purified VEGF-C were used to confirm the interaction of VEGF-C with KDR, a fact recently missed by others. The recombinant protein is going to be used in a large number of future experiments. The production of VEGF-B seems to be intrinsically difficult in non-mammalian cells. Although quantitatively satisfying results could not be obtained yet, the purified growth factor will be used in experiments to identify its receptor.10aVEGF-B10aVEGF-C1 aJeltsch, Michael uhttp://urn.fi/URN:NBN:fi-fe97734701905nas a2200217 4500008004100000245009100041210006900132260001400201300001500215490000800230520128700238100001401525700001201539700001301551700001501564700001601579700001501595700001401610700001501624856004801639 1997 eng d00aGenomic organization of human and mouse genes for vascular endothelial growth factor C0 aGenomic organization of human and mouse genes for vascular endot c1997/Oct/ a25176 - 830 v2723 aWe report here the cloning and characterization of human and mouse genes for vascular endothelial growth factor C (VEGF-C), a newly isolated member of the vascular endothelial growth factor/platelet-derived growth factor (VEGF/PDGF) family. Both VEGF-C genes comprise over 40 kilobase pairs of genomic DNA and consist of seven exons, all containing coding sequences. The VEGF homology domain of VEGF-C is encoded by exons 3 and 4. Exons 5 and 7 encode cysteine-rich motifs of the type C6C10CRC, and exon 6 encodes additional C10CXCXC motifs typical of a silk protein. A putative alternatively spliced rare RNA form lacking exon 4 was identified in human fibrosarcoma cells, and a major transcription start site was located in the human VEGF-C gene 523 base pairs upstream of the translation initiation codon. The upstream promoter sequences contain conserved putative binding sites for Sp-1, AP-2, and NF-kappaB transcription factors but no TATA box, and they show promoter activity when transfected into cells. The VEGF-C gene structure is thus assembled from exons encoding propeptides and distinct cysteine-rich domains in addition to the VEGF homology domain, and it shows both similarities and distinct differences in comparison with other members of the VEGF/PDGF gene family.1 aChilov, D1 aKukk, E1 aTaira, S1 aJeltsch, M1 aKaukonen, J1 aPalotie, A1 aJoukov, V1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/931213001376nas a2200241 4500008004100000245006300041210006200104260001400166300001300180490000800193520074000201100001500941700001700956700001400973700001200987700001300999700001501012700001401027700001601041700001401057700001501071856004801086 1997 eng d00aHyperplasia of lymphatic vessels in VEGF-C transgenic mice0 aHyperplasia of lymphatic vessels in VEGFC transgenic mice c1997/May/ a1423 - 50 v2763 aNo 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.1 aJeltsch, M1 aKaipainen, A1 aJoukov, V1 aMeng, X1 aLakso, M1 aRauvala, H1 aSwartz, M1 aFukumura, D1 aJain, R K1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/916201101810nas a2200229 4500008004100000245008100041210006900122260001400191300001500205490000700220520117000227100001401397700001301411700001301424700001501437700002201452700001101474700001501485700001701500700001501517856004801532 1997 eng d00aProteolytic processing regulates receptor specificity and activity of VEGF-C0 aProteolytic processing regulates receptor specificity and activi c1997/Jul/ a3898 - 9110 v163 aThe recently identified vascular endothelial growth factor C (VEGF-C) belongs to the platelet-derived growth factor (PDGF)/VEGF family of growth factors and is a ligand for the endothelial-specific receptor tyrosine kinases VEGFR-3 and VEGFR-2. The VEGF homology domain spans only about one-third of the cysteine-rich VEGF-C precursor. Here we have analysed the role of post-translational processing in VEGF-C secretion and function, as well as the structure of the mature VEGF-C. The stepwise proteolytic processing of VEGF-C generated several VEGF-C forms with increased activity towards VEGFR-3, but only the fully processed VEGF-C could activate VEGFR-2. Recombinant 'mature' VEGF-C made in yeast bound VEGFR-3 (K[D] = 135 pM) and VEGFR-2 (K[D] = 410 pM) and activated these receptors. Like VEGF, mature VEGF-C increased vascular permeability, as well as the migration and proliferation of endothelial cells. Unlike other members of the PDGF/VEGF family, mature VEGF-C formed mostly non-covalent homodimers. These data implicate proteolytic processing as a regulator of VEGF-C activity, and reveal novel structure-function relationships in the PDGF/VEGF family.1 aJoukov, V1 aSorsa, T1 aKumar, V1 aJeltsch, M1 aClaesson-Welsh, L1 aCao, Y1 aSaksela, O1 aKalkkinen, N1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/923380000565nas a2200205 4500008004100000245005800041210005600099260001400155300001200169490000800181100001400189700001700203700001500220700001600235700001600251700001300267700001600280700001500296856004800311 1997 eng d00aVascular endothelial growth factors VEGF-B and VEGF-C0 aVascular endothelial growth factors VEGFB and VEGFC c1997/Nov/ a211 - 50 v1731 aJoukov, V1 aKaipainen, A1 aJeltsch, M1 aPajusola, K1 aOlofsson, B1 aKumar, V1 aEriksson, U1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/936552402815nas a2200217 4500008004100000245013100041210006900172260001400241300001300255490000800268520214700276100001202423700001702435700002002452700001802472700001502490700001402505700001502519700001502534856004802549 1997 eng d00aVEGF and VEGF-C: specific induction of angiogenesis and lymphangiogenesis in the differentiated avian chorioallantoic membrane0 aVEGF and VEGFC specific induction of angiogenesis and lymphangio c1997/Aug/ a96 - 1090 v1883 aThe lymphangiogenic potency of endothelial growth factors has not been studied to date. This is partially due to the lack of in vivo lymphangiogenesis assays. We have studied the lymphatics of differentiated avian chorioallantoic membrane (CAM) using microinjection of Mercox resin, semi- and ultrathin sectioning, immunohistochemical detection of fibronectin and alpha-smooth muscle actin, and in situ hybridization with VEGFR-2 and VEGFR-3 probes. CAM is drained by lymphatic vessels which are arranged in a regular pattern. Arterioles and arteries are accompanied by a pair of interconnected lymphatics and form a plexus around bigger arteries. Veins are also associated with lymphatics, particularly larger veins, which are surrounded by a lymphatic plexus. The lymphatics are characterized by an extremely thin endothelial lining, pores, and the absence of a basal lamina. Patches of the extracellular matrix can be stained with an antibody against fibronectin. Lymphatic endothelial cells of differentiated CAM show ultrastructural features of this cell type. CAM lymphatics do not possess mediae. In contrast, the lymphatic trunks of the umbilical stalk are invested by a single but discontinuous layer of smooth muscle cells. CAM lymphatics express VEGFR-2 and VEGFR-3. Both the regular pattern and the typical structure of these lymphatics suggest that CAM is a suitable site to study the in vivo effects of potential lymphangiogenic factors. We have studied the effects of VEGF homo- and heterodimers, VEGF/PlGF heterodimers, and PlGF and VEGF-C homodimers on Day 13 CAM. All the growth factors containing at least one VEGF chain are angiogenic but do not induce lymphangiogenesis. PlGF-1 and PlGF-2 are neither angiogenic nor lymphangiogenic. VEGF-C is the first lymphangiogenic factor and seems to be highly chemoattractive for lymphatic endothelial cells. It induces proliferation of lymphatic endothelial cells and development of new lymphatic sinuses which are directed immediately beneath the chorionic epithelium. Our studies show that VEGF and VEGF-C are specific angiogenic and lymphangiogenic growth factors, respectively.1 aOh, S J1 aJeltsch, M M1 aBirkenhäger, R1 aMcCarthy, J E1 aWeich, H A1 aChrist, B1 aAlitalo, K1 aWilting, J uhttp://view.ncbi.nlm.nih.gov/pubmed/924551502083nas a2200205 4500008004100000245011700041210006900158260001400227300001400241490000800255520146100263100001201724700001901736700001301755700001701768700001501785700001401800700001501814856004801829 1996 eng d00aVEGF-C receptor binding and pattern of expression with VEGFR-3 suggests a role in lymphatic vascular development0 aVEGFC receptor binding and pattern of expression with VEGFR3 sug c1996/Dec/ a3829 - 370 v1223 aThe 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.1 aKukk, E1 aLymboussaki, A1 aTaira, S1 aKaipainen, A1 aJeltsch, M1 aJoukov, V1 aAlitalo, K uhttp://view.ncbi.nlm.nih.gov/pubmed/9012504