Lymphangiogenesis and Cancer MetastasisSlide Number 2Lymphatic System: simple factsHuman Lymphatic SystemPhysiology of lymphatic system Lymphatic flow Unique features of lymphatic vesselsLymphatic vs blood capillariesSlide Number 9Slide Number 10Lymphatic SpecificationLymphatic Vascular Development IIISlide Number 13Slide Number 14Pathological lymphangiogenesisSlide Number 16Slide Number 17Routes of Cancer MetastasisUCLA Small Animal Imaging Core:Luminescent Intensity Correlates with Extent of MetastasesSlide Number 21Slide Number 22Slide Number 23Slide Number 24Slide Number 25Soluble VEGF-R3 Blocks Metastasis to LN and LungLymphangiogenic Environment in the Prostate on Nodal Metastasis of LAPC-9 TumorsSlide Number 28Slide Number 29Slide Number 30Slide Number 31Slide Number 32Slide Number 33Slide Number 34Slide Number 35Slide Number 36Slide Number 37Slide Number 38Slide Number 39Lily Wu, M.D., Ph.D.Associate ProfessorDepartment of Molecular & Medical Pharmacology, Department of Urology,UCLA School of MedicineMCDB 224April 20, 2010Lymphangiogenesis and Cancer MetastasisToday’s Topics• Biology of lymphatic system• Lymphangiogenesis & metastasis• Review of a primary research articleLymphatic System: simple facts• exist in vertebrates, fish, amphibians, reptiles and mammals•in human, lymphatic vessels are found in all vascularizedtissues, except bone marrow and brain • functions to maintain tissue fluid homeostasis, immune cell trafficking, and intestinal lipid absorption. • returns approximately 1-2 liters of interstitial fluid back into the venous circulationMain%functions%:%%- to%maintain%fluid%balance%(collecting%interstitial%fluid,%macromolecules,% cells%from%tissue%and%return%to%the %blood)%- key%immune%function%(transport%lymphocytes,%antigen%presentation%in%lymph%nodes)%- to%absorb%lipids% from%the%intestine%and%transport%them%to%th e %bloo d%Human Lymphatic SystemOliver Nat Rev Imm 4:35, 2004Alitalo et al. Nature 438: 946, 2005Cueni1 & Detmar J Invest Derm 126:2167, 2006Tammela & Alitalo Cell 140:460, 2010Physiology of lymphatic system Pc =capillary pressure; Pt =tissue pressure; ʌS =plasma oncoticpressure; ʌW = tissue oncotic pressure; IFV = interstitial fluid volume; Kf = capillary filtration coefficient; ı solute reflection coefficient; Jv = transcapillary water fluxAdapted from M Witte et al. Can Met Rev 2006Tissue fluid homeostasis:• absence of lymphatic vessels is incompatible with life • lymphatic system likely evolves along side with blood circulatory system• blood pressure and other circulatory parameters forces plasma fluid to filtrate out of the capillary bed into the interstitial space•approx 90% extravasated fluid is reabsorbed by the venous capillaries, with 10% collected by the lymphaticsLymphatic flow • tissue fluid first collected into initial lymphatic capillaries, which are blind-ended, thin-walled vessels ~30-80P• from capillaries, lymph moves to precollectors and collecting vessels, filters through lymph nodes and collects into the thoracic ducts, which then drains into the venous circulation via the subclavian vein• propulsion of lymph is aided by intrinsic contractility of smooth muscle cells lining the collecting lymphatics, the bileaflet valves, contraction of skeletal muscles and arterial pulsation. Oliver Nat Rev Imm 4:35, 2004Alitalo et al. Nature 438: 946, 2005Unique features of lymphatic vessels• Discontinuous capillaries, seal by overlapping oak-leaf shaped LECs (A). • LECs connected to ECM by anchoring filaments, • During muscular contraction of tissue swelling, anchoring filaments can be taut, resulting in opening of lymphatic vessel lumen, and increased uptake of tissue fluid. • Valves prevents the back flow of lymph. Smooth muscle actin (red); lectin stained valve (green)Lymphatic capillaries are amendable for entry of macromolecules and cells• have larger diameter, • discontinuous, more flexible• lacks tight junction• lack pericytesLymphatic vs blood capillariesLymphatic (green), SMA (red, BV)Arrow, lymphatic valve; Arrowhead, blood vessel.Makinen et al. Cell Mol Life Sci 2007VEGF-CSabin,%1902,%pig%embryoRemarkable accurate description of origin of lymphatic system, >100 years agoHallmark Studies of Lymphatic SystemKaipainen, A. et al. PNAS 1995; Karkkainen, M. J. et al. Nature Genet. 2000.First identification lymphatic specific expression VEGFR3 and its role in certain forms of hereditary lymphoedema.Joukov V et al. EMBO J 1996First reported VEGF-C as a ligand for the Flt4 (VEGFR-3) and KDR (VEGFR-2) receptor tyrosine kinases..Wigle, J. T. & Oliver, G. Cell 1999The first report of targeted inactivation of prox1 that leads to embryos without any lymphatic vasculature.Skobe M et al. Nature Med. 2001; Stacker SA. et al. Nature Med 2001.These papers showed the connection of tumourlymphangiogenesis and metastasisKarkkainen et al. Nat Imm 2004Demonstration of VEGF-C haploinsufficiency and the sprouting of PROX1+LECs from embryonic veins is controlled by VEGFC.Differentiation of Lymphatic System in Mouse ModelOliver, Nat Rev Imm 4:35, 2004Lymphatic Specification(A) After differentiation from angioblasts, endothelial cells undergo arterial-venous (AV) specification.(B) VEGFR-3+ embryonic veins with a subset of LYVE-1+endothelial cells in the large central veins.(C) The transcription factor SOX18 is induced in the LYVE-1+LEC precursors. (Francois e al. Nature 2008; Irrthum et al. Am J Hum Genet 2003)(D) SOX18 induces Prox1, the first marker for LEC determination. At this time VEGFR-3 is downregulated in BEC, while LECs begin to express neuropilin-2, rendering them more responsive to VEGF-C signals, leading to formation of lymph sacs lateral to the central veins.(E) LECs begin to express podoplanin, which via the CLEC-2 receptor activates the Syk tyrosine kinase in platelets. This leads to platelet aggregation, which blocks lymphatico-venous connections and helps to separate the blood and lymphatic vascular systems. (Abtahian et al Science 2003)(F) Further growth of the lymphatic network, driven by VEGF-C/VEGFR-3. (Petrova et al EMBO J 2002; Karkkainen et al. Nat Imm 2004)(G)Further differentiation of lymphatic capillaries and collecting lymphatic vessels, including formation intralumenal valves, recruit smooth muscle cells, develop interendothelialjunctions, and produce a basement membrane.Lymphatic Vascular Development IIIMakinen et al. Cell Mol Life Sci 2007Petrova et al. Nat Med