ABSTRACT
Disruption of the precise balance of positive and negative molecular regulators of blood and lymphatic vessel growth can lead to myriad diseases. Although dozens of natural inhibitors of hemangiogenesis have been identified, an endogenous selective inhibitor of lymphatic vessel growth has not to our knowledge been previously described. We report the existence of a splice variant of the gene encoding vascular endothelial growth factor receptor-2 (Vegfr-2) that encodes a secreted form of the protein, designated soluble Vegfr-2 (sVegfr-2), that inhibits developmental and reparative lymphangiogenesis by blocking Vegf-c function. Tissue-specific loss of sVegfr-2 in mice induced, at birth, spontaneous lymphatic invasion of the normally alymphatic cornea and hyperplasia of skin lymphatics without affecting blood vasculature. Administration of sVegfr-2 inhibited lymphangiogenesis but not hemangiogenesis induced by corneal suture injury or transplantation, enhanced corneal allograft survival and suppressed lymphangioma cellular proliferation. Naturally occurring sVegfr-2 thus acts as a molecular uncoupler of blood and lymphatic vessels; modulation of sVegfr-2 might have therapeutic effects in treating lymphatic vascular malformations, transplantation rejection and, potentially, tumor lymphangiogenesis and lymphedema (pages 993-994).
Subject(s)
Lymphangiogenesis/genetics , Lymphangiogenesis/physiology , Lymphatic Vessels/physiology , Vascular Endothelial Growth Factor Receptor-2/genetics , Vascular Endothelial Growth Factor Receptor-2/physiology , Alternative Splicing , Animals , Animals, Newborn , Base Sequence , Cornea/blood supply , Cornea/growth & development , Cornea/metabolism , DNA, Complementary/genetics , Humans , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , Mice, Mutant Strains , Molecular Sequence Data , Vascular Endothelial Growth Factor C/antagonists & inhibitors , Vascular Endothelial Growth Factor C/physiology , Vascular Endothelial Growth Factor Receptor-2/deficiencyABSTRACT
Age-related macular degeneration (AMD), a leading cause of blindness worldwide, is as prevalent as cancer in industrialized nations. Most blindness in AMD results from invasion of the retina by choroidal neovascularisation (CNV). Here we show that the eosinophil/mast cell chemokine receptor CCR3 is specifically expressed in choroidal neovascular endothelial cells in humans with AMD, and that despite the expression of its ligands eotaxin-1, -2 and -3, neither eosinophils nor mast cells are present in human CNV. Genetic or pharmacological targeting of CCR3 or eotaxins inhibited injury-induced CNV in mice. CNV suppression by CCR3 blockade was due to direct inhibition of endothelial cell proliferation, and was uncoupled from inflammation because it occurred in mice lacking eosinophils or mast cells, and was independent of macrophage and neutrophil recruitment. CCR3 blockade was more effective at reducing CNV than vascular endothelial growth factor A (VEGF-A) neutralization, which is in clinical use at present, and, unlike VEGF-A blockade, is not toxic to the mouse retina. In vivo imaging with CCR3-targeting quantum dots located spontaneous CNV invisible to standard fluorescein angiography in mice before retinal invasion. CCR3 targeting might reduce vision loss due to AMD through early detection and therapeutic angioinhibition.
Subject(s)
Macular Degeneration/diagnosis , Macular Degeneration/therapy , Receptors, CCR3/antagonists & inhibitors , Receptors, CCR3/metabolism , Animals , Cell Movement , Cell Proliferation , Cells, Cultured , Chemokine CCL11/antagonists & inhibitors , Chemokine CCL11/metabolism , Chemokine CCL24/antagonists & inhibitors , Chemokine CCL24/metabolism , Chemokine CCL26 , Chemokines, CC/antagonists & inhibitors , Chemokines, CC/metabolism , Choroid/blood supply , Choroid/cytology , Choroid/metabolism , Choroidal Neovascularization/diagnosis , Choroidal Neovascularization/metabolism , Disease Models, Animal , Endothelial Cells/cytology , Endothelial Cells/metabolism , Humans , Inflammation , Leukocytes , Ligands , Macular Degeneration/metabolism , Mice , Mice, Inbred C57BL , Quantum Dots , Receptors, CCR3/analysis , Receptors, CCR3/genetics , Receptors, CCR3/immunology , Retina/drug effects , Retina/pathology , Vascular Endothelial Growth Factor A/antagonists & inhibitors , Vascular Endothelial Growth Factor A/immunologyABSTRACT
Neovascularization in response to tissue injury consists of the dual invasion of blood (hemangiogenesis) and lymphatic (lymphangiogenesis) vessels. We reported recently that 21-nt or longer small interfering RNAs (siRNAs) can suppress hemangiogenesis in mouse models of choroidal neovascularization and dermal wound healing independently of RNA interference by directly activating Toll-like receptor 3 (TLR3), a double-stranded RNA immune receptor, on the cell surface of blood endothelial cells. Here, we show that a 21-nt nontargeted siRNA suppresses both hemangiogenesis and lymphangiogenesis in mouse models of neovascularization induced by corneal sutures or hindlimb ischemia as efficiently as a 21-nt siRNA targeting vascular endothelial growth factor-A. In contrast, a 7-nt nontargeted siRNA, which is too short to activate TLR3, does not block hemangiogenesis or lymphangiogenesis in these models. Exposure to 21-nt siRNA, which we demonstrate is not internalized unless cell-permeating moieties are used, triggers phosphorylation of cell surface TLR3 on lymphatic endothelial cells and induces apoptosis. These findings introduce TLR3 activation as a method of jointly suppressing blood and lymphatic neovascularization and simultaneously raise new concerns about the undesirable effects of siRNAs on both circulatory systems.
