ABSTRACT
The importance of the lymphangiogenic factor VEGF-D and its receptor VEGFR-3 in early lymphatic development remains largely unresolved. We therefore investigated their role in Xenopus laevis tadpoles, a small animal model allowing chemicogenetic dissection of developmental lymphangiogenesis. Single morpholino antisense oligo knockdown of xVEGF-D did not affect lymphatic commitment, but transiently impaired lymphatic endothelial cell (LEC) migration. Notably, combined knockdown of xVEGF-D with xVEGF-C at suboptimal morpholino concentrations resulted in more severe migration defects and lymphedema formation than the corresponding single knockdowns. Knockdown of VEGFR-3 or treatment with the VEGFR-3 inhibitor MAZ51 similarly impaired lymph vessel formation and function and caused pronounced edema. VEGFR-3 silencing by morpholino knockdown, MAZ51 treatment, or xVEGF-C/D double knockdown also resulted in dilation and dysfunction of the lymph heart. These findings document a critical role of VEGFR-3 in embryonic lymphatic development and function, and reveal a previously unrecognized modifier role of VEGF-D in the regulation of embryonic lymphangiogenesis in frog embryos.
Subject(s)
Lymphangiogenesis/physiology , Vascular Endothelial Growth Factor D/physiology , Vascular Endothelial Growth Factor Receptor-3/physiology , Xenopus Proteins/physiology , Xenopus laevis/growth & development , Xenopus laevis/physiology , Animals , Gene Silencing , Larva/growth & development , Lymphangiogenesis/genetics , Lymphatic Vessels/abnormalities , Lymphatic Vessels/embryology , Oligodeoxyribonucleotides, Antisense/genetics , Vascular Endothelial Growth Factor D/antagonists & inhibitors , Vascular Endothelial Growth Factor D/genetics , Vascular Endothelial Growth Factor Receptor-3/antagonists & inhibitors , Vascular Endothelial Growth Factor Receptor-3/genetics , Xenopus Proteins/genetics , Xenopus laevis/embryology , Xenopus laevis/geneticsABSTRACT
OBJECTIVE: The endogenous role of the VEGF family member vascular endothelial growth factor-B (VEGF-B) in pathological angiogenesis remains unclear. METHODS AND RESULTS: We studied the role of VEGF-B in various models of pathological angiogenesis using mice lacking VEGF-B (VEGF-B(-/-)) or overexpressing VEGF-B(167). After occlusion of the left coronary artery, VEGF-B deficiency impaired vessel growth in the ischemic myocardium whereas, in wild-type mice, VEGF-B(167) overexpression enhanced revascularization of the infarct and ischemic border zone. By contrast, VEGF-B deficiency did not affect vessel growth in the wounded skin, hypoxic lung, ischemic retina, or ischemic limb. Moreover, VEGF-B(167) overexpression failed to enhance vascular growth in the skin or ischemic limb. CONCLUSIONS: VEGF-B appears to have a relatively restricted angiogenic activity in the ischemic heart. These insights might offer novel therapeutic opportunities.
Subject(s)
Coronary Vessels/metabolism , Ischemia/metabolism , Myocardial Ischemia/metabolism , Myocardium/metabolism , Neovascularization, Physiologic , Vascular Endothelial Growth Factor B/metabolism , Angiogenesis Inducing Agents/metabolism , Animals , Coronary Vessels/drug effects , Coronary Vessels/physiopathology , Disease Models, Animal , Gene Transfer Techniques , Genetic Therapy/methods , Hindlimb , Ischemia/pathology , Ischemia/physiopathology , Ischemia/therapy , Lung/blood supply , Mice , Mice, Inbred C57BL , Mice, Knockout , Mice, Nude , Muscle, Skeletal/blood supply , Myocardial Ischemia/pathology , Myocardial Ischemia/physiopathology , Myocardial Ischemia/therapy , Myocardium/pathology , Neovascularization, Physiologic/drug effects , Recombinant Proteins/metabolism , Retinal Vessels/metabolism , Skin/blood supply , Up-Regulation , Vascular Endothelial Growth Factor B/administration & dosage , Vascular Endothelial Growth Factor B/deficiency , Vascular Endothelial Growth Factor B/genetics , Vascular Endothelial Growth Factor Receptor-1/metabolismABSTRACT
Lymph vessels control fluid homeostasis, immunity and metastasis. Unraveling the molecular basis of lymphangiogenesis has been hampered by the lack of a small animal model that can be genetically manipulated. Here, we show that Xenopus tadpoles develop lymph vessels from lymphangioblasts or, through transdifferentiation, from venous endothelial cells. Lymphangiography showed that these lymph vessels drain lymph, through the lymph heart, to the venous circulation. Morpholino-mediated knockdown of the lymphangiogenic factor Prox1 caused lymph vessel defects and lymphedema by impairing lymphatic commitment. Knockdown of vascular endothelial growth factor C (VEGF-C) also induced lymph vessel defects and lymphedema, but primarily by affecting migration of lymphatic endothelial cells. Knockdown of VEGF-C also resulted in aberrant blood vessel formation in tadpoles. This tadpole model offers opportunities for the discovery of new regulators of lymphangiogenesis.
