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Vascular deficiency in ALK5 knock-in mice
14th IVBM Abstracts
VEGFR-1 stimulation. Mo from diabetic individuals display a baseline activation of several VEGFR-1-related signalling pathways. This may explain the lack of Mo responsiveness to further growth factor stimulation and impaired Mo chemotactic response in diabetes.
e137
3
Hubrecht Laboratory, Netherland Inst. for Dev. Biol., Utrecht, The Netherlands 4 Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands
The migration of endothelial cells plays an important role in many physiological and pathological conditions e.g. angiogenesis, arteriogenesis, restenosis and atherosclerosis. Many aspects of cell migration are well examined, including chemotaxis, protrusion of lamellipodia and filopodia via actin polymerization and stress fibre formation and the reversible development of focal adhesions which is regulated by Rho-GTPases. However, signal transduction pathways or receptors by which the migratory stimulus is transduced from the biochemical level into biomechanical action is still not completely understood. Recently we could show that the intermediate filament Vimentin is upregulated in endothelial cells with migratory phenotype compared to non-migrating endothelial cells. In further studies we suppressed the Vimentin expression in migrating endothelial cells using siRNA-technique. The lack of Vimentin lead to a decreased migration capacity of about 50% and to a decreased proliferation of about 40% compared to control-transfected cells. Vimentin suppression was accompanied by a dephosphorylation of vasodilator-stimulated Phosphoprotein (VASP). Our results show that Vimentin filaments are a functional cytoskeletal component in endothelial cell migration and proliferation and indicates a connection of the intermediate filament network to the VASP signaling pathway.
Angiogenesis is a multistep process, which is tightly regulated by inducers and inhibitors. Recently, genetic studies in mice and genetic evidence from human vascular disorders have demonstrated that TGF-beta superfamily plays important roles in determining the fate of vascular cells. TGF-beta superfamily regulates cellular processes by binding to specific heteromeric complexes of type I and type II serine/threonine kinase receptors. The type I receptor acts downstream of the type II receptors and propagates the TGF-beta signal into cells. There are two different TGF-beta type I receptors, activin receptor-like kinase (ALK)1 and ALK5. In most cells, TGF-beta signals via ALK5, phosphorylating Smad2 and Smad3, whereas TGF-beta appears to signal through two pathways in endothelial cells (ECs); either via ALK1, prominently expressed in ECs, through activation of Smad1 and Smad5 or via the conventional ALK5 pathway. ALK5 and ALK1 are known to inhibit and stimulate angiogenic responses, respectively. Multiple studies in mice indicate that TGF-beta is a potent mediator of angiogenesis. Mice lacking ALK1, ALK5 or TGF-beta type II receptor die at E10.5 due to defect of adequate angiogenesis. Moreover, neither TGF-beta/ALK1 nor TGF-beta/ALK5 signaling can be detected when ECs isolated from ALK5 knock-out mice are stimulated with TGF-beta. These and other results have revealed that ALK5 is indispensable for the activation of both TGF-beta/ALK1 and TGF-beta/ALK5 signalings in ECs. In order to elucidate molecular mechanisms that underlie the embryonic development via TGF-beta pathway, we made ALK5 knock-in mice, which are unable to activate conventional TGF-beta/ALK5 pathway, but keep the ability to potentiate TGF-beta/ALK1 signal. Although both ALK5 knock-in and knock-out mice show embryonic lethality, the vascular phenotype of ALK5 knock-in mice appears not as severe as that of ALK5 knock-out mice. At present, we are comparing the vessel formation in yolk sac from ALK5 knock-in mice with that from ALK5 knock-out and wildtype mice. This study will give us new insight on angiogenesis regulated by TGF-beta pathway.
doi:10.1016/j.vph.2006.08.360
doi:10.1016/j.vph.2006.08.361
B17.19
B17.20
Vascular deficiency in ALK5 knock-in mice
D-glucose
doi:10.1016/j.vph.2006.08.359
B17.18 Vimentin regulates endothelial cell migration, proliferation and VASP phosphorylation Natalie Obermeyer, Petra Tiede, Wulf D. Ito University Medical Center Schleswig-Holstein, Campus Luebeck, Luebeck, Germany
Fumiko Itoh1, Susumu Itoh1, Masatsugu Ema1, Marie-Jose Goumans2, Christine L. Mummery3, Satoru Takahashi1, Peter ten Dijke4, Mitsuyasu Kato1 1
Graduate School of Comprehensive Human Sciences, Univ of Tsukuba, Tsukuba, Japan 2 Heart Lung Centre, University of Medical Centre Utrecht, Utrecht, The Netherlands
increased l-arginine transport and nitric oxide synthesis through an autocrine mechanism involving TGF-β1 and TGF-β receptor II (TβRII) in human umbilical vein endothelium
Rodrigo Vasquez, Marcelo Farias, Rody San Martin, Paola Casanello, Luis Sobrevia CMPL, Obstet Gynecol, Pontificia Universidad Catolica de Chile, Santiago, Chile
VEGFR-1 stimulation. Mo from diabetic individuals display a baseline activation of several VEGFR-1-related signalling pathways. This may explain the lack of Mo responsiveness to further growth factor stimulation and impaired Mo chemotactic response in diabetes.
e137
3
Hubrecht Laboratory, Netherland Inst. for Dev. Biol., Utrecht, The Netherlands 4 Molecular Cell Biology, Leiden University Medical Center, Leiden, The Netherlands
The migration of endothelial cells plays an important role in many physiological and pathological conditions e.g. angiogenesis, arteriogenesis, restenosis and atherosclerosis. Many aspects of cell migration are well examined, including chemotaxis, protrusion of lamellipodia and filopodia via actin polymerization and stress fibre formation and the reversible development of focal adhesions which is regulated by Rho-GTPases. However, signal transduction pathways or receptors by which the migratory stimulus is transduced from the biochemical level into biomechanical action is still not completely understood. Recently we could show that the intermediate filament Vimentin is upregulated in endothelial cells with migratory phenotype compared to non-migrating endothelial cells. In further studies we suppressed the Vimentin expression in migrating endothelial cells using siRNA-technique. The lack of Vimentin lead to a decreased migration capacity of about 50% and to a decreased proliferation of about 40% compared to control-transfected cells. Vimentin suppression was accompanied by a dephosphorylation of vasodilator-stimulated Phosphoprotein (VASP). Our results show that Vimentin filaments are a functional cytoskeletal component in endothelial cell migration and proliferation and indicates a connection of the intermediate filament network to the VASP signaling pathway.
Angiogenesis is a multistep process, which is tightly regulated by inducers and inhibitors. Recently, genetic studies in mice and genetic evidence from human vascular disorders have demonstrated that TGF-beta superfamily plays important roles in determining the fate of vascular cells. TGF-beta superfamily regulates cellular processes by binding to specific heteromeric complexes of type I and type II serine/threonine kinase receptors. The type I receptor acts downstream of the type II receptors and propagates the TGF-beta signal into cells. There are two different TGF-beta type I receptors, activin receptor-like kinase (ALK)1 and ALK5. In most cells, TGF-beta signals via ALK5, phosphorylating Smad2 and Smad3, whereas TGF-beta appears to signal through two pathways in endothelial cells (ECs); either via ALK1, prominently expressed in ECs, through activation of Smad1 and Smad5 or via the conventional ALK5 pathway. ALK5 and ALK1 are known to inhibit and stimulate angiogenic responses, respectively. Multiple studies in mice indicate that TGF-beta is a potent mediator of angiogenesis. Mice lacking ALK1, ALK5 or TGF-beta type II receptor die at E10.5 due to defect of adequate angiogenesis. Moreover, neither TGF-beta/ALK1 nor TGF-beta/ALK5 signaling can be detected when ECs isolated from ALK5 knock-out mice are stimulated with TGF-beta. These and other results have revealed that ALK5 is indispensable for the activation of both TGF-beta/ALK1 and TGF-beta/ALK5 signalings in ECs. In order to elucidate molecular mechanisms that underlie the embryonic development via TGF-beta pathway, we made ALK5 knock-in mice, which are unable to activate conventional TGF-beta/ALK5 pathway, but keep the ability to potentiate TGF-beta/ALK1 signal. Although both ALK5 knock-in and knock-out mice show embryonic lethality, the vascular phenotype of ALK5 knock-in mice appears not as severe as that of ALK5 knock-out mice. At present, we are comparing the vessel formation in yolk sac from ALK5 knock-in mice with that from ALK5 knock-out and wildtype mice. This study will give us new insight on angiogenesis regulated by TGF-beta pathway.
doi:10.1016/j.vph.2006.08.360
doi:10.1016/j.vph.2006.08.361
B17.19
B17.20
Vascular deficiency in ALK5 knock-in mice
D-glucose
doi:10.1016/j.vph.2006.08.359
B17.18 Vimentin regulates endothelial cell migration, proliferation and VASP phosphorylation Natalie Obermeyer, Petra Tiede, Wulf D. Ito University Medical Center Schleswig-Holstein, Campus Luebeck, Luebeck, Germany
Fumiko Itoh1, Susumu Itoh1, Masatsugu Ema1, Marie-Jose Goumans2, Christine L. Mummery3, Satoru Takahashi1, Peter ten Dijke4, Mitsuyasu Kato1 1
Graduate School of Comprehensive Human Sciences, Univ of Tsukuba, Tsukuba, Japan 2 Heart Lung Centre, University of Medical Centre Utrecht, Utrecht, The Netherlands
increased l-arginine transport and nitric oxide synthesis through an autocrine mechanism involving TGF-β1 and TGF-β receptor II (TβRII) in human umbilical vein endothelium
Rodrigo Vasquez, Marcelo Farias, Rody San Martin, Paola Casanello, Luis Sobrevia CMPL, Obstet Gynecol, Pontificia Universidad Catolica de Chile, Santiago, Chile