RESUMEN
Angiotensin-II (Ang-II) is a well-established mediator of vascular remodeling. The multifunctional calcium-calmodulin-dependent kinase II (CaMKII) is activated by Ang-II and regulates Erk1/2 and Akt-dependent signaling in cultured smooth muscle cells in vitro. Its role in Ang-II-dependent vascular remodeling in vivo is far less defined. Using a model of transgenic CaMKII inhibition selectively in smooth muscle cells, we found that CaMKII inhibition exaggerated remodeling after chronic Ang-II treatment and agonist-dependent vasoconstriction in second-order mesenteric arteries. These findings were associated with increased mRNA and protein expression of smooth muscle structural proteins. As a potential mechanism, CaMKII reduced serum response factor-dependent transcriptional activity. In summary, our findings identify CaMKII as an important regulator of smooth muscle function in Ang-II hypertension in vivo.
Asunto(s)
Angiotensina II/metabolismo , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Miocitos del Músculo Liso/metabolismo , Remodelación Vascular/fisiología , Animales , Femenino , Masculino , Arterias Mesentéricas/metabolismo , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Músculo Liso Vascular/citología , Músculo Liso Vascular/metabolismo , ARN Mensajero/metabolismo , Vasoconstricción/fisiologíaRESUMEN
OBJECTIVE: Emerging evidence suggests that methionine oxidation can directly affect protein function and may be linked to cardiovascular disease. The objective of this study was to define the role of the methionine sulfoxide reductase A (MsrA) in models of vascular disease and identify its signaling pathways. APPROACH AND RESULTS: MsrA was readily identified in all layers of the vascular wall in human and murine arteries. Deletion of the MsrA gene did not affect atherosclerotic lesion area in apolipoprotein E-deficient mice and had no significant effect on susceptibility to experimental thrombosis after photochemical injury. In contrast, the neointimal area after vascular injury caused by complete ligation of the common carotid artery was significantly greater in MsrA-deficient than in control mice. In aortic vascular smooth muscle cells lacking MsrA, cell proliferation was significantly increased because of accelerated G1/S transition. In parallel, cyclin D1 protein and cdk4/cyclin D1 complex formation and activity were increased in MsrA-deficient vascular smooth muscle cell, leading to enhanced retinoblastoma protein phosphorylation and transcription of E2F. Finally, MsrA-deficient vascular smooth muscle cell exhibited greater activation of extracellular signal-regulated kinase 1/2 that was caused by increased activity of the Ras/Raf/mitogen-activated protein kinase signaling pathway. CONCLUSIONS: Our findings implicate MsrA as a negative regulator of vascular smooth muscle cell proliferation and neointimal hyperplasia after vascular injury through control of the Ras/Raf/mitogen-activated protein kinase kinase/extracellular signal-regulated kinase 1/2 signaling pathway.
Asunto(s)
Enfermedades de la Aorta/enzimología , Aterosclerosis/enzimología , Traumatismos de las Arterias Carótidas/enzimología , Eliminación de Gen , Metionina Sulfóxido Reductasas/deficiencia , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Neointima , Transducción de Señal , Trombosis/enzimología , Animales , Aorta/enzimología , Aorta/patología , Enfermedades de la Aorta/genética , Enfermedades de la Aorta/patología , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Aterosclerosis/genética , Aterosclerosis/patología , Arterias Carótidas/enzimología , Arterias Carótidas/patología , Traumatismos de las Arterias Carótidas/genética , Traumatismos de las Arterias Carótidas/patología , Ciclo Celular , Proteínas de Ciclo Celular/metabolismo , Movimiento Celular , Proliferación Celular , Células Cultivadas , Modelos Animales de Enfermedad , Femenino , Humanos , Hiperplasia , Masculino , Metionina Sulfóxido Reductasas/genética , Ratones Endogámicos C57BL , Ratones Noqueados , Miocitos del Músculo Liso/enzimología , Miocitos del Músculo Liso/patología , Trombosis/sangre , Trombosis/genética , Factores de Tiempo , Quinasas raf/metabolismo , Proteínas ras/metabolismoRESUMEN
BACKGROUND: Multifunctional calcium/calmodulin-dependent kinase II (CaMKII) is activated by angiotensin II (Ang II) in cultured vascular smooth muscle cells (VSMCs), but its function in experimental hypertension has not been explored. The aim of this study was to determine the impact of CaMKII inhibition selectively in VSMCs on Ang II hypertension. METHODS AND RESULTS: Transgenic expression of a CaMKII peptide inhibitor in VSMCs (TG SM-CaMKIIN model) reduced the blood pressure response to chronic Ang II infusion. The aortic depressor nerve activity was reset in hypertensive versus normotensive wild-type animals but not in TG SM-CaMKIIN mice, suggesting that changes in baroreceptor activity account for the blood pressure difference between genotypes. Accordingly, aortic pulse wave velocity, a measure of arterial wall stiffness and a determinant of baroreceptor activity, increased in hypertensive versus normotensive wild-type animals but did not change in TG SM-CaMKIIN mice. Moreover, examination of blood pressure and heart rate under ganglionic blockade revealed that VSMC CaMKII inhibition abolished the augmented efferent sympathetic outflow and renal and splanchnic nerve activity in Ang II hypertension. Consequently, we hypothesized that VSMC CaMKII controls baroreceptor activity by modifying arterial wall remodeling in Ang II hypertension. Gene expression analysis in aortas from normotensive and Ang II-infused mice revealed that TG SM-CaMKIIN aortas were protected from Ang II-induced upregulation of genes that control extracellular matrix production, including collagen. VSMC CaMKII inhibition also strongly altered the expression of muscle contractile genes under Ang II. CONCLUSIONS: CaMKII in VSMCs regulates blood pressure under Ang II hypertension by controlling structural gene expression, wall stiffness, and baroreceptor activity.
Asunto(s)
Angiotensina II/farmacología , Antihipertensivos/farmacología , Aorta/efectos de los fármacos , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/antagonistas & inhibidores , Hipertensión/tratamiento farmacológico , Músculo Liso Vascular/efectos de los fármacos , Presorreceptores/efectos de los fármacos , Remodelación Vascular/efectos de los fármacos , Animales , Aorta/fisiología , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/fisiología , Ecocardiografía , Hipertensión/inducido químicamente , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Músculo Liso Vascular/fisiopatología , Norepinefrina/sangre , Análisis de Secuencia por Matrices de Oligonucleótidos , Presorreceptores/fisiología , Remodelación Vascular/fisiologíaRESUMEN
Activation of the Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) and reactive oxygen species (ROS) promote neointimal hyperplasia after vascular injury. CaMKII can be directly activated by ROS through oxidation. In this study, we determined whether abolishing the oxidative activation site of CaMKII alters vascular smooth muscle cell (VCMC) proliferation, migration and apoptosis in vitro and neointimal formation in vivo. VSMC isolated from a knock-in mouse with oxidation-resistant CaMKIIδ (CaMKII M2V) displayed similar proliferation but decreased migration and apoptosis. Surprisingly, ROS production and expression of the NADPH oxidase subunits p47 and p22 were decreased in M2V VSMC, whereas superoxide dismutase 2 protein expression was upregulated. In vivo, after carotid artery ligation, no differences in neointimal size or remodeling were observed. In contrast to VSMC, CaMKII expression and autonomous activity were significantly higher in M2V compared to WT carotid arteries, suggesting that an autoregulatory mechanism determines CaMKII activity in vivo. Our findings demonstrate that preventing oxidative activation of CaMKII decreases migration and apoptosis in vitro and suggest that CaMKII regulates ROS production. Our study presents novel evidence that CaMKII expression in vivo is regulated by a negative feedback loop following oxidative activation.
Asunto(s)
Apoptosis/fisiología , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/metabolismo , Músculo Liso Vascular/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Animales , Proteína Quinasa Tipo 2 Dependiente de Calcio Calmodulina/genética , Arterias Carótidas/metabolismo , Proliferación Celular , Grupo Citocromo b/metabolismo , Femenino , Regulación de la Expresión Génica , Técnicas de Sustitución del Gen , Masculino , Ratones , Músculo Liso Vascular/citología , NADPH Oxidasas/metabolismo , Neointima/metabolismo , Oxidación-Reducción , Superóxido Dismutasa/metabolismoRESUMEN
OBJECTIVE: Sustained hemodynamic stress mediated by high blood flow promotes arteriogenesis, the outward remodeling of existing arteries. Here, we examined whether Ca²âº/calmodulin-dependent kinase II (CaMKII) regulates arteriogenesis. METHODS AND RESULTS: Ligation of the left common carotid led to an increase in vessel diameter and perimeter of internal and external elastic lamina in the contralateral, right common carotid. Deletion of CaMKIIδ (CaMKIIδ-/-) abolished this outward remodeling. Carotid ligation increased CaMKII expression and was associated with oxidative activation of CaMKII in the adventitia and endothelium. Remodeling was abrogated in a knock-in model in which oxidative activation of CaMKII is abolished. Early after ligation, matrix metalloproteinase 9 (MMP9) was robustly expressed in the adventitia of right carotid arteries of WT but not CaMKIIδ-/- mice. MMP9 mainly colocalized with adventitial macrophages. In contrast, we did not observe an effect of CaMKIIδ deficiency on other proposed mediators of arteriogenesis such as expression of adhesion molecules or smooth muscle proliferation. Transplantation of WT bone marrow into CaMKIIδ-/- mice normalized flow-mediated remodeling. CONCLUSION: CaMKIIδ is activated by oxidation under high blood flow conditions and is required for flow-mediated remodeling through a mechanism that includes increased MMP9 expression in bone marrow-derived cells invading the arterial wall.