ABSTRACT
AIMS: Mutations in the ALK1 gene, coding for an endothelial-specific receptor of the transforming growth factor-ß superfamily, are the underlying cause of hereditary haemorrhagic telangiectasia type 2, but are also associated with familial pulmonary hypertension (PH). We assessed the lung vasculature of mice with a heterozygous deletion of Alk1 (Alk1(+/-)) for disease manifestations and levels of reactive O(2) species (ROS) implicated in both disorders. METHODS AND RESULTS: Several signs of PH, including elevated right ventricular (RV) systolic pressure leading to RV hypertrophy, reduced vascular density, and increased thickness and outward remodelling of pulmonary arterioles, were observed in 8- to 18-week-old Alk1(+/-) mice relative to wild-type littermate controls. Higher ROS lung levels were also documented. At 3 weeks, Alk1(+/-) mice were indistinguishable from controls and were prevented from subsequently developing PH when treated with the anti-oxidant Tempol for 6 weeks, confirming a role for ROS in pathogenesis. Levels of NADPH oxidases and superoxide dismutases were higher in adults than newborns, but unchanged in Alk1(+/-) mice vs. controls. Prostaglandin metabolites were also normal in adult Alk1(+/-) lungs. In contrast, NO production was reduced, while endothelial NO synthase (eNOS)-dependent ROS production was increased in adult Alk1(+/-) mice. Pulmonary near resistance arteries from adult Alk1(+/-) mice showed less agonist-induced force and greater acetylcholine-induced relaxation; the later was normalized by catalase or Tempol treatment. CONCLUSION: The increased pulmonary vascular remodelling in Alk1(+/-) mice leads to signs of PH and is associated with eNOS-dependent ROS production, which is preventable by anti-oxidant treatment.
Subject(s)
Activin Receptors, Type I/deficiency , Blood Pressure , Hypertension, Pulmonary/metabolism , Lung/blood supply , Oxidative Stress , Pulmonary Artery/metabolism , Reactive Oxygen Species/metabolism , Activin Receptors, Type I/genetics , Activin Receptors, Type II , Age Factors , Animals , Antihypertensive Agents/pharmacology , Antioxidants/pharmacology , Arterioles/metabolism , Arterioles/physiopathology , Blood Pressure/drug effects , Catalase/metabolism , Cyclic N-Oxides/pharmacology , Dose-Response Relationship, Drug , Heterozygote , Hypertension, Pulmonary/genetics , Hypertension, Pulmonary/pathology , Hypertension, Pulmonary/physiopathology , Hypertension, Pulmonary/prevention & control , Hypertrophy, Right Ventricular/genetics , Hypertrophy, Right Ventricular/metabolism , Hypertrophy, Right Ventricular/physiopathology , Mice , Mice, Knockout , NADPH Oxidases/metabolism , Nitric Oxide/metabolism , Nitric Oxide Synthase Type III/metabolism , Oxidative Stress/drug effects , Prostaglandin-Endoperoxide Synthases/metabolism , Pulmonary Artery/pathology , Pulmonary Artery/physiopathology , Spin Labels , Superoxide Dismutase/metabolism , Vasodilation , Vasodilator Agents/pharmacology , Ventricular Function, Right , Ventricular PressureABSTRACT
OBJECTIVE: Loss-of-function mutations in genes coding for transforming growth factor-beta/bone morphogenetic protein receptors and changes in nitric oxide(*) (NO(*)) bioavailability are associated with hereditary hemorrhagic telangiectasia and some forms of pulmonary arterial hypertension. How these abnormalities lead to seemingly disparate pulmonary pathologies remains unknown. Endoglin (Eng), a transforming growth factor-beta coreceptor, is mutated in hereditary hemorrhagic telangiectasia and involved in regulating endothelial NO(*) synthase (eNOS)-derived NO(*) production and oxidative stress. Because some patients with pulmonary arterial hypertension harbor ENG mutations leading to haplo insufficiency, we investigated the pulmonary vasculature of Eng(+/-) mice and the potential contribution of abnormal eNOS activation to pulmonary arterial hypertension. METHODS AND RESULTS: Hemodynamic, histological, and biochemical assessments and x-ray micro-CT imaging of adult Eng(+/-) mice indicated signs of pulmonary arterial hypertension including increased right ventricular systolic pressure, degeneration of the distal pulmonary vasculature, and muscularization of small arteries. These findings were absent in 3-week-old Eng(+/-) mice and were attributable to constitutively uncoupled eNOS activity in the pulmonary circulation, as evidenced by reduced eNOS/heat shock protein 90 association and increased eNOS-derived superoxide ((*)O(2)(-)) production in a BH(4)-independent manner. These changes render eNOS unresponsive to regulation by transforming growth factor-beta/bone morphogenetic protein and underlie the signs of pulmonary arterial hypertension that were prevented by Tempol. CONCLUSIONS: Adult Eng(+/-) mice acquire signs of pulmonary arterial hypertension that are attributable to uncoupled eNOS activity and increased (*)O(2)(-) production, which can be prevented by antioxidant treatment. Eng links transforming growth factor/bone morphogenetic protein receptors to the eNOS activation complex, and its reduction in the pulmonary vasculature leads to increased oxidative stress and pulmonary arterial hypertension.
Subject(s)
Endothelium, Vascular/physiopathology , Hypertension, Pulmonary/physiopathology , Intracellular Signaling Peptides and Proteins/physiology , Oxidative Stress/physiology , Telangiectasia, Hereditary Hemorrhagic/physiopathology , Animals , Antioxidants/therapeutic use , Bone Morphogenetic Protein Receptors/metabolism , Cyclic N-Oxides/therapeutic use , Disease Models, Animal , Endoglin , Endothelium, Vascular/metabolism , Hypertension, Pulmonary/metabolism , Hypertension, Pulmonary/prevention & control , Intracellular Signaling Peptides and Proteins/genetics , Mice , Mice, Inbred C57BL , Mutation , Nitric Oxide Synthase Type III/metabolism , Reactive Oxygen Species/metabolism , Spin Labels , Telangiectasia, Hereditary Hemorrhagic/genetics , Telangiectasia, Hereditary Hemorrhagic/metabolism , Transforming Growth Factor beta/metabolismABSTRACT
Decreased endothelial NO synthase (eNOS)-derived NO bioavailability and impaired vasomotor control are crucial factors in cardiovascular disease pathogenesis. Hereditary hemorrhagic telangiectasia type 1 (HHT1) is a vascular disorder associated with ENDOGLIN (ENG) haploinsufficiency and characterized by venous dilatations, focal loss of capillaries, and arteriovenous malformations (AVMs). We report that resistance arteries from Eng+/- mice display an eNOS-dependent enhancement in endothelium-dependent dilatation and impairment in the myogenic response, despite reduced eNOS levels. We have found that eNOS is significantly reduced in endoglin-deficient endothelial cells because of decreased eNOS protein half-life. We demonstrate that endoglin can reside in caveolae and associate with eNOS, suggesting a stabilizing function of endoglin for eNOS. After Ca2+-induced activation, endoglin-deficient endothelial cells have reduced eNOS/Hsp90 association, produce less NO, and generate more eNOS-derived superoxide (O2-), indicating that endoglin also facilitates eNOS/Hsp90 interactions and is an important regulator in the coupling of eNOS activity. Treatment with an O2- scavenger reverses the vasomotor abnormalities in Eng(+/-) arteries, suggesting that uncoupled eNOS and resulting impaired myogenic response represent early events in HHT1 pathogenesis and that the use of antioxidants may provide a novel therapeutic modality.
