Ramzi El Accaoui

Oxidized CaMKII Triggers Atrial Fibrillation

Background —Atrial fibrillation is a growing public health problem without adequate therapies. Angiotensin II (Ang II) and reactive oxygen species (ROS) are validated risk factors for atrial fibrillation (AF) in patients, but the molecular pathway(s) connecting ROS and AF is unknown. The Ca 2+ /calmodulin-dependent protein kinase II (CaMKII) has recently emerged as a ROS activated proarrhythmic signal, so we hypothesized that oxidized CaMKIIδ(ox-CaMKII) could contribute to AF. Methods and Results —We found ox-CaMKII was increased in atria from AF patients compared to patients in sinus rhythm and from mice infused with Ang II compared with saline. Ang II treated mice had increased susceptibility to AF compared to saline treated WT mice, establishing Ang II as a risk factor for AF in mice. Knock in mice lacking critical oxidation sites in CaMKIIδ (MM-VV) and mice with myocardial-restricted transgenic over-expression of methionine sulfoxide reductase A (MsrA TG), an enzyme that reduces ox-CaMKII, were resistant to AF induction after Ang II infusion. Conclusions —Our studies suggest that CaMKII is a molecular signal that couples increased ROS with AF and that therapeutic strategies to decrease ox-CaMKII may prevent or reduce AF.Background— Atrial fibrillation (AF) is a growing public health problem without adequate therapies. Angiotensin II and reactive oxygen species are validated risk factors for AF in patients, but the molecular pathways connecting reactive oxygen species and AF are unknown. The Ca2+/calmodulin-dependent protein kinase II (CaMKII) has recently emerged as a reactive oxygen species–activated proarrhythmic signal, so we hypothesized that oxidized CaMKII&dgr; could contribute to AF. Methods and Results— We found that oxidized CaMKII was increased in atria from AF patients compared with patients in sinus rhythm and from mice infused with angiotensin II compared with mice infused with saline. Angiotensin II–treated mice had increased susceptibility to AF compared with saline-treated wild-type mice, establishing angiotensin II as a risk factor for AF in mice. Knock-in mice lacking critical oxidation sites in CaMKII&dgr; (MM-VV) and mice with myocardium-restricted transgenic overexpression of methionine sulfoxide reductase A, an enzyme that reduces oxidized CaMKII, were resistant to AF induction after angiotensin II infusion. Conclusions— Our studies suggest that CaMKII is a molecular signal that couples increased reactive oxygen species with AF and that therapeutic strategies to decrease oxidized CaMKII may prevent or reduce AF.

Osteoprotegerin Inhibits Aortic Valve Calcification and Preserves Valve Function in Hypercholesterolemic Mice

Background There are no rigorously confirmed effective medical therapies for calcific aortic stenosis. Hypercholesterolemic Ldlr −/− Apob 100/100 mice develop calcific aortic stenosis and valvular cardiomyopathy in old age. Osteoprotegerin (OPG) modulates calcification in bone and blood vessels, but its effect on valve calcification and valve function is not known. Objectives To determine the impact of pharmacologic treatment with OPG upon aortic valve calcification and valve function in aortic stenosis-prone hypercholesterolemic Ldlr −/− Apob 100/100 mice. Methods Young Ldlr −/− Apob 100/100 mice (age 2 months) were fed a Western diet and received exogenous OPG or vehicle (N = 12 each) 3 times per week, until age 8 months. After echocardiographic evaluation of valve function, the aortic valve was evaluated histologically. Older Ldlr −/− Apob 100/100 mice were fed a Western diet beginning at age 2 months. OPG or vehicle (N = 12 each) was administered from 6 to 12 months of age, followed by echocardiographic evaluation of valve function, followed by histologic evaluation. Results In Young Ldlr −/− Apob 100/100 mice, OPG significantly attenuated osteogenic transformation in the aortic valve, but did not affect lipid accumulation. In Older Ldlr −/− Apob 100/100 mice, OPG attenuated accumulation of the osteoblast-specific matrix protein osteocalcin by ∼80%, and attenuated aortic valve calcification by ∼ 70%. OPG also attenuated impairment of aortic valve function. Conclusions OPG attenuates pro-calcific processes in the aortic valve, and protects against impairment of aortic valve function in hypercholesterolemic aortic stenosis-prone Ldlr −/− Apob 100/100 mice.

Aortic valve sclerosis in mice deficient in endothelial nitric oxide synthase

Risk factors for fibrocalcific aortic valve disease (FCAVD) are associated with systemic decreases in bioavailability of endothelium-derived nitric oxide (EDNO). In patients with bicuspid aortic valve (BAV), vascular expression of endothelial nitric oxide synthase (eNOS) is decreased, and eNOS(-/-) mice have increased prevalence of BAV. The goal of this study was to test the hypotheses that EDNO attenuates profibrotic actions of valve interstitial cells (VICs) in vitro and that EDNO deficiency accelerates development of FCAVD in vivo. As a result of the study, coculture of VICs with aortic valve endothelial cells (vlvECs) significantly decreased VIC activation, a critical early phase of FCAVD. Inhibition of VIC activation by vlvECs was attenuated by N(G)-nitro-l-arginine methyl ester or indomethacin. Coculture with vlvECs attenuated VIC expression of matrix metalloproteinase-9, which depended on stiffness of the culture matrix. Coculture with vlvECs preferentially inhibited collagen-3, compared with collagen-1, gene expression. BAV occurred in 30% of eNOS(-/-) mice. At age 6 mo, collagen was increased in both bicuspid and trileaflet eNOS(-/-) aortic valves, compared with wild-type valves. At 18 mo, total collagen was similar in eNOS(-/-) and wild-type mice, but collagen-3 was preferentially increased in eNOS(-/-) mice. Calcification and apoptosis were significantly increased in BAV of eNOS(-/-) mice at ages 6 and 18 mo. Remarkably, these histological changes were not accompanied by physiologically significant valve stenosis or regurgitation. In conclusion, coculture with vlvECs inhibits specific profibrotic VIC processes. In vivo, eNOS deficiency produces fibrosis in both trileaflet and BAVs but produces calcification only in BAVs.

The Multifunctional Ca2+/Calmodulin-Dependent Kinase IIδ (CaMKIIδ) Regulates Arteriogenesis in a Mouse Model of Flow-Mediated Remodeling

Objective Sustained hemodynamic stress mediated by high blood flow promotes arteriogenesis, the outward remodeling of existing arteries. Here, we examined whether Ca2+/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.

Spontaneous Aortic Regurgitation and Valvular Cardiomyopathy in Mice

Objective— We studied the mechanistic links between fibrocalcific changes in the aortic valve and aortic valve function in mice homozygous for a hypomorphic epidermal growth factor receptor mutation (Wave mice). We also studied myocardial responses to aortic valve dysfunction in Wave mice. Approach and Results— At 1.5 months of age, before development of valve fibrosis and calcification, aortic regurgitation, but not aortic stenosis, was common in Wave mice. Aortic valve fibrosis, profibrotic signaling, calcification, osteogenic markers, lipid deposition, and apoptosis increased dramatically by 6 and 12 months of age in Wave mice. Aortic regurgitation remained prevalent, however, and aortic stenosis was rare, at all ages. Proteoglycan content was abnormally increased in aortic valves of Wave mice at all ages. Treatment with pioglitazone prevented abnormal valve calcification, but did not protect valve function. There was significant left ventricular volume overload, hypertrophy, and fetal gene expression, at all ages in Wave mice with aortic regurgitation. Left ventricular systolic function was normal until 6 months of age in Wave mice, but became impaired by 12 months of age. Myocardial transverse tubules were normal in the presence of left ventricular hypertrophy at 1.5 and 3 months of age, but became disrupted by 12 months of age. Conclusions— We present the first comprehensive phenotypic and molecular characterization of spontaneous aortic regurgitation and volume-overload cardiomyopathy in an experimental model. In Wave mice, fibrocalcific changes are not linked to valve dysfunction and are epiphenomena arising from structurally incompetent myxomatous valves.

Pituitary apoplexy after regadenoson myocardial perfusion scan

The high sensitivity and specificity of myocardial perfusion imaging has made it the preferred cardiac stress test for patients with abnormal resting EKG, history of coronary revascularization, or high likelihood of having coronary artery disease. Pharmacological myocardial perfusion imaging offers the additional benefit of testing patients that are unable to exercise or are on a beta-blocker. Various agents have been utilized to perform pharmacological nuclear stress testing. Adenosine and dipyridamole were the preferred pharmacological stress agents until recently. These agents are non-selective adenosine receptor agonists and have undesirable side effects by activation of A1, A2B, A3, and A4 receptors, which can give rise to atrioventricular blocks and bronchospasm leading to wheezing and shortness of breath. In 2008, the FDA approved regadenoson as an additional pharmacological stress agent. It is a selective A2A adenosine receptor agonist with weak affinity for A1, A2B, and A3 adenosine receptors. By activating A2A receptors, it causes coronary vasodilation. Hemodynamic effects of regadenoson include tachycardia and decrease in both systolic and diastolic blood pressure. Common side effects of regadenoson include dyspnea, headache, chest discomfort and flushing. In comparison to dipyridamole and adenosine, regadenoson showcases a favorable testing profile by its longer half-life, ease of administration, and less side effects. If needed, aminophylline can be administered to limit the side effects. The popularity of this drug came into threat in 2013, when FDA issued a black-box warning that regadenoson should not be used in patients with unstable angina or cardiovascular instability. The warning came in response to myocardial infarctions and deaths that were reported with regadenoson use. Recently, reports have associated the use of regadenoson with seizures. Despite these concerns, regadenoson continues to be used as a preferred pharmacological stress agent in the current age of nuclear stress testing. To date, only a single case of pituitary apoplexy has been reported during a nuclear stress test. Dipyridamole was the agent used in that study. Here we report the first case of pituitary apoplexy after using regadenoson during a pharmacological nuclear myocardial perfusion imaging.

Highly abnormal SPECT in a patient with left ventricular non-compaction and no evidence of obstructive coronary artery disease.

Left ventricular non-compaction (LVNC) is a rare congenital condition characterized by prominent trabeculae and deep recesses, best seen on cardiac MRI. Histologically, there is evidence of endomyocardial fibrosis. SPECT imaging in this condition is not well defined. We present a 68-year-old woman known to have LVNC (Figure 1) and non-obstructive coronary artery disease based on normal SPECT in 2007 (Figure 2) and cardiac catheterization in 2008. She presented with progressive exertional chest pain. Echocardiogram showed a wall motion abnormality. Repeat SPECT with Regadenoson Sestamibi was obtained for risk stratification and was abnormal. Stress images showed marked diffuse photopenia with the exception of the inferoseptum, while rest images showed uniform radiotracer uptake (Figure 3). This degree of abnormality raised concern for significant diffuse ischemia versus artifact. Cardiac CT angiogram was obtained promptly and showed no coronary atherosclerosis or stenosis (Figure 4). The presence of extensive perfusion defects on SPECT imaging has previously been described in patients with LVNC. The etiology of these perfusion defects is not well established and could be secondary to the interstitial fibrosis seen in the myocardium of patients with LVNC. Additional studies are warranted to better understand the etiology of this unusual finding.

Chronic vitamin K antagonist therapy and bleeding risk in ST elevation myocardial infarction patients

Objectives Acute management of ST elevation myocardial infarction (STEMI) patients on chronic vitamin K antagonist (VKA) therapy is uncertain. This study aims to estimate in-hospital major bleeding risk among STEMI patients on chronic VKA treated with primary percutaneous coronary intervention (PCI); and determine the relationship between bleeding and acute treatments stratified by international normalised ratio (INR) values. Methods We retrospectively examined 120 270 STEMI patients treated with primary PCI at 586 national registry hospitals (2007–2012). Results Overall, 3101 patients (2.6%) were on VKA which was associated with increased in-hospital major bleeding risk when compared with patients not on VKA (17.0%, vs 10.1%; adjusted OR 1.26, 95% CI 1.13 to 1.40). In patients on VKA, admission INR ≥2.0 was not associated with an increase in bleeding risk compared to INR <2.0. Patients on VKA were more likely to receive clopidogrel or bivalirudin within 24 h of presentation (acute), but less likely to receive prasugrel, heparin, or glycoprotein IIb/IIIa inhibitors (GPI). In those patients, acute GPI was associated with increased bleeding risk (adjusted OR 1.92, 95% CI 1.54 to 2.40) while bivalirudin was associated with decreased risk (adjusted OR 0.69, 95% CI 0.55 to 0.86); bleeding risk associated with heparin, bivalirudin, ADP-receptor blockers, or GPI was similar between INR ≥2.0 and <2.0. Conclusions In STEMI patients treated with primary PCI, chronic VKA therapy was associated with a significant increase in in-hospital major bleeding risk compared to no VKA therapy, irrespective of whether admission INR was ≥2.0 or not. In patients on VKA, GPI was associated with increased bleeding risk while bivalirudin was associated with decreased risk.