Lysyl oxidase expression in smooth muscle cells determines the level of intima calcification in hypercholesterolemia-induced atherosclerosis. / La expresión de la lisil oxidasa en las células musculares lisas determina el nivel de calcificación de la íntima en la aterosclerosis inducida por hipercolesterolemia.

INTRODUCTION: Cardiovascular calcification is an important public health issue with an unmeet therapeutic need. We had previously shown that lysyl oxidase (LOX) activity critically influences vascular wall smooth muscle cells (VSMCs) and valvular interstitial cells (VICs) calcification by affecting extracellular matrix remodeling. We have delved into the participation of LOX in atherosclerosis and vascular calcification, as well as in the mineralization of the aortic valve. METHODS: Immunohistochemical and expression studies were carried out in human atherosclerotic lesions and experimental models, valves from patients with aortic stenosis, VICs, and in a genetically modified mouse model that overexpresses LOX in CMLV (TgLOXCMLV). Hyperlipemia and atherosclerosis was induced in mice through the administration of adeno-associated viruses encoding a PCSK9 mutated form (AAV-PCSK9D374Y) combined with an atherogenic diet. RESULTS: LOX expression is increased in the neointimal layer of atherosclerotic lesions from human coronary arteries and in VSMC-rich regions of atheromas developed both in the brachiocephalic artery of control (C57BL/6J) animals transduced with PCSK9D374Y and in the aortic root of ApoE-/- mice. In TgLOXCMLV mice, PCSK9D374Y transduction did not significantly alter the enhanced aortic expression of genes involved in matrix remodeling, inflammation, oxidative stress and osteoblastic differentiation. Likewise, LOX transgenesis did not alter the size or lipid content of atherosclerotic lesions in the aortic arch, brachiocephalic artery and aortic root, but exacerbated calcification. Among lysyl oxidase isoenzymes, LOX is the most expressed member of this family in highly calcified human valves, colocalizing with RUNX2 in VICs. The lower calcium deposition and decreased RUNX2 levels triggered by the overexpression of the nuclear receptor NOR-1 in VICs was associated with a reduction in LOX. CONCLUSIONS: Our results show that LOX expression is increased in atherosclerotic lesions, and that overexpression of this enzyme in VSMC does not affect the size of the atheroma or its lipid content, but it does affect its degree of calcification. Further, these data suggest that the decrease in calcification driven by NOR-1 in VICs would involve a reduction in LOX. These evidences support the interest of LOX as a therapeutic target in cardiovascular calcification.

Upregulation of NOR-1 in calcified human vascular tissues: impact on osteogenic differentiation and calcification.

Cardiovascular calcification is a significant public health issue whose pathophysiology is not fully understood. NOR-1 regulates critical processes in cardiovascular remodeling, but its contribution to ectopic calcification is unknown. NOR-1 was overexpressed in human calcific aortic valves and calcified atherosclerotic lesions colocalizing with RUNX2, a factor essential for osteochondrogenic differentiation and calcification. NOR-1 and osteogenic markers were upregulated in calcifying human valvular interstitial cells (VICs) and human vascular smooth muscle cells (VSMCs). Gain- and loss-of-function approaches demonstrated that NOR-1 negatively modulates the expression of osteogenic genes relevant for the osteogenic transdifferentiation (RUNX2, IL-6, BMP2, and ALPL) and calcification of VICs. VSMCs from transgenic mice overexpressing NOR-1 in these cells (TgNOR-1VSMC) expressed lower basal levels of osteogenic genes (IL-6, BMP2, ALPL, OPN) than cells from WT littermates, and their upregulation by a high-phosphate osteogenic medium (OM) was completely prevented by NOR-1 transgenesis. Consistently, this was associated with a dramatic reduction in the calcification of both transgenic VSMCs and aortic rings from TgNOR-1VSMC mice exposed to OM. Atherosclerosis and calcification were induce in mice by the administration of AAV-PCSK9D374Y and a high-fat/high-cholesterol diet. Challenged-TgNOR-1VSMC mice exhibited decreased vascular expression of osteogenic markers, and both less atherosclerotic burden (assessed in whole aorta and lesion size in aortic arch and brachiocephalic artery) and less vascular calcification (assessed either by near-infrared fluorescence imaging or histological analysis) than WT mice. Our data indicate that NOR-1 negatively modulates the expression of genes critically involved in the osteogenic differentiation of VICs and VSMCs, thereby restraining ectopic cardiovascular calcification.

Amiodarone triggered Kounis syndrome complicated by refractory cardiac arrest rescued with VA-ECMO.

Key Clinical Message: An interesting case that shows an infrequent cause of cardiorespiratory arrest such as coronary vasospasm due to intravenous amiodarone induced Kounis syndrome. It highlights the usefulness of circulatory support with ECMO in the scenario of CPR. Abstract: A patient with atrial fibrillation was admitted for an elective electrical cardioversion. He was given an amiodarone bolus that triggered Kounis syndrome with cardiac arrest due to vasospasm requiring emergency coronary angiography with infusion of nitroglycerin. Due to following refractory shock and severe refractory hypoxemia required mechanical circulatory support with ECMO and inhaled nitric oxide with favorable evolution. Allergy to amiodarone was later confirmed.

Apolipoprotein J protects cardiomyocytes from lipid-mediated inflammation and cytotoxicity induced by the epicardial adipose tissue of diabetic patients.

Diabetic patients present increased volume and functional alterations in epicardial adipose tissue (EAT). We aimed to analyze EAT from type 2 diabetic patients and the inflammatory and cytotoxic effects induced on cardiomyocytes. Furthermore, we analyzed the cardioprotective role of apolipoprotein J (apoJ). EAT explants were obtained from nondiabetic patients (ND), diabetic patients without coronary disease (DM), and DM patients with coronary disease (DM-C) after heart surgery. Morphological characteristics and gene expression were evaluated. Explants were cultured for 24 h and the content of nonesterified fatty acids (NEFA) and sphingolipid species in secretomes was evaluated by lipidomic analysis. Afterwards, secretomes were added to AC16 human cardiomyocytes for 24 h in the presence or absence of cardioprotective molecules (apoJ and HDL). Cytokine release and apoptosis/necrosis were assessed by ELISA and flow cytometry. The EAT from the diabetic samples showed altered expression of genes related to lipid accumulation, insulin resistance, and inflammation. The secretomes from the DM samples presented an increased ratio of pro/antiatherogenic ceramide (Cer) species, while those from DM-C contained the highest concentration of saturated NEFA. DM and DM-C secretomes promoted inflammation and cytotoxicity on AC16 cardiomyocytes. Exogenous Cer16:0, Cer24:1, and palmitic acid reproduced deleterious effects in AC16 cells. These effects were attenuated by exogenous apoJ. Diabetic secretomes promoted inflammation and cytotoxicity in cardiomyocytes. This effect was exacerbated in the secretomes of the DM-C samples. The increased content of specific NEFA and ceramide species seems to play a key role in inducing such deleterious effects, which are attenuated by apoJ.

Intraoperative intra-aortic balloon pump insertion: step by step.

A 76-year-old patient with non-ST elevation myocardial infarction was admitted to our hospital. Coronary angiography revealed significant left main and two-vessel coronary artery disease. Preoperative testing indicated severe left ventricular dysfunction. The patient was scheduled for urgent off-pump coronary artery bypass grafting. Due to the low ejection fraction, an intra-aortic balloon pump was inserted in the operating theatre before sternotomy, to enhance the patient's haemodynamic stability during surgery. A 6 Fr introducer was inserted into the femoral artery under echocardiographic guidance. Using a 150-cm guidewire, the intra-aortic balloon catheter was advanced through the introducer to the descending thoracic aorta. The catheter's tip position, just distal to the origin of the left subclavian artery, was confirmed via transoesophageal echocardiography. The external part of the catheter was secured to the skin and connected to the balloon console. Therapy was initiated, and the inflation/deflation parameters were optimized. A double off-pump coronary artery bypass was performed via median sternotomy. The patient remained haemodynamically stable throughout the surgery, aided by the intra-aortic balloon pump, and careful volume and vasoactive management. The patient was extubated promptly, and the device was removed on the second postoperative day without complications.

Postpartum acute coronary syndrome due to intramural hematoma and coronary artery dissection.

A 38-year-old woman presented with acute coronary syndrome 9 days postpartum. Coronary catheterization showed an intramural hematoma of the left main artery (LM) and the left anterior descending artery (LAD). During angiography, dissection of the LM, with involvement of the LAD and circumflex artery occurred. The patient presented unstable hemodynamic status. After resuscitation, intra-aortic balloon pump was inserted. The patient underwent two-vessel off-pump surgical revascularization. She was discharged 10 days later.

Lysyl oxidase-dependent extracellular matrix crosslinking modulates calcification in atherosclerosis and aortic valve disease.

Extracellular matrix (ECM) is an active player in cardiovascular calcification (CVC), a major public health issue with an unmet need for effective therapies. Lysyl oxidase (LOX) conditions ECM biomechanical properties; thus, we hypothesized that LOX might impact on mineral deposition in calcific aortic valve disease (CAVD) and atherosclerosis. LOX was upregulated in calcified valves from two cohorts of CAVD patients. Strong LOX immunostaining was detected surrounding calcified foci in calcified human valves and atherosclerotic lesions colocalizing with RUNX2 on valvular interstitial cells (VICs) or vascular smooth muscle cells (VSMCs). Both LOX secretion and organized collagen deposition were enhanced in calcifying VICs exposed to osteogenic media. ß-aminopropionitrile (BAPN), an inhibitor of LOX, attenuated collagen deposition and calcification. VICs seeded onto decellularized matrices from BAPN-treated VICs calcified less than cells cultured onto control scaffolds; instead, VICs exposed to conditioned media from cells over-expressing LOX or cultured onto LOX-crosslinked matrices calcified more. Atherosclerosis was induced in WT and transgenic mice that overexpress LOX in VSMC (TgLOXVSMC) by AAV-PCSK9D374Y injection and high-fat feeding. In atherosclerosis-challenged TgLOXVSMC mice both atherosclerosis burden and calcification assessed by near-infrared fluorescence (NIRF) imaging were higher than in WT mice. These animals also exhibited larger calcified areas in atherosclerotic lesions from aortic arches and brachiocephalic arteries. Moreover, LOX transgenesis exacerbated plaque inflammation, and increased VSMC cellularity, the rate of RUNX2-positive cells and both connective tissue content and collagen cross-linking. Our findings highlight the relevance of LOX in CVC and postulate this enzyme as a potential therapeutic target for CVC.

Nuclear receptor NOR-1 (Neuron-derived Orphan Receptor-1) in pathological vascular remodelling and vascular remodelling. / El receptor nuclear NOR-1 (Neuron-derived Orphan Receptor-1) en el remodelado vascular patológico.

Vascular cells and their interaction with inflammatory cells and the immune system play a key role in pathological vascular remodeling. A large number of genes and proteins regulated in a coordinated manner by a small number of transcription factors are involved in this process. In recent years, research on a small subfamily of transcription factors, the NR4A subfamily, has had a major impact on our understanding of vascular biology. The NR4A1 (Nur77), NR4A2 (Nurr1) and NR4A3 (NOR-1) receptors are products of early response genes whose expression is induced by multiple pathophysiological and physical stimuli. Their wide distribution in different tissues and cells places them in the control of numerous processes such as cell differentiation, proliferation, survival and apoptosis, as well as inflammation and the metabolism of lipids and carbohydrates. This review analyzes the role of these receptors, particularly NOR-1, in pathological vascular remodeling associated with atherosclerosis, abdominal aortic aneurysm and pulmonary arterial hypertension.

Hemolytic Anemia After Surgical Mitral Valve Repair Treated With Transcatheter Edge-to-Edge Device.

An 80-year-old woman with mitral valve repair failure was admitted with hemolytic anemia secondary to the impact of a regurgitant jet on the annuloplasty ring. Transcatheter repair to treat both mitral regurgitation and hemolysis was favored because of surgical risk. Transcatheter edge-to-edge repair represents an alternative for treating hemolysis associated with mitral regurgitation. (Level of Difficulty: Advanced.).

ß2-adrenergic stimulation potentiates spontaneous calcium release by increasing signal mass and co-activation of ryanodine receptor clusters.

AIMS: It is unknown how ß-adrenergic stimulation affects calcium dynamics in individual RyR2 clusters and leads to the induction of spontaneous calcium waves. To address this, we analysed spontaneous calcium release events in green fluorescent protein (GFP)-tagged RyR2 clusters. METHODS: Cardiomyocytes from mice with GFP-tagged RyR2 or human right atrial tissue were subjected to immunofluorescent labelling or confocal calcium imaging. RESULTS: Spontaneous calcium release from single RyR2 clusters induced 91.4% ± 2.0% of all calcium sparks while 8.0% ± 1.6% were caused by release from two neighbouring clusters. Sparks with two RyR2 clusters had 40% bigger amplitude, were 26% wider, and lasted 35% longer at half maximum. Consequently, the spark mass was larger in two- than one-cluster sparks with a median and interquartile range for the cumulative distribution of 15.7 ± 20.1 vs 7.6 ± 5.7 a.u. (P < .01). ß2-adrenergic stimulation increased RyR2 phosphorylation at s2809 and s2815, tripled the fraction of two- and three-cluster sparks, and significantly increased the spark mass. Interestingly, the amplitude and mass of the calcium released from a RyR2 cluster were proportional to the SR calcium load, but the firing rate was not. The spark mass was also higher in 33 patients with atrial fibrillation than in 36 without (22.9 ± 23.4 a.u. vs 10.7 ± 10.9; P = .015). CONCLUSIONS: Most sparks are caused by activation of a single RyR2 cluster at baseline while ß-adrenergic stimulation doubles the mass and the number of clusters per spark. This mimics the shift in the cumulative spark mass distribution observed in myocytes from patients with atrial fibrillation.

CD69 expression on regulatory T cells protects from immune damage after myocardial infarction.

Increasing evidence has pointed to the important function of T cells in controlling immune homeostasis and pathogenesis after myocardial infarction (MI), although the underlying molecular mechanisms remain elusive. In this study, a broad analysis of immune markers in 283 patients revealed significant CD69 overexpression on Tregs after MI. Our results in mice showed that CD69 expression on Tregs increased survival after left anterior descending (LAD) coronary artery ligation. Cd69-/- mice developed strong IL-17+ γδT cell responses after ischemia that increased myocardial inflammation and, consequently, worsened cardiac function. CD69+ Tregs, by induction of AhR-dependent CD39 ectonucleotidase activity, induced apoptosis and decreased IL-17A production in γδT cells. Adoptive transfer of CD69+ Tregs into Cd69-/- mice after LAD ligation reduced IL-17+ γδT cell recruitment, thus increasing survival. Consistently, clinical data from 2 independent cohorts of patients indicated that increased CD69 expression in peripheral blood cells after acute MI was associated with a lower risk of rehospitalization for heart failure (HF) after 2.5 years of follow-up. This result remained significant after adjustment for age, sex, and traditional cardiac damage biomarkers. Our data highlight CD69 expression on Tregs as a potential prognostic factor and a therapeutic option to prevent HF after MI.

Systematic review and meta-analysis of surgical outcomes comparing mechanical valve replacement and bioprosthetic valve replacement in infective endocarditis.

BACKGROUND: Infective endocarditis (IE) is an infection involving either native or prosthetic heart valves, the endocardial surface of the heart or any implanted intracardiac devices. IE is a rare condition affecting 3-15 patients per 100,000 population. In-hospital mortality rates in patients with IE remain high at around 20% despite treatment advances. There is no consensus recommendation favoring either bioprosthetic valve or mechanical valve implantation in the setting of IE; patient age, co-morbidities and preferences should be considered selecting the replacement prosthesis. METHODS: A systematic review and meta-analysis of studies reporting the outcomes of patients undergoing bioprosthetic or mechanical valve replacement for infective endocarditis with data extracted for overall survival, valve reinfection rates and valve reoperation. RESULTS: Eleven relevant studies were identified, with 2,336 patients receiving a mechanical valve replacement and 2,057 patients receiving a bioprosthetic valve replacement. There was no significant difference for overall survival between patients treated with mechanical valves and those treated with bioprosthetic valves [hazard ratio (HR) 0.94, 95% confidence interval (CI): 0.73-1.21, P=0.62]. There was no significant difference in reoperation rates between patients treated with a bioprosthetic valve and those treated with a mechanical valve (HR 0.82, 95% CI: 0.34-1.98, P=0.66) and there was no significant difference in the rate of valve reinfection rates (HR 0.95, 95% CI: 0.48-1.89, P=0.89). CONCLUSIONS: The presence of infective endocarditis alone should not influence the decision of which type of valve prosthesis that should be implanted. This decision should be based on patient age, co-morbidities and preferences.

Use of the Perceval Sutureless Valve in Active Prosthetic Aortic Valve Endocarditis.

BACKGROUND: Surgical treatment of active prosthetic aortic valve endocarditis presents a challenge for cardiac surgeons because of tissue friability and destruction caused by infection. Sutureless prostheses, such as the Perceval S (LivaNova, Saluggia, Italy), have emerged as an option among the different surgical approaches for these complicated cases. METHODS: This study presents data from 9 patients who underwent aortic valve re-replacement with the Perceval S because of active prosthetic aortic valve endocarditis between January 2014 and August 2016. Hemodynamic performance (mean transprosthetic gradient and type of aortic regurgitation) was assessed intraoperatively after weaning from cardiopulmonary bypass, at discharge, and to 6 months postoperatively. RESULTS: After weaning from cardiopulmonary bypass, cases 1 and 3 through 6 had no or trivial aortic regurgitation, cases 7 and 8 presented with trivial to mild regurgitation, case 9 showed mild intraprosthetic regurgitation, and case 2 had mild periprosthetic regurgitation. Cases 4 and 7 died of septic shock and multiorgan failure in the perioperative period. In the remaining patients, severity of aortic regurgitation maintained practically invariable at discharge compared with intraoperative results. These 7 patients did well at 6-month follow-up, with good clinical and hemodynamic performance of the Perceval S prosthesis. The median of mean transprosthetic gradient was 11 mm Hg (interquartile range: 10 to 12 mm Hg). Only patient 2 showed mild periprosthetic regurgitation; patient 9 showed mild intraprosthetic insufficiency, and the remaining patients had no or trivial regurgitation. CONCLUSIONS: The sutureless Perceval S valve is a reasonable alternative for surgical treatment of prosthetic aortic valve endocarditis.