Impact of coronary artery disease on clinical outcomes after TAVR: Insights from the BRAVO-3 randomized trial.

OBJECTIVE: To determine the prognostic impact of coronary artery disease (CAD) in patients randomized to bivalirudin or unfractionated heparin (UFH) during transcatheter aortic valve replacement (TAVR). BACKGROUND: CAD is a common comorbidity among patients undergoing TAVR and studies provide conflicting data on its prognostic impact. METHODS: The Bivalirudin on Aortic Valve Intervention Outcomes-3 (BRAVO-3) randomized trial compared the use of bivalirudin versus UFH in 802 high-surgical risk patients undergoing transfemoral TAVR for severe symptomatic aortic stenosis. Patients were stratified according to the presence or absence of history of CAD as well as periprocedural anticoagulation. The coprimary endpoints were net adverse cardiac events (NACE; a composite of all-cause mortality, myocardial infarction, stroke, or major bleeding) and major Bleeding Academic Research Consortium (BARC) bleeding ≥3b at 30 days postprocedure. RESULTS: Among 801 patients, 437 (54.6%) had history of CAD of whom 223 (51.0%) received bivalirudin. There were no significant differences in NACE (adjusted odds ratio [OR]: 1.04; 95% confidence interval [CI]: 0.69-1.58) or BARC ≥ 3b bleeding (adjusted OR: 0.84; 95% CI: 0.51-1.39) in patients with vs without CAD at 30 days. Among CAD patients, periprocedural use of bivalirudin was associated with similar NACE (OR: 0.80; 95% CI: 0.47-1.35) and BARC ≥ 3b bleeding (OR: 0.64; 95% CI: 0.33-1.25) compared with UFH, irrespective of history of CAD (p-interaction = 0.959 for NACE; p-interaction = 0.479 for major bleeding). CONCLUSION: CAD was not associated with a higher short-term risk of NACE or major bleeding after TAVR. Periprocedural anticoagulation with bivalirudin did not show any advantage over UFH in patients with and without CAD.

Myeloid-Derived Growth Factor Protects Against Pressure Overload-Induced Heart Failure by Preserving Sarco/Endoplasmic Reticulum Ca2+-ATPase Expression in Cardiomyocytes.

BACKGROUND: Inflammation contributes to the pathogenesis of heart failure, but there is limited understanding of inflammation's potential benefits. Inflammatory cells secrete MYDGF (myeloid-derived growth factor) to promote tissue repair after acute myocardial infarction. We hypothesized that MYDGF has a role in cardiac adaptation to persistent pressure overload. METHODS: We defined the cellular sources and function of MYDGF in wild-type (WT), Mydgf-deficient (Mydgf-/-), and Mydgf bone marrow-chimeric or bone marrow-conditional transgenic mice with pressure overload-induced heart failure after transverse aortic constriction surgery. We measured MYDGF plasma concentrations by targeted liquid chromatography-mass spectrometry. We identified MYDGF signaling targets by phosphoproteomics and substrate-based kinase activity inference. We recorded Ca2+ transients and sarcomere contractions in isolated cardiomyocytes. Additionally, we explored the therapeutic potential of recombinant MYDGF. RESULTS: MYDGF protein abundance increased in the left ventricular myocardium and in blood plasma of pressure-overloaded mice. Patients with severe aortic stenosis also had elevated MYDGF plasma concentrations, which declined after transcatheter aortic valve implantation. Monocytes and macrophages emerged as the main MYDGF sources in the pressure-overloaded murine heart. While Mydgf-/- mice had no apparent phenotype at baseline, they developed more severe left ventricular hypertrophy and contractile dysfunction during pressure overload than WT mice. Conversely, conditional transgenic overexpression of MYDGF in bone marrow-derived inflammatory cells attenuated pressure overload-induced hypertrophy and dysfunction. Mechanistically, MYDGF inhibited G protein-coupled receptor agonist-induced hypertrophy and augmented SERCA2a (sarco/endoplasmic reticulum Ca2+-ATPase 2a) expression in cultured neonatal rat ventricular cardiomyocytes by enhancing PIM1 (Pim-1 proto-oncogene, serine/threonine kinase) expression and activity. Along this line, cardiomyocytes from pressure-overloaded Mydgf-/- mice displayed reduced PIM1 and SERCA2a expression, greater hypertrophy, and impaired Ca2+ cycling and sarcomere function compared with cardiomyocytes from pressure-overloaded WT mice. Transplanting Mydgf-/- mice with WT bone marrow cells augmented cardiac PIM1 and SERCA2a levels and ameliorated pressure overload-induced hypertrophy and dysfunction. Pressure-overloaded Mydgf-/- mice were similarly rescued by adenoviral Serca2a gene transfer. Treating pressure-overloaded WT mice subcutaneously with recombinant MYDGF enhanced SERCA2a expression, attenuated left ventricular hypertrophy and dysfunction, and improved survival. CONCLUSIONS: These findings establish a MYDGF-based adaptive crosstalk between inflammatory cells and cardiomyocytes that protects against pressure overload-induced heart failure.

Novel self-expanding ALLEGRA transcatheter aortic valve for native aortic stenosis and degenerated bioprosthesis.

OBJECTIVES: To investigate the safety and efficacy of the ALLEGRA valve in routine use. BACKGROUND: The ALLEGRA aortic valve is a self-expanding transcatheter heart valve (THV) with bovine pericardial tissue and was CE approved in March 2017. Its unique design was developed to provide low prosthesis gradients. METHODS: We analyzed patients receiving an ALLEGRA THV between May 2017 and March 2021 at our center for treatment of aortic valve stenosis or degenerated valve prosthesis. Hemodynamic results and clinical outcome according to the Valve Academic Research Consortium-2 consensus criteria were evaluated at discharge and three months post transcatheter aortic valve replacement (TAVR) procedure. 93 patients with a mean age of 82.5 ± 4.8 years and a median EuroScore II of 4.7 ± 3.4 were treated, 15 of them were valve-in-valve procedures. RESULTS: Implantation was successful in 97.8% (91/93) and VARC-2 defined device success was achieved in 94.6% (88/93). In-hospital all-cause mortality was 2.2% (2/93). Life-threatening bleeding, major vascular complications and strokes were 3.2% (3/93), 2.2% (2/93) and 3.2% (3/93), respectively. Paravalvular leakage was none to trace in 60.4%, mild in 38.5% and moderate in 1.1%. Permanent pacemaker implantation in pacemaker naive patients was necessary in 9.5% (8/84). Mean gradient at discharge was 8.2 ± 4.3 mmHg for all patients; 7.1 ± 2.6 mmHg in patients treated for stenosis of the native aortic valve and 13.8 ± 6.3 mmHg in patients treated valve-in-valve. CONCLUSIONS: The ALLEGRA THV provides excellent hemodynamic results and a good safety profile with a low complication rate.

Impact of anemia on short-term outcomes after TAVR: A subgroup analysis from the BRAVO-3 randomized trial.

OBJECTIVES: To determine the prognostic impact of anemia in patients randomized to bivalirudin or unfractionated heparin (UFH) during transcatheter aortic valve replacement (TAVR). BACKGROUND: Whether the periprocedural use of bivalirudin as compared with UFH in anemic patients undergoing TAVR has an impact on outcomes remains unknown. METHODS: The BRAVO-3 trial compared the use of bivalirudin versus UFH in 802 high risk patients undergoing transfemoral TAVR for severe symptomatic aortic stenosis. Patients were stratified according to the presence (defined as hemoglobin levels <13 g/dl in men and <12 g/dl in women) or absence of anemia. The primary outcomes were net adverse cardiac events (NACE; a composite of all-cause mortality, myocardial infarction, stroke, or bleeding) and major bleeding (Bleeding Academic Research Consortium ≥3b) at 30 days. RESULTS: Among 798 patients with available baseline hemoglobin levels, 427 (54%) were anemic of whom 221 (52%) received bivalirudin. There were no significant differences in NACE and major bleeding at 30 days between patients with and without anemia, irrespective of the type of anticoagulant used (pinteraction  = 0.71 for NACE, pinteraction  = 1.0 for major bleeding). However, anemic patients had a higher risk of major vascular complications (adjusted OR 2.43, 95% CI 1.42-4.16, p = 0.001), and acute kidney injury (adjusted OR 1.74, 95% CI 1.16-2.59, p = 0.007) compared to non-anemic patients at 30 days. CONCLUSIONS: Anemia was not associated with a higher risk of NACE or major bleeding at 30 days after TAVR without modification of the treatment effects of periprocedural anticoagulation with bivalirudin versus UFH.

Mid-term results of interventional closure of patent foramen ovale with the Occlutech Figulla® Flex II Occluder.

BACKGROUND: Patients with a patent foramen ovale (PFO) who suffered from stroke, TIA or peripheral paradoxical embolism are at substantial risk for recurrent neurologic events and in need for secondary prevention. Interventional closure of PFO has been performed for over 20 years. Numerous devices have been developed and used for treatment. We investigated PFO closure with the third generation Occlutech Figulla® Flex II Occluder device. METHODS: Between 2012 and 2015 57 patients (mean age 47.3 ± 1.5 years) who had suffered from a thromboembolic event of unknown cause underwent transcatheter PFO closure with the Occlutech Figulla® Flex II Occluder at our department. 68.4 % of all patients had suffered from cryptogenic stroke, while TIA had occurred in 28.1 %. Almost all patients were diagnosed with an atrial septum aneurysm (90.9 %) and a severe right-to-left shunt grade 3: >20 microbubbles (92.0 %). Follow-up was done 6 months post intervention by clinical examination and transesophageal contrast echocardiography. RESULTS: No major periprocedural or in-hospital complication occurred. Closure was sufficient with no residual right-to-left shunt in 94.4 % of all patients at 6 months post implantation and only minimal residual shunt in three cases. There were no thrombotic formations associated to the occluder device. Atrial fibrillation occurred in one patient and a recurrent cerebral ischemic event was seen in one patient, who suffered from another TIA. CONCLUSIONS: The Occlutech Figulla® Flex II Occluder device and its delivery system is safe and provides sufficient closure of PFO in patients who suffered from cryptogenic stroke, TIA or paradoxical peripheral embolism.

Multicenter comparison of transcatheter aortic valve implantation with the self-expanding ACURATE neo2 versus Evolut PRO transcatheter heart valves.

BACKGROUND: New-generation self-expanding transcatheter aortic heart valves (THV) were designed to overcome technical constraints of their preceding generations. We sought to compare the efficacy and safety of the self-expanding ACURATE neo2 (Neo2) versus Evolut PRO (PRO) devices. METHODS: Seven hundred nine patients undergoing transfemoral transcatheter aortic valve implantation (TAVI) with either Neo2 (n = 496) or PRO (n = 213) were included. Propensity score matching (PSM) was performed to account for differences in baseline characteristics. In-hospital and 30-day clinical outcomes were evaluated according to Valve Academic Research Consortium-3 criteria. RESULTS: Baseline characteristics were comparable between both groups after PSM (Neo2: n = 155, Evolut Pro: n = 155). Technical success rates were high in both groups (Neo2: 94.8% vs PRO: 97.4%; p = 0.239). Need for permanent pacemaker implantation was less frequent with Neo2 compared with PRO (7.5% vs 20.6%; p = 0.002), whereas major vascular complications were more frequent with Neo2 (Neo2: 11.6% vs PRO: 4.5%; p = 0.022). Intended valve performance at discharge was high in both groups without relevant differences among groups (Neo2: 97.4% vs. 95.3%; p = 0.328). CONCLUSIONS: Short-term outcomes after TAVI using latest-generation self-expanding THV were excellent, with overall low rates of adverse events. However, Neo2 was associated with lower pacemaker rates and reduced the prevalence of moderate-severe paravalvular leakage. Transprosthetic gradients after TAVI were higher with Neo2 compared with PRO.

Impairment of endothelial progenitor cell function and vascularization capacity by aldosterone in mice and humans.

AIMS: Hyperaldosteronism is associated with vascular injury and increased cardiovascular events. Bone marrow-derived endothelial progenitor cells (EPCs) play an important role in endothelial repair and vascular homeostasis. We hypothesized that hyperaldosteronism impairs EPC function and vascularization capacity in mice and humans. METHODS AND RESULTS: We characterized the effects of aldosterone and mineralocorticoid receptor (MR) blockade on EPC number and function as well as vascularization capacity and endothelial function. Treatment of human EPC with aldosterone induced translocation of the MR and impaired multiple cellular functions of EPC, such as differentiation, migration, and proliferation in vitro. Impaired EPC function was rescued by pharmacological blockade or genetic ablation of the MR. Aldosterone protein kinase A (PKA) dependently increased reactive oxygen species formation in EPC. Aldosterone infusion in mice impaired EPC function, EPC homing to vascular structures and vascularization capacity in a MR-dependent but blood pressure-independent manner. Endothelial progenitor cells from patients with primary hyperaldosteronism compared with controls of similar age displayed reduced migratory potential. Impaired EPC function was associated with endothelial dysfunction. MR blockade in patients with hyperaldosteronism improved EPC function and arterial stiffness. CONCLUSION: Endothelial progenitor cells express a MR that mediates functional impairment by PKA-dependent increase of reactive oxygen species. Normalization of EPC function may represent a novel mechanism contributing to the beneficial effects of MR blockade in cardiovascular disease prevention and treatment.

Right Ventricular Function Improves Early After Percutaneous Mitral Valve Repair in Patients Suffering From Severe Mitral Regurgitation.

Background: Percutaneous mitral valve edge-to-edge procedure (PMVR) using the MitraClip® system (Abbot Vascular, CA) is an established therapy for severe mitral regurgitation (MR) in patients judged inoperable or at high surgical risk. Besides determining exercise capacity, right ventricular (RV) function has prognostic value in heart failure and after cardiac surgery. We therefore investigated the impact of PMVR on RV function in patients with severe MR. Methods and Results: Sixty-three patients undergoing PMVR at our department were prospectively enrolled. Transthoracic echocardiography was performed before, early (2-12d) after PMVR and after 3 months, including advanced echocardiographic analyses such as 3D imaging and strain analyses. At baseline, all patients presented with advanced heart failure symptoms. Etiology of MR was more often secondary and, if present, left ventricular (LV) dysfunction was predominantly caused by ischemic cardiomyopathy. PMVR substantially reduced MR to a grade ≤ 2 in most patients. Echocardiographic assessment revealed a largely unchanged LV systolic function early after PMVR, while in contrast RV function substantially improved after PMVR [3D RV EF (%): pre 33.7% [27.4; 39.6], post 40.0% [34.5; 46.0] (p < 0.01 vs. pre), 3 months 42.8% [38.3; 48.1] (p < 0.01 vs. pre); 2D RV GLS (%): pre -12.9% [-14.5; -10.5], post -16.0% [-17.9; -12.6] (p < 0.01 vs. pre), 3 months -17.2% [-21.7; -14.9] (p < 0.01 vs. pre)]. Factors that attenuated RV improvement were larger ventricular volumes, lower LV function, secondary MR, and a higher STS score (all p < 0.05). Conclusion: By using advanced echocardiographic parameters, we discovered an early improvement of RV function after PMVR that is preserved for months, independent from changes in LV function. Improvement of RV function was less pronounced in patients presenting with an advanced stage of heart failure and a higher burden of comorbidities reflected by the STS score.

Multicenter comparison of latest-generation balloon-expandable versus self-expanding transcatheter heart valves: Ultra versus Evolut.

BACKGROUND: Direct comparisons of latest-generation balloon-expandable versus self-expanding transcatheter heart valves (THV) are scarce. To compare outcomes after transcatheter aortic valve replacement (TAVR) with SAPIEN 3 Ultra (Ultra) versus Evolut R or Pro (Evolut) THVs. METHODS: 1612 consecutive patients undergoing TAVR with either Ultra (n = 616) or Evolut (n = 996) were included. After propensity score matching (PSM), 467 and 205 matched pairs were identified in the entire cohort and with latest-generation THVs, respectively. Outcomes were investigated up to 30 days after TAVR. RESULTS: After PSM, baseline characteristics were comparable in the entire cohort (n = 934). Device success (92.7% vs. 87.6%; p = 0.011) and need for permanent pacemaker implantation (PPI) (15.2% vs. 8.4%; p = 0.002) were higher for Evolut compared with Ultra. Elevated gradients (≥20 mm Hg) were less frequent (1.6% vs. 10.4%; p < 0.001), whereas rates of ≥ moderate paravalvular leakage (PVL II+) were more frequent for Evolut compared with Ultra (3.7% vs. 1.3%; p = 0.019). With latest-generation THVs (n = 410), device success was comparable (93.2% vs. 89.8%; p = 0.216), whereas the need for PPI was higher for Evolut Pro compared with Ultra (15.6% vs. 9.8%; p = 0.075). Elevated gradients were less frequent (0% vs. 8%; p < 0.001), whereas rates of PVL II+ were more frequent for Evolut compared with Ultra (5.4% vs. 1.5%; p = 0.028). CONCLUSIONS: Device success rates were high with both THV platforms with low rates of adverse events up to 30 days after TAVR. Compared with Ultra, Evolut was associated with higher pacemaker rates as well as PVL II+, but with less elevated gradients.

Conditional overexpression of neuronal nitric oxide synthase is cardioprotective in ischemia/reperfusion.

BACKGROUND: We previously demonstrated that conditional overexpression of neuronal nitric oxide synthase (nNOS) inhibited L-type Ca2+ channels and decreased myocardial contractility. However, nNOS has multiple targets within the cardiac myocyte. We now hypothesize that nNOS overexpression is cardioprotective after ischemia/reperfusion because of inhibition of mitochondrial function and a reduction in reactive oxygen species generation. METHODS AND RESULTS: Ischemia/reperfusion injury in wild-type mice resulted in nNOS accumulation in the mitochondria. Similarly, transgenic nNOS overexpression caused nNOS abundance in mitochondria. nNOS translocation into the mitochondria was dependent on heat shock protein 90. Ischemia/reperfusion experiments in isolated hearts showed a cardioprotective effect of nNOS overexpression. Infarct size in vivo was also significantly reduced. nNOS overexpression also caused a significant increase in mitochondrial nitrite levels accompanied by a decrease of cytochrome c oxidase activity. Accordingly, O(2) consumption in isolated heart muscle strips was decreased in nNOS-overexpressing nNOS(+)/αMHC-tTA(+) mice already under resting conditions. Additionally, we found that the reactive oxygen species concentration was significantly decreased in hearts of nNOS-overexpressing nNOS(+)/αMHC-tTA(+) mice compared with noninduced nNOS(+)/αMHC-tTA(+) animals. CONCLUSION: We demonstrated that conditional transgenic overexpression of nNOS resulted in myocardial protection after ischemia/reperfusion injury. Besides a reduction in reactive oxygen species generation, this might be caused by nitrite-mediated inhibition of mitochondrial function, which reduced myocardial oxygen consumption already under baseline conditions.

Cardiac iron concentration in relation to systemic iron status and disease severity in non-ischaemic heart failure with reduced ejection fraction.

AIMS: Low cardiac iron levels promote heart failure in experimental models. While cardiac iron concentration (CI) is decreased in patients with advanced heart failure with reduced ejection fraction (HFrEF), CI has never been measured in non-advanced HFrEF. We measured CI in left ventricular (LV) endomyocardial biopsies (EMB) from patients with non-advanced HFrEF and explored CI association with systemic iron status and disease severity. METHODS AND RESULTS: We enrolled 80 consecutive patients with non-ischaemic HFrEF with New York Heart Association class II or III symptoms and a median (interquartile range) LV ejection fraction of 25 (18-33)%. CI was 304 (262-373) µg/g dry tissue. CI was not related to immunohistological findings or the presence of cardiotropic viral genomes in EMBs and was not related to biomarkers of systemic iron status or anaemia. Patients with CI in the lowest quartile (CIQ1 ) had lower body mass indices and more often presented with heart failure histories longer than 6 months than patients in the upper three quartiles (CIQ2-4 ). CIQ1 patients had higher serum N-terminal pro-B-type natriuretic peptide levels than CIQ2-4 patients [3566 (1513-6412) vs. 1542 (526-2811) ng/L; P = 0.005]. CIQ1 patients also had greater LV end-diastolic (P = 0.001) and end-systolic diameter indices (P = 0.003) and higher LV end-diastolic pressures (P = 0.046) than CIQ2-4 patients. CONCLUSION: Low CI is associated with greater disease severity in patients with non-advanced non-ischaemic HFrEF. CI is unrelated to systemic iron homeostasis. The prognostic and therapeutic implications of CI measurements in EMBs should be further explored.

Improvement in left ventricular remodeling by the endothelial nitric oxide synthase enhancer AVE9488 after experimental myocardial infarction.

BACKGROUND: Reduced endothelial nitric oxide (NO) bioavailability contributes to the progression of heart failure. In this study, we investigated whether the transcription enhancer of endothelial NO synthase (eNOS) AVE9488 improves cardiac remodeling and heart failure after experimental myocardial infarction (MI). METHODS AND RESULTS: Starting 7 days after coronary artery ligation, rats with MI were treated with placebo or AVE9488 (25 ppm) as a dietary supplement for 9 weeks. AVE9488 therapy versus placebo substantially improved left ventricular (LV) function, reduced LV filling pressure, and prevented the rightward shift of the pressure-volume curve. AVE9488 also attenuated the extent of pulmonary edema, reduced LV fibrosis and myocyte cross-sectional area, and prevented the increases in LV gene expression of atrial natriuretic factor, brain natriuretic peptide, and endothelin-1. eNOS protein levels and calcium-dependent NOS activity were decreased in the surviving LV myocardium from placebo MI rats and normalized by AVE9488. The beneficial effects of AVE9488 on LV dysfunction and remodeling after MI were abrogated in eNOS-deficient mice. Aortic eNOS protein expression and endothelium-dependent NO-mediated vasorelaxation were significantly enhanced by AVE9488 treatment after infarction, whereas increased vascular superoxide anion formation was reduced. Moreover, AVE9488 prevented the marked depression of circulating endothelial progenitor cell levels in rats with heart failure after MI. CONCLUSIONS: Long-term treatment with the eNOS enhancer AVE9488 improved LV remodeling and contractile dysfunction after MI. Molecular alterations, circulating endothelial progenitor cell levels, and endothelial vasomotor dysfunction were improved by AVE9488. Pharmacological interventions designed to increase eNOS-derived NO constitute a promising therapeutic approach for the amelioration of postinfarction ventricular remodeling and heart failure.

Multidrug resistance protein-1 affects oxidative stress, endothelial dysfunction, and atherogenesis via leukotriene C4 export.

BACKGROUND: We recently showed that the multidrug resistance related protein-1 (MRP1) is important for the management of oxidative stress in vascular cells. However, the underlying mechanism and the in vivo relevance of these findings remain elusive. We hypothesize that inside-outside transport of leukotriene C(4) (LTC(4)) via MRP1 is a substantial proatherogenic mechanism in the vasculature. To test this hypothesis, we investigated the effects of MRP1 inhibition and LTC(4) receptor blockade (Cys-LT1 receptor) in vitro and in vivo. METHODS AND RESULTS: MRP1 is expressed abundantly in vascular smooth muscle cells (VSMCs). Pharmacological inhibition of MRP1 via MK571 reduces angiotensin II-induced reactive oxygen species release by 59% (L012 fluorescence) in VSMCs. The release of reactive oxygen species after angiotensin II stimulation also is inhibited by blockade of the Cys-LT1 receptor with montelukast. Incubation of VSMCs with recombined LTC(4) causes enhanced rates of reactive oxygen species and proliferation in wild-type and MRP1(-/-) VSMCs. Accordingly, the LTC(4) release in the cell culture supernatant of MRP1(-/-) VSMCs is significantly decreased compared with wild-type cells. To extend our observations to the in vivo situation, atherosclerosis-prone apolipoprotein E-deficient mice on a high-cholesterol diet were treated with placebo, the MRP1 inhibitor MK571, or the Cys-LT1 receptor inhibitor montelukast for 6 weeks. Treatment with MK571 or montelukast reduced vascular reactive oxygen species production, significantly improved endothelial function, and ameliorated atherosclerotic plaque generation by 52% and 61%, respectively. CONCLUSIONS: These findings indicate that MRP1 and LTC(4) exert proatherosclerotic effects and that both MRP1 and LTC(4) are potentially promising targets for atheroprotective therapy.

Regulation of tetrahydrobiopterin biosynthesis by shear stress.

An essential cofactor for the endothelial NO synthase is tetrahydrobiopterin (H4B). In the present study, we show that in human endothelial cells, laminar shear stress dramatically increases H4B levels and enzymatic activity of GTP cyclohydrolase (GTPCH)-1, the first step of H4B biosynthesis. In contrast, protein levels of GTPCH-1 were not affected by shear. Shear did not change protein expression or activity of the downstream enzymes 6-pyruvoyl-tetrahydropterin synthase and sepiapterin reductase and decreased protein levels of the salvage enzyme dihydrofolate reductase. Oscillatory shear only modestly affected H4B levels and GPTCH-1 activity. We also demonstrate that laminar, but not oscillatory shear stress, stimulates phosphorylation of GTPCH-1 on serine 81 and that this is mediated by the alpha prime (alpha') subunit of casein kinase 2. The increase in H4B caused by shear is essential in allowing proper function of endothelial NO synthase because GPTCH-1 blockade with 2,4-diamino-6-hydroxypyrimidine during shear inhibited dimer formation of endothelial NO synthase, increased endothelial cell superoxide production, and prevented the increase in NO production caused by shear. Thus, shear stress not only increases endothelial NO synthase levels but also stimulates production of H4B by markedly enhancing GTPCH-1 activity via casein kinase 2-dependent phosphorylation on serine 81. These findings illustrate a new function of casein kinase 2 in the endothelium and provide insight into regulation of GTPCH-1 activity.

Impact of diabetes mellitus on short term vascular complications after TAVR: Results from the BRAVO-3 randomized trial.

AIMS: The impact of diabetes mellitus (DM) on clinical outcomes after transcatheter aortic valve replacement (TAVR) remains unclear. The aim of this study was to investigate the impact of DM on short-term clinical outcomes after TAVR in a large randomized trial population. METHODS AND RESULTS: BRAVO-3 trial randomized 802 patients undergoing trans-femoral TAVR to procedural anticoagulation with bivalirudin or unfractionated heparin. The study population was divided according to the presence of DM, and further stratified according to the use of insulin. Net adverse cardiovascular outcomes (NACE - death, myocardial infarction (MI), stroke or major bleeding by Bleeding Academic Research Consortium (BARC) type 3b or above) was the primary outcome in-hospital and at 30-days. Of the total 802 randomized patients, 239 (30%) had DM at baseline, with 87 (36%) being treated with insulin. At 30-days, DM patients experienced numerically higher rates of net adverse cardiovascular events (16.3% vs. 14.4%, p=0.48) and acute kidney injury (19.7% vs. 15.1%, p=0.11), while non-DM (NDM) patients had numerically higher rates of cerebrovascular accidents (3.6% vs. 1.7%, p=0.22). After multivariable adjustment, DM patients had higher odds of vascular complications at 30-days (OR 1.57, p=0.03) and life-threatening bleeding both in-hospital (OR 1.50, p=0.046) and at 30-days (OR 1.50, p=0.03) with the excess overall risk primarily attributed to the higher rates observed among non-insulin dependent DM patients. CONCLUSIONS: Patients with DM had higher adjusted odds of vascular and bleeding complications up to 30-days post-TAVR. Overall, there was no significant association between DM and early mortality following TAVR.

Endothelial nitric oxide synthase uncoupling impairs endothelial progenitor cell mobilization and function in diabetes.

Uncoupling of the endothelial nitric oxide synthase (eNOS) resulting in superoxide anion (O(2)(-)) formation instead of nitric oxide (NO) causes diabetic endothelial dysfunction. eNOS regulates mobilization and function of endothelial progenitor cells (EPCs), key regulators of vascular repair. We postulate a role of eNOS uncoupling for reduced number and function of EPC in diabetes. EPC levels in diabetic patients were significantly reduced compared with those of control subjects. EPCs from diabetic patients produced excessive O(2)(-) and showed impaired migratory capacity compared with nondiabetic control subjects. NOS inhibition with N(G)-nitro-l-arginine attenuated O(2)(-) production and normalized functional capacity of EPCs from diabetic patients. Glucose-mediated EPC dysfunction was protein kinase C dependent, associated with reduced intracellular BH(4) (tetrahydrobiopterin) concentrations, and reversible after exogenous BH(4) treatment. Activation of NADPH oxidases played an additional but minor role in glucose-mediated EPC dysfunction. In rats with streptozotocin-induced diabetes, circulating EPCs were reduced to 39 +/- 5% of controls and associated with uncoupled eNOS in bone marrow. Our results identify uncoupling of eNOS in diabetic bone marrow, glucose-treated EPCs, and EPCs from diabetic patients resulting in eNOS-mediated O(2)(-) production. Subsequent reduction of EPC levels and impairment of EPC function likely contributes to the pathogenesis of vascular disease in diabetes.

Role of the multidrug resistance protein-1 in hypertension and vascular dysfunction caused by angiotensin II.

OBJECTIVE: Human endothelial cells use the multidrug resistance protein-1 (MRP1) to export glutathione disulfide (GSSG). This can promotes thiol loss during states of increased glutathione oxidation. We investigated how MRP1 modulates blood pressure and vascular function during angiotensin II-induced hypertension. METHODS AND RESULTS: Angiotensin II-induced hypertension altered vascular glutathione flux by increasing GSSG export and decreasing vascular levels of glutathione in wild-type (FVB) but not in MRP1-/- mice. Aortic endothelium-dependent vasodilatation was reduced in FVB after angiotensin II infusion, but unchanged in MRP1-/- mice. Aortic superoxide (O2*-) production and expression of several NADPH oxidase subunits were increased by angiotensin II in FVB. These effects were markedly blunted in MRP1-/- vessels. The increase in O2*- production in FVB vessels caused by angiotensin II was largely inhibited by L-NAME, suggesting eNOS uncoupling. Accordingly, aortic tetrahydrobiopterin and levels of NO were decreased by angiotensin II in FVB but were unchanged in MRP1-/-. Finally, the hypertension caused by angiotensin II was markedly blunted in MRP1-/- mice (137+/-4 versus 158+/-6 mm Hg). CONCLUSION: MRP1 plays a crucial role in the genesis of multiple vascular abnormalities that accompany hypertension and its presence is essential for the hypertensive response to angiotensin II.

Endothelial dysfunction in heart failure.

Endothelial dysfunction crucially contributes to the development of impaired coronary and systemic perfusion as well as reduced exercise capacity in patients with congestive heart failure, with fundamental impact on morbidity and mortality. Reduced bioavailability of nitric oxide (NO) and abundant formation of reactive oxygen species (ROS) within the vascular wall are the key determinants in endothelial dysfunction. The imbalance between NO and ROS mainly results from neurohumoral activation associated with heart failure. As endothelial derived NO is a major endogenous modulator of platelet function, reduced intravascular bioactivity of NO contributes to platelet activation, adhesion and thromboembolic events in heart failure. Treatment with angiotensin converting enzyme (ACE) inhibitors, angiotensin and aldosterone antagonists, and statins beneficially modulates endothelial dysfunction in heart failure. All these therapies increase NO bioactivity by either modulation of ROS generation, thereby preventing the interaction of superoxide anions with NO, and/or increasing endothelial NO synthase (eNOS) expression/activity. AVE9488, a novel eNOS transcription enhancer, attenuates cardiac remodeling and endothelial dysfunction in rats after large myocardial infarction. Endothelial progenitor cell (EPC) levels and their mobilization are regulated by eNOS. After myocardial infarction in rats, EPC levels and formation of endothelial colony forming units are markedly reduced. AVE 9488, ACE or HMG-CoA reductase inhibition result in significant increases in EPC levels, and beneficial effects on bone marrow molecular alterations after myocardial infarction.

The role of the multidrug resistance protein-1 in modulation of endothelial cell oxidative stress.

Glutathione (GSH) is the major source of intracellular sulfhydryl groups. Oxidized GSH (GSSG) can be recycled to GSH by the GSH reductase or exported from the cell. The mechanism by which GSSG is exported and the consequence of its export from endothelial cells has not been defined previously. We found that human endothelial cells express the multidrug resistance protein-1 (MRP1) and use this as their major exporter of GSSG. Oscillatory shear stress, which is known to stimulate endothelial cell production of reactive oxygen species, decreased intracellular GSH. In contrast, laminar shear significantly increased intracellular GSH. Oscillatory shear also caused a robust export of GSSG that was prevented by the MRP1 inhibitor MK571 and by MRP1 small interfering RNA. MRP1 inhibition prevented the decline in intracellular GSH, preserved the intracellular GSH Nernst potential, and reduced apoptosis caused by oscillatory shear. In aortas of hypertensive mice, endothelial disulfide export was doubled, and this was prevented by MK571 and was not observed in aortas of hypertensive MRP1-/- mice. Further, the altered endothelium-dependent vasodilatation caused by hypertension was ameliorated in MRP1-/- mice. GSSG export by MRP1 leads to a perturbation of endothelial redox state and ultimately endothelial cell apoptosis. Endothelial MRP1 may provide a novel therapeutic target for prevention of vascular disease.

Early determinants of H2O2-induced endothelial dysfunction.

Reactive oxygen species (ROS) can stimulate nitric oxide (NO(*)) production from the endothelium by transient activation of endothelial nitric oxide synthase (eNOS). With continued or repeated exposure, NO(*) production is reduced, however. We investigated the early determinants of this decrease in NO(*) production. Following an initial H(2)O(2) exposure, endothelial cells responded by increasing NO(*) production measured electrochemically. NO(*) concentrations peaked by 10 min with a slow reduction over 30 min. The decrease in NO(*) at 30 min was associated with a 2.7-fold increase in O(2)(*-) production (p < 0.05) and a 14-fold reduction of the eNOS cofactor, tetrahydrobiopterin (BH(4), p < 0.05). Used as a probe for endothelial dysfunction, the integrated NO(*) production over 30 min upon repeated H(2)O(2) exposure was attenuated by 2.1-fold (p = 0.03). Endothelial dysfunction could be prevented by BH(4) cofactor supplementation, by scavenging O(2)(*-) or peroxynitrite (ONOO(-)), or by inhibiting the NADPH oxidase. Hydroxyl radical (()OH) scavenging did not have an effect. In summary, early H(2)O(2)-induced endothelial dysfunction was associated with a decreased BH(4) level and increased O(2)(*-) production. Dysfunction required O(2)(*-), ONOO(-), or a functional NADPH oxidase. Repeated activation of the NADPH oxidase by ROS may act as a feed forward system to promote endothelial dysfunction.