Marc W. Merx

Nitrite reductase activity of myoglobin regulates respiration and cellular viability in myocardial ischemia-reperfusion injury

The nitrite anion is reduced to nitric oxide (NO•) as oxygen tension decreases. Whereas this pathway modulates hypoxic NO• signaling and mitochondrial respiration and limits myocardial infarction in mammalian species, the pathways to nitrite bioactivation remain uncertain. Studies suggest that hemoglobin and myoglobin may subserve a fundamental physiological function as hypoxia dependent nitrite reductases. Using myoglobin wild-type (+/+) and knockout (−/−) mice, we here test the central role of myoglobin as a functional nitrite reductase that regulates hypoxic NO• generation, controls cellular respiration, and therefore confirms a cytoprotective response to cardiac ischemia-reperfusion (I/R) injury. We find that myoglobin is responsible for nitrite-dependent NO• generation and cardiomyocyte protein iron-nitrosylation. Nitrite reduction to NO• by myoglobin dynamically inhibits cellular respiration and limits reactive oxygen species generation and mitochondrial enzyme oxidative inactivation after I/R injury. In isolated myoglobin+/+ but not in myoglobin−/− hearts, nitrite treatment resulted in an improved recovery of postischemic left ventricular developed pressure of 29%. In vivo administration of nitrite reduced myocardial infarction by 61% in myoglobin+/+ mice, whereas in myoglobin−/− mice nitrite had no protective effects. These data support an emerging paradigm that myoglobin and the heme globin family subserve a critical function as an intrinsic nitrite reductase that regulates responses to cellular hypoxia and reoxygenation. myoglobin knockout mice

Sepsis and the Heart

Sepsis is generally viewed as a disease aggravated by an inappropriate immune response encountered in the afflicted individual. As an important organ system frequently compromised by sepsis and always affected by septic shock, the cardiovascular system and its dysfunction during sepsis have been studied in clinical and basic research for more than 5 decades. Although a number of mediators and pathways have been shown to be associated with myocardial depression in sepsis, the precise cause remains unclear to date. There is currently no evidence supporting global ischemia as an underlying cause of myocardial dysfunction in sepsis; however, in septic patients with coexistent and possibly undiagnosed coronary artery disease, regional myocardial ischemia or infarction secondary to coronary artery disease may certainly occur. A circulating myocardial depressant factor in septic shock has long been proposed, and potential candidates for a myocardial depressant factor include cytokines, prostanoids, and nitric oxide, among others. Endothelial activation and induction of the coagulatory system also contribute to the pathophysiology in sepsis. Prompt and adequate antibiotic therapy accompanied by surgical removal of the infectious focus, if indicated and feasible, is the mainstay and also the only strictly causal line of therapy. In the presence of severe sepsis and septic shock, supportive treatment in addition to causal therapy is mandatory. The purpose of this review is to delineate some characteristics of septic myocardial dysfunction, to assess the most commonly cited and reported underlying mechanisms of cardiac dysfunction in sepsis, and to briefly outline current therapeutic strategies and possible future approaches.

Sustained Benefits in Vascular Function Through Flavanol-Containing Cocoa in Medicated Diabetic Patients: A Double-Masked, Randomized, Controlled Trial

OBJECTIVES Our goal was to test feasibility and efficacy of a dietary intervention based on daily intake of flavanol-containing cocoa for improving vascular function of medicated diabetic patients. BACKGROUND Even in fully medicated diabetic patients, overall prognosis is unfavorable due to deteriorated cardiovascular function. Based on epidemiological data, diets rich in flavanols are associated with a reduced cardiovascular risk. METHODS In a feasibility study with 10 diabetic patients, we assessed vascular function as flow-mediated dilation (FMD) of the brachial artery, plasma levels of flavanol metabolites, and tolerability after an acute, single-dose ingestion of cocoa, containing increasing concentrations of flavanols (75, 371, and 963 mg). In a subsequent efficacy study, changes in vascular function in 41 medicated diabetic patients were assessed after a 30-day, thrice-daily dietary intervention with either flavanol-rich cocoa (321 mg flavanols per dose) or a nutrient-matched control (25 mg flavanols per dose). Both studies were undertaken in a randomized, double-masked fashion. Primary and secondary outcome measures included changes in FMD and plasma flavanol metabolites, respectively. RESULTS A single ingestion of flavanol-containing cocoa was dose-dependently associated with significant acute increases in circulating flavanols and FMD (at 2 h: from 3.7 +/- 0.2% to 5.5 +/- 0.4%, p < 0.001). A 30-day, thrice-daily consumption of flavanol-containing cocoa increased baseline FMD by 30% (p < 0.0001), while acute increases of FMD upon ingestion of flavanol-containing cocoa continued to be manifest throughout the study. Treatment was well tolerated without evidence of tachyphylaxia. Endothelium-independent responses, blood pressure, heart rate, and glycemic control were unaffected. CONCLUSIONS Diets rich in flavanols reverse vascular dysfunction in diabetes, highlighting therapeutic potentials in cardiovascular disease.

Statin Treatment After Onset of Sepsis in a Murine Model Improves Survival

Background—HMG-CoA-reductase inhibitors have been shown to exhibit pronounced immunomodulatory effects independent of lipid lowering. We have recently demonstrated that pretreatment with simvastatin profoundly improves survival in a cecal ligation and perforation (CLP) model of sepsis. Here, we studied whether treatment with simvastatin after onset of sepsis-induced hemodynamic alterations is beneficial and whether prolonged survival can also be achieved with other statins. Methods and Results—Mice were rendered septic by CLP. At 6 hours after sepsis induction, when profound hemodynamic alterations were manifest, treatment with atorvastatin, fluvastatin, pravastatin, simvastatin, or placebo was initiated. Except for fluvastatin (27±2.3 hours), survival time was extended from 23±1.2 hours for placebo-treated mice to 37±3.6 hours for simvastatin-treated, to 40±4.2 hours for atorvastatin-treated, and to 39±3.9 hours for pravastatin-treated mice. This profound improvement is based on the preservation of cardiac function and hemodynamic status in statin-treated animals, both of which are severely impaired in untreated CLP mice. As underlying mechanisms, improved susceptibility to endothelial nitric oxide synthase stimulation and reduced endothelial adhesion of leukocytes could be demonstrated after statin treatment. Conclusions—Well established in the treatment of lipid disorders and coronary artery disease, statins harbor the additional and novel potential of effective sepsis treatment. This benefit extends to several but not all statins tested.

HMG-CoA Reductase Inhibitor Simvastatin Profoundly Improves Survival in a Murine Model of Sepsis

Background—HMG-CoA reductase inhibitors, such as simvastatin, have been shown to exhibit pronounced immunomodulatory effects independent of lipid lowering but to date have not been used to treat severe inflammatory disease such as sepsis. We thus approached the question of whether treatment with simvastatin might improve cardiovascular function and survival in sepsis. Methods and Results—Mice treated with simvastatin and rendered septic by cecal ligation and perforation (CLP) show a mean survival time close to 4 times the value found in untreated mice. This dramatic improvement is based on a complete preservation of cardiac function and hemodynamic status, which are severely impaired in untreated CLP mice [eg, 20 hours after CLP, cardiac output declined from 1.24±0.09 to 0.87±0.11 mL · min−1 · g−1 in untreated mice (P <0.005; n=12), while remaining unaltered (1.21±0.08 mL · min−1 · g−1 at baseline and 1.15±0.1 mL · min−1 · g−1 20 hours after CLP, P =NS, n=12) in CLP mice treated with simvastatin]. Untreated CLP mice remained refractory to β-stimulation, whereas the responsiveness to dobutamine was restored by treatment with simvastatin. Susceptibility of coronary flow to endothelial nitric oxide synthase (eNOS) stimulation by bradykinin was close to 3 times as pronounced in untreated CLP mice as in untreated sham-operated mice, indicating a high level of eNOS activation secondary to sepsis. In addition, treatment with simvastatin reversed inflammatory alterations in CLP mice, namely, increased monocyte adhesion to endothelium. Conclusions—Simvastatin, which is well established in the treatment of lipid disorders and coronary artery disease, might have the additional potential of being an effective agent in sepsis treatment.

Brain natriuretic peptide predicts right heart failure in patients with acute pulmonary embolism

BACKGROUND Plasma levels of brain natriuretic peptide (BNP) are increased in patients with left heart failure. In patients with severe pulmonary embolism (PE), primary right ventricular (RV) dysfunction is frequent. Little is known about BNP secretion in acute RV failure. METHODS We prospectively studied 50 consecutive patients with confirmed PE (age range, 57 +/- 19 years; 36 men). PE was confirmed with pulmonary angiography, spiral computed tomography, or echocardiography and subsidiary analyses. On admission, echocardiography and BNP measurements were performed in all patients. RESULTS Patients without RV dysfunction had significantly lower BNP levels than patients with RV dysfunction (55 +/- 69 pg/mL vs 340 +/- 362 pg/mL, P <.001). There was a significant correlation between RV end-diastolic diameter and BNP (r = 0.43, P <.05). BNP discriminated patients with or without RV dysfunction (area under the receiver operating characteristic curve, 0.78; 95% CI, 0.64-0.92). A BNP >90 pg/mL was associated with a risk ratio of 28.4 (95% CI, 3.22-251.12) for the diagnosis of RV dysfunction. All patients without LV systolic dysfunction who had syncope necessitating cardiopulmonary resuscitation had normal BNP levels. Patients with RV dysfunction had significantly more in-hospital complications (cardiogenic shock, inotropic therapy, mechanical ventilation). However, BNP levels were not predictive of mortality or in-hospital complications. CONCLUSIONS BNP levels are frequently increased in patients with PE who have RV dysfunction, whereas patients without RV dysfunction show reference range BNP levels in the absence of left ventricular dysfunction. In acute PE, BNP elevation is highly predictive of RV dysfunction, but not of in-hospital complications and mortality.

Taurine transporter knockout depletes muscle taurine levels and results in severe skeletal muscle impairment but leaves cardiac function uncompromised

Taurine is the most abundant free amino acid in heart and skeletal muscle. In the present study, the effects of hereditary taurine deficiency on muscle function were examined in taurine transporter knockout (taut−/−) mice. These mice show an almost complete depletion of heart and skeletal muscle taurine levels. Treadmill experiments demonstrated that total exercise capacity of taut−/− mice was reduced by >80% compared with wild‐type controls. The decreased performance of taut−/− mice correlated with increased lactate levels in serum during exercise. Surprisingly, cardiac function of taut−/− mice as assessed by magnetic resonance imaging, echocardiography, and isolated heart studies showed a largely normal phenotype under both control and stimulated conditions. However, analysis of taut−/− skeletal muscle revealed electromyographic abnormalities. 1H nuclear magnetic resonance spectroscopy of tissue extracts showed that in the heart of taut−/− mice the lack of taurine was compensated by the up‐regulation of various organic solutes. In contrast, a deficit of >10 mM in total organic osmolyte concentration was found in skeletal muscle. The present study identifies taurine transport as a crucial factor for the maintenance of skeletal muscle function and total exercise capacity, while cardiac muscle apparently can compensate for the loss of taurine.

Transthoracic Echocardiography Using Second Harmonic Imaging Diagnostic Alternative to Transesophageal Echocardiography for the Detection of Atrial Right to Left Shunt in Patients With Cerebral Embolic Events

OBJECTIVES We sought to evaluate whether transthoracic contrast echocardiography using second harmonic imaging (SHI) is a diagnostic alternative to transesophageal contrast echocardiography (TEE) for the detection of atrial right to left shunt. BACKGROUND Paradoxic embolism is considered to be the major cause of cerebral ischemic events in young patients. Contrast echocardiography using TEE has proven to be superior to transthoracic echocardiography (TTE) for the detection of atrial shunting, SHI is a new imaging modality that enhances the visualization of echocardiographic contrast agents. METHODS We evaluated 111 patients with an ischemic cerebral embolic event for the presence of atrial right to left shunt using an intravenous (IV) contrast agent in combination with three different echocardiographic imaging modalities: 1) TTE using fundamental imaging (FI); 2) TTE using SHI; and 3) TEE. The severity of atrial shunting and the duration of contrast visibility within the left heart chambers were evaluated for each imaging modality. Image quality was assessed separately for each modality by semiquantitative scoring (0 = poor to 3 = excellent). Presence of atrial right to left shunt was defined as detection of contrast bubbles in the left atrium within the first three cardiac cycles after contrast appearance in the right atrium either spontaneously or after the Valsalva maneuver. RESULTS A total of 57 patients showed evidence of atrial right to left shunt with either imaging modality. Fifty-one studies were positive with TEE, 52 studies were positive with SHI, and 32 were positive with FI (p<0.001 for FI vs. SHI and TEE). The severity of contrast passage was significantly larger using SHI (61.6+/-80.2 bubbles) compared to FI (53.7+/-69.6 bubbles; p<0.005 vs. SHI) but was not different compared to TEE (43.9+/-54.3 bubbles; p = NS vs. SHI). The duration of contrast visibility was significantly longer for SHI (17.4+/-12.4 s) compared to FI (13.1+/-9.7 s; p<0.001) and TEE (11.9+/-9.6 s; p<0.02). Mean image quality improved significantly from FI (1.5+/-0.8) to SHI (2.0+/-0.8; p<0.001 vs. FI) and TEE (2.5+/-0.7; p<0.001 vs. SHI). CONCLUSIONS In combination with IV contrast injections, TEE and SHI have a comparable yield for the detection of atrial right to left shunt. Both modalities may miss patients with atrial shunting. In young patients with an unexplained cerebrovascular event and no clinical evidence of cardiac disease, a positive SHI study may obviate the need to perform a TEE study to search for cardiac sources of emboli.

Double-Edged Role of the CXCL12/CXCR4 Axis in Experimental Myocardial Infarction

OBJECTIVES Here we assess the intrinsic functions of the chemokine receptor CXCR4 in remodeling after myocardial infarction (MI) using Cxcr4 heterozygous (Cxcr4(+/-)) mice. BACKGROUND Myocardial necrosis triggers complex remodeling and inflammatory changes. The chemokine CXCL12 has been implicated in protection and therapeutic regeneration after MI through recruiting angiogenic outgrowth cells, improving neovascularization and cardiac function, but the endogenous role of its receptor CXCR4 is unknown. METHODS MI was induced by ligation of the left descending artery. Langendoff perfusion, echocardiography, quantitative immunohistochemistry, flow cytometry, angiogenesis assays, and cardiomyocyte analysis were performed. RESULTS After 4 weeks, infarct size was reduced in Cxcr4(+/-) mice compared with wild-type mice and in respective bone marrow chimeras compared with controls. This was associated with altered inflammatory cell recruitment, decreased neutrophil content, delayed monocyte infiltration, and a predominance of Gr1(low) over classic Gr1(high) monocytes. Basal coronary flow and its recovery after MI were impaired in Cxcr4(+/-)mice, paralleled by reduced angiogenesis, myocardial vessel density, and endothelial cell count. Notably, no differences in cardiac function were seen in Cxcr4(+/-)mice compared with wild-type mice. Despite defective angiogenesis, Cxcr4(+/-) mouse hearts showed no difference in CXCL12, vascular endothelial growth factor or apoptosis-related gene expression. Electron microscopy revealed lipofuscin-like lipid accumulation in Cxcr4(+/-) mouse hearts and analysis of lipid extracts detected high levels of phosphatidylserine, which protect cardiomyocytes from hypoxic stress in vitro. CONCLUSIONS CXCR4 plays a crucial role in endogenous remodeling processes after MI, contributing to inflammatory/progenitor cell recruitment and neovascularization, whereas its deficiency limits infarct size and causes adaptation to hypoxic stress. This should be carefully scrutinized when devising therapeutic strategies involving the CXCL12/CXCR4 axis.

Age-dependent endothelial dysfunction is associated with failure to increase plasma nitrite in response to exercise

Age-dependent alterations of the vessel wall may predispose older individuals to increased cardiovascular pathology. Aging is associated with an impaired bioactivity of nitric oxide (NO). Plasma nitrite reflects NO-synthase activity under fasting conditions and is an important storage pool of NO. To test the hypothesis that aging is associated with an impaired capacity of the vasculature to increase plasma nitrite during exercise, 29 young and 28 old healthy individuals (25 ± 1 years and 58 ± 2 years; P < 0.001) without major cardiovascular risk factors were enrolled. Exercise stress was similar in both groups. Baseline nitrite did not differ (107 ± 8 vs. 82 ± 10 nmol/l, young vs. old; n.s.) although a trend toward higher nitrite levels in young individuals was seen. In young subjects, exercise increased plasma nitrite by 38 ± 7% (P < 0.001) compared to only 13 ± 8% (P = n.s.) in older subjects. L-NMMA blocked increases of nitrite. Endothelial function, as defined by flow–mediated-dilation (FMD) of the brachial artery via ultrasound, was impaired in older subjects (5.4 ± 0.4% vs. 6.7 ± 0.3%; P < 0.01). Multivariate analysis showed that age (P = 0.007), BMI (P = 0.010), and LDL (P = 0.021) were independent predictors of nitrite increase. The fact that aging is associated with an impaired capacity of the vasculature to adequately increase nitrite to physiological stimuli may contribute to attenuated maintenance and further deterioration of vascular homeostasis with aging.