Myron A. Waclawiw

Nitrite reduction to nitric oxide by deoxyhemoglobin vasodilates the human circulation.

Nitrite anions comprise the largest vascular storage pool of nitric oxide (NO), provided that physiological mechanisms exist to reduce nitrite to NO. We evaluated the vasodilator properties and mechanisms for bioactivation of nitrite in the human forearm. Nitrite infusions of 36 and 0.36 μmol/min into the forearm brachial artery resulted in supra- and near-physiologic intravascular nitrite concentrations, respectively, and increased forearm blood flow before and during exercise, with or without NO synthase inhibition. Nitrite infusions were associated with rapid formation of erythrocyte iron-nitrosylated hemoglobin and, to a lesser extent, S-nitroso-hemoglobin. NO-modified hemoglobin formation was inversely proportional to oxyhemoglobin saturation. Vasodilation of rat aortic rings and formation of both NO gas and NO-modified hemoglobin resulted from the nitrite reductase activity of deoxyhemoglobin and deoxygenated erythrocytes. This finding links tissue hypoxia, hemoglobin allostery and nitrite bioactivation. These results suggest that nitrite represents a major bioavailable pool of NO, and describe a new physiological function for hemoglobin as a nitrite reductase, potentially contributing to hypoxic vasodilation.

Prognostic Value of Coronary Vascular Endothelial Dysfunction

Background—Whether patients at increased risk can be identified from a relatively low-risk population by coronary vascular function testing remains unknown. We investigated the relationship between coronary endothelial function and the occurrence of acute unpredictable cardiovascular events (cardiovascular death, myocardial infarction, stroke, and unstable angina) in patients with and without coronary atherosclerosis (CAD). Methods and Results—We measured the change in coronary vascular resistance (&Dgr;CVR) and epicardial diameter with intracoronary acetylcholine (ACh, 15 &mgr;g/min) to test endothelium-dependent function and sodium nitroprusside (20 &mgr;g/min) and adenosine (2.2 mg/min) to test endothelium-independent vascular function in 308 patients undergoing cardiac catheterization (132 with and 176 without CAD). Patients underwent clinical follow-up for a mean of 46±3 months. Acute vascular events occurred in 35 patients. After multivariate analysis that included CAD and conventional risk factors for atherosclerosis, &Dgr;CVR with ACh (P =0.02) and epicardial constriction with ACh (P =0.003), together with increasing age, CAD, and body mass index, were independent predictors of adverse events. Thus, patients in the tertile with the best microvascular responses with ACh and those with epicardial dilation with ACh had improved survival by Kaplan-Meier analyses in the total population, as did those in the subset without CAD. Similar improvement in survival was also observed when all adverse events, including revascularization, were considered. Endothelium-independent responses were not predictive of outcome. Conclusions—Epicardial and microvascular coronary endothelial dysfunction independently predict acute cardiovascular events in patients with and without CAD, providing both functional and prognostic information that complements angiographic and risk factor assessment.

Association between Prior Cytomegalovirus Infection and the Risk of Restenosis after Coronary Atherectomy

BACKGROUND Restenosis occurs commonly after coronary angioplasty and atherectomy, but the causes of restenosis are poorly understood. Recently, it has been found that cytomegalovirus (CMV) DNA is present in restenotic lesions from atherectomy specimens. This and other evidence suggest that CMV may have a role in the process of restenosis. METHODS We prospectively studied 75 consecutive patients undergoing directional coronary atherectomy for symptomatic coronary artery disease. Before atherectomy was performed, we measured blood levels of anti-CMV IgG antibodies to determine whether previous exposure to CMV increased the risk of restenosis, as determined by coronary angiography performed six months after atherectomy. RESULTS After atherectomy, the mean (+/- SD) minimal luminal diameter of the target vessel was greater in the 49 patients who were seropositive for CMV than in the 26 patients who were seronegative (3.18 +/- 0.51 mm vs. 2.89 +/- 0.45 mm, P=0.01). After six months, however, the seropositive patients had a greater reduction in the luminal diameter (1.24 +/- 0.83 mm vs. 0.68 +/- 0.69 mm, P = 0.003), resulting in a significantly higher rate o restenosis in the seropositive patients (43 percent vs. 8 percent, P = 0.002). In a multivariable logistic-regression model, CMV seropositivity and the CMV titer were independently predictive of restenosis (odds ratios, 12.9 and 8.1, respectively). There was no evidence of acute infection, since the titer of anti-CMV IgG antibodies did not increase over time and tests for anti-CMV IgM antibodies were negative in all patients. CONCLUSIONS Prior infection with CMV is strong independent risk factor for restenosis after coronary atherectomy. If confirmed, these findings may help identify patients at risk for restenosis.

Imipramine in patients with chest pain despite normal coronary angiograms

BACKGROUND Ten to 30 percent of patients undergoing cardiac catheterization because of chest pain are found to have normal coronary angiograms. Because these patients may have a visceral pain syndrome unrelated to myocardial ischemia, we investigated whether drugs that are useful in chronic pain syndromes might also be beneficial in such patients. METHODS Sixty consecutive patients underwent cardiac, esophageal, psychiatric, and pain-sensitivity testing and then participated in a randomized, double-blind, placebo-controlled three-week trial of clonidine at a dose of 0.1 mg twice daily (20 patients), imipramine at a dose of 50 mg nightly with a morning placebo (20 patients), or placebo twice daily (20 patients); this treatment phase was compared with an identical period of twice-daily placebo for all patients (placebo phase). RESULTS Thirteen (22 percent) of the 60 patients had ischemic-appearing electrocardiographic responses to exercise, 22 of the 54 tested (41 percent) had abnormal esophageal motility, 38 of 60 (63 percent) had one or more psychiatric disorders, and 52 of 60 (87 percent) had their characteristic chest pain provoked by right ventricular electrical stimulation or intracoronary infusion of adenosine. During the treatment phase, the imipramine group had a mean (+/- SD) reduction of 52 +/- 25 percent in episodes of chest pain, the clonidine group had a reduction of 39 +/- 51 percent, and the placebo group a reduction of 1 +/- 86 percent, all as compared with the placebo phase of the trial. Only the improvement with imipramine was statistically significant (P = 0.03). Repeat assessment of sensitivity to cardiac pain while the patients were receiving treatment showed significant improvement only in the imipramine group (P = 0.01). The response to imipramine did not depend on the results of cardiac, esophageal, or psychiatric testing at base line, or on the change in the psychiatric profile during the course of the study, which generally improved in all three study groups. CONCLUSIONS Imipramine improved the symptoms of patients with chest pain and normal coronary angiograms, possibly through a visceral analgesic effect.

Effects of Hormone-Replacement Therapy on Fibrinolysis in Postmenopausal Women

Background Plasma levels of plasminogen-activator inhibitor type 1 (PAI-1), an essential inhibitor of fibrinolysis in humans, increase in women after menopause, and this may contribute to the risk of cardiovascular disease. We studied the effects of hormone-replacement therapy on PAI-1 levels. Methods In a randomized, crossover study, we investigated the effects of oral conjugated estrogen (0.625 mg per day) in 30 postmenopausal women and transdermal estradiol (0.1 mg per day) in 20 postmenopausal women, either alone or in combination with medroxyprogesterone acetate (2.5 mg daily) for one month, on plasma PAI-1 antigen levels. Degradation products of cross-linked fibrin (D-dimer) were measured in serum as an index of fibrinolysis. Results PAI-1 levels were inversely associated with D-dimer levels at base line (r = -0.540, P = 0.002). Conjugated estrogen, both alone and in combination with medroxyprogesterone acetate, reduced mean (±SD) plasma levels of PAI-1 from 32±34 ng per milliliter to 14±10 ng per m...

Hydroxycarbamide in very young children with sickle-cell anaemia: a multicentre, randomised, controlled trial (BABY HUG)

BACKGROUND Sickle-cell anaemia is associated with substantial morbidity from acute complications and organ dysfunction beginning in the first year of life. Hydroxycarbamide substantially reduces episodes of pain and acute chest syndrome, admissions to hospital, and transfusions in adults with sickle-cell anaemia. We assessed the effect of hydroxycarbamide therapy on organ dysfunction and clinical complications, and examined laboratory findings and toxic effects. METHODS This randomised trial was undertaken in 13 centres in the USA between October, 2003, and September, 2009. Eligible participants had haemoglobin SS (HbSS) or haemoglobin Sβ(0)thalassaemia, were aged 9-18 months at randomisation, and were not selected for clinical severity. Participants received liquid hydroxycarbamide, 20 mg/kg per day, or placebo for 2 years. Randomisation assignments were generated by the medical coordinating centre by a pre-decided schedule. Identical appearing and tasting formulations were used for hydroxycarbamide and placebo. Patients, caregivers, and coordinating centre staff were masked to treatment allocation. Primary study endpoints were splenic function (qualitative uptake on (99)Tc spleen scan) and renal function (glomerular filtration rate by (99m)Tc-DTPA clearance). Additional assessments included blood counts, fetal haemoglobin concentration, chemistry profiles, spleen function biomarkers, urine osmolality, neurodevelopment, transcranial Doppler ultrasonography, growth, and mutagenicity. Study visits occurred every 2-4 weeks. Analysis was by intention to treat. The trial is registered with ClinicalTrials.gov, number NCT00006400. FINDINGS 96 patients received hydroxycarbamide and 97 placebo, of whom 83 patients in the hydroxycarbamide group and 84 in the placebo group completed the study. Significant differences were not seen between groups for the primary endpoints (19 of 70 patients with decreased spleen function at exit in the hydroxycarbamide group vs 28 of 74 patients in the placebo group, p=0·21; and a difference in the mean increase in DTPA glomerular filtration rate in the hydroxycarbamide group versus the placebo group of 2 mL/min per 1·73 m(2), p=0·84). Hydroxycarbamide significantly decreased pain (177 events in 62 patients vs 375 events in 75 patients in the placebo group, p=0·002) and dactylitis (24 events in 14 patients vs 123 events in 42 patients in the placebo group, p<0·0001), with some evidence for decreased acute chest syndrome, hospitalisation rates, and transfusion. Hydroxyurea increased haemoglobin and fetal haemoglobin, and decreased white blood-cell count. Toxicity was limited to mild-to-moderate neutropenia. INTERPRETATION On the basis of the safety and efficacy data from this trial, hydroxycarbamide can now be considered for all very young children with sickle-cell anaemia. FUNDING The US National Heart, Lung, and Blood Institute; and the National Institute of Child Health and Human Development.

Design of the multicenter study of hydroxyurea in sickle cell anemia

The Multicenter Study of Hydroxyurea in Sickle Cell Anemia is a randomized double-blind placebo-controlled trial to test whether hydroxyurea can reduce the rate of painful crises in adult patients who have at least three painful crises per year. The sample size of 299 patients yields at least 90% power to detect a 50% or greater reduction in crisis rate. Dosage starts at 15 mg/kg/day and is titrated to the patients maximum tolerated dose up to 35 mg/kg/day. Placebo dosage is titrated in similar fashion to maintain blinding. Attempts are made to ascertain medical contacts for at least 2 years after study entry. The Core Laboratory, Treatment Distribution Center, and Data Coordinating Center collaborate to provide standardized monitoring for toxicity and dose adjustments. The Core Laboratory also reduces the possibility of inadvertent unmasking of treatment assignment during review of hematologic data in clinical centers. An independent Crisis Review Committee classifies clinical events to assure that outcome evaluations are standardized and unbiased by knowledge of treatment assignments. The Data and Safety Monitoring Board assures scientific integrity of the study, as well as the safety and ethical treatment of study patients. We expect the study to determine whether or not treatment with hydroxyurea can offer significant clinical benefit to patients with the most common hereditary disorder among African-Americans in the United States.

Predisposition to Atherosclerosis by Infections Role of Endothelial Dysfunction

Background—Several microorganisms have been implicated in the pathogenesis of atherosclerosis. We hypothesized that infections may predispose to atherosclerosis by inflicting endothelial injury. Methods and Results—Of 375 patients undergoing coronary angiography, 218 had assessment of endothelial function using intracoronary acetylcholine (ACH) and of endothelium-independent function with sodium nitroprusside and adenosine. Immunoglobulin-G antibody titers to cytomegalovirus, Chlamydia pneumoniae, Helicobacter pylori, hepatitis A virus, and herpes simplex virus-1 were measured. Pathogen burden was defined as the number of positive antibodies. Although positive serology to individual pathogens tended to be associated with increased incidence of coronary arteriosclerosis (CAD), the pathogen burden correlated with the presence of CAD, even after adjustment for risk factors (OR 1.3; 95% CI, 1.05 to 1.6, P =0.018). Moreover, the severity of CAD was independently associated with the pathogen burden (P =0.001). Pathogen burden was an independent predictor of endothelial dysfunction, determined as the percent change in coronary vascular resistance in response to ACH (P =0.009) but not the responses to sodium nitroprusside or adenosine. Pathogen burden was also an independent determinant of endothelial function in the subgroup with angiographically normal coronary arteries. Conclusions—The immunoglobulin-G antibody response to multiple pathogens (pathogen burden) is an independent risk factor for endothelial dysfunction and the presence and severity of CAD. Endothelial dysfunction provides the crucial link by which pathogens may contribute to atherogenesis.

Contribution of endothelium-derived nitric oxide to exercise-induced vasodilation.

BackgroundEndothelium-derived nitric oxide is an important modulator of resting vascular tone in animals and humans. However, the contribution of nitric oxide to exercise-induced vasodilation is unknown. Methods and ResultsThe effect of NG-monomethyl-L-arginine (L- NMMA), an inhibitor of nitric oxide synthesis, on exercise-induced vasodilation was studied in 18 healthy subjects (mean ± SD, 40 ± 10 years; 10 women). Acetylcholine was used to test the efficacy of L-NMMA in inhibiting stimulation of nitric oxide synthesis and sodium nitroprusside to test the specificity of L-NMMA in inhibiting endothelium-dependent vasodilation. Intermittent handgrip exercise and infusions of acetylcholine and sodium nitroprusside were performed during intra-arterial infusion of 5% dextrose (control) and L-NMMA (4 to 16 μmol/min). Forearm blood flow was determined by strain-gauge plethysmography. Forearm oxygen extraction was measured from arterial and venous oxygen saturations. In a separate study, 10 subjects performed exercise during infusions of 5% dextrose, L-arginine (the substrate for nitric oxide production), and D-arginine (the stereoisomer that is not a substrate for nitric oxide production). L-NMMA reduced exercise blood flow by 7 ± 13% (P = .04), increased exercise resistance by 18 ± 20% (P = .02), and increased exercise oxygen extraction by 16 ± 17% (P < .001). The degree of inhibition of acetylcholine-induced vasodilation with L-NMMA correlated positively with the degree of reduction in exercise blood flow (r = .55, P = .02). The highest dose of L-NMMA (16 μmol/min) produced the greatest effect; exercise blood flow was reduced by 11 ± 14% (P = .03), and vascular resistance increased by 26 ± 23% (P = .005). L-NMMA did not affect the forearm vasodilation produced by sodium nitroprusside. Exercise blood flow, resistance, and oxygen extraction were not significantly modified by infusions of either L- or D-arginine. ConclusionsInhibition of nitric oxide synthesis reduces exercise-induced vasodilation in the human forearm, indicating that nitric oxide plays a role in exercise-induced vasodilation. Increased availability of nitric oxide substrate does not enhance exercise-induced vasodilation in healthy subjects. These findings have important implications for disease states in which endothelium-derived nitric oxide production is impaired.

Effects of total pathogen burden on coronary artery disease risk and C-reactive protein levels

Infection and inflammation have been suggested to play roles in coronary artery disease (CAD). We hypothesized that: (1) CAD risk is associated with the aggregate number of pathogens (pathogen burden), and (2) increased pathogen burden is associated with elevated levels of C-reactive protein (CRP), a marker of inflammation. We evaluated 233 patients for CAD. Blood samples from each patient were tested for immunoglobulin-G (IgG) antibodies to cytomegalovirus (CMV), Chlamydia pneumoniae, hepatitis A virus (HAV), herpes simplex virus type 1 (HSV-1) and HSV type 2 (HSV-2), and for the CRP levels. Of the 233 study subjects, 68% had evidence of CAD by coronary angiography. Although the prevalence of seropositivity for each pathogen tended to be higher in the patients with CAD than those without, only the association between CAD and seropositivity to HAV was significant in multivariate analysis. Over 75% of study subjects had been exposed to > or =3 of the 5 pathogens tested, and analysis determined that increasing pathogen burden was significantly associated with increasing CAD risk, even after adjustment for traditional CAD risk factors. The prevalence of CAD was 48%, 69%, and 85% in individuals with antibodies to < or =2 pathogens, to 3 or 4 pathogens, and to 5 pathogens, respectively. A similar association between increasing pathogen burden and CRP levels was also found. The pathogen burden remained a significant predictor of CRP levels after multivariate analysis. Our data suggest that infection does play a role in the genesis of atherosclerosis. However, the risk posed by infection is related to the pathogen burden that may contribute to CAD through inflammatory responses.