William A. Clark

Isoproterenol-induced myocardial fibrosis in relation to myocyte necrosis.

Treatment of rats with the β-adrenergic agonist isoproterenol results in cardiac hypertrophy, myocyte necrosis, and interstitial cell fibrosis. Our objectives in this study have been to examine whether hypertrophy and fibrosis occur in a compensatory and reparative response to myocyte loss or whether either process may be occurring independently of myocyte loss and thus be a reactive response to adrenergic hormone stimulation. We have examined this question by evaluating each of these responses in rats treated with different doses and forms of isoproterenol administration. Myocyte necrosis was evaluated using in vivo labeling with monoclonal antimyosin for identification of myocytes with permeable sarcolemma, which was indicative of irreversible injury. Myocardial fibrosis was evaluated by morphometric point counting of Gomori-stained tissue sections and by assessment of the stimulation of fibroblast proliferation by determination of increased levels of DNA synthesis. Stimulation of fibroblast DNA synthesis was determined from DNA specific radioactivities and radioautography after pulse labeling with [3H]thymidine. The evidence provided by this study suggests that the degree and timing of myocardial hypertrophy does not follow the course of myocyte loss and, thus, appears to be either a response to altered cardiac loading or a reactive response to β-adrenergic hormone stimulation rather than a compensation for myocyte loss. Myocardial fibrosis, on the other hand, appears to be more closely related to myocyte necrosis with respect to collagen accumulation in the same areas of the heart, its dose-response relation to the amount of isoproterenol administered, and the timing of increased DNA synthesis, or fibroblast proliferation, after myocyte loss.

Physiologic Versus Pathologic Hypertrophy and the Pressure-Overloaded Myocardium

The myocardium consists of myocytes and capillaries embedded in a connective tissue matrix. Myocardial mass, which is predominantly a function of myocyte size, is determined by systolic tension: when systolic pressure is gradually elevated above the normal range, mass will increase. The hypertrophic process is a continuum consisting of subtle transitions that take place within the muscular, collagenous, and vascular compartments; these transitions, however, need not be temporarily concordant. We would identify three phases to the hypertrophic process. First, there is an evolutionary phase, whereby the structural and biochemical remodeling of the various compartments of the myocardium is in transition, with each compartment having its own rate of adjustment. During this evolutionary phase, myocardial contractility, as reflected by stress-length and stress-velocity relations, may or may not be normal, but ventricular pump function and O2 delivery are preserved. Second, there is a physiologic phase during which the structural and biochemical remodeling of the compartments reaches a coordinated balance. The myocardial stress-length relation and ventricular function are each normal, but rate-dependent indices of contractility may be abnormal. During the physiologic phase of hypertrophy, the remodeled myocardium will revert to normal when the abnormal loading condition is removed. Finally, there is a pathologic phase. In this phase, compartment remodeling is no longer balanced (e.g., the ratio of structural versus maintenance proteins), and length and rate-dependent indices of myocardial contractility are depressed. Ventricular pump function is also abnormal in the pathologic phase: consequently, O2 delivery to the tissues is impaired. This imbalance in O2 demand and supply may be apparent at rest in more advanced expressions of disease or may appear during the physiologic stress of exercise in less severe disease. In the latter case, the patients aerobic capacity is reduced to the extent that it can be used to grade the severity of heart failure and to predict the cardiac reserve. During the pathologic phase of hypertrophy, the structural and biochemical remodeling of the myocardium may be irreversible, although this may not be the case for each compartment. Finally, it is important to distinguish cardiac (or myocardial) failure from the clinical syndrome of congestive heart failure. The latter arises from congested organs and hypoperfused tissues; its clinical manifestations are dependent on the activation of the adrenergic nervous and renin-angiotensin-aldosterone systems and the presence of a salt-avid kidney. Congestive heart failure is a late clinical feature of chronic pressure overload and pathologic hypertrophy.

Intermittent inotropic therapy in an outpatient setting: a cost-effective therapeutic modality in patients with refractory heart failure.

Patients with intractable heart failure (New York Heart Association [NYHA] class III and IV) who were receiving maximal conventional treatment were enrolled in an outpatient program that included inotropic infusions, intensive patient education, and close follow-up. The effects of this approach to therapy were evaluated on (1) the number of hospital admissions, (2) length of stay, and (3) number of emergency room visits during the ensuing year. These data were compared with similar data from the year before entry in the program for each patient. Thirty-six patients with stable NYHA class III and IV heart failure received milrinone or dobutamine to manage chronic heart failure in an outpatient setting. The cause of heart failure was ischemic heart disease in 12, idiopathic in 11, hypertension in 8, and pulmonary hypertension in 5. Four patients received dobutamine and 32 patients received milrinone. The mean period of observation was 294 days. For the period before entry in the program, patients had 21 emergency room visits, 75 admissions, and 528 days spent in the hospital. After enrollment, patients had 10 emergency room visits, 34 admissions, and 150 days spent in the hospital. In conclusion, this therapeutic regimen reduced the number of hospital admissions, days spent in the hospital, and emergency room visits. Our study supports the concept that the use of intermittent inotropic therapy in the outpatient setting plays an important role in managing this severely ill group of patients.

Effect of long-term β-blockade on aortic root compliance in patients with Marfan syndrome

Abstract Background This study was undertaken to assess the effect of long-term β-blockade on the aortic root stiffness index and distensibility in patients with Marfan syndrome. Methods Aortic root stiffness index and distensibility were calculated according to the formulas of Stefanadis and Hirai, respectively, with 2-dimensional guided M-mode echocardiogram before and after an average of 26 months of atenolol administration. Results Twenty-three asymptomatic patients were studied (11 men and 12 women, aged 31 ± 14.2 years). The follow-up was 4 ± 2.2 years. The dose of atenolol was individualized (mean 43.5 ± 21.6 mg/d). Heart rate decreased from 79 ± 9 beats/min to 64 ± 9 beats/min ( P = .01), and systolic blood pressure decreased from 124 ± 13 mm Hg to 114 ± 2 mm Hg ( P = .01). Distensibility increased from 1.85 ± 0.70 × 10 –6 cm 2 /dynes –1 to 2.21 ± 0.76 × 10-6 cm 2 /dynes –1 ( P = .02), and the stiffness index decreased from 9.68 ± 3.78 to 8.85 ± 3.15 ( P = .2). Two groups of responses to treatment were identified. Compared with baseline values 15 (65%) patients who responded to treatment had increased distensibility and decreased stiffness index of the aortic root ( P = .05). Eight patients (35%) who did not respond to treatment had no significant change. Body weight >91 kg and baseline end-diastolic aortic root diameter >40 mm were significantly associated with no response ( P = .05). Two patients in the nonresponding group had echocardiographic progression of aortic insufficiency. Conclusions There was a heterogeneous response in the aortic root elastic properties after long-term treatment with atenolol in asymptomatic patients with Marfan syndrome. Stiffness index and distensibility are more likely to respond when the baseline end-diastolic aortic root diameter is

Characterization of adrenoceptor involvement in skeletal and cardiac myotoxicity induced by sympathomimetic agents: Toward a new bioassay for β-blockers

Excessive levels of catecholamines have long been known to be cardiotoxic, but less well known are their toxic effects on skeletal muscle. By using an antimyosin monoclonal antibody and quantitative methods to measure the extent of myocyte necrosis, and by employing modulators of adrenoceptors (ARs), including clenbuterol, bupranolol, propranolol, bisoprolol, atenolol, ICI-118551, phenoxybenzamine, prazosin, and yohimbine, the involvement of ARs in isoproterenol-induced myotoxicity was characterized. In the myocardium, the toxic effects were predominantly mediated via the &bgr;1-ARs. In the soleus muscle, it was almost solely via the &bgr;2-ARs. Myotoxicity was also observed in the myocardium when challenged with the &bgr;2-AR agonist clenbuterol. This was found to be mediated via sympathetic presynaptic &bgr;2-ARs, leading to enhanced release of norepinephrine. This effect was abolished by prior treatment with reserpine. The skeletal muscle was found to be more sensitive to the myotoxic effects than cardiac muscle at lower doses of &bgr;-AR agonists. These experiments introduce a new way of assaying &bgr;-AR antagonists by classifying them according to their ability to prevent catecholamine-induced myotoxicity. Further research along these lines may deepen understanding of which &bgr;-blockers work best in heart failure therapy.

Dose-dependent separation of the hypertrophic and myotoxic effects of the β2-adrenergic receptor agonist clenbuterol in rat striated muscles.

Muscle growth in response to large doses (milligrams per kilogram) of β2‐adrenergic receptor agonists has been reported consistently. However, such doses may also induce myocyte death in the heart and skeletal muscles and hence may not be safe doses for humans. We report the hypertrophic and myotoxic effects of different doses of clenbuterol. Rats were infused with clenbuterol (range, 1 μg to 1 mg.kg−1) for 14 days. Muscle protein content, myofiber cross‐sectional area, and myocyte death were then investigated. Infusions of ≥10 μg.kg−1.d−1 of clenbuterol significantly (P < 0.05) increased the protein content of the heart (12%–15%), soleus (12%), plantaris (18%–29%), and tibialis anterior (11%–22%) muscles, with concomitant myofiber hypertrophy. Larger doses (100 μg or 1 mg) induced significant (P < 0.05) myocyte death in the soleus (peak 0.2 ± 0.1% apoptosis), diaphragm (peak 0.15 ± 0.1% apoptosis), and plantaris (peak 0.3 ± 0.05% necrosis), and significantly increased the area fraction of collagen in the myocardium. These data show that the low dose of 10 μg.kg−1.d−1 can be used in rats to investigate the anabolic effects of clenbuterol in the absence of myocyte death. Muscle Nerve, 2006

Dose-dependent apoptotic and necrotic myocyte death induced by the β2-adrenergic receptor agonist, clenbuterol

We have investigated the dose‐ and time‐dependency of myocyte apoptosis and necrosis induced by the β2‐adrenergic receptor agonist, clenbuterol, with the aim of determining whether myocyte apoptosis and necrosis are two separate processes or a continuum of events. Male Wistar rats were administered subcutaneous injections of clenbuterol, and immunohistochemistry was used to detect myocyte‐specific apoptosis and necrosis. Myocyte apoptosis peaked 4 h after, and necrosis 12 h after, clenbuterol administration. In the soleus, peak apoptosis (5.8 ± 2.0%; P < 0.05) was induced by 10 μg and peak necrosis (7.4 ± 1.7%; P < 0.05) by 5 mg.kg−1 clenbuterol. Twelve hours after clenbuterol administration, 73% of damaged myocytes labeled as necrotic, 27% as apoptotic and necrotic, and 0% as purely apoptotic. Administrations of clenbuterol (10 μg.kg−1) at 48‐h intervals induced cumulative myocyte death over 8 days. These data show that the phenotype of myocyte death is dependent on the magnitude of the insult and the time at which it is investigated. Only very low doses induced apoptosis alone; in most cases apoptotic myocytes lysed and became necrotic and the magnitude of necrosis was greater than that of apoptosis. Thus, it is important to investigate both apoptotic and necrotic myocyte death, contrary to the current trend of only investigating apoptotic cell death. Muscle Nerve, 2005

Electrocardiographic patterns of patients with echocardiographically determined biventricular hypertrophy.

Abstract The numerous criteria proposed for the electrocardiographic (ECG) diagnosis of biventricular hypertrophy (BVH) suffer from inadequate correlative data. We used two-dimensional (2D) echocardiography to identify BVH and analyzed the ECG patterns in these patients. The study group had 69 such patients with BVH and the control group had 22 patients with isolated left ventricular hypertrophy (LVH) demonstrated by 2D echocardiography. The electrocardiograms were analyzed for the presence of established criteria used in the diagnosis of LVH and right ventricular hypertrophy (RVH). Of the 69 patients in the study group, 17 (25%) had EGG findings of BVH, 25 (36%) had LVH, and 14 (20%) had RVH. An S wave in V5/V6 of >7 mm was most the frequent finding in the 17 patients with BVH on the electrocardiogram. The sensitivity of ECG criteria for BVH was 24.6%, specificity was 86.4%, and positive predictive value was 85%. This study reemphasizes the difficulty of ECG diagnosis of BVH. The electrocardiogram has a low sensitivity but satisfactory specificity and positive predictive accuracy for BVH.