Myocardial connexin43 expression in left ventricular hypertrophy resulting from aortic regurgitation.

Intercellular conduction in the working myocardium of the mammalian heart is mediated by gap junctions composed of connexin43 or 45. Recently, it has been shown that myocardial connexin expression is malleable and may be altered with disease. To better understand myocardial conduction in left ventricular hypertrophy resulting from volume overload, we used indirect immunofluorescence microscopy to examine cardiac connexin43 expression in 10 New Zealand white rabbits with surgically induced aortic regurgitation (AR) and in 10 age-matched sham-operated controls. Animals were sacrificed at approximately 1 month or > or =2.5 years after operation. All AR animals developed eccentric hypertrophy; none evidenced heart failure. The heart-to-body weight ratios for the 1 month AR and control groups were 2.9+/-0.8 vs 1.8+/-0.2 g/kg (p < or = 0.01) while ratios for the > or =2.5 year AR and control groups were 2.4+/-0.3 vs 1.9+/-0.3 (p < or = 0.05). No significant differences in posterior wall thickness were found among any of the groups. Although the overall pattern of connexin43-like immunoreactivity was similar for all four groups, staining in the I month AR animals tended to be less than that of age-matched controls; staining was increased in the > or =2.5 year AR animals and was greater than control (p < 0.05), in which staining did not change with animal age. This disease duration-related increase differs from the long-term decrease in connexin43 expression associated with other forms of heart disease and suggests that alterations in connexin expression may play a role in the rhythm abnormalities commonly seen in AR.

Myocardial collagen in cardiac hypertrophy resulting from chronic aortic regurgitation.

Myocardial fibrosis and abnormal myocardial collagen content are common in many forms of pathological cardiac hypertrophy, including that mediated by pressure overload. Recently, in an experimental animal model of chronic aortic regurgitation (AR), we found a strong relation between myocardial fibrosis and congestive heart failure development. To determine if these fibrotic lesions are composed of collagen, as they are in pressure overload, and to determine if potential preventive therapies should be developed similarly in both diseases, we assessed left ventricular collagen content at three time points after AR induction. Moderate to severe AR was induced in 19 New Zealand White rabbits by inserting a catheter through the carotid artery to perforate the aortic valve leaflets. Animals were killed (1) when they showed echocardiographically discernible systolic dysfunction or (2) if normal cardiac function continued, either early (1 month) or late (>3 years) after operation. Fourteen age-matched, sham-operated controls and seven normal unoperated rabbits also were studied. Collagen concentrations were determined biochemically by hydroxyproline measurement. Fibrosis was measured histologically using Mason's trichrome stain and the fibrous collagen-specific stain, Picro-Sirius Red. Our results show an age-related increase in left ventricular collagen concentration with no specific increase among animals with evidence of fibrosis. We conclude that, unlike pressure overload, volume overload produces fibrotic lesions not composed predominantly of excess collagen and that the therapy needed to prevent fibrosis may be different in these conditions. Further study is needed to define the chemical characteristics of the fibrous lesions and the pathophysiological importance of this finding.

Protein turnover in compensated chronic aortic regurgitation.

We recently demonstrated dynamic alterations in protein turnover 3 days and 1 month after surgical induction of aortic regurgitation (AR). To characterize protein synthesis and degradation during the long-term plateau phase, we performed [3H]-leucine infusion 2.5 years after induction of AR in 10 New Zealand White rabbits and 12 sham-operated controls. Protein fractional synthesis rates were obtained by analyses of plasma and protein hydrolysates, growth rates from protein concentration and heart weight measurements, and degradation rates by subtraction of growth from synthesis rates. AR (regurgitant fraction 25 +/- 11%) caused a 57% increase in left ventricular (LV) weight in comparison with controls (7.4 +/- 1.7 vs. 4.7 +/- 0.6 g, p < 0.001) and no evidence of heart failure. Although concentrations of total cardiac protein, myosin heavy chain and actin were similar, the enlarged AR hearts had increased amounts of total cardiac protein (1,009 +/- 312 vs. 682 +/- 120 mg/LV, p < 0.05), myosin heavy chain (148 +/- 91 vs. 81 +/- 29 mg/LV, p < 0.05), and actin (73 +/- 42 vs. 44 +/- 16 mg/LV, p < 0.06). Individual protein fractional synthesis and degradation rates were closely balanced. However, myosin fractional synthesis rates were 152% (p < 0.01) greater than those of total cardiac protein in AR animals, while only 52% (p < 0.05) greater in controls (AR vs. controls, p = 0.05). Variations in actin turnover between AR and control animals did not attain statistical significance. Myosin and actin fractional synthesis rates correlated closely in AR rabbits (R = 0.81, p < 0.02), but not among controls (R = 0.41, NS). Thus, selective alterations in myofibrillar protein turnover contribute to the maintenance of increased myofibrillar protein content in the 'compensatory' LV hypertrophy of chronic AR.

Heart failure due to chronic experimental aortic regurgitation.

Previously reported experimental models of aortic regurgitation generally have manifested normal systolic performance and have not developed heart failure [Magid et al. Am. J. Physiol. 263 (Heart Circ. Physiol. 32): H226-H233, 1992]. To determine whether more severe chronic experimental aortic regurgitation would generate systolic malperformance, heart failure, and emulate the human disease process, 11 New Zealand White rabbits underwent surgical induction of aortic regurgitation and 5 control animals underwent sham operation. Doppler echocardiography was performed serially for up to 3 yr, and pathological studies were performed at necropsy. Left ventricular internal dimension at end diastole increased 80% (P < 0.00002) and left ventricular weight increased 250% (P < 0.0002) in aortic regurgitant rabbits (regurgitant fraction 52 +/- 13%) compared with baseline values. Six of 11 aortic regurgitant animals died with pathological evidence of congestive heart failure at 1.5 +/- 0.8 yr postoperatively; 2 of these developed severe systolic malperformance, manifest as fractional shortenings of 15 and 19% at 1.6 and 1.7 yr, respectively. Five of 11 aortic regurgitant animals survived until killed at 2.9 +/- 0.1 yr. Thus moderate-to-severe chronic aortic regurgitation in rabbits frequently results in heart failure and systolic dysfunction and may usefully model chronic aortic regurgitation and heart failure in humans.

Suppression of protein degradation in progressive cardiac hypertrophy of chronic aortic regurgitation.

BACKGROUND: The heart adapts to the volume overload of aortic regurgitation with dilation and hypertrophy. The development of left ventricular hypertrophy at the protein level is a dynamic process resulting from an imbalance between cardiac protein synthesis and degradation. The objective of the present study was to determine in vivo the relative contributions of cardiac protein synthesis and degradation to the progressive hypertrophy that occurs in response to chronic aortic regurgitation and to compare these with responses earlier in the course of this stress. METHODS AND RESULTS: Continuous intravenous infusions of [3H]-leucine were administered 3 days and 1 month after surgical induction of aortic regurgitation and sham operation in rabbits. Total cardiac protein and myosin heavy chain fractional synthesis rates were obtained by analysis of plasma and protein hydrolysate data using [14C]-dansyl chloride assays. Left ventricular growth rates were determined from serial echocardiographic and postmortem left ventricular weight and protein concentration measurements; protein degradation rates were determined by subtraction of growth rates from synthesis rates. CONCLUSIONS: In comparison with sham-operated control rabbits, protein fractional synthesis rates were increased at 3 days but not at 1 month after induction of aortic regurgitation Progressive cardiac hypertrophy occurring at 1 month was caused by a decrease in protein fractional degradation rates. An increase in protein synthesis contributes only to the early phase of hypertrophy caused by acute aortic regurgitation, whereas progressive eccentric hypertrophy in chronic volume overload is due to suppression of protein degradation.

Myofibrillar protein turnover in cardiac hypertrophy due to aortic regurgitation.

We recently demonstrated that total cardiac protein and myosin heavy chain fractional synthesis rates were not increased during the progressive cardiac hypertrophy that occurred 1 month following induction of aortic regurgitation. The increase in total cardiac protein and myosin heavy chain observed after 1 month of chronic volume overload was caused by a decrease in protein fractional degradation rates. The objective of the present study was to determine in vivo the relative contributions of protein synthesis and degradation of a variety of individual myofibrillar protein constituents, other than myosin heavy chain, to the left ventricular hypertrophic response to chronic aortic regurgitation. Intravenous infusions of [3H]-leucine were administered 3 days and 1 month following surgical induction of aortic regurgitation and sham operation in rabbits, and actin, myosin light chains 1 and 2, alpha-actinin and desmin fractional synthesis rates were obtained by analysis of plasma and protein hydrolysate data using [14C]-dansyl chloride assays. Individual myofibrillar protein growth rates were determined from protein concentration and serial echocardiographic and postmortem left ventricular weight measurements; protein degradation rates were determined by subtraction of growth rates from synthesis rates. Individual myofibrillar protein content increased most rapidly during the 1st week and progressively increased at a slower rate between 1 week and 1 month, in parallel with increases in left ventricular weight. In comparison with sham-operated controls, individual myofibrillar protein fractional synthesis rates were consistently increased at 3 days but not at 1 month. Progressive myocyte hypertrophy occurring at 1 month was caused by a decrease in myofibrillar protein fractional degradation rates. Increased myofibrillar protein synthesis contributed only to the early phase of myocyte hypertrophy while progressive hypertrophy in chronic aortic regurgitation was due to suppression of myofibrillar protein degradation.

Left ventricular diastolic and systolic performance during chronic experimental aortic regurgitation.

To study the time course of left ventricular structural and functional responses to chronic aortic regurgitation, aortic regurgitation was surgically induced in rabbits, and Doppler echocardiography was performed preoperatively and serially postoperatively for up to 2.5 yr. Twenty-five New Zealand White rabbits underwent surgical induction of aortic regurgitation and 13 control animals underwent sham operation. Left ventricular endocardial and epicardial surfaces were digitized from M-mode echocardiograms to measure the rates of change of cavity dimensions and wall thicknesses during diastolic relaxation and systolic contraction. Aortic regurgitant animals developed left ventricular dilatation and eccentric hypertrophy that remained relatively stable throughout the follow-up period. Compared with baseline values, left ventricular mass increased 120% and left ventricular internal dimension at end diastole increased 40%, whereas posterior wall thickness at end diastole and fractional shortening remained relatively stable. Left ventricular diastolic performance was enhanced at 6 mo after operation, a finding associated with increased volume load and heart rate following induction of aortic regurgitation. Diastolic performance was then reduced at 12 mo after operation and demonstrated no further decline throughout the remainder of the follow-up period. In contrast, left ventricular systolic performance was not altered following operation and remained preserved until the final assessment at up to 2.5 yr. Thus alterations in diastolic performance occurred without impairment of systolic performance during long-term follow-up of chronic experimental aortic regurgitation.

High-frequency analysis of the signal-averaged ECG. Correlation with left ventricular mass in rabbits.

Standard electrocardiographic (ECG) criteria have exhibited poor correlation with left ventricular mass and poor sensitivity for left ventricular hypertrophy at acceptable levels of specificity. To assess the ability of the high-frequency filtered signal-averaged ECG to improve ECG correlation with left ventricular mass, signal-averaged orthogonal lead recordings in 29 normal rabbits and seven rabbits with left ventricular hypertrophy due to chronic aortic regurgitation were compared with left ventricular mass corrected for body weight. Voltage of the vector QRS complex was integrated over the total duration of the QRS after separate filtering with standard frequency (0-100 Hz) low-pass and high-frequency (44 Hz) high-pass filters. Measurement of individual X, Y, and Z lead R and S wave voltage was performed on averaged, standard frequency filtered complexes, and the maximal spatial vector magnitude was determined from the standard frequency filtered vectors. Voltage of the 44 Hz high-pass filtered vector QRS complex integrated over the total duration of the QRS (high-frequency vector integral) correlated closely with indexed left ventricular mass (r = 0.84, p less than 0.0001), significantly better than the correlation of standard frequency vector integral or maximal spatial vector magnitude voltages (r = 0.35 and r = 0.61, each p less than 0.01 vs high-frequency vector integral) and the correlation of orthogonal lead X R wave or lead Y S wave voltages (r = 0.55 and r = 0.37, respectively, each p less than 0.01 vs high-frequency vector integral).(ABSTRACT TRUNCATED AT 250 WORDS)

Hypertrophic and functional response to experimental chronic aortic regurgitation.

Aortic regurgitation was induced by retrograde perforation of an aortic valve cusp under hemodynamic guidance in 12 New Zealand White rabbits. Regurgitant fraction was documented by electromagnetic flow probe and six sham-operated animals served as controls. Two-dimensional, M-mode and Doppler echocardiography was performed pre-operatively and serially post-operatively for 3 to 6 months. Animals with aortic regurgitation developed progressive left ventricular dilatation and eccentric hypertrophy. Left ventricular internal dimension at end-diastole and left ventricular mass were increased from baseline values by 41 and 94% (P less than 0.001), respectively; fractional shortening was stable while end-systolic stress increased 50% (P less than 0.01. Thus, acutely induced aortic regurgitation in rabbits results in a chronic model which may be appropriate for stimulation of the hypertrophic response to aortic regurgitation in humans.

Heart rate variability and sudden death secondary to coronary artery disease during ambulatory electrocardiographic monitoring.

Data are analyzed from 5 patients who died suddenly during ambulatory electrocardiographic monitoring. Three of the patients were also assessed in terms of 2 recently developed indexes of heart rate (HR) variability. One of these, the standard deviation of RR intervals during successive 5-minute segments averaged over 24 hours, has been reported to be a putative index of vagal tone. Comparisons were made with HR variability findings in 20 normal volunteers. Sudden death was due to ventricular tachycardia degenerating into ventricular fibrillation in all cases. Both early (3 patients) and late cycle (2 patients) ventricular premature complexes initiated the terminal dysrhythmia. An increased density of ventricular ectopic activity was noted in the hour before onset of ventricular fibrillation. HR variability as measured by the standard deviation was significantly (p less than 0.01) lower in the patients who died suddenly (30 +/- 10 ms) than in the normal subjects (76 +/- 14 ms). These findings support suggestions that HR variability analysis may be useful in identifying patients at a higher risk of sudden death.

Protein synthesis and degradation during starvation-induced cardiac atrophy in rabbits.

To determine the relative importance of protein degradation in the development of starvation-induced cardiac atrophy, in vivo fractional synthetic rates of total cardiac protein, myosin heavy chain, actin, light chain 1, and light chain 2 were measured in fed and fasted rabbits by continuous infusion of [3H] leucine. In addition, the rate of left ventricular protein accumulation and loss were assessed in weight-matched control and fasted rabbits. Rates of total cardiac protein degradation were then estimated as the difference between rates of synthesis and growth. Fasting produced left ventricular atrophy by decreasing the rate of left ventricular protein synthesis (34.8 +/- 1.4, 27.3 +/- 3.0, and 19.3 +/- 1.2 mg/day of left ventricular protein synthesized for 0-, 3-, and 7-day fasted rabbits, respectively). Inhibition of contractile protein synthesis was evident by significant reductions in the fractional synthetic rates of all myofibrillar protein subunits. Although fractional rates of protein degradation increased significantly within 7 days of fasting, actual amounts of left ventricular protein degraded per day were unaffected. Thus, prolonged fasting profoundly inhibits the synthesis of new cardiac protein, including the major protein constituents of the myofibril. Both this inhibition in new protein synthesis as well as a smaller but significant reduction in the average half-lives of cardiac proteins are responsible for atrophy of the heart in response to fasting.

Cardiac protein synthesis and degradation during thyroxine-induced left ventricular hypertrophy.

Assessment of cardiac protein metabolism in thyroxine-induced left ventricular hypertrophy requires measurements of both protein synthesis and degradation. In vivo protein degradative rates can best be measured as the difference between rates of protein synthesis and growth. Accordingly, rates of left ventricular protein accumulation were determined in growing rabbits, and in animals administered intravenous L-thyroxine (200 micrograms X kg-1 X day-1) for up to 15 days. Left ventricular protein fractional synthetic rates in euthyroid and thyroxine-treated rabbits were measured by continuous infusion of [3H]leucine (200 mu Ci/h X 6 h), and results converted to milligrams protein synthesized and degraded per day. Thyroxine administration produced left ventricular hypertrophy by increasing the rate of total protein synthesis (35.7 +/- 2.0, 71.0 +/- 7.0, and 62.6 +/- 4.0 mg of left ventricular protein synthesized per day for 0-, 3-, and 9-day, thyroxine-treated rabbits, respectively). However, the increased rate of total protein synthesis was greater than the measured rate of total protein accumulation (8.1 vs. 15.9 mg protein/day for euthyroid and thyroxine-treated animals), indicating that left ventricular protein degradative rates were increased as well. These studies indicate that accelerated proteolysis may be important in the molecular and architectural remodeling of the rapidly hypertrophying heart during thyrotoxicosis.

The long-term increase of baseline and reflexly augmented levels of human vagal-cardiac nervous activity induced by scopolamine.

We tested the hypothesis that transdermal scopolamine increases vagal-cardiac nervous outflow over the long term in 16 healthy young men. Twenty-four hours after application of one scopolamine patch, the average RR interval was increased by 13% and the average standard deviation of the RR interval (taken as an index of the level of vagal-cardiac nervous activity) was increased by 31%. Baroreceptor-cardiac reflex responsiveness (as reflected by prolongation of RR interval provoked by graded neck suction) also was increased substantially. These findings suggest that vagal-cardiac nervous activity can be augmented pharmacologically in man on a long-term basis. Since vagal outflow influences cardiac electrical properties in an important way, these findings may have therapeutic implications.