Phosphorus nuclear magnetic resonance spectroscopy (P-31 NMR) has been used to assess dynamic aspects of the metabolism of phosphorus-containing compounds in intact cells, organs, and animals. This review describes the NMR experiment and the kinds of information the P-31 NMR spectrum provides for intact, functioning cardiac and skeletal muscles. The P-31 NMR spectrum not only identifies which phosphorus-containing compounds are present in high concentration, namely adenosine 5'-triphosphate (ATP) and creatine phosphate, but also provides information about their chemical environment (including pH) and intracellular distribution. The method is quantitative and nondestructive and permits repetitive measurements in an intact functioning organ. For the perfused heart, it is possible to manipulate the chemical and gaseous composition of the perfusate and to define the effects of, for example, ischemia and reperfusion on the metabolism of ATP and creatine phosphate in the same sample. Using saturation-transfer NMR techniques, it is also possible to measure rates of certain reactions, including creatine kinase and adenylate kinase, in the intact cell. NMR can also be used as an imaging modality.
Several intracellular enzymes have been shown to have altered total activity or isoenzyme composition in cardiac hypertrophy. This study tests the hypothesis that the accumulation of the fetal-type (BB + MB) creatine kinase (CK) isoenzymes in hypertrophied adult myocardium is related to an increase in blood pressure. Consideration was made for the location, size, and hemodynamic load of the myocytes. By using the two-kidney, one-clip (2K1C) rat model of renal hypertension with and without hydralazine treatment, CK (total and isoenzyme), lactate dehydrogenase, and citrate synthase activities and myocyte size were measured. An increased heart weight/body weight ratio occurred in both untreated 2K1C rats (4.15 +/- 0.09) and hydralazine-treated 2K1C rats (4.12 +/- 0.13) as compared with control rats (3.25 +/- 0.10). Blood pressure was high only in untreated 2K1C rats (196 +/- 9 mm Hg), as compared with hydralazine-treated 2K1C rats (142 +/- 6 mm Hg) and control rats (135 +/- 3 mm Hg). Myocytes were isolated from five ventricular regions: left ventricular epicardial and endocardial free wall, left and right halves of the interventricular septum, and right ventricular free wall. Regional differences in normal and hypertrophied myocardium were demonstrated for morphological and biochemical parameters, with the greatest changes occurring in left ventricular endocardium. The shift in CK isoenzyme expression toward accumulating more BB + MB was greater in "hypertensive hypertrophy" (untreated 2K1C rats) than in "nonhypertensive hypertrophy" (hydralazine-treated 2K1C rats). Calculations incorporating isolated myocyte volume showed that the cellular content of total CK remained the same during the hypertrophic process, accounting for a decrease in the tissue activity. Measurement of lactate dehydrogenase and citrate synthase activities suggests that hypertrophied myocardium has relatively higher glycolytic capacity and that this effect is exacerbated in the presence of high blood pressure. We conclude that increased blood pressure is more closely linked to the fetal CK isoenzyme shift than is hypertrophy alone.
Myocardial hypertrophy was produced in the dog by volume overload, secondary to hypertension, and pressure overload to left or right ventricles and in the rat by pressure overload to the left or right ventricles, by elevating thyroxine-levels and secondary to spontaneous hypertension in order to test whether there are changes in the creatine kinase system in hypertrophied heart. Although there was little or no change in total creatine kinase activity, there were changes in the distribution of the creatine kinase isozymes. In the dog, a 4-10-fold increase in the tissue content of MB-creatine kinase was observed for heart chambers with a 40-90% increase in the ratio of ventricular weight to body weight. In the rat, MB-creatine kinase also accumulated in hypertrophied ventricles. For the spontaneously hypertensive rat, the correlation between increased fetal creatine kinase isozymes and increased ratio of ventricular weight to body weight was excellent (r approximately 0.92). During the transition from compensated hypertrophy to failure in the spontaneously hypertensive rat, there is a 50% decrease in mitochondrial creatine kinase activity. P-31 NMR magnetization transfer experiments suggest that flux through the creatine kinase reaction is 3-fold lower than normal in these failing hearts. These results show that there are changes in the distribution of the creatine kinase isozymes in hypertrophied heart and suggest that one of these changes characterizes compensated hypertrophy (increased fetal-type creatine kinase isozymes) while another characterizes the transition to failure (decreased mitochondrial creatine kinase).
The purpose of this study was to determine if biochemical, functional, and ultrastructural abnormalities persist in nonnecrotic postischemic myocardium salvaged by coronary reperfusion. Anesthetized dogs were subjected to 15 min of occlusion of the left anterior descending (LAD) coronary artery followed by 3 days of reperfusion. Biopsies were obtained for measurement of adenosine 5'-triphosphate (ATP) and creatine phosphate (CP) nmol/mg protein), and regional function was evaluated using sonomicrometry. Myocardial ATP concentration after 15 min of occlusion was 37 +/- 1 nmol/mg cardiac protein in nonischemic subendocardium and 19 +/- 2 nmol/mg in ischemic subendocardium. After the hearts underwent 90 min and 72 h of reperfusion, ATP remained significantly depressed in reperfused subendocardium with values of 25 +/- 5 and 29 +/- 2 nmol/mg, respectively (P less than 0.05 and P less than 0.01 compared with the nonischemic zone in which ATP remained normal). CP levels fell during ischemia but returned to normal by 90 min of reperfusion. Percent systolic shortening of myocardial segments fell from +18 +/- 1% (active shortening) to -13 +/- 2% (passive lengthening) during ischemia and was still significantly depressed at +11 +/- 1% (P less than 0.05 vs. preocclusion) at 72 h of reperfusion. Histological examination showed no necrosis, but ultrastructural abnormalities were present. Therefore brief periods of myocardial ischemia are not associated with necrosis but result in functional, biochemical, and ultrastructural abnormalities, which are present for at lest 3 days after coronary reperfusion.
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