Myocardial Ribonuclease Activity in Heart Failure
We recently observed that degradation of RNA isolated from myocardium of dogs with heart failure (HF) was abnormally increased. Accordingly, we tested the hypothesis that, compared with healthy myocardium, ribonuclease (RNase) activity was increased in HF occurring due to various etiologies and in various species, including idiopathic and ischemic cardiomyopathies in humans, idiopathic dilated cardiomyopathy in dogs, rapid ventricular pacing in dogs, furazolidone toxicosis in turkeys, and hypothermia in chickens. Activity was determined using a fluorometric, kinetic assay for homogenized myocardium diluted 83-fold with physiologic saline containing 10mg/ml ethidium bromide and yeast RNA as a substrate. The mean activity of myocardial extracts was increased in 6 failing compared with 7 nonfailing human hearts (34.6 ± 6.9 and 62.3 ± 19.1 units/sec), 14 failing compared with 11 nonfailing dog hearts (15.0 ±4.1 and 38.3 ± 6.4 units/esc), 14 failing compared with 13 nonfailing turkey hearts (30.1 ±4.7 and 39.3 ± 4.4units/sec), and 9 failing compared with 5 nonfailing chicken hearts (17.0 ± 3.5 and 25.7 ± 5.4units/sec) In all P < 0.0001). These data indicate that in HF produced from various causes in various species, there is increased myocardial RNase activity. We conclude that the increased RNA degradation that occurs in HF can be attributed to increased RNase activity.
It is generally accepted that there is decreased mRNA content for various key proteins involved in metabolism and Calcium (Ca2+) homeostasis [1,2]. However, it has not been determined to what extent the decrease is attributable to decreased RNA stability compared with decreased gene expression. In a recent study of myocardial mRNA content in dogs with end-stage heart failure (HF) due to idiopathic dilated cardiomyopathy (IDCM) or rapid ventricular pacing (RVP), we observed abnormally increased degradation of the RNA and abnormally increased total and ribosomal RNA . We hypothesized that this was attributable to increased myocardial RNase activity and that it was a consistent feature of HF, regardless of etiology or the species of animal affected.
We used a fluorescent kinetic assay  to determine RNase activity in supernatants of centrifuged myocardial homogenates. Ethidium bromide (EtBr) was used as the RNA dye because it forms a fluorescent complex with nonhydrolyzed RNA but fails to fluoresce in the presence of hydrolyzed RNA. Thus RNase activity can be determined by observing the change in fluorescence over time as a standard amount of RNA is hydrolyzed by the myocardial extract.
We determined RNase activity in nonfailing myocardium from people, dogs, turkey, and chickens, and in myocardium from people with IDCM or ischemic cardiomyopathy (ISCM), from dogs with HF due to IDCM or RVP, from turkeys with furazolidone cardiomyopathy, and from broiler chickens with HF produced by hypothermia and a high-protein diet.
KeywordsIschemic Cardiomyopathy RNase Activity Idiopathic Dilate Cardiomyopathy Decrease Gene Expression Rapid Ventricular Pace
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