Untargeted metabolomics analysis of ischemia-reperfusion-injured hearts ex vivo from sedentary and exercise-trained rats.

Introduction: The effects of exercise on the heart and its resistance to disease are well-documented. Recent studies have identified that exercise-induced resistance to arrhythmia is due to the preservation of mitochondrial membrane potential. Objectives: To identify novel metabolic changes that occur parallel to these mitochondrial alterations, we performed non-targeted metabolomics analysis on hearts from sedentary and exercise-trained rats challenged with isolated heart ischemia-reperfusion injury (I/R). Methods: Eight-week old Sprague-Dawley rats were treadmill trained 5 days/week for 6 weeks (exercise duration and intensity progressively increased to 1 h at 30 m/min up a 10.5% incline, 75-80% VO2max). The recovery of pre-ischemic function for sedentary rat hearts was 28.8 ± 5.4% (N = 12) compared to exercise trained hearts, which recovered 51.9% ± 5.7 (N = 14) (p < 0.001). Results: Non-targeted GC-MS metabolomics analysis of (1) sedentary rat hearts; (2) exercise-trained rat hearts; (3) sedentary rat hearts challenged with global ischemia-reperfusion (I/R) injury; and (4) exercise-trained rat hearts challenged with global I/R (10/group) revealed 15 statistically significant metabolites between groups by ANOVA using Metaboanalyst (p < 0.001). Enrichment analysis of these metabolites for pathway-associated metabolic sets indicated a > 10-fold enrichment for ammonia recycling and protein biosynthesis. Subsequent comparison of the sedentary hearts post-I/R and exercise-trained hearts post-I/R further identified significant differences in three metabolites (oleic acid, pantothenic acid, and campesterol) related to pantothenate and CoA biosynthesis (p ≤ 1.24E-05, FDR ≤ 5.07E-4). Conclusions: These studies shed light on novel mechanisms in which exercise-induced cardioprotection occurs in I/R that complement both the mitochondrial stabilization and antioxidant mechanisms recently described. These findings also link protein synthesis and protein degradation (protein quality control mechanisms) with exercise-linked cardioprotection and mitochondrial susceptibility for the first time in cardiac I/R.

Exercise-Induced Alterations in Skeletal Muscle, Heart, Liver, and Serum Metabolome Identified by Non-Targeted Metabolomics Analysis.

BACKGROUND: The metabolic and physiologic responses to exercise are increasingly interesting, given that regular physical activity enhances antioxidant capacity, improves cardiac function, and protects against type 2 diabetes. The metabolic interactions between tissues and the heart illustrate a critical cross-talk we know little about. METHODS: To better understand the metabolic changes induced by exercise, we investigated skeletal muscle (plantaris, soleus), liver, serum, and heart from exercise trained (or sedentary control) animals in an established rat model of exercise-induced aerobic training via non-targeted GC-MS metabolomics. RESULTS: Exercise-induced alterations in metabolites varied across tissues, with the soleus and serum affected the least. The alterations in the plantaris muscle and liver were most alike, with two metabolites increased in each (citric acid/isocitric acid and linoleic acid). Exercise training additionally altered nine other metabolites in the plantaris (C13 hydrocarbon, inosine/adenosine, fructose-6-phosphate, glucose-6-phosphate, 2-aminoadipic acid, heptadecanoic acid, stearic acid, alpha-tocopherol, and oleic acid). In the serum, we identified significantly decreased alpha-tocopherol levels, paralleling the increases identified in plantaris muscle. Eleven unique metabolites were increased in the heart, which were not affected in the other compartments (malic acid, serine, aspartic acid, myoinositol, glutamine, gluconic acid-6-phosphate, glutamic acid, pyrophosphate, campesterol, phosphoric acid, creatinine). These findings complement prior studies using targeted metabolomics approaches to determine the metabolic changes in exercise-trained human skeletal muscle. Specifically, exercise trained vastus lateralus biopsies had significantly increased linoleic acid, oleic acid, and stearic acid compared to the inactive groups, which were significantly increased in plantaris muscle in the present study. CONCLUSIONS: While increases in alpha-tocopherol have not been identified in muscle after exercise to our knowledge, the benefits of vitamin E (alpha-tocopherol) supplementation in attenuating exercise-induced muscle damage has been studied extensively. Skeletal muscle, liver, and the heart have primarily different metabolic changes, with few similar alterations and rare complementary alterations (alpha-tocopherol), which may illustrate the complexity of understanding exercise at the organismal level.

Comparison of Estimations Versus Measured Oxygen Consumption at Rest in Patients With Heart Failure and Reduced Ejection Fraction Who Underwent Right-Sided Heart Catheterization.

Cardiac output during right-sided heart catheterization is an important variable for patient selection of advanced therapies (cardiac transplantation and left ventricular assist device implantation). The Fick method to determine cardiac output is commonly used and typically uses estimated oxygen consumption (VO2) from 1 of 3 published empirical formulas. However, these estimation equations have not been validated in patients with heart failure and reduced ejection fraction (HFrEF). The objectives of the present study were to determine the accuracy of 3 equations for estimating VO2 compared with direct measurement of VO2 and determine the extent clinically significant error occurred in calculating cardiac output of patients with HFrEF. Breath-by-breath measurements of VO2 from 44 patients who underwent cardiac catheterization (66% men; age, 65 ± 11 years, left ventricular ejection fraction, 22 ± 6%) were compared with the derived estimations of LaFarge and Miettinen, Dehmer et al, and Bergstra et al. Single-sample t tests found only the mean difference between the estimation of LaFarge and Miettinen and the measured VO2 to be nonsignificant (-10.3 ml/min ± 6.2 SE, p = 0.053). Bland-Altman plots demonstrated unacceptably large limits of agreement for all equations. The rate of ≥25% error in the equations by LaFarge and Miettinen, Dehmer et al, and Bergstra et al occurred in 11%, 23%, and 45% of patients, respectively. Misclassification of cardiac index derived from each equation for 2 clinically important classifications: cardiogenic shock-21%, 23%, and 32% and hypoperfusion-16%, 16%, and 25%; respectively. In conclusion, these findings do not support the use of these empiric formulas to estimate the VO2 at rest in patients with HFrEF who underwent right-sided heart catheterization.

Exercise alters the regulation of myocardial Na(+)/H(+) exchanger-1 activity.

The myocardial Na(+)/H(+) exchanger-1 (NHE1) plays a major role in regulation of intracellular pH, and its upregulation has been implicated in increased ischemia-reperfusion injury and other pathologies. Hydrogen peroxide (H2O2) increases NHE1 activity acutely via ERK1/2 signaling. Chronic strenuous exercise upregulates NHE1 in skeletal muscle, but we hypothesize this will not occur in the heart, because exercise creates a cardioprotective phenotype. NHE1 activity and its regulation by H2O2 were examined at physiological pH using isolated cardiomyocytes from female Sprague-Dawley rats exercised on a treadmill for 5 wk (E; n = 11). Compared with sedentary (S; n = 15), E displayed increases (P < 0.05) in heart-to-body weight ratio (6.8%) and plantaris mitochondria content (89%). NHE1 activity (acid efflux rate following an acid load) was 209% greater in E (0.65 ± 0.12 vs. 2.01 ± 0.29 fmol/min). The difference was attributed primarily to greater cell volume (22.2 ± 0.6 vs. 34.3 ± 1.1 pl) and intracellular pH-buffering capacity (33.94 ± 1.59 vs. 65.82 ± 5.20 mM/pH unit) of E myocytes. H2O2 stimulation (100 µM) raised NHE1 activity significantly less in E (45%) than S (167%); however, activity remained 185% greater in E. ERK1/2 inhibition abrogated the increases. H2O2-stimulated ERK1/2 phosphorylation levels normalized to total ERK1/2 were similar between groups. Content of NHE1 and activities of H2O2 scavengers were also similar. We observed that intracellular pH-buffering capacity differences between groups became progressively less with declining pH, which may be an exercise-induced cardioprotective adaptation to lower NHE1 activity during certain pathological situations. We conclude that strenuous endurance exercise increases myocardial NHE1 activity at physiological pH, which would likely enhance cardiac performance under physiological conditions.

Myocardial Na+/H+ exchanger-1 (NHE1) content is decreased by exercise training

The effect of exercise training on myocardial Na+/H+ exchanger-1 (NHE1) protein expression was examined. Adult female Sprague–Dawley rats were randomly divided into sedentary (S; n = 8) and exercised (E; n = 9) groups. Twenty-four hours after the last exercise bout, hearts were weighed and connected to an isolated perfused working heart apparatus for evaluation of cardiac functional performance. Heart weight and heart weight/body weight from E rats was significantly increased by 7.1 and 7.2 % (P < 0.05), respectively, compared with S hearts. The E hearts displayed 15 % greater cardiac output and 35 % external cardiac work compared with the S group at both low and high workloads (P < 0.05 for both parameters). Left ventricular tissue from the same hearts was homogenized and NHE1 and Na+/Ca2+ exchanger (NCX) content determined by Western blotting. E hearts had a 38 % (P < 0.001) reduction in NHE1 content related to S hearts, and there was no difference in NCX content between groups. Cytochrome c oxidase activity in plantaris increased by 100 % (P < 0.05) and was assessed as a marker of mitochondria content and to verify training status. Our data indicate that exercise training at an intensity that results in cardiac hypertrophy and improved performance is accompanied by decreased NHE1 content in heart (AU)

Synergistic effect of exercise and statins on femoral strength in rats.

It is now widely recognized that in order to optimize bone health in the later years, bone healthy behaviors should begin at a young age and continue throughout life. Prescribed orally to lower lipid levels in adults of all ages, statins have also been shown to stimulate bone formation in vitro by promoting bone morphogenic protein-2 (BMP-2) activity and to stimulate bone formation in vivo. Weight bearing exercise is well known to stimulate bone formation through a mechanism whereby mechanical loading is 'sensed' by the mechano-sensors leading to a cascade of events involving the activation of osteoblasts. For individuals with high cholesterol levels, both of these interventions are recommended throughout adult life. Since statins and exercise stimulate bone formation via different mechanisms, we hypothesized that exercise in combination with oral simvastatin synergistically increases bone mineral density and strength. Mature adult female, Sprague Dawley rats were divided into 4 groups: control (n=9), statin only (n=8), exercise only (n=11), and statin plus exercise (n=11). Simvastatin was given to the two groups at a dose of 10 mg/kg/day in standard rat chow for the entire 5 week period. All rats ate the same mass of food. The two exercise groups ran on a treadmill with progressively greater speeds and time, ending on week 5 at 30 m/min for 60 min. After 5 weeks, rats were euthanized, and excised femurs were scanned for areal bone mineral density (BMD) and tested by three point bending to obtain the following performance measures: maximum force (strength), stiffness, and work-to-fracture. Only the group treated with statins and exercise showed a positive effect on the biomechanical performance of the femurs. Compared to controls, this group had increased maximum force, stiffness, moment of inertia, and BMD. Linear regression analysis revealed that the increased performance was related to increased BMD. We conclude that the combination of oral statins and appropriate exercise increases bone strength better than either individual treatment and may provide optimal protection against osteoporosis.

Myocardial Na+/H+ exchanger-1 (NHE1) content is decreased by exercise training.

The effect of exercise training on myocardial Na(+)/H(+) exchanger-1 (NHE1) protein expression was examined. Adult female Sprague-Dawley rats were randomly divided into sedentary (S; n = 8) and exercised (E; n = 9) groups. Twenty-four hours after the last exercise bout, hearts were weighed and connected to an isolated perfused working heart apparatus for evaluation of cardiac functional performance. Heart weight and heart weight/body weight from E rats was significantly increased by 7.1 and 7.2 % (P < 0.05), respectively, compared with S hearts. The E hearts displayed 15 % greater cardiac output and 35 % external cardiac work compared with the S group at both low and high workloads (P < 0.05 for both parameters). Left ventricular tissue from the same hearts was homogenized and NHE1 and Na(+)/Ca(2+) exchanger (NCX) content determined by Western blotting. E hearts had a 38 % (P < 0.001) reduction in NHE1 content related to S hearts, and there was no difference in NCX content between groups. Cytochrome c oxidase activity in plantaris increased by 100 % (P < 0.05) and was assessed as a marker of mitochondria content and to verify training status. Our data indicate that exercise training at an intensity that results in cardiac hypertrophy and improved performance is accompanied by decreased NHE1 content in heart.

Reactive oxygen species are not a required trigger for exercise-induced late preconditioning in the rat heart.

Reactive oxygen species (ROS) have been reported to play a primary role in triggering the cardioprotective adaptations by some preconditioning procedures, but whether they are required for exercise-induced preconditioning is unclear. Thus in this study we used the free radical scavenger N-(2-mercaptopropionyl)glycine (MPG) to test the hypothesis that ROS is the trigger for exercise-induced preconditioning of the heart against ischemia-reperfusion injury. Male F344 rats were assigned to four groups: sedentary (SED, n = 7), SED/MPG (100 mg/kg ip daily for 2 days, n = 12), exercised on a treadmill for 2 days at 20 m/min, 6° grade, for 60 min (RUN, n = 7), and RUN/MPG with 100 mg/kg MPG injected 15 min before exercise (n = 10). Preliminary experiments verified that MPG administration maintained myocardial redox status during the exercise bout. Twenty-four hours postexercise or MPG treatment isolated perfused working hearts were subjected to global ischemia for 22.5 min followed by reperfusion for 30 min. Recovery of myocardial external work (percentage of preischemic systolic pressure times cardiac output) for SED (50.4 ± 4.5) and SED/RUN (54.7 ± 6.6) was similar and improved in both exercise groups (P < 0.05) to 77.9 ± 3.0 in RUN and 76.7 ± 4.5 in RUN/MPG. A 2 × 2 ANOVA also revealed that exercise decreased lactate dehydrogenase release from the heart during reperfusion (marker of cell damage) without MPG effects or interactions. Expression of the cytoprotective protein inducible heat shock protein 70 increased by similar amounts in the left ventricles of RUN and RUN/MPG compared with sedentary groups (P < 0.05). We conclude that ROS are not a necessary trigger for exercise-induced preconditioning in rats.

Effect of N-2-mercaptopropionyl glycine on exercise-induced cardiac adaptations.

The purpose of this study was to test the hypothesis that exercise-induced cardiac adaptations would be attenuated by the free radical scavenger N-2-mercaptopropionyl glycine (MPG). Male Sprague-Dawley rats were divided into four groups (n = 9-13 per group) for 3-4 wk: sedentary (S), S+MPG (100 mg/kg ip daily), exercised on a treadmill (E) (60 min/day, 5 days/wk, at a speed of 20 m/min up a 6° grade in a 6°C room), or E+MPG given 10 min prior to exercise. Additional rats (n = 55) were used to determine acute exercise effects on myocardial redox state [nonprotein nonglutathione sulfhydryls (NPNGSH)] and PI3K/Akt signaling pathway activation. Compared with S, NPNGSH levels were 48% lower in E (P < 0.05) and unchanged in E+MPG (P > 0.05). MPG also attenuated exercise-induced activation of the signaling proteins Akt and S6. Hearts from the 4-wk groups were weighed, and cardiac function was evaluated using an isolated perfused working heart preparation. Similar increases (P < 0.05) in both exercised groups were observed for heart weight and heart weight-to-body weight ratio. Cardiac function improved in E vs. S, as indicated by greater (P < 0.05) external work performed (cardiac output × systolic pressure) and efficiency of external work (work/Vo(2)). MPG prevented these exercise-induced functional improvements. Skeletal muscle mitochondria content increased to similar levels in E and E+MPG. This study provides evidence that free radicals do not play an essential role in the development of exercise-induced cardiac hypertrophy; however, they appear to be involved in functional cardiac adaptations, which may be mediated through the PI3K/Akt pathway.

Rapamycin, but not resveratrol or simvastatin, extends life span of genetically heterogeneous mice.

Rapamycin was administered in food to genetically heterogeneous mice from the age of 9 months and produced significant increases in life span, including maximum life span, at each of three test sites. Median survival was extended by an average of 10% in males and 18% in females. Rapamycin attenuated age-associated decline in spontaneous activity in males but not in females. Causes of death were similar in control and rapamycin-treated mice. Resveratrol (at 300 and 1200 ppm food) and simvastatin (12 and 120 ppm) did not have significant effects on survival in male or female mice. Further evaluation of rapamycin's effects on mice is likely to help delineate the role of the mammalian target of rapamycin complexes in the regulation of aging rate and age-dependent diseases and may help to guide a search for drugs that retard some or all of the diseases of aging.

Effect of storage conditions on lactate dehydrogenase released from perfused hearts.

BACKGROUND: Release of lactate dehydrogenase (LDH) is a widely used marker of tissue damage. A review of the literature indicates that after collection of coronary effluents from hearts, a variety of storage conditions have been utilized prior to measurement of LDH activity. The purpose of this study was to determine whether storage conditions reported in the literature affect LDH activity. METHODS: Isolated rat hearts perfused with Krebs-Henseleit buffer (KHB) were subjected to 30 min of global ischemia followed by normoxic reperfusion to generate tissue damage and elevate LDH release. Coronary effluents were collected and stored at 4 degrees C, 23 degrees C, -20 degrees C, and -80 degrees C prior to measurement of LDH activity. RESULTS: After 24 h no decline in LDH occurred at 4 degrees C storage (P>0.05), but a 42% decline occurred at 23 degrees C (P<0.05). After 48 h, activity declined (P<0.05) 11% at 4 degrees C and 98% at 23 degrees C. Frozen storage resulted in a 40% loss at -80 degrees C and a 79% loss at -20 degrees C (P<0.05). CONCLUSIONS: Short-term storage in Krebs-Henseleit buffer for 24 h at 4 degrees C does not affect LDH activity and frozen storage should be avoided.

Exercise-induced cardioprotection: endogenous mechanisms.

It is now well established that exercise can result in cardioprotection against ischemia-reperfusion (I-R) injury; however, the adaptations within the heart that provide the protection are still in doubt. The cytoprotective proteins receiving the most attention to date are antioxidant enzymes and heat shock proteins. The extent of I-R injury is dependent on the interactions of several events, including energy depletion, metabolite accumulation, oxidant stress, and calcium overload. Adaptations that directly influence any of these could affect I-R outcome. Thus, the exercise-induced cardioprotective phenotype is likely to include additional cytoprotective proteins beyond antioxidant enzymes or heat shock proteins. In this review, we will consider evidence for some of these in the cytosol, mitochondria, and sarcolemma of the cardiomyocyte. We will not consider potentially important adaptations within vascular tissue or the autonomic nervous system. Results of recent studies support the hypothesis that exercise leads to cardioprotective adaptations that are unique from other forms of preconditioning against I-R injury.

Exercise training decreases rat heart mitochondria free radical generation but does not prevent Ca2+-induced dysfunction.

Exercise provides cardioprotection against ischemia-reperfusion injury, a process involving mitochondrial reactive oxygen species (ROS) generation and calcium overload. This study tested the hypotheses that isolated mitochondria from hearts of endurance-trained rats have decreased ROS production and improved tolerance against Ca(2+)-induced dysfunction. Male Fischer 344 rats were either sedentary (Sed, n = 8) or endurance exercise trained (ET, n = 11) by running on a treadmill for 16 wk (5 days/wk, 60 min/day, 25 m/min, 6 degrees grade). Mitochondrial oxidative phosphorylation measures were determined with glutamate-malate or succinate as substrates, and H(2)O(2) production and permeability transition pore (PTP) opening were determined with succinate. All assays were carried out in the absence and presence of calcium. In response to 25 and 50 microM CaCl(2), Sed and ET displayed similar decreases in state 3 respiration, respiratory control ratio, and ADP:O ratio. Ca(2+)-induced PTP opening was also similar. However, H(2)O(2) production by ET was lower than Sed (P < 0.05) in the absence of calcium (323 +/- 12 vs. 362 +/- 11 pmol.min(-1).mg protein(-1)) and the presence of 50 microM CaCl(2) (154 +/- 3 vs. 197 +/- 7 pmol.min(-1).mg protein(-1)). Rotenone, which blocks electron flow from succinate to complex 1, reduced H(2)O(2) production and eliminated differences between ET and Sed. Mitochondrial superoxide dismutase and glutathione peroxidase were not affected by exercise. Catalase activity was extremely low but increased 49% in ET (P < 0.05). In conclusion, exercise reduces ROS production in myocardial mitochondria through adaptations specific to complex 1 but does not improve mitochondrial tolerance to calcium overload.

Improved postischemic function following acute exercise is not mediated by nitric oxide synthase in the rat heart.

The mediators of acute exercise-induced preconditioning against ischemia-reperfusion injury are not understood. This study assesses the role of nitric oxide synthase (NOS), a reported mediator of other forms of preconditioning. Male Fischer 344 rats were divided into five groups (n = 6-7): sedentary (Sed); exercised 2 days on a treadmill at 20 m/min, 6 degrees grade, for 60 min (Run); sedentary, perfused with 100 microM N(omega)-nitro-l-arginine methyl ester hydrochloride (l-NAME) to inhibit NOS (Sed/L-N); exercised, perfused with l-NAME (Run/L-N); and exercised in a 4 degrees C environment, perfused with l-NAME (CRun/L-N). Twenty-four hours following exercise, isolated, perfused working hearts were subjected to 22.5 min of global ischemia plus 30 min of normoxic reperfusion. Left ventricle contents of several putative preconditioning mediators were determined. Postischemic recovery of cardiac output times systolic pressure was better in Run than Sed (78.4 vs. 50.2% of preischemia, P < 0.05). Inhibition of NOS did not abrogate the improved recovery in the exercise groups or alter recovery in Sed. All exercise groups also displayed improved myocardial efficiency (cardiac output times systolic pressure/oxygen consumption) postischemia and less lactate dehydrogenase release (P < 0.05). l-NAME appeared to lower lactate dehydrogenase release independent of exercise. The only change in antioxidant enzyme activity was a decrease in manganese superoxide dismutase in CRun/L-N (P < 0.05). Heat shock protein 72 expression increased only in Run and Run/L-N and endothelial NOS only in CRun/L-N (P < 0.05). Acute exercise-induced preconditioning of the Fischer 344 rat heart is not mediated by NOS and does not require increases in heat shock protein 72 or antioxidant enzymes.

Effects of caloric restriction and exercise training on skeletal muscle histochemistry in aging Fischer 344 rats.

The purpose of this study was to determine the effects of calorie restriction and exercise on hindlimb histochemistry and fiber type in Fischer 344 rats as they advanced from adulthood through senescence. At 10 months of age, animals were divided into sedentary fed ad libitum, exercise (18 m/min, 8% grade, 20 min/day, 5 days/week) fed ad libitum, and calorie restricted by alternate days of feeding. Succinic dehydrogenase, myosin adenosine triphosphatase (mATPase at pH 9.4), nicotine adenonine dinucleotide reductase, and Periodic Acid Shiff histochemical stains were performed on plantaris and soleus muscles. The results indicated that aging resulted in a progressive decline in plantaris Type I muscle fiber in sedentary animals, while exercise resulted in maintenance of these fibers. The percent of plantaris Type II fibers increased between 10 and 24 months of age. Exercise also resulted in a small, but significant, increase in the percentage of plantaris Type IIa fibers at 24 months of age. The soleus fiber distribution for Type I fibers was unaffected by increasing age in all groups of animals. The implications of these results suggest the implementation of exercise as a lifestyle modification as early as possible.

Myocardial heat shock protein 70 expression in young and old rats after identical exercise programs.

Synthesis of inducible heat shock protein 70 (HSP70) is impaired in aged animals following acute stresses including exercise. In this study we determined whether aging affects expression of this cytoprotective protein following chronic exercise participation. Male Fischer 344 rats, final ages 6 and 24 months, exercised identically for 10 weeks on a treadmill (15 degrees incline, 15 m/min for up to 60 minutes, 5 days/week). In 6-month-old animals, exercise increased HSP70 in heart (44%), liver (216%), and skeletal muscle (126%) (p <.05 vs sedentary). In 24-month-old animals, exercise increased HSP70 in muscle (69%), but not in heart or liver. In heart, antioxidant enzyme activities and HSP70 messenger RNA were measured and found to be unaffected by exercise at both ages. Our results indicate an age-related decrease in HSP70 production in heart and liver following chronic exercise. Furthermore, the aged heart does not increase its antioxidant enzyme defenses to compensate for the HSP70 deficit.

Habitual low-intensity exercise does not protect against myocardial dysfunction after ischemia in rats.

BACKGROUND: It is well established that participation in a chronic exercise program can reduce coronary heart disease (CHD) risk factors and improve myocardial tolerance to ischemia-reperfusion (I-R) injury. Low-intensity exercise programs are known to be effective in reducing CHD risk factors in humans and rats, but whether similar programs are of sufficient intensity to improve intrinsic tolerance to I-R injury has not been established. Thus, the purpose of this study is to determine whether low-intensity exercise provides self-protection to the heart against I-R injury. METHODS: Male, Sprague-Dawley rats were exercised on a treadmill at an intensity of 55-60% VO2max, 40 min/day, 5 days/week for 16 weeks. Reperfusion injury following 20 min of global ischemia was evaluated using the isolated perfused working heart model. Left ventricular content of the cytoprotective protein heat shock protein 70 (HSP70) was determined by Western blotting. RESULTS: The exercise program elevated HSP70 2.7-fold, but did not provide enhanced protection following 20 min of ischemia. Final post-ischemic recovery of cardiac external work was 63+/-9% of pre-ischemic value in the sedentary group (n=9) and 51+/-11% in the exercising group (n=9) (P>0.05). Post-ischemic lactate dehydrogenase release was also similar between groups and the magnitude of release was low, consistent with stunning. CONCLUSIONS: Regular exercise at 55-60% VO2max is below the threshold intensity necessary to induce intrinsic cardioprotection against I-R injury. Furthermore, elevated myocardial HSP70 is not necessarily a marker of improved protection against dysfunction associated with stunning.

Effect of exercise training on the ability of the rat heart to tolerate hydrogen peroxide.

OBJECTIVE: The purpose of this study was to determine whether exercise training could precondition the myocardium against hydrogen peroxide (H(2)O(2))-induced damage. METHODS: Male Fischer 344 rats ran on a treadmill for 9 weeks (60 min/day, 22 m/min, 6 degrees grade, 5 days/week). Isolated perfused working hearts from exercise trained (ET, n=8) and sedentary (SED, n=10) animals were perfused with 150 microM H(2)O(2). RESULTS: Pre-H(2)O(2) baseline values for cardiac external work (COxSP), coronary flow (CF), and lactate dehydrogenase (LDH) release were similar between groups. At 5 min of H(2)O(2), COxSP was unchanged from baseline but CF was increased 30% in SED and 46% in ET (P<0.05 vs. SED). COxSP began to decline similarly thereafter in both groups, dropping to 20% of baseline at 20 min. CF in ET remained higher than SED throughout (P<0.05). LDH leakage remained near baseline during the first 15 min of H(2)O(2) exposure, but was elevated (P<0.05) 72% in SED and 40% in ET after 20 min, and was 2.2-fold greater in SED than ET (P<0.05) after 25 min. Heat shock protein 70 was 2.1-fold greater in ET than SED (P<0.05), but ET did not change catalase and glutathione peroxidase. CONCLUSIONS: The results of this study indicate that chronic moderate exercise will enhance coronary flow and attenuate the development of myocardial injury when exposed to H(2)O(2), but will not affect H(2)O(2)-induced decrease in pump function.

Exercise improves postischemic function in aging hearts.

Exercise improves cardioprotection against ischemia-reperfusion in young animals but has not been investigated in older animals, which represent the population most likely to suffer an ischemic event. Therefore, we sought to determine the effects of aging on exercise-induced cardioprotection. Young, middle-aged, and old (4, 12, and 21 mo old) male Fischer 344 rats ran 60 min at 70-75% of maximum oxygen consumption. Twenty-four hours postexercise, isolated perfused working hearts underwent 22.5 min of global ischemia and then 30 min of recovery (reperfusion). Compared with sedentary rats (n = 8-9 rats/group), recovery of function (cardiac output x systolic pressure) improved after exercise (n = 9 rats/group) by 40% at 4 mo, 78% at 12 mo, and 59% at 21 mo. Exercise increased inducible heat shock protein 70 expression 105% at 4 mo but only 27% at 12 mo and 24% at 21 mo. Catalase activity progressively increased with age (P < 0.05) and was increased by exercise at 4 mo (26%) and 21 mo (19%). Manganese superoxide dismutase activity was increased by exercise only at 21 mo (45%). No exercise-related change in any antioxidant enzyme was observed at 12 mo. We conclude that exercise can enhance cardioprotection regardless of age, but the cardioprotective protein phenotype changes with age.

Effect of body temperature during exercise on skeletal muscle cytochrome c oxidase content.

This study determined the role of body temperature during exercise on cytochrome-c oxidase (CytOx) activity, a marker of mitochondrial content, and mitochondrial heat shock protein 70 (mtHSP70), which is required for import of nuclear-coded preproteins. Male, 10-wk-old, Sprague-Dawley rats exercised identically for 9 wk in ambient temperatures of 23 degrees C (n = 10), 8 degrees C with wetted fur (n = 8), and 4 degrees C with wetted fur and fan (n = 7). These conditions maintained exercising core temperature (T(c)) at 40.4, 39.2, or 38.0 degrees C (resting temperature), respectively. During weeks 3-9, exercisers ran 5 days/wk up a 6% grade at 20 m/min for 60 min. Animals were housed at 23 degrees C. Gastrocnemius CytOx activity in T(c)=38.0 degrees C (83.5 +/- 5.5 microatoms O x min(-1) x g wet wt(-1)) was greater than all other groups (P < 0.05), exceeding sedentary (n = 7) by 73.2%. T(c) of 40.4 and 39.2 degrees C also were higher than sedentary by 22.4 and 37.4%, respectively (P < 0.05). Quantification of CytOx content verified that the increased activity was due to an increase in protein content. In extensor digitorum longus, a nonactive muscle, CytOx was not elevated in T(c) = 38.0 degrees C. mtHSP70 was significantly elevated in gastrocnemius of T(c) = 38.0 degrees C compared with sedentary (P < 0.05) but was not elevated in extensor digitorum longus (P > 0.05). The data indicate that decreasing exercise T(c) may enhance mitochondrial biogenesis and that mtHSP70 expression is not dependent on temperature.