Efeitos do treinamento físico e do destreinamento sobre propriedades moleculares e mecânicas de cardiomiócitos isolados de ratos normotensos e hipertensos

Abstract

Effects of exercise training and detraining on the molecular and mechanical properties of isolated cardiomyocytes from normotensive and hypertensive rats. Introduction. The endurance training induces benefits on cardiac function that are related to cellular and subcellular adaptations. Studies show that endurance training increases the expression of transient intracellular global Ca2+ ([Ca2+]i) regulatory proteins, the [Ca2+]i transient and contractility of isolated cardiomyocytes from the left ventricle of normotensive rats, although detraining reverses these adaptions. However, the effects of endurance training and detraining on these parameters in hypertensive rat hearts, are not known. Aims. To investigate the effects of low-intensity endurance training and detraining on cardiovascular parameters and molecular and mechanical properties of isolated cardiomyocytes from the left ventricle of normotensive and hypertensive rats. Materials and Methods. Male wistar rats [initial weight of 385.3 ± 6.6 g (mean ± standard error of mean) and systolic blood pressure (SBP) of 110.7 ± 0.5 mmHg] and SHR [Spontaneously Hypertensive Rats (initial weight of 357.5 ± 4.1 g and 178.4 ± 1.3 SBP mm Hg)] with 16 weeks of age, were randomly distributed into 8 groups (n = 8): NC8 and HC8 - normotensive and hypertensive controls for 8 weeks; NT8 and HT8 - normotensive and hypertensive trained at 50-60% of maximal exercise capacity on a treadmill for 8 weeks; NC12 and HC12 - normotensive and hypertensive controls for 12 weeks; NDT and HDT - normotensive and hypertensive trained at 50-60% of maximal exercise capacity on a treadmill for 8 weeks and detrained for 4 weeks. The resting heart rate (RHR) and SBP were measured before and after treatments, using a sensor and tail plethysmography, respectively. The total exercise time until fatigue (TTF) was determined by a maximal exercise capacity test. After treatments, the euthanasia of animals was performed by cervical dislocation followed by thoracotomy. The heart was removed, washed and cannulated via the aorta on an infusion system, and the left ventricular cardiomyocytes were isolated using enzymatic digestion (collagenase). To determine the [Ca2+]i transient, isolated cardiomyocytes from the left ventricle were incubated with the Ca2+ fluorescent indicator Fluo-4 acetoxymethyl ester (Fluo-4 AM) and placed inside an experimental chamber mounted on a confocal microscope. The cell contractility was determined using the technique of changing the length of cardiomyocytes using an edge detection system, with the aid of an inverted microscope. For both experiments, we used field electrical stimulation (20 V, 1 Hz, ~ 25 ° C). The expression of [Ca2+]i regulatory proteins of the left ventricle was measured by 21 Western Blot, and the markers of pathologic cardiac hypertrophy by real-time polymerase chain reaction (RT-PCR). Results. Low-intensity endurance training augmented the TTF (NC8, 11.4 ± 1.5 min vs. NT8, 22.5 ± 1.4 min; HC8, 11.7 ± 1.4 min vs. HT8, 24.5 ± 1.3 min; P ≤ 0.05), reduced RHR (NT8initial, 340 ± 8 bpm vs. NT8final, 322 ± 10 bpm; HT8initial, 369±8 bpm vs. HT8final, 344 ± 10 bpm; P ≤ 0.05), and SBP in SHR animals (HC8, 178 ± 3 mmHg vs. HT8, 161 ± 4 mmHg; P ≤ 0.05). HC8 rats showed a slower [Ca2+]i transient (Tpeak, 83.7 ± 1.8 ms vs. 71.7 ± 2.4 ms; T50%decay, 284.0 ± 4.3 ms vs. 264.0 ± 4.1 ms; P<0.05) and cell contractility (Vshortening, 86.1 ± 6.7 µm/s vs. 118.6 ± 6.7 µm/s; Vrelengthening, 57.5 ± 7.4 µm/s vs. 101.3 ± 7.4 µm/s; P ≤ 0.05), and higher expression of ANF (300%; P ≤ 0.05), skeletal α-actin (250%; P ≤ 0.05) and a decreased α/β-MHC ratio (70%; P ≤ 0.05) compared to NC8. Exercise training increased [Ca2+]i transient (NC8, 2.39 ± 0.06 F/F0 vs. NT8, 2.72 ± 0.06 F/F0; HC8, 2.28 ± 0.05 F/F0 vs. HT8, 2.82 ± 0.05 F/F0; P ≤ 0.05), cell contractility (NC8, 7.4 ± 0.3 % vs. NT8, 8.4 ± 0.3 %; HC8, 6.8 ± 0.3 % vs. HT8, 7.8 ± 0.3 %; P ≤ 0.05), and normalized the expression of ANF, skeletal α-actin, and the α/βMHC ratio in HT8 rats. Furthermore, augmented the expression of sarcoplasmic reticulum Ca2+ ATPase (SERCA2a) (NC8, 0.93 ± 0.15 vs. NT8, 1.49 ± 0.14; HC8, 0.83 ± 0.13 vs. HT8, 1.32 ± 0.14; P<0.05), phosphorylation of phospholamban at serine 16 (PLBser16) (NC8, 0.89 ± 0.18 vs. NT8, 1.23 ± 0.17; HC8, 0.77 ± 0.17 vs. HT8, 1.32 ± 0.16; P<0.05), and reduced the ratio total phospholamban (PLBt)/SERCA2a (NC8, 1.21 ± 0.19 vs. NT8, 0.50 ± 0.21; HC8, 1.38 ± 0.17 vs. HT8, 0.66 ± 0.21; P<0.05) in the left ventricle of trained animals. However, all these adaptations returned to sedentary values within 4 weeks of detraining in both SHR and normotensive animals. Conclusion. The low-intensity endurance training induces beneficial adaptations on the molecular and mechanical properties of isolated cardiomyocytes from the left ventricle of normotensive and hypertensive rats. Detraining for a period of 4 weeks, reversed all these benefits.