ABC | Volume 112, Nº1, January 2019

Original Article Shekarforoush & Naghii Cardioprotection by whole-body vibration Arq Bras Cardiol. 2019; 112(1):32-37 Methods Male Wistar rats weighing 250 to 300 g (10-12 weeks old) were obtained from the animal house of Shiraz University of Medical Sciences and housed under standard conditions, with free access to food and water. The investigation was approved by the University Ethics Committee in accordance with the Guide for the Care and Use of Laboratory Animals. Experimental designs A total of 24 rats were randomly assigned to 1 of 3 treatment groups (control vs. two experimental groups) by picking numbers out of a hat. The sample size (n) was established based on studies that evaluated the effects of exercise against myocardial IR injury. 16,17 Animals in the vibration groups were placed in a compartment attached to a vibration platform (Crazy Fit Massager/Model: YD 1002, Union Brilliant Group Co., LTD, Fujian, China). The vibration training consisted of a 5-min cycle on day 1, followed by an extra 5-min cycle each time for the next five sessions in the first week and then each rat was exposed to vertical sinusoidal vibration for 30 min per session (3 × 10 min cycles), 6 days a week for one week (WBV1 group) or 3 weeks (WBV3 group). The animals were given 1–2 min rest break between the cycles. The vibration was performed at mode 1 with amplitude of 1–10 mm and at a frequency of 10–50 Hz. The speed of mode 1 in each cycle increased gradually and then decreased with the same trend within each time period. The control animals remained in their cages and were placed over the vibration platform, without vibration treatment. Each training session was performed between 8.30–10.00 A.M. 18 Surgical procedure The protocol used has been thoroughly described in detail in our previous publication. 12 Briefly, 24 hours after the last training session, the animals were anesthetized and ventilated with room air enriched with oxygen at a rate of 70 breaths per min. A standard limb lead II electrocardiogram was monitored and recorded throughout the experiment. Catheters were inserted into the left carotid artery and tail vein for monitoring of blood pressure and infusion of Evans blue solution, respectively. After the thoracotomy, a 6-0 silk suture was passed around the left anterior descending coronary artery (LAD). Following a stabilization period of 20 min, the LAD was occluded for 30 min of ischemia and released for 120 min of reperfusion. Rectal temperature was continuously monitored and maintained at 37 ± 0.5°C. Determination of infarct size and area at risk At the end of reperfusion, the LAD was reoccluded and 1 mL of 2% solution of Evans Blue dye (Sigma, St. Louis, MO) was injected into the tail vein to identify the non-perfused area, also known as area at risk (AAR), from the perfused area. The rats were then killed with a pentobarbital overdose and their hearts were excised and frozen for one hour. The atria and right ventricle were removed, and the left ventricle was cut into transverse slices of 2 mm thickness from the apex to the base. Tissue samples were then incubated with a 1% solution of 2,3,5 triphenyltetrazolium chloride (Sigma)] for 20 min at 37°C, and subsequently fixed in 10% phosphate-buffered formalin for one hour. Viable myocardium was stained red by triphenyltetrazolium chloride, whereas necrotic myocardium appeared as pale yellow. In each slice, areas at risk and infarcted areas were determined by computerized planimetry using an image analysis software (Image Tool, University of Texas, San Antonio, TX). Infarct size (IS) was expressed as percentage of the AAR (IS/AAR). 12 Assessment of ventricular arrhythmias Ischemia-induced ventricular arrhythmias were determined in accordance with the Lambeth conventions 19 including ventricular ectopic beat as premature ventricular complexes (PVC), ventricular tachycardia (VT) as a run of four or more consecutive ventricular premature beats at a rate faster than the resting sinus rate, and ventricular fibrillation (VF) as a signal for which individual QRS deflection can no longer be distinguished from one another. Complex forms (bigeminy and salvos) were added to PVC count and not analyzed separately. In order to determine the incidence of VT and VF, they were recorded as either occurring or not occurring during the first 30 min of ischemia in each group. Statistical analyses Unless stated otherwise, the results were expressed as Mean ± SD. All data were processed with the SPSS 16.0 statistical package for Windows version. The normality of distributions was verified by the Kolmogorov-Smirnov test. Fisher exact test (Chi-square) was used to analyze the incidence of VT and VF. Analysis of baseline, ischemia, and reperfusion HR and BP was done by repeated measures analysis of variance (ANOVA). The other data were analyzed using one-way ANOVA and then significant differences were examined by Tukey’s post‑hoc test. Differences between the groups were considered significant at a level of p < 0.05. Results Hemodynamic parameters Table 1 summarizes the hemodynamic data. There were no significant differences at baseline values for heart rate (HR) and mean arterial blood pressure (MBP) among the groups. Ischemia caused a marked reduction in blood pressure without any significant effect on the HR in the groups. MBP was nearly restored to the baseline level during the reperfusion period. Infarct size Figure 1 shows AAR and IS following 30 min of regional ischemia and 120 min of reperfusion. There was no marked difference in AAR/LV ratio among the groups (p = 0.92). Infarct size was 47 ± 5% in the control group. WBV1 or WBV3 resulted in a smaller infarct size, i.e. 39 ± 5% and 37 ± 5% (p = 0.047 and p = 0.009 vs. the controls), respectively. 33