ABC | Volume 112, Nº2, February 2019

Original Article Arq Bras Cardiol. 2019; 112(2):154-162 Naderi et al Garlic and exercise increase cardiac angiogenesis significantly (p < 0.001) the myocardial miR-126 expression in diabetic rats compared to the diabetes group. Moreover, the Diabetes+Garlic+Exercise group had significantly higher level of miR-126 expression compared to the garlic treatment group (p < 0.05) and the just voluntary exercise group (p < 0.01) in diabetic animals. Effects of garlic and voluntary exercise on miR-210 in the myocardium As shown in Figure 2, the expression of miR-210 significantly increased (p < 0.001) in animals with diabetes compared with the control group. Treatment with garlic (p < 0.01), voluntary exercise (p < 0.01), or both combined reduced significantly (p < 0.001) the myocardial miR-210 expression in diabetic rats compared to the diabetes group. The combined Garlic+Voluntary Exercise group significantly lowered miR‑210 expression compared to the Diabetes+Exercise (p < 0.05) and Diabetes+Garlic (p < 0.01) groups Effect of garlic and voluntary exercise on angiogenesis in the myocardium Immunostaining with CD31 marker was performed for the assessment of angiogenesis in the transversal section of the ventricles at their midportion. Brown stained tissues show CD-31 immunostained endothelial cells. Figure 4 shows the scores for staining intensity, which are as follows: 0 (<10%); 1 (10% to 25%); 2 (25% to 50%); 3 (50% to 75%) or 4 (75% to 100%). As shown in Figures 3 and 4, statistical analysis of our immunohistochemical study revealed that angiogenesis decreased significantly (p < 0.01) in the diabetes group compared to the control group. Six weeks of garlic treatment, voluntary exercise, or a combination thereof in the diabetes groups increased significantly (p < 0.001) the angiogenesis in their left ventricle compared to the diabetes group (Figure 3 and 4). Combined garlic consumption and exercise in diabetic animals induced more angiogenesis compared to garlic alone and exercise alone, though the difference was not significant. Effect of garlic and voluntary exercise on serum lipid profile Lipid profile alterations in different groups are shown in Table 2. The induction of diabetes in the animals increased significantly (p < 0.001) the serum TGs and LDL levels while lowering serum HDL and HDL/LDL compared to the control animals. Voluntary exercise reduced significantly (p < 0.05) the serum triglycerides levels in the diabetes group comparedwith the control group. Sixweeks of garlic treatment alone or with voluntary exercise decreased significantly (p < 0.01) the triglycerides levels in the animals with diabetes. In these, serumLDL levels decreased significantly (p < 0.001) after garlic alone and exercise alone or a combination thereof. However, serumHDL level was significantly increased (p<0.001) by garlic treatment, voluntary exercise, or a combination thereof in diabetic rats. Furthermore, the HDL:LDL ratio was significantly higher (p < 0.001) in the Diabetes+Garlic, Diabetes+Exercise and Diabetes+Garlic+Exercise groups compared with diabetes group. Discussion The present study has shown that the induction of diabetes impaired serum lipid profile, decreased myocardial angiogenesis and miR-126 expression, and increased myocardial expression of miR-210. However, the treatment with garlic alone, voluntary exercise alone or both combined ameliorated these effects in the myocardium of diabetic animals. Interestingly, treating diabetic rats simultaneously with garlic and voluntary exercise had an additional effect on the cardiac expression of miR-126 and miR-210. In line with our study, research has shown that diabetes leads to an impaired function of early endothelial progenitor cells, which results in a reduced capacity of neovascularisation and angiogenesis in the myocardium of diabetic rats. 23 VEGF, as an inducer of angiogenesis, is a highly specific mitogen for endothelial cells. 24 It is well-known that the expression of VEGF-A and its receptors decreases in the myocardium of diabetic rats and humans. 25 However, the actual process of VEGF and angiogenesis reduction in the diabetic heart has not been fully elucidated. Figure 2 – Real-time quantitative PCR analysis of miR-210 in the heart tissue of experimental groups. The values represent means ± S.E.M for 7 animals. ***p < 0.001 vs control group, $$ p < 0.01 and $$$ p < 0.001 vs diabetes group, & p < 0.05 vs Diabetes + Exercise group, and ## p < 0.01 vs Diabetes + Garlic group. 3.5 2.5 1.5 0.5 1.0 0.0 3.0 2.0 *** $$ $$ $$$&## MiR-210: rno-miR-191 (Fold change) Control Diabetes Diabetes+Garlic Diabetes+Exercise Diabetes+Garlic+Exercise 157