Physical Exercise Promotes a Reduction in Cardiac Fibrosis in the Chronic Indeterminate Form of Experimental Chagas Disease

Abstract

Chagas disease (CD), caused by the protozoan Trypanosoma cruzi, is a neglected tropical disease and a health problem in Latin America. Etiological treatment has limited effectiveness in chronic CD; thus, new therapeutic strategies are required. The practice of physical exercises has been widely advocated to improve the quality of life of CD patients. The most frequent clinical CD manifestation is the chronic indeterminate form (CIF), and the effect of physical exercises on disease progression remains unknown. Here, in a CIF model, we aimed to evaluate the effect of physical exercises on cardiac histological, parasitological, mitochondrial, and oxidative metabolism, electro and echocardiographic profiles, and immunological features. To establish a CIF model, BALB/c and C57BL/6 mice were infected with 100 and 500 trypomastigotes of the Y T. cruzi strain. At 120 days postinfection (dpi), all mouse groups showed normal PR and corrected QT intervals and QRS complexes. Compared to BALB/c mice, C57BL/6 mice showed a lower parasitemia peak, mortality rate, and less intense myocarditis. Thus, C57BL/6 mice infected with 500 parasites were used for subsequent analyses. At 120 dpi, a decrease in cardiac mitochondrial oxygen consumption and an increase in reactive oxygen species (ROS) were detected. When we increased the number of analyzed mice, a reduced heart rate and slightly prolonged corrected QT intervals were detected, at 120 and 150 dpi, which were then normalized at 180 dpi, thus characterizing the CIF. Y-infected mice were subjected to an exercise program on a treadmill for 4 weeks (from 150 to 180 dpi), five times per week in a 30-60-min daily training session. At 180 dpi, no alterations were detected in cardiac mitochondrial and oxidative metabolism, which were not affected by physical exercises, although ROS production increased. At 120 and 180 dpi, comparing infected and non-infected mice, no differences were observed in the levels of plasma cytokines, indicating that a crucial biomarker of the systemic inflammatory profile was absent and not affected by exercise. Compared with sedentary mice, trained Y-infected mice showed similar parasite loads and inflammatory cells but reduced cardiac fibrosis. Therefore, our data show that physical exercises promote beneficial changes that may prevent CD progression.

Keywords: Chagas disease; ROS; chronic indeterminate form; cytokines; experimental model; mitochondrial metabolism; oxidative metabolism; physical exercise.

Figure 1

T. cruzi infection induces splenomegaly and heart inflammation in Y-infected BALB/c mice. BALB/c and C57BL/6 mice were infected with 100 or 500 bloodstream trypomastigotes (Y strain). (A) Experimental design. (B) Parasitemia and survival curves. (C) Relative spleen weight. (D) ECG records showing PR and QTc intervals and the QRS complex (ms). (E) Relative heart weight. (F) Inflammatory cells in heart tissue at 120 dpi. Data are represented as means ± SEM. Data represent three to four mice per group. Significant differences between infected and uninfected groups. *p < 0.05; **p < 0.01 ***p < 0.001 (Student’s t-test; Kruskal–Wallis Dunn’s posttest; one-way ANOVA, Tukey posttest).

Figure 2

T. cruzi infection does not change body weight but reduces muscular strength in C57BL/6 mice. Mice were infected with 500 trypomastigotes (Y strain). (A) Experimental design. (B) Body weight. (C) Muscular strength. The data are represented as means ± SEM. Data represent eight to nine mice per group. Significant differences between infected and uninfected groups. **p < 0.01. Significant differences between infected groups at 120 and 180 dpi. ^$p > 0.05 (Test t; one-way ANOVA, Tukey posttest; Kruskal–Wallis Dunn’s posttest; two-way ANOVA, Sidak’s posttest).

Figure 3

T. cruzi infection does not induce electrical alterations but leads to ECHO abnormalities in C57BL/6 mice at 180 dpi. Absence of electrical alterations in C57BL/6-infected mice at 180 dpi. (A) ECG records show the average heart rate (beats per minute, bpm), P wave duration (ms), variation in PR and QTc intervals, and QRS complex (ms). (B) Representative ECG register segments. (C) ECHO records show the % left ventricular ejection fraction (LVEF), stroke volume (µL), % FAC, LV area (mm²), and RV area (mm²). (D) Representative ECHO images of ventricular areas. The data are represented as means ± SEM. Data represent eight to nine mice per group. Significant differences between the untrained and trained groups. ^#p < 0.05. Significant differences between infected and uninfected groups. *p < 0.05; ***p < 0.001; ****p < 0.0001 (Kruskal–Wallis Dunn’s posttest).

Figure 4

Parasite persistence and inflammation in heart tissue of trained Y-infected mice. (A) Relative heart weight. (B) Parasite load at 180 dpi. (C) Quantitative analysis of inflammatory infiltrates. (D) Representative sections stained with H&E (ND = not detected). The data are represented as means ± SEM. Data represent nine mice per group. Significant differences between infected and uninfected groups. *p < 0.05; ***p < 0,001. (Mann–Whitney; Kruskal–Wallis test, Dunn’s posttest).

Figure 5

Effects of T. cruzi infection and physical exercise on mitochondrial oxygen consumption and ROS production in heart tissue of C57BL/6 mice. (A, B) Oxygen consumption at (A) 120 dpi and (B) 180 dpi. (C) Representative sections stained with DHE. (D) Densitometric analysis of DHE staining. White arrows indicate DHE staining. The data are represented as means ± SEM. Data represent six to eight mice per group. Significant differences between infected and uninfected groups *p < 0.05; **p < 0.01 (Mann–Whitney; Kruskal–Wallis test, Dunn’s posttest).

Figure 6

T. cruzi infection causes mild splenomegaly but does not induce a systemic inflammatory profile in C57BL/6 mice. (A) Relative spleen weight. (B) Representative dot plots of Y-infected C57BL/6 mice at 180 dpi and inserted dot plots of Colombian-infected C57BL/6 mice at 150 dpi. The data are represented as means ± SEM. Data represent six to eight mice per group *p < 0,05 (Mann–Whitney; two-way ANOVA Sidak’s posttest; Kruskal–Wallis test, Dunn’s posttest).

Figure 7

Physical exercise induce a reduction in collagen deposition in the hearts of C57BL/6-infected mice. (A) Representative sections stained with Picrosirius red. Black arrows indicate positive staining for Picrosirius red. (B) Densitometric analysis of Picrosirius red staining. (C) Correlations of the QTc interval and collagen deposition. The data are represented as means ± SEM. Data represent six to eight mice per group. Significant differences between infected and uninfected groups. ***p < 0.001 and ****p < 0.0001; significant differences between the untrained and trained groups, ^####p < 0.0001; significant differences comparing infected groups at 120 dpi and 180 dpi ^$p < 0.0001 (Mann–Whitney; two-way ANOVA, Sidak’s posttest; Kruskal–Wallis test, Dunn’s posttest).