Cholinergic Stimulation Improves Oxidative Stress and Inflammation in Experimental Myocardial Infarction

Figures

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  Figure 1. CD68 + cell count (Total macrophages). The macrophages were stained in brown, and the total area occupied by these cells was measured. Both I and I + PY groups showed dense infiltration of CD68 + cell within the infarcted and peri-infarcted zone with no statistical difference (P = 0.9, Kruskal-Wallis test). As expected, both groups were statistically different from the Sham group (P < 0.05, Kruskal-Wallis test completed by Dunn’s multiple comparison test). Values are expressed as mean ± SEM. (Immunohistochemistry, DAB, scale bar 400 μm).

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  Figure 2. M2 macrophage (CD206+) cell count. The M2 macrophages were stained in brown, and the total area occupied by these cells was measured. Sham animals presented no M2 macrophages; I + PY group showed higher infiltration of CD206 + cell within the infarcted and peri-infarcted zone when compared to the I group (P = 0.04, Kruskal-Wallis test). The I group and I + PY were statistically different from the Sham group (P = 0.003 and P < 0.0001, respectively, Kruskal-Wallis). Values are expressed as mean ± SEM. (Immunohistochemistry, DAB, scale bar 400 μm).

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  Figure 3. Treg cells (FOXP3+ lymphocytes) cell count. Treg cells were stained in brown, and the total area occupied by these cells was measured. Sham animals presented no Treg cells; I + PY group showed a higher infiltration of FOXP3+ cells within the infarcted and peri-infarcted zone when compared to I groups (P = 0.007, Kruskal-Wallis test). As expected, I group and I + PY were statistically different from the Sham group (P < 0.0001, Kruskal-Wallis test). Values are expressed as mean ± SEM. (Immunohistochemistry, DAB, scale bar 400 μm).

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  Figure 4. Antioxidant enzyme activity. Superoxide dismutase activity (SOD) (A), Catalase (CAT) (B), and GPX activity (C) were significantly lower in the infarcted rats (I group) when compared to sham animals (S group). Infarcted rats treated with PY (I + PY) presented a significantly higher activity of all anti-oxidant enzymes when compared to the untreated rats. Values are expressed as mean ± SEM. *P < 0.05 vs Sham; # P < 0.05 vs I (n = 7) for each group (two-way ANOVA test with Tukey as post hoc).

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  Figure 5. Lipid peroxidation and protein oxidation. Lipid peroxidation, obtained by measurement of TBARS, increased similarly in both infarcted groups (I and I + PY) t when compared to the S group (A). Protein oxidation, obtained by measurement of carbonyl groups (carbonyls) was significantly higher in the I group when compared to S group. However, in the I + PY group carbonyls were significantly lower when compared to I group, reaching values similar to those of the S group. (B). Values are expressed as mean ± SEM. *P < 0.05 vs Sham; # P < 0.05 vs I (n = 7) for each group, (two-way ANOVA test with Tukey as post hoc).

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  Figure 6. Cytokine concentrations. When compared to the S group, the I group, presented significantly higher concentrations of the pro-inflammatory cytokines IL-1β (A), IL-6 (B) and TNF-α (C), and anti-inflammatory cytokine IL-10 (D). Conversely, I + PY animals presented significantly lower values when compared to I animals. Furthermore, no differences in cytokines concentrations were observed when I + PY was compared to S group. Values are expressed as mean ± SEM. *P < 0.05 vs Sham; # P < 0.05 vs I (n = 7) for each group (two-way ANOVA test with Tukey as post hoc).