Figure 2

Carbon monoxide prevents mitochondrial swelling and depolarization. A. Mitochondria were pre-treated with 10 μM of CO for 15 minutes at RT, swelling measurements were performed after 6.25 μM and 12.5 μM of Ca²⁺ or 300 μM of atractyloside addition into isolated mitochondria. The decrease on absorbance was assessed at 540 nm, at 37°C for 30 minutes and is presented as representative graphics of experiments, which were done in triplicates and repeated, at least, three times. B. Mitochondrial depolarization was assessed by rhodamine 123 fluorescence change (λ_exc: 485 nm, λ_em: 535 nm), at 37°C, for 30 min, in the absence or presence of CO at 10 μM and atractyloside at 300 μM or Ca²⁺ at 6.25 or 12.5 μM. C. left panel Quantitative expression of mitochondrial swelling measured at 15 minutes of incubation and right panel, 28 minutes of incubation. The effect of 12.5 μM of Ca²⁺ (left panel) or Atra 300 μM (right panel) was normalized as 100% of swelling. All values are mean ±SD (error bars), n = 3; *p < 0.05, compared to control mitochondria; **p < 0.05, compared to Ca²⁺ 6.25 μM (left panel) or Atra 300 μM (right panel) treated mitochondria; ***p < 0.05, compared to Ca²⁺ treated mitochondria. D. Quantification of mitochondrial depolarization experiments performed in triplicates and repeated, at least, three times. left panel, quantitative expression of mitochondrial depolarization measured at 15 minutes of incubation and right panel, 28 minutes of incubation. The effect of 6.25 μM of Ca²⁺ (left panel) or Atra 300 μM (right panel) was normalized to 100% of swelling. All values are mean ± SD (error bars), n = 3; *p < 0.05, compared to control mitochondria; **p < 0.05, compared to Ca²⁺ 6.25 μM (left panel) or Atra 300 μM (right panel) treated mitochondria.