Figure 3. Xanthine Oxidase-derived superoxide mediates the increase of leak in cardiomyocytes.

(A) Representative images of H₂DCF fluorescence in response to ROS in WT or NOS1^−/− cardiomyocytes in the presence or the absence of 100 μmol/L oxypurinol at 23°C or 37°C (Bar, 10 μm). (B) Averaged ROS production in WT or NOS1^−/− cardiomyocytes in the presence or the absence of oxypurinol 100 μmol/L at 23°C or 37°C. Values were normalized by the maximal signal for each strain in the presence of H₂O₂ (catalase-treated cardiomyocytes were used as negative control). * p <0.05, ** p < 0.01 and *** p < 0.001 (-)Oxy vs. +Oxy-treated cardiomyocytes, Student's t-test. (C) Dependence of SR Ca²⁺ leak with temperature in NOS1^−/− cardiomyocytes in the absence or the presence of 100 μmol/L oxypurinol [at matched SR Ca²⁺ load ≈ 75 μmol/L] (** p < 0.01 (-)Oxy vs. +Oxy-treated NOS1^−/−, Two-way ANOVA). (D) SR Ca²⁺ load-leak relationship in non-treated and oxypurinol-treated NOS1^−/− cardiomyocytes at 23°C and (E) at 37°C († p< 0.05 (-)Oxy vs. +Oxy NOS1^−/−; exponential growth fitting).