Fig. 6: Increased Hint1 levels in cardiomyocytes reduces mitochondrial respiratory capacity.
a Representative western blots (left) and quantitation (right) of Hint1 protein levels in heart tissues of 12-week-old sham-operated WT mice and at 2, 4, and 8 weeks (W) after TAC in WT mice (sham: n = 8, other groups: n = 6 each). Hsc70 served as the loading control. Protein intensity values of sham-operated heart tissue were set to 1. b Representative immunoblotting of HA-tagged Hint1 protein in H9c2-mock and H9c2-Hint1 cells. Hsc70 served as the loading control. This experiment was repeated independently three times with similar results. c Representative TMRM fluorescence images of H9c2-mock and H9c2-Hint1 cells (scale bar, 50 μm). d, e Quantification of TMRM (d) and MitoSOX Red (e) fluorescence intensity in H9c2-mock and H9c2-Hint1 cells (n = 4 per group). Values derived from H9c2-mock cells were set at 1. f Mitochondrial oxygen consumption rate (OCR) in H9c2-mock and H9c2-Hint1 cells (n = 7 biological replicates per group). g Quantification of basal respiration, maximal respiration, ATP production, spare respiratory capacity, proton leak, non-mitochondrial respiration, and coupling efficiency in H9c2-mock and H9c2-Hint1 cells (n = 7 biological replicates per group). Data show mean ± SD and all points for a, d, e, g. For f, data show mean ± SD. Statistical significance was determined by two-sided unpaired Student’s t-test (d, e, g) or one-way ANOVA with Sidak’s post hoc test (a). †p < 0.0001, n.s; not significant, between groups.
