Age-related Mitochondrial Function Analysis

Graphing and statistical analysis were performed using Microsoft Office Excel and GraphPad Prism for OS X (GraphPad Software, San Diego, California USA). For all statistical tests, alpha levels were set to p<0.05. *p<0.05 for post hoc tests or direct comparisons unless described otherwise. IC₅₀ for ROS and membrane potential in isolated mitochondria was determined using nonlinear regression [Inhibitor] vs. Response – variable slope (four parameters) equation, and EC₅₀ for OCR was determined using [Agonist] vs normalized dose response. For comparisons across a range of ADP concentrations or frequencies for muscle contraction, young vs old, young vs young acute, and old vs old acute were compared by two-way RM ANOVA with Sidak’s post hoc test. Old vs old acute vs old chronic were compared with mixed-effects model (REML) with Tukey’s post hoc test. For heart function, pre- and post-ELAM treatment was compared by paired two-tailed student’s t-test. Gastrocnemius fibers ROS production was compared by unpaired Welch’s t-test, because the variances were significantly different. Principal component analysis was performed using ClustVis as previously described (Metsalu and Vilo, 2015 (link); Pharaoh et al., 2019 (link)). Main effect significant results are described in text and on graph. Plots depict mean ± standard deviation.