Figure 9.

Metformin inhibits mitochondrial respiration in SCs. (a) Oxygen consumption rate (OCR) of the Mito Stress Test conducted with a Seahorse Extracellular Flux Analyzer. OCR is expressed in pmol of O₂ consumed per minute, normalized over the number of cells in the assay well. Oligomycin (Oligom.), FCCP, and rotenone/antimycin (Rot/Ant) were sequentially added to the well to perturb the electron transport chain machinery. (b–d) Bar plot graphs of mitochondrial functional parameters: (b) basal respiration is the delta between the basal OCR and the lowest plateau reached upon Rot/Ant injection, (c) maximal respiration is inferred as the delta between Rot/Ant and FCCP maximal plateau, and (d) ATP production is referred to as the delta between the basal and the oligomycin injection. Statistical analysis was performed through Student's t-test (^∗∗∗∗ p < 0.0001, n = 4). (e) Western blot analysis of Tom20 protein levels in control and metformin-treated SCs after 2, 4, 6, and 8 days of treatment (t2, t4, t6, and t8, respectively). Vinculin was used as a loading control. (f) Quantitation graph of Tom20 protein levels monitored by western blot in four independent cell isolations and biological replicates (n = 4). Statistical significance was evaluated by the ANOVA test (^∗∗ p < 0.01).