Figure 6.

CCG-203971 and CCG-232601 regulate mitochondrial bioenergetics. WI-38 cells were treated with varying concentrations of CCG-203971 and CCG-232601 for 24 h. (A) Seahorse XF Cell Mito Stress Test results of control (0.5% DMSO treated) and CCG molecule-treated cells showing mean ± SD normalized to equal number of cells. Oligomycin was injected to inhibit ATP-linked mitochondrial respiration as a measure of mitochondrial ATP production. To determine maximal respiration, the mitochondrial uncoupler FCCP was injected. To determine the non-mitochondrial respiration to the OCR, rotenone was injected, inhibiting mitochondrial respiration. Quantification of basal respiration, maximal respiration, and spare respiratory capacity calculated from Seahorse XF Cell Mito Stress Test. (B) Glycolytic rate assay shown as mean ± SD normalized to equal number of cells. Rotenone/Antimycin A mix was injected to block mitochondrial activity and 2-DG (to inhibit glycolysis) as an internal control. Graphs show the calculated glycolytic parameters of basal and compensatory glycolysis. (C) Real-time ATP rate assay in WI-38 cells. On treatment with CCG molecules, there is reduction in total ATP produced from oxidative phosphorylation, while ATP generated from glycolysis was increased compared to control cells. Error bars represent mean ± SD (n = 3). * p < 0.05; *** p < 0.001; **** p  <  0.0001 by one-way ANOVA with Tukey’s method.