Figure 2.

TNFα increases oxidative phosphorylation in quiescent astrocytes. (A) Astrocytes were left untreated (white symbols) or stimulated with TNFα (10 ng/mL for 24 h) (red) and then the realtime oxygen consumption rate (OCR) was measured using the Seahorse XF analyzer. Data are normalized to total protein (μg) present in the wells at the end of the analysis and represent at least 3 independent experiments. (B) Combined OCR measurements from technical replicates in 3 separate experiments are shown as mean ± SEM for the replicates; each symbol represents one well. P-values were calculated using mixed model nested t-tests. (C) Astrocytes were left untreated (white symbols) or stimulated with TNFα (10 ng/mL for 24 h) (red) and then the realtime extracellular acidification rate (ECAR) was measured. Data are normalized to total protein (μg) per well and represent at least 3 independent experiments. (D) Combined ECAR measurements from technical replicates in 2 separate experiments are shown as mean ± SEM for the replicates; each symbol represents one well. P-values were calculated using mixed model nested t-tests. (E) The contribution of glycolytic or mitochondrial ATP production was analyzed by Seahorse in quiescent cells stimulated with TNFα (10 ng/mL for 24 h) (red) or vehicle (DMSO; white). Data are normalized to total protein and shown as mean ± SEM from at least 6–8 replicates in two separate experiments. P-values were calculated using mixed model nested t-tests. (F) The ratio of mitochondrial ATP production rate to glycolytic ATP production rate was calculated in at least 8 technical replicates in 3 separate experiments and shown as mean ± SEM for the replicates. P-value was calculated using mixed model nested t-tests.