Figure 3.

Metabolic profile analysis of resistant and sensitive cells and tumor pH. In vitro oxygen consumption rate (OCR) and extracellular acidification rate (ECAR) of resistant and sensitive cells, measured using Seahorse XF-96 Instrument. Metabolic data are presented as mean ± SD. (A) ECAR measurements in response to 5.55 mmol/l glucose stimulation indicate basal glycolytic activity of cells. Sensitive cells are significantly more glycolytic than resistant cells. (B) The increase in ECAR of cells in response to treatment with 1 μmol/l oligomycin minus basal glycolytic activity indicates the glycolytic reserve of cells. Sensitive cells have significantly reduced glycolytic reserve compared resistant cells.(C) Basal OCR measurements show significantly higher oxidative phosphorylation flux in resistant cells compared to sensitive cells. (D) OCR contributing to production of ATP during oxidative phosphorylation is measured by the difference of basal OCR and the OCR of cells after treatment with 1 μmol/l oligomycin, a mitochondrial ATP synthase inhibitor. The amount of OCR contributing to the production of ATP by oxidative phosphorylation is significantly higher in resistant cells compared to sensitive cells. (E) The OCR/ECAR ratio of cells during basal metabolism indicates that sensitive cells are significantly more glycolytic than resistant cells. (F) Intratumoral pH measurements of subcutaneous tumors using pH electrodes shows that increased glycolytic activity of sensitive cells, PC3M_S, contributes to a more acidic tumor microenvironment than resistant tumors, B16-F10_R. pH data are presented as mean of independent measurements (B16-F10_Rn = 10; PC3M_Sn = 5) ± SEM. *P < .05; ****P < .0001. _R, resistant; _S, sensitive.