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Fig. 1 | BMC Biology

Fig. 1

From: Proton export upregulates aerobic glycolysis

Fig. 1

Over-expression of CA-IX in MCF-7 breast cancer cells increases glycolytic metabolism in vitro. A Overall survival Kaplan–Meier curve in ER-positive luminal B breast cancer comparing low and high CA9 gene expression n = 226 (kmplot.com). Statistical analysis using Log-rank P test p = 0.0433. B Immunoblotting of protein lysates from MCF-7 cells transfected with empty vector (MOCK-2) or Ca9 vector (M1 and M6). Proteins from total cell extracts were immunoblotted for CA-IX, CA-II, CA-XII, and B-actin (loading control). C Representative immunocytochemistry images of CA-IX protein expression in MCF-7, MOCK-2, and CA-IX clones M1 and M6. CA-IX clones (M1 & M6) exhibit CA-IX membrane staining, whereas MOCK-2 and parental MCF-7 cells do not. DAPI nuclear stain (blue), wheat germ agglutinin membrane stain (green), and CA-IX stain (red). D Glucose induced proton production rate (PPR) using the Seahorse extracellular flux analyzer, measured post glucose injection (N = 8 biological replicates per group). E Glucose uptake of cells in each group (n = 3 biological replicates) over 24 h, measured as luminescence generated using Glucose Uptake-Glo assay (Promega). F Lactate measured in extracellular media after 24 h using Sigma kit (n = 3 biological replicates per group). G Basal oxygen consumption rate (OCR) measured using the Seahorse extracellular flux analyzer in 5.8 mM glucose concentration (N = 8 biological replicates per group). D–G Data are shown as mean ± SD, statistical analysis using ordinary one-way ANOVA. *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001

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