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. Author manuscript; available in PMC: 2020 Jan 8.
Published in final edited form as: Cell Metab. 2018 Aug 30;29(1):141–155.e9. doi: 10.1016/j.cmet.2018.08.007

Figure 3. Glycogen fuels glycolysis to promote proliferation and invasion.

Figure 3.

(A) TYK-nu cells were pretreated (1 hr) with the glycogen phosphorylase inhibitor CP-91149 followed by stimulation with conditioned media (CM) from primary human CAFs for 16 hr. Glycolysis was measured using the Seahorse SF96 Extracellular Flux Analyzer. Left, extracellular acidification rates (ECAR) are shown normalized to cell number. ‘a’, ba sal glycolysis (no glucose); ‘b’, glycolysis (stimulated with glucose); ‘c’, glycolytic capacity (stimulated with oligomycin). Right panel shows the average of the 3 glycolysis time points for each group. Values are from 4 independent experiments (n=15/group). Comparisons were made to CAF CM using a paired, two-tailed t test (*p<0.05, **p<0.01). (B) ATP/ADP ratios in TYK-nu cells treated with CAF or control CM pretreated with or without CP-91149, an inhibitor of glycogen phosphorylase. Values are mean + SEM from 3 independent experiments (n=4/group). Comparisons were made to CAF CM using a paired, two-tailed t test (*p<0.05). (C) Steady state 12C metabolites in TYK-nu cells stimulated with control CM or CAF CM for 6 hr (n=3/group). Key metabolites are shown along with a schematic depicting the central pathways (* p<0.05, **p<0.01, ***p<0.001). (D) ROS level in GFP-labeled SKOV3ip1 cancer cells with or without CAF co-culture and CP-91149. Left, representative images of ROS staining (red). Right, quantification of 3 independent experiments. Values are mean + SEM. Samples were compared to OvCa-GFP+CAF using an unpaired, two-tailed t test (* p<0.05, **p<0.01). (E) Proliferation of TYK-nu cells exposed to CAF CM with or without the glycogen phosphorylase inhibitor CP-91149. Values are mean + SEM from 4 independent experiments (n=6/group). Comparisons were made to CAF CM using a Two-way ANOVA (* p<0.05, **p<0.01). (F) Invasion of TYK-nu cells exposed to CAF CM with or without the glycogen phosphorylase inhibitor CP-91149. Data is representative of 3 independent experiments. Values are mean + SEM (n=3/group). ***p<0.001.