Fig. 1.

Mitochondrial volume, MMP, ATP, l-lactate, and ROS generation in epithelial–stromal unit (indirect keratinocytes–fibroblasts co-culture system). a Graphic representation of the indirect co-culture model system used in the study. Either NOFs or CAFs (representing the stromal compartment) were co-cultured with NOKs or OSCCs (representing the epithelial compartment) in six-well plates with transwell membranes with pore size 0.4 μm. Flow cytometric analysis of different epithelial–stromal units for specific MMP (b) and their mitochondrial volume (c). Increase of mitochondrial volume but decrease of MMP was observed from NOFs + NOKs unit, NOFs + OSCCs unit, CAFs +NOKs unit, CAFs + OSCCs unit. d Measurement of intracellular ATP in four different epithelial–stromal unit using luminescent assay showed no significant differences among these four units. e, f Measurement of l-lactate and ROS activity in these different epithelial–stromal units showed statistically significant increased lactate production in the units containing OSCC cells and no significant differences in ROS activity among the four units. The data points were presented from six different patient sets in b–d. The data points were presented from the replicates on the averages of the six different patient combinations in e and f. All points represent mean values and SEM. Independent experiments were performed more than three times. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001; Wilcoxon signed-rank test was used in b–d; paired t test was used in e and f