Fig. 6
Enhanced lactate production is mediated by IL-8/DAPK1/PK axis. A. Quantification of Huh-7 lactate production in PFK-158 and control group; determination after 24 h of culture; the protein concentration was used to standardize lactate production. B. Quantification of HCCLM3 (2 × 105 cells/cm2) lactate production in PFK-158 and control group; determination after 24 h of culture; the protein concentration was used to standardize lactate production. C. Quantification of Huh-7 and HCCLM3 PK activity; control and IL-8 KD or OE HCC cells were intervened with PFK-158 (10 µM for 24 h); the protein concentration was used to standardize PK activities. D. Spearman correlation between IL-8 and DAPK1 expression in HCC; IHC and Image J were used for quantification; n = 65. E. The protein expression of DAPK1 in the IL-8 KD and control Huh-7 cells; The protein expression of DAPK1 in Huh-7 cells with different cell densities. F. The protein expression of DAPK1 in IL-8 OE and control HCCLM3 cells. G. The protein expressions of DAPK1 and PKM2 in DAPK1 KD and control Huh-7/HCCLM3 IL-8 OE cells. H. Quantification of lactate production and PK activity in DAPK1 KD and control Huh-7 cells of different cell densities; determination after 24 h of culture. I. Quantification of lactate production and PK activity in DAPK1 KD and control HCCLM3 IL-8 OE cells (2 × 105 cells/cm2) of different cell densities; determination after 24 h of culture. J. Quantification of ECAR in Huh-7 and HCCLM3 cells with PFK-158, DAPK1 KD, and IL-8 KD/OE. K. Effect of coculture with DAPK1 KD and control Huh-7 cells (1 × 105 cells/cm2) on Tregs polarization in PBMCs; IL-8 expression of DAPK1 KD and control Huh-7 cells (1 × 105 cells/cm2); determination after 24 h of culture. L. Effect of coculture with DAPK1 KD and control IL-8 OE HCCLM3 cells (2 × 105 cells/cm2) on Tregs polarization in PBMCs; IL-8 expression of DAPK1 KD and control IL-8 OE HCCLM3 cells (2 × 105 cells/cm2); determination after 24 h of culture. The data represent the mean ± S.D. of three independent experiments. Mann–Whitney U test or t-tests were used to compare the means of two groups. ANOVA were used for comparison among three groups. *P < 0.05; **P < 0.01; ***P < 0.001. Abbreviations: KD, knockdown; OE, overexpression; PBMCs, peripheral blood mononuclear cells; PK, pyruvate kinase