Fig. 2.

Induction of Snail mediates lactate-induced chemoresistance. a and b 24 h after transfection with either GPR81 siRNA (a) or GPR81 cDNA (b), cells were stimulated first with LA (10 mM, 20 mM) for 3 h and then with 25 μM etoposide for additional 36 h. The blots have been probed with antibodies against Cleaved-PARP, GPR81, Snail. c The cells were pre-treated with MCT1 inhibitor, a-cyano-4-hydroxycin-namate (CHC) or DMSO as control followed by further combined treatment of LA and etoposide. In contrast to Snail levels, western blot demonstrates a significant decrease in levels of Cleaved-PARP in CHC-treated group. d and e Western blot analysis of Cleaved-PARP, p-ATM, ATM, Snail, Puma in A549 and H1299 cells at day 3 after infection with Snail cDNA (d) or Snail siRNA (e) and then further treated with LA and etoposide described in the legend to Fig. 1e. f The cells were pre-treated with TGF-β inhibitor, LY2157299, and then with the same treatment as Fig. 2c. g The A549 cells were first transfected with Snail cDNA plasmid. At 48 h post-transfection, cells were treated with etoposide in the presence and absence of LA followed by MTT assay. Corresponding EC₅₀values for etoposide, Etoposide+Snail and Etoposide+LA are 19.39 μM, 35.31 μM and 28.28 μM respectively. (*p<0.05, **p<0.01 for difference of overexpressed Snail from etoposide-treated cells, ^#p<0.05 for difference of LA-treated from etoposide-treated cells by ANOVA with Dunnett’s correction for multiple comparisons, ns means no statistical difference)