Fig. 3
PSC-CM induces resistance to gemcitabine in PCCs but does not affect gemcitabine uptake. PCCs seeded on 96-well plates were incubated with SFM or PSC-CM for 24 h prior to incubation with (a, b) gemcitabine (10 μM) for 48 h or with (c) transport buffer containing [3H]-gemcitabine for 4 h. For (a), cell viability was determined using the MTT assay. *p < 0.05, **p < 0.01 for control vs gemcitabine; #p < 0.05, ##p < 0.01 and $p < 0.05, $$p < 0.01 for SFM vs PSC-CM in control and gemcitabine groups, respectively. b The table indicates gemcitabine-induced cytotoxicity in percentage, and PSC-CM-induced resistance to gemcitabine, calculated by relative reduction in cytotoxicity between SFM and PSC-CM. *p < 0.05, **p < 0.01 comparing SFM with PSC-CM. For c, gemcitabine uptake was determined by scintillation counting. Data are mean ± SEM of triplicate determinations. d The PCCs were lysed and proteins subjected to immunoblotting using anti-pERK1/2, anti-ERK1/2, anti-PI3K, anti-pAKT, and anti-AKT antibodies. GAPDH was used as a loading control. PCC, pancreatic cancer cell; PSC, pancreatic stellate cell; PSC-CM, PSC-conditioned medium; SFM, serum-free DMEM