Fig. 3From: Elevated circulatory levels of leptin and resistin impair therapeutic efficacy of dacarbazine in melanoma under obese stateMolecular events associated with leptin and resistin induced impaired outcome of DTIC therapy in melanoma cells. a, b A375 cells were treated with leptin or resistin at a concentration of 100 ng/ml in DMEM containing 1% FBS for 48 h as described in the “Materials and methods” section. Thereafter, cycloheximide (100 μg/ml) treatment was given for the indicated time points. Representative immunoblot of FASN (a) and Cav-1 (b) in A375 cells treated with leptin or resistin respectively. c, d Bar graph showing the quantitation of band intensity of FASN and Cav-1 immunoblots. e Rhodamine-123 efflux assay in A375 cells treated with leptin (upper panel) or resistin (lower panel). A375 (human melanoma) cells were plated in 12-well plates. After 24 h, cells were treated with 100 ng/ml of recombinant leptin in DMEM containing 1% FBS for 48 h. Thereafter, these cells were subjected to Rh-123 efflux assay via flow cytometry. f, g RT-PCR (f) and immunoblotting (g) analysis of MDR and P-gp respectively in A375 cells treated with resistin. h, i RT-PCR (h) and immunoblotting (i) analysis of HSP90 in A375 cells treated with leptin. j, k Representative image showing the long-term survival of A375 cells grown in the presence or absence of leptin (j) or resistin (k) together with inhibitors. The results are given as means ± standard error of the mean. All the experiments were performed three times. Statistical analysis was performed using two-tailed unpaired Student’s t test for c and d. *p < 0.05, **p < 0.001; Ctrl, control; Lep, leptin; Res, resistin; Chx, cycloheximide; Ceru or C, cerulenin; GA or G, geldanamycinBack to article page