Figure 4.

PANC-1 clones and heterogeneity with respect to deTD and circadian rhythmicity. (A) The indicated PANC-1 clones were subjected to deTD culture for 5 days in normal growth medium supplemented with 100 ng/mL IFNγ, 25 ng/mL IL1β and 50 ng/mL TNFα (TDC-IIT). Following lysis and RNA extraction, INS and NEUROG3 expression was determined by qPCR and C_t values normalized with those for GAPDH. The assay shown (means ± SD of three replicates) is representative of three experiments. Significant differences (p < 0.05) are marked by asterisks (*). (B) Bmal1::Luc bioluminescence reporter assays in parental (par) PANC-1 cells and subclones. Each clone is presented separately. For PANC-1, each applied synchronization treatment is depicted in a different color, as indicated (dexa: black; forskolin: blue; horse serum shock: red). The 24 h baseline subtracted bioluminescence data are shown as mean ± SEM. Data are pooled from two independent experiments with 4–8 replicates each. Dampened sine curves were fitted for rhythmic clones for rhythm parameter determination (Figure S7).