Figure 3.

Influence of CK2 inhibition on insulin expression and secretion. MIN6 cells were treated with 1 or 10 µM SGC-CK2-1 (SGC) or 1 or 10 µM CX-4945 (CX) for 24 h. Control cells were incubated with an equal volume of the vehicle DMSO. (a) Cells were harvested, and total RNA was isolated using the QIAzol lysis reagent. The mRNA amount of insulin was determined using qRT-PCR. After normalization to GAPDH, the amount of insulin mRNA of control-treated cells was set 100% and the amount of treated mRNA cells calculated in reference to it. (b) Cells were harvested, proteins were extracted, and equal amounts were loaded on a 12.5% SDS polyacrylamide gel and blotted onto a PVDF membrane. A representative immunoblot analysis of MIN6 cell extracts for the detection of proinsulin is shown; the detection of α-tubulin served as loading control. Signals for proinsulin from three independent experiments were analyzed by a densitometric scan and normalized to the arbitrary amount of the loading control α-tubulin. The relative mean amount of proinsulin +/− SD for the different treatments is shown as bar graphs. (c) After a glucose stimulus, secreted insulin was determined in the cell culture supernatant with the insulin ELISA kit. Statistical analysis in all experiments was performed as described in “Material and Methods”. * Statistical significance was accepted with a p-value of at least p < 0.05.