Fig. 1. Levels of Cx36 mRNA and protein are decreased by glucose in insulin-secreting cells. (A) Quantitative RT-PCR analysis showed a decrease of Cx36 mRNA in insulin-secreting cell lines and pancreatic rat islets after a 24 hour incubation at high (20 mM) versus low (2 mM) glucose concentration. Rat islets, ßTC3 cells and MIN6-B1 cells display a 40% decrease in Cx36 transcript whereas the INS-1 and INS-1E cell lines present a 60% decrease in Cx36 mRNA level. Data are means ± s.e.m. of four independent experiments. Relative Cx36 transcripts levels were normalized to the ribosomal protein L27. (B) Northern blots showed a dose-dependent decrease of Cx36 mRNA in INS-1 cells after 24 hours. By contrast, the level of the macrophage inhibitory factor (MIF) encoding mRNA, used as a positive control, is increased by glucose. Quantitative assessment of Cx36 mRNA confirmed that glucose induces a 60% reduction of Cx36 transcript expression. Data are means ± s.e.m. of five independent experiments. (C) Western-blot analysis normalized with tubulin and ß-actin showed a dose-dependent decrease of Cx36 in INS-1 cells after 24 hours. Densitometric analysis indicated that Cx36 protein levels were decreased by 70% in presence of a high glucose concentration. Data are means ± s.e.m. of six independent experiments. ^*, P<0.05; ^**, P<0.01; ^***, P<0.001.