Fig. 6.
Expression of Dnmt genes in islets and beta cells. (a) The transcription of several Dnmt genes was measured in islets from NOD mice at the indicated ages and normalised to levels in islets from 3-week-old mice. (b) Beta cells and islet-infiltrating lymphocytes were sorted from islets of 6-week-old NOD mice. The levels of Dnmt gene transcription in beta cells (black bars) was normalised to levels in islet-infiltrating lymphocytes (white bars). Data are the means ± SEM of 3 experiments, each with 4 mice. (c) The expression of Dnmt genes was determined by qRT-PCR in beta cells from islets cultured in IL-6 plus IL-1β (horizontal stripes) or IFN-γ plus IL-1β (diagonal stripes). (d, e) Islets from 4-week-old NOD mice were transfected with siRNA against Dnmt3a (grey bars) or scrambled control siRNA (siControl; black bars) 24 h prior to cytokine treatment with IFN-γ plus IL-1β for 48 h. Levels of Ins1 and Ins2 expression and Dnmt3a mRNA in beta cells were then analysed by qRT-PCR. (d) Methylation of CpG sites in Ins2 exon 1 was determined by Sanger sequencing. (e) χ2 test, p<0.0001. (a–d) ANOVA with Tukey’s multiple comparison tests: *p<0.05, ****p<0.0001. (c, d) Data were normalised to beta cells from islets cultured in control medium. (d, e) Data are the means ± SEM from three experiments, each with eight mice