Fig. 4: Intrauterine hyperglycaemia inhibited the expression of Ppargc1α in foetal skeletal muscle through CREB/PGC1A signalling.
A ChIP sequence heatmap of genome-wide binding of H3K27ac showed that H3K27ac were highly enriched in GDM male foetal muscle around transcription starting sites. The rows showed the normalised unique tag counts for one H3K27ac binding event (ChIP peak) of the union, ordered by signalling strength; B H3K27ac occupancy at the Ppargc1a locus showed no significant difference between two groups determined by ChIP-seq analysis in foetal mouse skeletal muscle; C Schematic representation of selected loci for epigenetic experiments; D H3K4me3 ChIP-qPCR in foetal male skeletal muscle did not reveal difference in the promoter and enhancer regions of Ppargc1α between two groups (n = 5, multiple t tests); E DNA methylation of Ppargc1α at one CpG site showed no difference between groups (n = 6, two-tailed t test); F Representative western blotting images and analysis showed PGC1A, OXPHOS and pCREB were consistently down-regulated in GDM foetal muscle (n = 9, two-tailed t test); G ChIP-qPCR showed pCREB was fewer bound at the CRE element of Ppargc1α, Pck and G6pc in GDM foetal male skeletal muscle (nCTR= 4, nGDM = 6, multiple t tests); H Heatmap of transcription level of Pck and G6pc showed they were both down-regulated in GDM foetal muscle according to RNA-seq (n = 5); I Representative mRNA levels of genes mentioned above in C2C12 were inhibited to some extent after CREB signalling inhibition (n = 4, multiple t tests); J Representative protein levels of CREB/PGC1A/OXPHOS signalling also demonstrated to be lower in C2C12 after CREB signalling inhibition (n = 3, two-tailed t tests); K Genes mentioned above were stimulated to transcript in C2C12 after CREB signalling stimulation (n = 3, multiple t tests). The data are expressed as the mean ± SEM. Significance of the differences: *p < 0.05, **p < 0.01, ***p < 0.001.
