Intrauterine hyperglycemia during late gestation caused mitochondrial dysfunction in skeletal muscle of male offspring through CREB/PGC1A signaling

Abstract

Background: Maternal diabetes mellitus can influence the development of offspring. Gestational diabetes mellitus (GDM) creates short-term intrauterine hyperglycemic environment for offspring, leading to insulin resistance in skeletal muscle, but the long-term effect and specific mechanism involved in skeletal muscle dysfunction in offspring remains to be clarified. Methods: Pregnant mice were divided into two groups: the GDM group was intraperitoneally injected with 100mg/kg streptozotocin on gestational days (GD) 6.5 and 12.5, while the control (CTR) group was treated with vehicle buffer. Only pregnant mice whose random blood glucose higher than 16.8mmol/L from GD13.5 will be regarded as GDM group. The growth of offspring was monitored and the glucose tolerance test was traced at different periods. Body composition analysis and immunohistochemical methods were used to evaluate the development of lean mass at 8 weeks. Transmission electron microscopy was utilized to observe the morphology inside skeletal muscle at 8 weeks and fetus. Genes and protein expression associated with mitochondrial biogenesis and oxidative metabolism were investigated. We also co-analyzed RNA sequencing and proteomics to explore its mechanism. Chromatin immunoprecipitation and bisulfite-converted DNA methylation detection were performed to explain the phenomenon. Results: Short-term intrauterine hyperglycemia inhibited the growth and reduced the lean mass of male offspring. The myofiber composition in GDM offspring male tibialis anterior muscle turned into glycolytic type. The morphology and function of mitochondria in skeletal muscle of GDM male offspring were destroyed, and co-analysis of RNA sequencing and proteomics of fetal skeletal muscle showed mitochondrial element and lipid oxidation were consistently impaired. Ex vivo and in vitro myoblast experiments also demonstrated that high glucose impeded mitochondrial organization and function, transcription of genes associated with mitochondrial biogenesis and oxidative metabolism was decrease at 8 weeks and fetal period. The protein and mRNA levels of Ppargc1a in male offspring were decreased at fetus (CTR vs GDM, 1.004 vs 0.665, p=0.002), 6 weeks (1.018 vs 0.511, p=0.023) and 8 weeks (1.006 vs 0.596, p=0.018) in skeletal muscle. In addition, CREB phosphorylation was restrained, with fewer activated pCREB protein binding to CRE element of Ppargc1a (1.042 vs 0.681, p=0.037), Pck(1.091 vs 0.432, p=0.014) and G6pc (1.118 vs 0.472, p=0.027), resulting in less transcription. Interestingly, we found sarcopenia and mitochondrial dysfunction could even be inherited by the next generation. Conclusions: Short-term intrauterine hyperglycemia reduced lean mass in male offspring significantly, and disrupted the organization and function of the mitochondrion in skeletal muscle which contributed to insulin resistance and glucose intolerance. Fetal exposure to hyperglycemia decreased phosphorylated CREB and reduced transcription of Ppargc1a. Abnormal mitochondrion was also observed in the F2 generation, which might be transmitted through aberrant gametes.