Previously, the paternal contribution to programming of metabolic disease was thought to be minimal. It is now accepted that paternal obesity can contribute to offspring metabolic risk via non-genetic mechanisms.  However, the identity and mechanisms of this non-genetic inheritance are poorly understood. We previously reported altered glucose metabolism in offspring of obese male rats, an effect inherited through non-genetic means. Here we examined whether transgenerational defects can be transmitted to F2 offspring. 

Control F1 male rats from obese (ob) (30% heavier at time of mating) and control (con) F0 fathers  were mated with control females to generate F2 offspring.  F2 animals were fed either a control diet (CD) or challenged with a high fat (HFD) diet; increased body weight was observed in those from obese versus con F0. Liver and tibialis showed 25% increased triglyceride content in F2 siblings fed HFD versus CD (p<0.05). In both tissues, HFD consumption in F2 from obF0 resulted in an increased expression of gene involved in lipogenesis (Srebf1, Fasn) and triglyceride synthesis (Dgat1-2). In liver, genes involved in mitochondrial function (Mct-Cox genes) and biogenesis (Sirt1-Nrf genes) were down-regulated by grand-paternal obesity, with an interactive effect of HFD consumption in F2. Reduced expression of genes involved in lipid β-oxidation (Hadh, Acat1) was observed in liver and muscle from HFD fed F2 from ob-F0.

We also investigated sperm DNA methylation in this cohort. A small (0.25%) but significant increase in 5-methylcytosine was detected in obese rat sperm with LCMS. Examination of the repetitive fraction of the genome with MBD-Seq and pyrosequencing revealed that LINE and IAP retrotransposon DNA methylation states in sperm were not affected by obesity but centromeric satellite repeats had an approximate 8% increase in methylation. Examination of muscle, liver and sperm of F1 offspring revealed normal DNA methylation levels.  These data demonstrate the efficiency of the transgenerational epigenetic reprogramming mechanisms for normalising paternally-inherited DNA methylation levels.