Paternal chronic folate supplementation induced the transgenerational inheritance of acquired developmental and metabolic changes in chickens

Abstract

Increasing evidence indicates that paternal diet can result in metabolic changes in offspring, but the definite mechanism remains unclear in birds. Here, we fed breeder cocks five different diets containing 0, 0.25, 1.25, 2.50 and 5.00 mg kg^-1 folate throughout life. Paternal folate supplementation (FS) was beneficial to the growth and organ development of broiler offspring. Most importantly, the lipid and glucose metabolism of breeder cocks and broiler offspring were affected by paternal FS, according to biochemical and metabolomic analyses. We further employed global analyses of hepatic and spermatozoal messenger RNA (mRNA), long non-coding RNA (lncRNA) and micro RNA (miRNA). Some key genes involved in the glycolysis or gluconeogenesis pathway and the PPAR signalling pathway, including PEPCK, ANGPTL4 and THRSP, were regulated by differentially expressed hepatic and spermatozoal miRNAs and lncRNAs in breeder cocks and broiler offspring. Moreover, the expression of ANGPTL4 could also be regulated by differentially expressed miRNAs and lncRNAs in spermatozoa via competitive endogenous RNA (ceRNA) mechanisms. Overall, this model suggests that paternal folate could transgenerationally regulate lipid and glucose metabolism in broiler offspring and the epigenetic transmission may involve altered spermatozoal miRNAs and lncRNAs.

Keywords: PPAR signalling pathway; folate; glycolysis or gluconeogenesis; lncRNA; miRNA; paternal transgenerational epigenetics.

Figure 1.

Transgenerational developmental changes in breeder cocks and their broiler offspring induced by dietary FS in breeder cocks. (a,b) Effects on BW. (c,d) Effects on organ indexes. (e–g) Effects on intestinal morphology. (h) Effects on relative caecum length. Superscripts (a and b) denote significant differences in the same index. (Online version in colour.)

Figure 2.

Altered hepatic metabolomic profiles and transcriptomics profiles of breeder cocks and broiler offspring induced by dietary FS in breeder cocks. (a–c) Altered hepatic metabolomic profiles of breeder cocks and 1-day-old and 21-day-old broiler offspring; the upregulated metabolites are depicted in red, whereas the downregulated metabolites are depicted in green. (d–f) Significantly enriched metabolic pathways based on the differential metabolites. (g,h) Significantly enriched pathways in breeder cocks and 1-day-old broiler offspring based on hepatic differentially expressed genes. (i) Hepatic co-differentially expressed genes between breeder cocks and broiler offspring. (Online version in colour.)

Figure 3.

Transgenerational regulatory roles of miRNAs and lncRNAs in regulating lipid and glucose metabolism in response to paternal FS. (a,b) Significantly enriched pathways based on targets of hepatic differentially expressed miRNAs (DEmiRs) in breeder cocks and broiler offspring. (c) Co-enriched pathways based on targets of hepatic DEmiRs between breeder cocks and broiler offspring. (d–g) Significantly enriched pathways based on targets of spermatozoal DEmiRs between the CON and other four FS groups. (h) Co-enriched pathways based on targets of spermatozoal DEmiRs among four different compared groups between the CON and other four FS groups. (i,j) Significantly enriched pathways based on targets of hepatic DElRs in breeder cocks and 1-day-old broiler offspring. (k) Significantly enriched pathways based on targets of spermatozoal DElRs between the CON and other four FS groups of breeder cocks. (Online version in colour.)

Figure 4.

Integrated omics analysis based on the identified differentially expressed spermatozoal miRNA and lncRNAs of breeder cocks and the hepatic mRNAs of their offspring broilers. (a) The ceRNA network among differentially expressed hepatic mRNAs of offspring broilers and the spermatozoal miRNA and lncRNAs of breeder cocks. (b) The expression of PC-5p-30232_97 and PC-3p-448507_5 in primary chicken hepatocytes after the transfection of these miRNA mimics and mimic-NTC. (c) The expression of PEPCK and ANGPTL4 in primary chicken hepatocytes after the transfection of PC-5p-30232_97 and PC-3p-448507_5. (d) Luciferase activities in luciferase reporter assays when PEPCK or ANGPTL4 3′UTR was co-transfected in 293T cells with a PC-5p-30232_97 or PC-3p-448507_5 mimic, respectively. (Online version in colour.)