Environmental constraints during early life result in phenotypic changes that can be associated with increased disease risk in later life. This suggests persistent alteration of gene transcription. DNA methylation, which is largely established in utero, provides a causal mechanism by which unbalanced prenatal nutrition results in such altered gene expression. We investigated the effect of unbalanced maternal nutrition on the methylation status and expression of the glucocorticoid receptor (GR) and peroxisomal proliferator-activated receptor (PPAR) genes in rat offspring after weaning. Dams were fed a control protein (C; 180 g/kg protein plus 1 mg/kg folic acid), restricted protein (R; 90 g/kg casein plus 1 mg/kg folic acid), or restricted protein plus 5 mg/kg folic acid (RF) diet throughout pregnancy. Pups were killed 6 d after weaning (n = 10 per group). Gene methylation was determined by methylation-sensitive PCR and mRNA expression by semiquantitative RT-PCR. PPARα gene methylation was 20.6% lower (P < 0.001) and expression 10.5-fold higher in R compared with C pups. GR gene methylation was 22.8% lower (P < 0.05) and expression 200% higher (P < 0.01) in R pups than in C pups. The RF diet prevented these changes. PPARγ methylation status and expression did not differ among the groups. Acyl-CoA oxidase expression followed that of PPARα. These results show that unbalanced prenatal nutrition induces persistent, gene-specific epigenetic changes that alter mRNA expression. Epigenetic regulation of gene transcription provides a strong candidate mechanism for fetal programming.

Reproduction Nutrition Development, formerly Annales de Biologie Animale Biochimie Biophysique

The extent to which women of reproductive age are able to convert then−3 fatty acid α-linolenic acid (ALNA) to eicosapentaenoic acid (EPA), docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA) was investigatedin vivoby measuring the concentrations of labelled fatty acids in plasma for 21 d following the ingestion of [U−¹³C]ALNA (700 mg). [¹³C]ALNA excursion was greatest in cholesteryl ester (CE) (224 (SEM 70) μmol/l over 21 d) compared with triacylglycerol (9-fold), non-esterified fatty acids (37-fold) and phosphatidylcholine (PC, 7-fold). EPA excursion was similar in both PC (42 (SEM 8) μmol/l) and CE (42 (SEM 9) μmol/l) over 21 d. In contrast both [¹³C]DPA and [¹³C]DHA were detected predominately in PC (18 (SEM 4) and 27 (SEM 7) μmol/l over 21 d, respectively). Estimated net fractional ALNA inter-conversion was EPA 21 %, DPA 6 % and DHA 9 %. Approximately 22 % of administered [¹³C]ALNA was recovered as¹³CO₂on breath over the first 24 h of the study. These results suggest differential partitioning of ALNA, EPA and DHA between plasma lipid classes, which may facilitate targeting of individualn−3 fatty acids to specific tissues. Comparison with previous studies suggests that women may possess a greater capacity for ALNA conversion than men. Such metabolic capacity may be important for meeting the demands of the fetus and neonate for DHA during pregnancy and lactation. Differences in DHA status between women both in the non-pregnant state and in pregnancy may reflect variations in metabolic capacity for DHA synthesis.

OBJECTIVE: Fixed genomic variation explains only a small proportion of the risk of adiposity. In animal models, maternal diet alters offspring body composition, accompanied by epigenetic changes in metabolic control genes. Little is known about whether such processes operate in humans. RESEARCH DESIGN AND METHODS: Using Sequenom MassARRAY we measured the methylation status of 68 CpGs 5′ from five candidate genes in umbilical cord tissue DNA from healthy neonates. Methylation varied greatly at particular CpGs: for 31 CpGs with median methylation ≥5% and a 5–95% range ≥10%, we related methylation status to maternal pregnancy diet and to child’s adiposity at age 9 years. Replication was sought in a second independent cohort. RESULTS: In cohort 1, retinoid X receptor-α (RXRA) chr9:136355885+ and endothelial nitric oxide synthase (eNOS) chr7:150315553+ methylation had independent associations with sex-adjusted childhood fat mass (exponentiated regression coefficient [β] 17% per SD change in methylation [95% CI 4–31], P = 0.009, n = 64, and β = 20% [9–32], P < 0.001, n = 66, respectively) and %fat mass (β = 10% [1–19], P = 0.023, n = 64 and β =12% [4–20], P = 0.002, n = 66, respectively). Regression analyses including sex and neonatal epigenetic marks explained >25% of the variance in childhood adiposity. Higher methylation of RXRA chr9:136355885+, but not of eNOS chr7:150315553+, was associated with lower maternal carbohydrate intake in early pregnancy, previously linked with higher neonatal adiposity in this population. In cohort 2, cord eNOS chr7:150315553+ methylation showed no association with adiposity, but RXRA chr9:136355885+ methylation showed similar associations with fat mass and %fat mass (β = 6% [2–10] and β = 4% [1–7], respectively, both P = 0.002, n = 239). CONCLUSIONS: Our findings suggest a substantial component of metabolic disease risk has a prenatal developmental basis. Perinatal epigenetic analysis may have utility in identifying individual vulnerability to later obesity and metabolic disease.

The capacity for conversion of α-linolenic acid (ALNA) ton−3 long-chain polyunsaturated fatty acids was investigated in young men. Emulsified [U−¹³C]ALNA was administered orally with a mixed meal to six subjects consuming their habitual diet. Approximately 33 % of administered [¹³C]ALNA was recovered as¹³CO₂on breath over the first 24 h. [¹³C]ALNA was mobilised from enterocytes primarily as chylomicron triacylglycerol (TAG), while [¹³C]ALNA incorporation into plasma phosphatidylcholine (PC) occurred later, probably by the liver. The time scale of conversion of [¹³C]ALNA to eicosapentaenoic acid (EPA) and docosapentaenoic acid (DPA) suggested that the liver was the principal site of ALNA desaturation and elongation, although there was some indication of EPA and DPA synthesis by enterocytes. [¹³C]EPA and [¹³C]DPA concentrations were greater in plasma PC than TAG, and were present in the circulation for up to 7 and 14 d, respectively. There was no apparent¹³C enrichment of docosahexaenoic acid (DHA) in plasma PC, TAG or non-esterified fatty acids at any time point measured up to 21 d. This pattern of¹³Cn−3 fatty acid labelling suggests inhibition or restriction of DHA synthesis downstream of DPA. [¹³C]ALNA, [¹³C]EPA and [¹³C]DPA were incorporated into erythrocyte PC, but not phosphatidylethanolamine, suggesting uptake of intact plasma PC molecules from lipoproteins into erythrocyte membranes. Since the capacity of adult males to convert ALNA to DHA was either very low or absent, uptake of pre-formed DHA from the diet may be critical for maintaining adequate membrane DHA concentrations in these individuals.

Prenatal nutritional constraint induces an altered metabolic phenotype in the offspring which in humans confers an increased risk of non-communicable disease. Feeding a protein-restricted (PR) diet to pregnant rats causes hypomethylation of specific gene promoters in the offspring and alters the phenotype. We investigated how altered epigenetic regulation of the hepatic glucocorticoid receptor (GR) 1₁₀promoter is induced in the offspring. Rats were fed a control (180 g casein/kg) or a PR (90 g casein/kg) diet throughout pregnancy, and chow during lactation. Offspring were killed at postnatal day 34 (n5 per maternal dietary group). Methylation-sensitive PCR showed that GR1₁₀promoter methylation was 33 % lower (P < 0·001) and GR expression 84 % higher (P < 0·05) in the PR offspring. Reverse transcription–PCR showed that DNA methyltransferase-1 (Dnmt1) expression was 17 % lower (P < 0·05) in PR offspring, while Dnmt3a/b and methyl binding domain protein-2 expression was not altered. Thus hypomethylation of the GR1₁₀promoter may result from lower capacity to methylate hemimethylated DNA during mitosis. Histone modifications which facilitate transcription were increased at the GR1₁₀promoter (147–921 %,P < 0·001), while those that suppress methylation were decreased (54 %,P < 0·01) or similar to controls. In human umbilical cord (n15), there was a 2-fold difference between the highest and lowest level of GR1-C_Totalpromoter methylation. Dnmt1, but not Dnmt3a, expression predicted 49 % (P = 0·003) of the variation in GR1-C_Totalpromoter methylation. These findings suggest that induction in the offspring of altered epigenetic regulation of the hepatic GR1₁₀promoter, and hence metabolic phenotype, may be due to reduced Dnmt1 expression.