Higher Hepatic miR-29 Expression in Undernourished Male Rats During the Postnatal Period Targets the Long-Term Repression of IGF-1.

A nutritional mismatch in postnatal life of low birth weight offspring increases the risk of developing the metabolic syndrome. Moreover, this is associated with decreased hepatic Igf1 expression, leading to impaired growth and metabolism. Previously, we have demonstrated that the timing of nutritional restoration in perinatal life can differentially program hepatic gene expression. Although microRNAs also play an important role in silencing gene expression, to date, the impact of a nutritional mismatch in neonatal life on their long-term expression has not been evaluated. Given the complementarity of miR-29 to the 3' untranslated region of Igf1, we examined how protein restoration in maternal protein restriction rat offspring influences hepatic miR-29 and Igf1 expression in adulthood. Pregnant Wistar rats were designated into 1 of 4 dietary regimes: 20% protein (control), 8% protein during lactation only (LP-Lact), 8% protein during gestation only (LP1) or both (LP2). The steady-state expression of hepatic miR-29 mRNA significantly increased in LP2 offspring at postnatal day 21 and 130, and this was inversely related to hepatic Igf1 mRNA and body weight. Interestingly, this reciprocal association was stronger in LP-Lact offspring at postnatal day 21. Functional relevance of this in vivo relationship was evaluated by transfection of miR-29 mimics in neonatal Clone 9 rat hepatoma cells. Transfection with miR-29 suppressed Igf1 expression by 12 hours. Collectively, these findings implicate that nutritional restoration after weaning (post liver differentiation) in maternal protein restriction rat offspring fails to prevent long-term impaired growth, in part, due to miR-29 suppression of hepatic Igf1 expression.

Enhanced trimethylation of histone h3 mediates impaired expression of hepatic glucose 6-phosphatase expression in offspring from rat dams exposed to hypoxia during pregnancy.

Given that hepatic glucose 6-phosphatase (G6Pase, involved in gluconeogenesis) has been demonstrated to be altered long term in animal models of intrauterine growth restriction (IUGR), we hypothesized that hypoxia in utero may regulate G6Pase expression via epigenetic mechanisms. To address this further, a rat model of maternal hypoxia leading to IUGR and impaired liver growth was utilized. In the 12-month-old male offspring of pregnant rat dams exposed to 11.5% atmospheric oxygen from gestational day (gd) 15 to gd 21, nonfasting glucose was lower in association with decreased hepatic G6Pase messenger RNA and protein levels. This was concomitant with enhanced methylation of histone H3 [K9] surrounding the promoter of G6Pase. Moreover, when McA-RH7777 hepatoma cells were exposed to various concentrations of oxygen for 48 hours, we observed an oxygen-dependent decrease in G6Pase expression associated with enhanced histone H3 [K9] methylation. Collectively, these results indicate that hypoxia directly and indirectly impairs G6Pase expression through enhanced methylation of histone H3 [K9].

Nutritional mismatch in postnatal life of low birth weight rat offspring leads to increased phosphorylation of hepatic eukaryotic initiation factor 2 α in adulthood.

OBJECTIVE: Epidemiological studies have established that low birth weight offspring, when faced with a nutritional mismatch in postnatal life, have an increased risk of developing the metabolic syndrome. Our laboratory and others have demonstrated that maternal protein restriction (MPR) leads to high cholesterol and insulin resistance in the offspring due to impaired liver function, though the underlying molecular mechanisms remain elusive. Recent in vitro studies have associated decreased phosphorylation of Akt1 (Serine 473), a marker of insulin sensitivity, with increased phosphorylation of eukaryotic initiation factor (eIF)-2α (Serine 51), a key regulator of protein translation attenuation. The main aim of the study was to determine whether nutritional mismatch in MPR offspring leads to elevated phospho-eIF2α (Ser51) levels in the liver. MATERIALS/METHODS: To investigate if this occurs long-term in MPR offspring, pregnant Wistar rats were fed a control (20%) protein diet (control) or a low (8%) protein diet during pregnancy and postnatal life (LP1), or during pregnancy and lactation (LP2). RESULTS: At postnatal day 130, LP2 offspring exhibited increases in hepatic phosphorylation of eIF2α (Ser51) concomitant with decreases in the phosphorylation of Akt1 (Ser473), while LP1 offspring exhibited the converse relationship. Interestingly, in embryonic day 19 livers derived from control or MPR pregnancy, no changes in eIF2α (Ser51) or Ak1 (Ser473) phosphorylation were observed. CONCLUSION: Collectively, our data provide robust evidence that phosphorylation of eIF2α (Ser51) is inversely correlated with phosphorylated Akt1 (Ser473) in vivo. Moreover, this study demonstrates that this inverse relationship is adversely influenced in these MPR offspring by a mismatch in the postnatal nutritional environment.

Maternal protein restriction leads to enhanced hepatic gluconeogenic gene expression in adult male rat offspring due to impaired expression of the liver X receptor.

Epidemiological studies demonstrate that the link between impaired fetal development and glucose intolerance in later life is exacerbated by postnatal catch-up growth. Maternal protein restriction (MPR) during pregnancy and lactation in the rat has been previously demonstrated to lead to impaired glucose tolerance in adulthood, however the effects of protein restoration during weaning on glucose homeostasis are largely unknown. Recent in vitro studies have identified that the liver X receptor α (LXRα) maintains glucose homeostasis by inhibiting critical genes involved in gluconeogenesis including G6pase (G6pc), 11ß-Hsd1 (Hsd11b1) and Pepck (Pck1). Therefore, we hypothesized that MPR with postnatal catch-up growth would impair LXRα in vivo, which in turn would lead to augmented gluconeogenic LXRα-target gene expression and glucose intolerance. To examine this hypothesis, pregnant Wistar rats were fed a control (20%) protein diet (C) or a low (8%) protein diet during pregnancy and switched to a control diet at birth (LP). At 4 months, the LP offspring had impaired glucose tolerance. In addition, LP offspring had decreased LXRα expression, while hepatic expression of 11ß-HSD1 and G6Pase was significantly higher. This was concomitant with decreased binding of LXRα to the putative LXRE on 11ß-Hsd1 and G6pase. Finally, we demonstrated that the acetylation of histone H3 (K9,14) surrounding the transcriptional start site of hepatic Lxrα (Nr1h3) was decreased in LP offspring, suggesting MPR-induced epigenetic silencing of the Lxrα promoter. In summary, our study demonstrates for the first time the important role of LXRα in mediating enhanced hepatic gluconeogenic gene expression and consequent glucose intolerance in adult MPR offspring.

Permanent implications of intrauterine growth restriction on cholesterol homeostasis.

Susceptibility to disease begins during fetal life, and adverse events in utero are a critical factor in determining quality of life and overall health. In fact, up to 50% of metabolic syndrome diseases can be attributed to an adverse in utero environment. However, the mechanisms linking impaired fetal development to augmented cholesterol, an important clinical risk factor characterizing the metabolic syndrome and cardiovascular disease, remain elusive. This review discusses the latest research in the fetal programming of cholesterol homeostasis from both clinical studies and animal models. It also underscores the role of the placenta as an important mediator in cholesterol homeostasis during pregnancy and uncovers some of the molecular mechanisms underlying how the homeostatic mechanisms in liver may be impaired in fetal and postnatal life due to undernutrition and/or hypoxia.

Maternal protein restriction elevates cholesterol in adult rat offspring due to repressive changes in histone modifications at the cholesterol 7alpha-hydroxylase promoter.

Adverse events in utero, such as intrauterine growth restriction (IUGR), can permanently alter epigenetic mechanisms leading to the metabolic syndrome, which encompasses a variety of symptoms including augmented cholesterol. The major site for cholesterol homeostasis occurs via the actions of hepatic cholesterol 7α-hydroxylase (Cyp7a1), which catabolizes cholesterol to bile acids. To determine whether posttranslational histone modifications influence the long-term expression of Cyp7a1 in IUGR, we used a protein restriction model in rats. This diet during pregnancy and lactation led to IUGR offspring with decreased liver to body weight ratios, followed by increased circulating and hepatic cholesterol levels in both sexes at d 21 and exclusively in the male offspring at d 130. The augmented cholesterol was associated with decreases in the expression of Cyp7a1. Chromatin immunoprecipitation revealed that this was concomitant with diminished acetylation and enhanced methylation of histone H3 lysine 9 [K9,14], markers of chromatin silencing, surrounding the promoter region of Cyp7a1. These epigenetic modifications originate in part due to dietary-induced decreases in fetal hepatic Jmjd2a expression, a histone H3 [K9] demethylase. Collectively, these findings suggest that the augmented cholesterol observed in low-protein diet-derived offspring is due to permanent repressive posttranslational histone modifications at the promoter of Cyp7a1. Moreover, this is the first study to demonstrate that maternal undernutrition leads to long-term cholesterol dysregulation in the offspring via epigenetic mechanisms.