Effects of Isochronal Induced Feed Restriction With Transition Period On mRNA Abundance of The Hepatic Genes Related to Lipid Metabolism in Fat-Tailed Ewes

The process of fat mobilization during the transition period requires deep re-orchestration of the energy indices, and understanding its mechanism has generated considerable interest among the TP-related studies. The present study aims to validate the effect of feed restriction and TP on the mRNA abundance of hepatic genes related to fat metabolism in fat-tailed sheep. Twenty pregnant ewes with the age of 40.8 ± 6.2 (mean ± standard error) month were randomly assigned to Control (n=10) or Restriction (n=10), and investigated from week -5 to 5 relative to parturition. Control animals received 100% DM during the trial. Restriction animals received 100% DM through weeks -5, -1, 1 and 5 and were fed with 50, 65, and 80% DM in the weeks -4, -3, -2 and 2,3, and 4, respectively. On the third week of experiment (65%) during both pre and post-partum, the hepatic tissue was biopsied, and the mRNA load of the fatty acid synthase, acetyl-CoA carboxylase, carnitine palmitoyltransferase (CPT) 1, CPT2, and acyl-CoA synthase long-chain family member-1 genes was quantified by the TaqMan qPCR technique. Data were analyzed using the Mixed Model procedure of SAS. The mRNA abundance of the target genes was not influenced by feed restriction, during the pre and postpartum periods. Parturition suppressed the mRNA abundance of target genes in both groups. It can be concluded that the liver of the fat-tailed sheep would have a higher capacity for the metabolism of free fatty acids mobilization during the feed insufficiency and the challenging period of transition.

Neonatal thyroxine activation modifies epigenetic programming of the liver

The type 2 deiodinase (D2) in the neonatal liver accelerates local thyroid hormone triiodothyronine (T3) production and expression of T3-responsive genes. Here we show that this surge in T3 permanently modifies hepatic gene expression. Liver-specific Dio2 inactivation (Alb-D2KO) transiently increases H3K9me3 levels during post-natal days 1–5 (P1–P5), and results in methylation of 1,508 DNA sites (H-sites) in the adult mouse liver. These sites are associated with 1,551 areas of reduced chromatin accessibility (RCA) within core promoters and 2,426 within intergenic regions, with reduction in the expression of 1,363 genes. There is strong spatial correlation between density of H-sites and RCA sites. Chromosome conformation capture (Hi-C) data reveals a set of 81 repressed genes with a promoter RCA in contact with an intergenic RCA ~300 Kbp apart, within the same topologically associating domain (χ2 = 777; p < 0.00001). These data explain how the systemic hormone T3 acts locally during development to define future expression of hepatic genes.

Growth Hormone Pulses and Liver Gene Expression Are Differentially Regulated by the Circadian Clock Gene Bmal1

In this study, we found that loss of the circadian clock gene Bmal1 causes disruptions throughout the growth hormone (GH) axis, from hepatic gene expression to production of urinary pheromones and pheromone-dependent behavior. First, we show that Bmal1 knockout (KO) males elicit reduced aggressive responses from wild-type (WT) males and secrete lower levels of major urinary proteins (MUPs); however, we also found that a liver-specific KO of Bmal1 (liver-Bmal1-KO) produces a similar reduction in MUP secretion without a defect in aggressive behavior, indicating that the decrease in elicited aggression arises from another factor. We then shifted our investigation to determine the cause of MUP dysregulation in Bmal1 KO animals. Because the pulse pattern of GH drives sexually dimorphic expression of hepatic genes including MUPs, we examined GH pulsatility. We found that Bmal1 KO males have a female-like pattern of GH release, whereas liver-Bmal1-KO mice are not significantly different from either WT or Bmal1 KO. Since differential patterns of GH release regulate the transcription of many sexually dimorphic genes in the liver, we then examined hepatic gene transcription in Bmal1 KO and liver-Bmal1-KO mice. We found that while some female-predominant genes increase in the Bmal1 KO, there was no decrease in male-predominant genes, and little change in the liver-Bmal1-KO. We also found disrupted serum insulin growth factor 1 (IGF-1) and liver Igf1 messenger RNA in the Bmal1 KO mice, which may underlie the disrupted GH release. Overall, our findings differentiate between GH-pulse–driven and circadian-driven effects on hepatic genes, and the functional consequences of altered GH pulsatility.

Maternal intake of alpha-lipoic acid prevents development of symptoms associated with a fructose-rich diet in the male offspring in Wistar rats

The hypothesis was that maternal intake of the antioxidant alpha-lipoid acid (ALA), during the developmental period of the hypothalamic orexigenic neurons, causes a permanent beneficial effect in offspring metabolism. Pregnant Wistar rats were fed with standard diet (food) + ALA (0.4% wt/wt) from day 14 of gestation to day 20 of lactation (n = 4) or food (n = 4). At 3 months of age, male offspring born from ALA-fed rats or controls (CT) were randomly assigned to be fed with food + 10% fructose solution in drinking water (F) or food + tap water (C), resulting in four groups: ALAF, ALAC, CTF, and CTC (n = 5/group). Food intake and body weight (BW) were measured twice a week for 31 days. Metabolites’ levels in blood, mRNA expressions of Npy, Agrp (hypothalamus), Fasn, Srebf1, Ppard, and Pparg (liver), and the antioxidant capacity of the liver were determined. Results significance was set at p < 0.05. Average BW gain, daily BW gain, and intraabdominal fat tissue at necropsy were higher in CTF group followed by CTC, ALAF, and ALAC groups. There were no differences between groups in Kcal intake per day. mRNA expressions of hypothalamic and hepatic genes and plasmatic levels of glucose and triglycerides were higher in CTF group followed by ALAF, CTC, and ALAC groups. Fructose intake affected the oxidative capacity of the liver, but this effect was not observed in the ALAF group. In conclusion, maternal ALA intake protected the adult offspring to develop metabolic symptoms associated with high fructose in the drinking water.

Direct differentiation of Dental Pulp Pluripotent-like Stem Cells differentiation into Hepatocyte-like Cells

Liver diseases is a major cause of morbidity and mortality. Dental pulp pluripotent-like stem cells (DPPSC) are a considerable promise in tissue engineering and regenerative medicine as a source of tissue-specific cells, therefore the aim of this study is to demonstrate their ability to generate functional hepatocyte-like cells in vitro. Cells were cultured in serum free-differentiation media supplemented with growth factors and cytokines to direct the differentiation into hepatocytes in a process that recapitulate liver development. At day 5, the differentiated DPPSC cells expressed the endodermal markers FOXA1 and FOXA2. Then, the cells were derived into hepatic lineage generating hepatocyte-like cells. In addition to the associated morphological changes, the cells expressed the hepatic genes HNF6, AFP. The terminally differentiated hepatocyte-like cells expressed the liver functional proteins albumin and CYP3A4. In this study, we report an efficient serum free protocol to differentiate DPPSCs into functional hepatocyte like cells. Our approach promotes the use of DPPSCs, as new source of stem cells, for prospective use in liver regenerative medicine.

NEONATAL THYROXINE ACTIVATION MODIFIES EPIGENETIC PROGRAMMING OF THE LIVER

In the neonatal liver, a peak of type 2 deiodinase (D2) activity accelerates local T3 production and the expression of thyroid hormone (TH)-responsive genes. Here we show that this acute increase in T3 signaling permanently modifies hepatic gene expression. Liver-specific Dio2 inactivation (Alb-D2KO) transiently increased H3K9me3 levels during post-natal days 1-5 (P1-P5) in discrete chromatin areas, and methylation of 1,508 DNA sites (H-sites) that remained in the adult mouse liver. These sites were associated with 1,551 areas of reduced chromatin accessibility (RCA; Atac-seq) within core promoters and 2,426 within intergenic regions, with reduction in the expression of 1,525 genes (RNA-seq). There was strong correlation between H-sites and RCA sites (r=0.85; p<0.0002), suggesting a cause-effect relationship. The analysis of chromosome conformation capture (Hi-C) data revealed a set of 57 repressed genes that have a promoter RCA in close contact with an intergenic RCA ~300 Kbp apart, including Foxa2 that plays an important role during development. Thus, the post-natal surge in hepatic D2 activity and TH-signaling prevents discrete DNA methylation and modifies the transcriptome of the adult mouse. This explains how the systemic T3 hormone acts locally during development to define future chromatin accessibility and expression of critically relevant hepatic genes.

Hepatic pyruvate carboxylase expression differed prior to hyperketonemia onset in transition dairy cows

Fatty acids (FA) provide an energy source to the liver during negative energy balance; however, when FA influx is excessive, FA can be stored as liver lipids or incompletely oxidized to β-hydroxybutyrate (BHB). The objectives of this study were to quantify plasma and liver FA profiles and hepatic gene expression in cows diagnosed with hyperketonemia (HYK; BHB ≥ 1.2 mM) or not (nonHYK; BHB < 1.2 mM) to determine a relationship between FA profile and expression of hepatic genes related to oxidation and gluconeogenesis. Production parameters, blood samples (-28, -3, 1, 3, 5, 7, 9, 11, and 14 d relative to parturition; n = 28 cows), and liver biopsies (1, 14, and 28 d postpartum; n = 22 cows) were collected from Holstein cows. Cows were retrospectively grouped as HYK or nonHYK based on BHB concentrations in postpartum blood samples. Average first positive test (BHB ≥ 1.2 mM) was 9 ± 5 d (± SD). Cows diagnosed with HYK had greater C18:1 and lower C18:2 plasma proportions. Liver FA proportions of C16:0 and C18:1 were related to proportions in plasma, but C18:0 and C18:2 were not. Some interactions between plasma FA and HYK on liver FA proportion suggests that there may be preferential use depending upon metabolic state. Cows diagnosed with HYK had decreased pyruvate carboxylase (PC) expression, but no difference at 1 d postpartum in either cytosolic or mitochondrial isoforms of phosphoenolpyruvate carboxykinase (PCK). The increased PC to PCK ratios in nonHYK cows suggests the potential for greater hepatic oxidative capacity, coinciding with decreased circulating BHB. Interestingly, FA, known regulators of PC expression, were not correlated with PC expression at 1 d postpartum. Taken together, these data demonstrate that HYK cows experience a decrease in the ratio of hepatic PC to PCK at 1 day postpartum prior to HYK diagnosis which, on average, manifested a week later. The differential regulation of PC involved in HYK diagnosis may not be completely due to shifts in FA profiles and warrants further investigation.