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Gata4 is a member of the zinc finger GATA transcription factor family and is required for liver development during the embryonic stage. Gata4 expression is repressed during NAFLD progression, however how it functions in this situation remains unclear. Here, Gata4 was deleted specifically in hepatocytes via Cre recombinase driven by the Alb promoter region. Under a high-fat diet (HFD) or methionine and choline deficient diet (MCD), Gata4 knockout (KO) male, but not female, mice displayed more severe NAFLD or NASH, evidenced by increased steatosis, fibrosis, as well as a higher NAS score and serum ALT level. The Gata4KO male liver exposed to a HFD or MCD had a reduced ratio of pACC/ACC, similar to the Gata4KO hepatocytes treated with palmitic acid. More cell apoptosis, which is associated with activated JNK signaling and inhibited NFκB signaling, was observed in the Gata4KO male liver and isolated hepatocytes. However, the inflammatory status in the Gata4KO male liver was similar to the control liver. Importantly, lower activation of AKT signaling in the liver, which is consistent with de-sensitized insulin signaling in isolated hepatocytes, was found in the Gata4KO male. In summary, our data demonstrated that loss of Gata4 in hepatocytes promoted NAFLD progression in male mice.

Background & aims Pregnant women may transmit their metabolic phenotypes to their offspring, enhancing the risk for nonalcoholic fatty liver disease (NAFLD); however, the molecular mechanisms remain unclear. Methods Prior to pregnancy female mice were fed either a maternal normal-fat diet (NF-group, "no effectors"), or a maternal high-fat diet (HF-group, "persistent effectors"), or were transitioned from a HF to a NF diet before pregnancy (H9N-group, "effectors removal"), followed by pregnancy and lactation, and then offspring were fed high-fat diets after weaning. Offspring livers were analysed by functional studies, as well as next-generation sequencing for gene expression profiles and DNA methylation changes. Results The HF, but not the H9N offspring, displayed glucose intolerance and hepatic steatosis. The HF offspring also displayed a disruption of lipid homeostasis associated with an altered methionine cycle and abnormal one-carbon metabolism that caused DNA hypermethylation and L-carnitine depletion associated with deactivated AMPK signalling and decreased expression of PPAR-α and genes for fatty acid oxidation. These changes were not present in H9N offspring. In addition, we identified maternal HF diet-induced genes involved in one-carbon metabolism that were associated with DNA methylation modifications in HF offspring. Importantly, the DNA methylation modifications and their associated gene expression changes were reversed in H9N offspring livers. Conclusions Our results demonstrate for the first time that maternal HF diet disrupted the methionine cycle and one-carbon metabolism in offspring livers which further altered lipid homeostasis. CpG islands of specific genes involved in one-carbon metabolism modified by different maternal diets were identified.

While the correlation between diabetes during pregnancy and birth defects is well-established, how hyperglycemia causes developmental abnormalities remains unclear. In this study, we developed a novel “hyperglycemic” chicken embryonic model by administrating various doses of glucose to fertilized eggs at embryonic stages HH16 or HH24. When the embryos were collected at HH35, the LD50 was 1.57 g/Kg under HH16 treatment and 0.93 g/Kg under HH24 treatment, indicating that “hyperglycemic” environments can be lethal for the embryos. When exposed to a dose equal to or higher than 1 g/Kg glucose at HH16 or HH24, more than 40% of the surviving chicken embryos displayed heart defects and/or limb defects. The limb defects were associated with proliferation defects of both the wing and leg buds indicated by reduced numbers of p-H3S10 labeled cells. These limb defects were also associated with ectopic apoptosis in the leg bud and expression changes of key apoptotic genes. Furthermore, glucose treatment induced decreased expression of genes involved in Shh-signaling, chondrogenesis, and digit patterning in the limb bud. In summary, our data demonstrated that a high-glucose environment induces congenital heart and limb defects associated with disrupted cell proliferation and apoptosis, possibly through depressed Shh-signaling.

Novel progress has been made to understand the adverse pathophysiology in the pancreas of offspring exposed to over-nutrition in utero. Our study is the first to evaluate whether the adverse effects of maternal over-nutrition on offspring β-cell function are reversible or preventable through pre-conception maternal diet interventions. Herein, offspring mice were exposed in-utero to one of the following: maternal normal-fat diet (NF-group), maternal high-fat diet (HF-group), or maternal diet transition from a HF to NF diet 9 weeks before pregnancy (H9N-group). Offspring mice were subjected to post-weaning HF-diet for 12 weeks. HF offspring, but not H9N, displayed glucose intolerance and insulin resistance. HF male offspring had enlarged islet β-cells with reduced β-cell density, whereas, H9N male offspring did not show these changes. Co-immunofluorescent (Co-IF) staining of glucose transporter 2 (Glut2) and insulin (Ins) revealed significantly more Glut2(+)Ins(−) cells, indicative of insulin degranulation, in HF male offspring, but not H9N. In addition, Co-IF of insulin and p-H3S10 indicated that β-cells of HF male offspring, but not H9N, had proliferation defects, likely due to inhibited protein kinase B (AKT) phosphorylation. In summary, our study demonstrates that maternal H9N diet effectively prevents functional deterioration of β-cells seen in HF male offspring by avoiding β-cell proliferation defects and de-granulation.

OBJECTIVES: Pregnant women may transmit their metabolic phenotype to offspring, enhancing the risk for non-alcoholic fatty liver disease (NAFLD) in the next generation; however, the molecular mechanisms remain unclear. In the present study, we hypothesized that the maternal HF diet disrupts one-carbon metabolism with a consequent methylation change in the offspring liver that contributes to a worsen NAFLD induced by postweaning high-fat (HF) diet. METHODS: To test our hypothesis, we used offspring mice exposed to either maternal HF diet (HF group), or an early transition to a maternal NF diet before pregnancy (H9N group), or maternal methyl donor supplement (H1S or H2S group), comparing to the offspring with normal-fat diet (NF group). RESULTS: The HF offspring displayed obesity, glucose intolerance and hepatic steatosis, the H9N offspring avoided all, while the H1S and H2S offspring released hepatic steatosis only. Data from the gene expression study suggested a disruption of pathways involved in the one-carbon metabolism in the HF offspring with a restoration in the H9N, the H1S and the H2S offspring. Furthermore, we showed a disruption of methionine cycle associated with DNA hypermethylation and a de-production of L-carnitine and PPAR-α in the HF, but not the H9N offspring. However, the H1S and H2S diet only normalized the methionine cycle and restored L-carnitine and PPAR-α content, but not rescued the DNA hypermethylation in offspring liver. Thus, we proved that the maternal HF diet disrupted the methionine cycle and further led to methylation changes for mitochondrial dysfunction and fatty acid oxidation responsible for the fatty liver, rather than for the global DNA hypermethylation, which might contribute to the obesity and glucose intolerance. CONCLUSIONS: Our study provides a novel mechanism to understand the lipid metabolism and epigenetics in transgeneration. More importantly, it suggested to us a potentially effective diet intervention strategy to reduce the transgeneration risk for NAFLD. FUNDING SOURCES: This project is supported by grants from the National Institutes of Health. This work is supported by the USDA National Institute of Food and Agriculture.

Introduction: Hyperglycemic conditions achieved during pregnancy have been shown to have detrimental effects to fetal development and increase the prevalence of childhood comorbidities. However, the mechanisms in which diabetic pregnancies affect placental development and subsequently contribute to adverse health effects on the mother and offspring remain unclear. Research design and methods: Streptozotocin was used to induce gestational diabetes in mice. In this model, hyperglycemia was established at embryonic day 3.5 (E3.5). Pregnancy mass was collected at E10.5, E12.5, E14.5, and E16.5 for different assessments. Results: Both placental and embryonic weights were found to be significantly elevated at E16.5. At E14.5, a significantly larger junctional zone with increased number of glycogen trophoblasts was found in the placentas from hyperglycemic pregnancies (HG group) compared to the placentas from normoglycemic pregnancies (NG group). Importantly, the HG placenta exhibited decreased trophoblast giant cell (TGC) association and TUNEL+ cells, and increased expression of α-SMA on the spiral artery, suggesting arterial remodeling was impacted. Moreover, the interhemal membrane of the labyrinth layer, was found to be thicker in the HG placentas. Furthermore, hyperglycemia resulted in more offspring congenital defects, which were associated with a thicker interhemal membrane. Conclusions: Together, these results suggest that gestational diabetes perturbs proper placental development and function, specifically spiral artery remodeling and angiogenesis, thereby negatively impacting embryonic development.