Insulin Stimulates Goose Liver Cell Growth by Activating PI3K-AKT-mTOR Signal Pathway.

BACKGROUND/AIMS: Recent studies have suggested a crucial role for PI3K-Akt-mTOR pathway in regulating cell proliferation, so we hypothesize that insulin acts goose hepatocellular growth by PI3K-Akt-mTOR signal pathway. Because the physiological status of liver cells in vitro is different from that in vivo, a simplified cell model in vitro was established. METHODS: Goose primary hepatocytes were isolated and incubated in either no addition as a control or insulin or PI3K-Akt-mTOR pathway inhibitors or co-treatment with glucose and PI3K-Akt-mTOR pathway inhibitors; Then, cell DNA synthesis and cell cycle analysis were detected by BrdU-incorporation Assay and Flow cytometric analysis; the mRNA expression and protein expression of factors involved in the cell cycle were determined by Real-Time RT-PCR, ELISA, and western blot. RESULTS: Here we first showed that insulin evidently increased the cell DNA synthesis, the mRNA level and protein content of factors involved in the cell proliferation of goose primary hepatocytes. Meanwhile, insulin evidently increased the mRNA level and protein content of factors involved in PI3K-Akt-mTOR pathway. However, the up-regulation of insulin on cell proliferation was decreased significantly by the inhibitors of PBK-Akt-mTOR pathway, LY294002, rapamycin or NVP-BEZ235. CONCLUSION: These findings suggest that PI3K-Akt-mTOR pathway plays an essential role in insulin-regulated cell proliferation of goose hepatocyte.

Research Note: Integrative analysis of transcriptome and gut microbiome reveals foie gras capacity difference between cage and floor rearing systems.

To explore the differences in foie gras performance between geese raised in cages and on the ground, we conducted an integrative analysis of liver transcriptome and gut microbial metagenomes. The results showed extremely significant differences in the liver weight (P < 0.01) and liver lipid accumulation of FRS and CRS groups. The levels of triglyceride (TG), high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) of CRS were significantly higher than those of FRS (P < 0.05). Transcriptome analysis showed that 3,917 upregulated and 1,395 downregulated genes were identified, and lipid metabolism pathway and fatty acid metabolism were significantly enriched. Analysis of cecum microbiota revealed that several inflammation-related bacteria (including Gallibacterium, Escherichia-Shigella, Desulfovibrio, Alistipes, and Fournierella) were enriched in CRS, while beneficial bacteria (including Lactobacillus, Limosilactobacillus, and Ligilactobacillus) were significantly enriched in FRS. In conclusion, CRS was better than FRS in foie gras production, which was more conducive to lipid deposition in the goose liver.

Multi-omics reveals goose fatty liver formation from metabolic reprogramming.

To comprehensively provide insight into goose fatty liver formation, we performed an integrative analysis of the liver transcriptome, lipidome, and amino acid metabolome, as well as peripheral adipose tissue transcriptome analysis using samples collected from the overfed geese and normally fed geese. Transcriptome analysis showed that liver metabolism pathways were mainly enriched in glucolipid metabolism, amino acid metabolism, inflammation response, and cell cycle; peripheral adipose tissue and the liver cooperatively regulated liver lipid accumulation during overfeeding. Liver lipidome patterns obviously changed after overfeeding, and 157 different lipids were yielded. In the liver amino acid metabolome, the level of Lys increased after overfeeding. In summary, this is the first study describing goose fatty liver formation from an integrative analysis of transcriptome, lipidome, and amino acid metabolome, which will provide a whole new dimension to understanding the mechanism of goose fatty liver formation.

Lipidomics analysis reveals new insights into the goose fatty liver formation.

Our previous study described the mechanism of goose fatty liver formation from cell culture and transcriptome. However, how lipidome of goose liver response to overfeeding is unclear. In this study, we used the same batch of geese (control group and corn flour overfeeding group) to explore the lipidome changes and underlying metabolic mechanisms of goose fatty liver formation. Liquid chromatography-mass spectrometry (LC-MS) was provided to lipidome detection. Liver lipidomics profiles analysis was performed by principal component analysis (PCA), partial least squares-discriminant analysis (PLS-DA) and orthogonal partial least squares-discriminant analysis (OPLS-DA), different lipids were identified and annotated, and the enriched metabolic pathways were showed. The results of PCA, PLS-DA, and OPLS-DA displayed a clear separation and discrimination between control group and corn flour overfeeding group. Two hundred and fifty-one different lipids were yielded, which were involved in triglyceride (TG), diglyceride (DG), phosphatidic acids (PA), phosphatidylinositols (PI), phosphatidylethanolamines (PE), phosphatidylcholines (PC), lyso-phosphatidylcholines (LPC), monogalactosylmonoacylglycerol (MGMG), sphingolipids (SM), ceramides (Cer), and hexaglycosylceramides (Hex1Cer). Different lipids were enriched in glycerophospholipid metabolism, glycerolipid metabolism, phosphatidylinositol signaling system, inositol phosphate metabolism, glycosylphosphatidylinositol (GPI)-anchor biosynthesis and sphingolipid metabolism. In conclusion, this is the first report describing the goose fatty liver formation from lipidomics, this study might provide some insights into the underlying glucolipid metabolism disorders in the process of fatty liver formation.

Integrative analysis of transcriptome and lipidome reveals fructose pro-steatosis mechanism in goose fatty liver.

To further explore the fructose pro-steatosis mechanism, we performed an integrative analysis of liver transcriptome and lipidome as well as peripheral adipose tissues transcriptome analysis using samples collected from geese overfed with maize flour (control group) and geese overfed with maize flour supplemented with 10% fructose (treatment group). Overfeeding period of the treatment group was significantly shorter than that of the control group (p < 0.05). Dietary supplementation with 10% fructose induced more severe steatosis in goose liver. Compared with the control group, the treatment group had lower in ceramide levels (p < 0.05). The key differentially expressed genes (DEGs) (control group vs. treatment group) involved in liver fatty acid biosynthesis and steroid biosynthesis were downregulated. The conjoint analysis between DEGs and different lipids showed that fatty acid biosynthesis and steroid biosynthesis were the highest impact score pathways. In conclusion, fructose expedites goose liver lipid accumulation maximization during overfeeding.

Influence of different types of sugar on overfeeding performance-part of meat quality.

Previous research in our lab showed that 10% glucose, 10% fructose, and 10% sucrose can induce lipid deposition in goose fatty liver formation process more efficiently. However, whether the overfeeding diet supplement with sugar can affect the meat quality is unclear. The aim of this research was to estimate the meat quality of geese overfed with overfeeding diet adding with different types of sugar. The results indicated there were no significant differences in the diameter of muscle fiber, the muscle fiber density, pH0, pH24, the meat color, the cooking loss, the drip loss, the shear force and the dry matter in breast muscle and thigh muscle between corn flour groups and three sugars groups (P > 0.05). The crude fat content of breast muscle in fructose group was significantly higher than that in sucrose group (P < 0.05); the inosinic acid content of leg muscle in fructose group was significantly higher than that in the sucrose group (P < 0.05); the ratios of essential amino acids to total amino acids (EAA/TAA) in the breast muscle of maize flour group, fructose group, sucrose group and glucose group were 42%, 35%, 32% or 34%;57%, 64%, 64%, and 62%, respectively; the ratios of essential amino acids to total amino acids in leg muscle of maize flour group, fructose group, sucrose group and glucose group were 31%, 33%, 35%, and 34%, respectively. The contents of C16:1 and C18:1 n-9c in breast muscle in fructose group were significantly higher than that in sucrose group (P < 0.05). Compared with maize flour group, the contents of C18:0 and C20:0 were lower in leg muscle of sugar group (P < 0.05). Compared with the maize flour group, the activities of hydrogen peroxide (H2O2) and glutathione peroxidase (GSH-PX) in breast muscle were higher than those of sucrose group (P < 0.05), the total antioxidant capacity (T-AOC) levels in breast muscle was higher than that of fructose group and sucrose group (P < 0.05). Cluster analysis and principal component analysis (PCA) showed that there was no difference in meat quality between maize flour and sugar group. In conclusion, the overfeeding with maize flour supplement with 10% sugar had no evident influence on the meat quality.

Study on the effect of different types of sugar on lipid deposition in goose fatty liver.

Early research in our lab indicated that the effect of glucose, fructose and sucrose on the levels of triacylglycerol, and inflammatory factor was significantly different, and it is speculated that the regulatory mechanism of lipid deposition by different type of sugar in the liver is different. In order to explore lipid deposition difference mediated by different types of sugar (glucose, fructose, and sucrose) in goose fatty liver formation, this experiment was performed from cell culture, overfeeding experiment, and transcriptome analysis at 3 levels. Cell culture experiment results indicated that the levels of intracellular triglyceride, total cholesterol, and lipid content of fructose and sucrose treatment were significantly higher than those of glucose treatment (P < 0.05). In slaughter performance, the liver weight, the ratio of liver weight to body weight, feed conversion ratio (liver weight/feed consumption) were better in sucrose overfeeding group (P < 0.05). In addition, the liver of the sucrose overfeeding group contained a lot of unsaturated fatty acids, especially (n-3) polyunsaturated fatty acids (P < 0.05). Transcriptome analysis shown that the peroxisome proliferators-activated receptor (PPAR) signaling pathway is highly enriched in the fructose and sucrose overfeeding groups; cell cycle, and DNA replication pathways were highly enriched in the glucose overfeeding group. In conclusion, due to the decrease of lipids outward transportation and the anti-inflammation of unsaturated fatty acids, fructose, and sucrose have better ability to induce steatosis in goose fatty liver formation.

The Regulation of Lipid Deposition by Insulin in Goose Liver Cells Is Mediated by the PI3K-AKT-mTOR Signaling Pathway.

BACKGROUND: We previously showed that the fatty liver formations observed in overfed geese are accompanied by the activation of the PI3K-Akt-mTOR pathway and an increase in plasma insulin concentrations. Recent studies have suggested a crucial role for the PI3K-Akt-mTOR pathway in regulating lipid metabolism; therefore, we hypothesized that insulin affects goose hepatocellular lipid metabolism through the PI3K-Akt-mTOR signaling pathway. METHODS: Goose primary hepatocytes were isolated and treated with serum-free media supplemented with PI3K-Akt-mTOR pathway inhibitors (LY294002, rapamycin, and NVP-BEZ235, respectively) and 50 or 150 nmol/L insulin. RESULTS: Insulin induced strong effects on lipid accumulation as well as the mRNA and protein levels of genes involved in lipogenesis, fatty acid oxidation, and VLDL-TG assembly and secretion in primary goose hepatocytes. The stimulatory effect of insulin on lipogenesis was significantly decreased by treatment with PI3K-Akt-mTOR inhibitors. These inhibitors also rescued the insulin-induced down-regulation of fatty acid oxidation and VLDL-TG assembly and secretion. CONCLUSION: These findings suggest that the stimulatory effect of insulin on lipid deposition is mediated by PI3K-Akt-mTOR regulation of lipogenesis, fatty acid oxidation, and VLDL-TG assembly and secretion in goose hepatocytes.

RETRACTED: Role of mammalian sirtuin 1 (SIRT1) in lipids metabolism and cell proliferation of goose primary hepatocytes.

This article has been retracted: please see Elsevier Policy on Article Withdrawal (http://www.elsevier.com/locate/withdrawalpolicy). This article has been retracted at the request of the Authors. It has come to the attention of the corresponding author that there are two errors in Section 3.1 of the Results section titled "Effect of overfeeding on gene expression and enzyme activity of several genes in liver". The first error is that the article contains the wrong number of overfeeding days. The second error is that there are incorrect correlations between liver weight, lipids content in live and plasma metabolic substrates because of the wrong overfeeding days. The authors take responsibility for them and apologize to the readership of Molecular and Cellular Endocrinology.