The Study Of Goose Hepatocellular Steatosis Induced By Several Factors

The lipids deposition in liver can be affected by many factors. High energy food, glucose, insulin, cholesterol and LXRa agnonist all can induce hepatic steatosis in mammals. The feature of hepatic steatosis in waterfowl is different from that in mammals, and the mechanism of hepatic steatosis in waterfowls remains unclear yet. In this study, after overfeeding carbohydrate to Landes goose and Sichuan Landes goose, we measured the plasma level of glucose, insulin and cholesterol; lipid content in liver, hepatic TG content and mRNA expression of genes involved in the lipids metabolism in vivo; also measured the effect of glucose, insulin, cholesterol and LXRa agnonist on activities of lipogeneic enzymes FAS and ACC, total TG accumulation, intracellular and extracellular TG content and extracellular VLDL content, mRNA level of related genes in vitro; at last, we measured the SREBP-1 nuclear protein content using western blot andand the binding of SREBP-1 nuclear protein and SRE probe of ACCa gene using EMSA. There rsults are as followed:(1) Overfeeding induced the lipids deposition in goose liver, and the lipid deposition intensity of Lands goose was higher than that of Sichuan white goose; the increase of the plasma level of insulin and cholesterol induced by overfeeding showed a difference between the two breeds, and the increase of Lands goose was bigger than that of Sichuan white goose (P<0.05); the plasma glucose concentration increased in Landes goose after overfeeding, but decreased in Sichuan white goose (P<0.05). After overfeeding, the mRNA level of genes (ACCa, FAS, SREBP-1 and ChREBP)involved in the lipogenesis decreased in the two breeds, and the gene expression of LPL and LXRa increased (P＜0.05); the mRNA level of genes (MTTP) related with assembly and secretion of VLDL-TG decreased (P＜0.05); the change of mRNA level induced by overfeeding is different in the two breeds, and the decease of mRNA level of SREBP-1, ChREBP and MTTP in Lands goose is bigger than that in Sichuan white goose; the increase of mRNA level of LPL gene in Lands goose is bigger than that in Sichuan white goose.(2) Glucose and insulin cultured with hepatocytes separately evidently increase the activity of lipogenic enzyme FAS and ACC, mRNA level of lipogenic genes (FAS, ACCa, SREBP-1, ChREBP, LXRa and SCD1), and intracellular TG content (P<0.05). There was evident synergetic effect when glucose and insulin cultured together (P<0.05). In addition,5 mmol/L glucose and 50 nmol/L insulin cultured alone had no significant effect on the SREBP-1 nuclear protein levels and the binding of SREBP-1 nuclear protein with a target gene ACCa probe, but 5 mmol/L glucose and 50 nmol/L insulin cultured together significantly increased the binding (P<0.05). Glucose could increase the extracellular concentration of TG and VLDL (P<0.05), and evidently increase the mRNA level of genes related with VLDL-TG synthesis and secretion. Glucose and insulin cultured together with goose primary hepatocytes increased the mRNA level of DGAT1, DGAT2 and MTTP genes (P＜0.05), and the induction of high concentration of insulin and glucose was higher than the effect of low concentration of insulin and glucose (P<0.05); Glucose and insulin cultured together had synergetic effect on gene level of apoB, and the effect of low concentration of insulin and glucose was higher than the effect of high concentration of insulin and glucose(P<0.05); the effect of glucose and insulin cultured together on extracellelμlar TG and VLDL concentration was also higher than the effect when cultured with glucose and insulin seperatly (P<0.05).(3) The regulation mode of intracellular and extrcellular TG level, total TG level and extrcellular cholesterol level by cholesterol was similar with the regulation of mRNA level of lipogenic genes (FAS, ACCa, SREBP-1, ChREBP, LXRa and SCD1) and the genes involved in the assembly and secretion of VLDL-TG (DGAT2, MTTP and FoxO1).10μg/mL cholesterol up-regulated the level of the detected indexes; the induction by 20μg/mL cholesterol is most evident, and 30μg/mL cholesterol showed an inhibiting role (P＜0.05). The regulation of gene expression of DGAT1 and apoB by cholesterol is different from the regulation of other genes. Cholesterol could increase the mRNA level of DGAT1 gene in a dose-dependent manner, and had no evident effect on mRNA level of apoB gene (P>0.05).(4) LXR agonist (T0901317) increased intracellular TG content and total TG levels, activity of FAS and ACC, mRNA level of genes involved in lipogensis(FAS, ACCa, SREBP-1, ChREBP, LXRa and SCD1), and also increased the gene expression of LPL and DGAT1 genes in a dose-dependent manner. In addition, T0901317 could increase the concentration of SREBP-1 nuclear protein and the binding of SREBP-1 nuclear protein and its target gene ACCa probe. The regulation of extracellular concentration of TG and VLDL by T0901317 was similar with that of gene expression of MTTP, DGAT2, FoxO1 and apoB. With the increase of T0901317, the induction increased. But when the concentration of T0901317 reached 10μmol/L, the induction to gene expression decreased, and the gene level after the treatment of 10μmol/L T0901317 was lower than that in 1μmol/L T0901317.Conclusion:Overfeeding, culture with glucose, insulin, cholesterol and LXR agonist (T0901317) could induce the lipids deposition in goose hepatocytes by affecting the fatty acid and TG synthesis and VLDL-TG assembly and secretion, then could lead to hepatic steatosis; The induction of fatty acid synthesis by glucose, insulin and LXR agonist (T0901317) can be indirectly achieved by SREBP-1 activting transcription of its target gene ACCa.