| PART Ⅰ THE IMPROVEMENT OF JAZF1 ON THE FORMATION OF AGING-INDUCED HEPATIC STEATOSISObjective: To investigate the effects of JAZF1 on the formation of aging-induced hepatic steatosis,and to explore the involved molecular mechanism.Methods: 8-week-old JAZF1 transgenic mice(JAZF1-Tg mice,n=15)and the age-matched C57 BL / 6J mice(WT mice,n=15) were fed on a normal chow diet for three periods(3 months, 6 months, 12 months) and was randomly selected five mice to sacrifice at the 3, 6, 12 months of the feeding period. Basic physiological metabolic indexes and m RNA and protein levels involved in the de novo synthesis and oxidation of fatty acid in the liver of each group were measured.Results: The hepatic TG accumulation was significantly increasedwith age(P <0.05); Compared with the age-matched WT mice, the hepatic TG accumulation in JAZF1-Tg mice was significantly reduced(P <0.05),and the serum TG and TC in the JAZF1-Tg mice fed for 12 months were significantly reduced(P <0.05); Compared with the age-matched WT mice,the m RNA levels of genes(SREBP-1c, SCD-1, FAS, ACC-1)involved in the de novo synthesis of fatty acid in the liver of JAZF1-Tg mice were significantly downregulated(P <0.05), while the m RNA levels of genes(PPAR-α, CPT-1) associated with fatty acid oxidation did not change significantly(p> 0.05); m RNA and protein levels of JAZF1 were downregulated with age, while the m RNA and protein levels of SREBP-1were upregulated with age;Conclusion: m RNA and protein levels of JAZF1 in the liver were downregulated with age, and JAZF1 overexpression improves the formation of aging-induced hepatic steatosis.PART Ⅱ THE IMPROVEMENT OF JAZF1 ON THE DEVELOPMENT AND PROGRESS OF FATTY LIVER INDUCED BY HIGH-FAT DIET.Objective: To investigate the effects of JAZF1 on the development and progress of fatty liver induced by high-fat diet, and to explore the involved molecular mechanism.Methods: 8-week-old JAZF1-Tg mice(n=30) and the age-matched WT mice(n=30) were randomly divided into two groups,respectively. One group was fed on a high fat diet(HFD), the other was fed on a normal chow diet(NC). Continuous feeding for 12 months, each group was randomly selected five mice to sacrifice at the 3, 6, 12 months of the feeding period.;Basic physiological metabolic indexes and m RNA and protein levels involved in the de novo synthesis and oxidation of fatty acid in the liver of each group were measured. AMPK / ACC signals in the liver of each period were analyzed.Results: The results of HE staining showed that the hepatic steatosis in the WT mice fed on a high-fat diet gradually increased, from small amount of visible lipid droplets at the first stage to diffuse steatosis, even to the emergence of bullous steatosis and infiltration of inflammatory cell at the last stage. However, compared with the age-matched WT mice fed onthe same high-fat diet, the results of basal metabolic indexs, liver TG and TC content, HE staining and oil red O staining showed that hepatic steatosis in the JAZF1-Tg mice were significantly improved. The m RNA levels of genes(SREBP-1c, SCD-1, FAS, ACC-1) involved in the de novo synthesis of fatty acid in the liver of JAZF1-Tg mice were significantly reduced, and protein expression of SREBP-1 and FAS were also significantly reduced(P <0.05). While the m RNA levels of genes(PPAR-α, CPT-1)associated with fatty acid oxidation, to a large extent, did not change significantly(p> 0.05).Compared with WT mice, AMPK signals were activated by increasing phosphorylation of.AMPK and ACC in the liver of JAZF1-Tg mice fed for 3 months and 6 months.Conclusion: JAZF1 overexpression can improve the development and progress of fatty liver induced by high-fat diet through reducing hepatic de nove lipogenesis.PART III ROLE AND RELATED MECHANISMS OF JAZF1 ON REDUCING HEPATOCYTE LIPIDACCUMULATIONObjective: To investigate the role of JAZF1 on hepatocyte lipid accumulation induced by FFAs, and to explore the involved molecular mechanism.Methods: Steatosis cell models were induced by 1m M of FFAs(PA:OA = 2: 1 ratio) in primary murine hepatocytes(PMH cells) and human hepatoma Hep G2 cells. m RNA and related proteins of the genes involved in de novo synthesis of fatty acid(SREBP-1c, SCD-1,FAS, ACC-1) were detected to explore the role of JAZF1 on hepatocyte lipid accumulation by JAZF1 overexpression virus(Ad-JAZF1) and JAZF1 inhibition virus(sh JAZF1) respectively;AMPK / ACC signals in the steatosis cell models treated with Ad-JAZF1 or sh JAZF1 were analyzed. At the same time,SREBP-1c gene promoter was generated to assess the regulation of JAZF1 on SREBP-1c gene promoter.by dual-luciferase reporter assay.Results: Steatosis cell models were successfully induced in PMH and Hep G2 cells; In both steatosis cells, compared with Ad-GFP group,JAZF1 overexpression in Ad-JAZF1 group reduced lipid droplet accumulation induced by FFAs, and the m RNA levels of genes involved in hepatocytesde novo synthesis(SREBP-1c, SCD-1, FAS, ACC-1) and related proteins(SREBP-1, FAS) were significantly lower(P <0.05); While compared with HK group,JAZF1 inhibition in sh JAZF1 group increased lipid droplet accumulation induced by FFAs, and the m RNA levels of genes involved in hepatocytes de novo synthesis(SREBP-1c, SCD-1, FAS, ACC-1) and related proteins(SREBP-1, FAS) were significantly up-regulated(P <0.05);Compared with Ad-GFP group, phosphorylation levels of AMPK and ACC were significantly induced in the two steatosis hepatocytes treated with Ad-JAZF1 virus; while compared with the HK group, phosphorylation levels of AMPK and ACC were significantly decreased in Hep G2 steatosis cells treated with sh JAZF1 virus; Although compared with the HK group,AMPK phosphorylation was not significantly decreased in PMH steatosis cells treated with sh JAZF1 virus, but the ACC phosphorylation was significantly decreased(P <0.05). In the basal state, JAZF1 overexpression had no effect on the luciferase activity of SREBP-1c gene promoter by dual luciferase reporter assay( p> 0.05), but in the state stimulated by insulin, JAZF1 overexpression significantly reduced the luciferase activity of SREBP-1c promoter induced by insulin(p <0.05).Conclusion:JAZF1 plays a significant role on reducing hepatocyte lipid accumulation induced by FFAs through activating AMPK signal. AndJAZF1 can significantly reduced the activity of SREBP-1c promoter induced by insulin. |
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