4G, H), relative tgh expression is down-regulated (Fig. 5A). Conversely, in H2O2-treated larvae the expression of tgh was increased (Fig. 5J). Down-regulating TGH activity level by E600 (Fig. 5D) to the level in GMP synthetases850

Obeticholic Acid ic50 mutant larvae (Fig. 5B) was sufficient to induce hepatic steatosis (Fig. 5E-I), supporting the hypothesis that reduced tgh expression is responsible for hepatic steatosis in GMP synthetases850 mutant larvae. A previous study indicated that hepatic TG levels in Tgh-null mice were not statistically different from those in wild-type mice.[5] In mice, Tgh also acts in white adipose tissues[5] and it is likely that the absence of hepatic steatosis in tgh-null mice is due to decreased fatty acids delivery to the liver from adipose tissue, since isolated Tgh-null hepatocytes in culture accumulate more exogenous lipids than wild-type hepatocytes.[4] In contrast, zebrafish white adipose tissues only develop after 12 dpf,[29] potentially explaining why suppressing Tgh activity was sufficient to induced hepatic steatosis at 7 dpf. In GMP synthetases850 mutant larvae, expression of genes involved in de novo lipogenesis (srebp1, acc1, agapt, and fads2), β-oxidation (aco, cpt1, cyp4a10, and echs1) or lipid uptake (cd36) Dinaciclib in vivo are not significantly changed at 6 dpf (Supporting Fig. 11). These data also support the hypothesis that reduced tgh expression is responsible for hepatic steatosis in GMP synthetases850

mutant larvae. Under physiological conditions, ROS produced by β-oxidation of triglyceride-derived free fatty acids may provide feedback to influence Tgh activity, adjusting lipid dynamics in hepatocytes. ROS are recognized to play important roles in host defense, especially in the innate immune response of leukocytes to pathogens,[10] although the excessive production of ROS frequently results in inflammatory responses in many tissues, including the liver. In the liver, the two-hit model has been proposed for the transition of hepatic

steatosis to more severe NASH, in which the first hit is hepatic steatosis and the second hit is ROS-mediated inflammation.[30] Our data provide genetic evidence that physiological ROS levels are also necessary for the prevention of hepatic steatosis in zebrafish larvae (Fig. 6). The ability Phosphatidylinositol diacylglycerol-lyase of H2O2 to rescue hepatic steatosis in GMP synthetases850 mutant, Rac1 inhibitor-treated, and Tg (fabp10:GFP-DNRac1)lri4 larvae (Figs. 4, 6) further supports this idea. Our data do not, however, conflict with the current two-hit model or the notion that excess ROS production is pathological; rather, we propose that a reduction in physiological levels of ROS can be equally pathogenic to increased levels of ROS. These data suggest that proposed antioxidant supplementation for the treatment of NAFLD[31] would require careful dosage control to ensure that ROS levels are not reduced below their physiologically normal levels.