Untersuchungen zur Bedeutung des Transkriptionsfaktors Nrf2 für den Energiemetabolismus
Zusammenfassung der Projektergebnisse
The nuclear factor erythroid 2-related factor (Nrf2) is a transcription factor that functions as the key controller of the redox homeostatic gene regulatory network. Beyond its known function to protect cells against oxidative damage, there is evidence that it might also play an important role in protecting against metabolic dysregulation in chronic liver diseases. Therefore, the aim of this project was to analyze the link between metabolic processes in the liver and Nrf2-mediated antioxidative mechanisms. In order to induce a NASH, mice were fed with a high fat nutrition. We found that overall liver and microvesicular fatty droplets accumulation was very similar over time in Nrf2-/- mice compared to WT mice. Interestingly however body weight gain was significantly delayed in Nrf2-/- mice, which became evident as early as three weeks after starting the high fat feeding. Increased weight gain was associated with a more prominent abdominal white adipose tissue in WT mice in comparison to Nrf2-/- mice. Moreover, a detailed analysis of the hepatic lipid compounds 10 weeks after HFD treatment revealed a considerably stronger increase of hepatic triglycerides and higher levels of serum insulin as marker of glucose resistance in WT animals compared to Nrf2-/- mice after 10 weeks of HFD treatment. To further understand the underlying mechanism for the diminished hepatic TG accumulation in Nrf2-/- mice, RNA microarray analysis were performed. Direct comparison of gene expression profiles of WT and Nrf2-/- mice revealed 1364 genes that were differentially regulated. Using the functional annotation tool from DAVID analysis wizard, these genes were clustered into categories from which the most significant were involved in drug metabolism or metabolism of xenobiotics by cytochrome P450. Both clusters contained genes that are regulated in a Nrf2-dependent manner. Furthermore, loss of Nrf2 suppressed the upregulation of acyl-CoA thioesterases (Acots) that catalyze the hydrolysis of acyl-CoAs to the corresponding free fatty acid and coenzyme A. To determine the impact of Nrf2 on cellular energy homeostasis, mitochondrial function was analyzed in primary Nrf2-/- and WT hepatocytes using the XF Analyzer. Under untreated conditions loss of Nrf2 displayed no influence on the ATP-coupled respiration rate (oligomycin sensitive fraction) or the leak rate (oligomycin insensitive fraction) as well as on the maximal uncoupled rate induced by FCCP. To further investigate how elevated lipid concentrations affect mitochondrial respiration and ATP production, isolated Nrf2-/- and WT hepatocytes were either pretreated with 5 mM metformin or directly treated with 0.4 mM oleic acid. Metformin alone had no impact on the mitochondrial respiration, whereas oleic acid treatment was able to enhance oxygen consumption rate (OCR). While this effect was moderate in WT hepatocytes, loss of Nrf2 significantly increased OCR in comparison to the untreated control group. Together, our data reveal that activation of Nrf2 alters antioxidant, energy consumption-, and metabolism-related gene expression in the liver and delays diet induced obesity. Thus, the Keapl-Nrf2 system is an interesting target to prevent diet-induced obesity.
