Abstract
Background and AimAtractyloside (ATR), a mitochondrial uncoupler, is known for its specific inhibition of mitochondrial oxidative phosphorylation. Previous studies have reported that moderate mitochondrial uncoupling effect is beneficial to increase the decomposition and clearance of hepatic lipid, prevent the occurrence of fatty liver diseases. Moreover, the beneficial effects of mitochondrial uncouplers on type 2 diabetes and metabolic syndromes have been consistently observed. The present study investigated the effect of ATR on steatosis level of HepG2 cells treated with free fatty acid (FFA).MethodsIntracellular triglyceride level and Oil Red O staining were assessed, the mitochondrial adaptation and ADP/ATP ratio were analyzed, the protein level of AMPK, mTOR and LC3B, autophagic flux, and the co-localization of LC3B with lipid droplets was performed.ResultsATR treatment inhibited the activity of mitochondrial respiratory chain complexes I and IV, decreased the mitochondrial membrane potential, and increased the ADP/ATP ratio in the FFA-treated cells. Furthermore, ATR increased the gene expression and protein level of LC3B and promoted the autophagic flux processing from early autophagosome to late autolysosome by increasing the protein level of AMPKα and decreasing the protein level of mTOR. An increased number of autophagosomes (LC3B) was also observed in the lipid droplets. ATR treatment accelerated lipid degradation in the FFA-treated cells, and the lowest lipid content was observed in the cell group with 7.5 μM ATR.ConclusionLow concentrations (2.5, 5, and 7.5 μM) of ATR treatment could activate autophagy to accelerate the degradation of TGs in steatosis HepG2 cells; the mechanism may be related to the activation of the AMPK/mTOR pathway induced by the increased ADP/ATP ratio. In addition, the ideal concentration of ATR for improving steatotic HepG2 cells was 7.5 μM.
Highlights
Fatty liver disease (FLD) induced by the accumulation of excessive lipids in hepatocytes may lead to steatohepatitis and even cirrhosis accompanied with obesity, hyperglycemia, and other metabolic syndromes (Neuschwander-Tetri, 2017)
The present study investigated the effect of ATR on steatosis level of HepG2 cells treated with free fatty acid (FFA)
ATR treatment inhibited the activity of mitochondrial respiratory chain complexes I and IV, decreased the mitochondrial membrane potential, and increased the ADP/ATP ratio in the FFA-treated cells
Summary
Fatty liver disease (FLD) induced by the accumulation of excessive lipids in hepatocytes may lead to steatohepatitis and even cirrhosis accompanied with obesity, hyperglycemia, and other metabolic syndromes (Neuschwander-Tetri, 2017). A novel class of liver-targeted mitochondrial uncoupling effects has been proven to increase hepatocellular energy expenditure and reverse metabolic and hepatic steatosis (Demine et al, 2019). The common characteristics of these new pharmacological treatments include ATP deprivation, activation of the energy-sensing enzyme of cells (i.e., adenosine 5monophosphate activated protein kinase or AMPK) (Smith et al, 2016), and increased level of autophagy (Zhang X. et al, 2018). Pharmacological treatment targeting mitochondrial energy expenditure to induce a moderate level of autophagy may be advantageous for preventing or reversing hepatic steatosis and the associated metabolic complications. Previous studies have reported that moderate mitochondrial uncoupling effect is beneficial to increase the decomposition and clearance of hepatic lipid, prevent the occurrence of fatty liver diseases. The present study investigated the effect of ATR on steatosis level of HepG2 cells treated with free fatty acid (FFA)
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