Investigating Inflammatory Pathways in Alcohol-Associated Liver Disease

Alcohol-associated liver disease (ALD) is a global health issue, with limited treatment options. The pathogenic mechanisms involve many biochemical pathways and processes. Some of the key factors include iron overload, reactive oxygen species (ROS) production and mitochondrial dysregulation. This project aimed to investigate the role of alcohol and iron in inducing liver damage and toxicity.

HepG2 (VL-17A) cells were treated with alcohol (200 mM, 300 mM, and 350 mM) with/without iron (50 μM) over 72 hrs and markers of oxidative damage, cell death and mitochondrial function were assessed. The protective effects of antioxidants and nanoformulations were also measured. Subsequent studies also investigated the association between frequency of drinking, liver iron and liver fat in a UK BioBank cohort.

Results show 350 mM ethanol led to a 50% decrease (p<0.0001) in cell viability at 72 hrs and a significant increase in ROS at 30 mins (p=0.0027). At 72 hrs a substantial number of cells were late apoptosis (44%) (p=0.0153) as well as a 47% reduction (p=0.0160) in mitochondrial membrane potential after 350 mM ethanol treatment. Treatment with iron produced similar effects, whereby, 350 mM ethanol + 50 μM iron decreased cell viability by 63% at 72 hrs (p=0.0448) and increased ROS by 125% (p=0.0014) at 2 hrs. Pretreatment with 10 μM curcumin nanoformulations increased viability by 78% (p=0.0405), as well as reducing ROS by up 51% (p=0.0013). In the UK BioBank cohort, increased frequency of drinking showed significant associations with liver iron and liver fat (p<0.0001).

In summary, alcohol alone or in combination with iron is associated with significant liver injury, which can be ameliorated with antioxidants and nanoformulations. In addition, UK BioBank data showed that frequency of drinking was associated with higher levels of liver iron. Future studies can target antioxidant-based formulations in the prevention of mitochondrial damage due to ethanol and iron.