• HCC
• Insulin resistance
• Liver disease
• liver transplantation
• NAFLD

The liver is a key organ in whole body metabolism and waste handling and small-molecule metabolites have been proven to be excellent markers of liver function. Hepatic lipotoxicity due to elevated free fatty acid (FFA), inflammatory adipokines, impaired β-oxidation, together with insulin resistance (IR), lead to metabolic dysregulation, mitochondrial impairment and oxidative stress. These events have a significant role also in determining hepatocyte damage and result in profound changes in hepatic inflammation and gene expression, leading not only to fatty liver disease (FLD), but also to fibrosis, apoptosis and ultimately to hepatocellular carcinoma (HCC). The aims of this thesis were to identify, by “omics” Mass Spectrometry (MS) techniques, non-invasive biomarkers of hepatic dysfunction and tissue damage in subjects with different degree of liver disease from fatty liver, to fibrosis to HCC and to analyze if these biomarkers could change after liver transplantation.
The thesis was developed in three work packages (WPs). In WP1 I have analyzed if metabolomics indexes could be used as markers of the severity of non alcoholic fatty liver disease (NAFLD). By analyzing plasma samples of 45 subjects with biopsy proven NAFLD, I have identified indexes of de novo lipogenesis and amino acid metabolism associated with the severity of liver disease. I have also demonstrated that adipose tissue (Adipo-IR), hepatic (Hep-IR) and peripheral (HOMA-IR) insulin resistance indexes were increased in those with higher degree of steatosis and/or fibrosis. In the same WP I have developed a new metabolic index (GSG_index = glutamate/(serine+glycine)) that was associated with worsening of liver fibrosis. This metabolic index is the expression of increased oxidative stress/glutathione and TCA cycle activities and is increased proportionally to fibrosis.
In WP2, I evaluated if the indexes discovered in WP1, and associated with liver fibrosis in NAFLD, were also altered in subjects with HCC. Adipo-IR and HOMA-IR were increased proportionally to the severity of liver disease. Also the DNL and the new GSG index were increased in patients with HCC compared to those with NAFLD.
In WP3 I have analyzed if and how metabolomic markers of chronic liver disease change after liver transplantation (LT). I have analyzed the plasma metabolites of 22 subjects before LT and during follow up at 3, 6, and 12 months after surgery. I have found that after LT there was an improvement in Adipo-IR and HOMA-IR already 3 months after surgery and this improvement was maintained at 6 and 12 months. In addition, I have observed an improvement in fatty acid composition and aromatic amino acid concentrations but no changes in DNL and GSG indexes.
In conclusion, severity of liver disease is strongly associated with several biomarkers discovered through metabolomics analysis and with the degree of insulin resistance, not only at the level of the liver but also of muscle and adipose tissue. In addition, I identified a set of metabolomic indexes able to mark differences in degree of liver fibrosis and presence of HCC, indicating metabolic dysfunction as one of the major risk factor for liver disease.