Non Alcoholic Fatty Liver Disease: non invasive markers of severity and new experimental treatments

Non Alcoholic Fatty Liver Disease (NAFLD) is a worldwide increasing disease but still many questions about its evolution, the need of a screening and the availability of effective specific treatments are open. Aims of this PhD project were: 1) the evaluation of NAFLD natural history in a subgroup of NAFLD affected diabetic patients enrolled during the daily clinical activity of a splenohepatology ecoDoppler laboratory in order to identify, if present, predictive factors of “evolutive NAFLD”; 2) the experimental evaluation, in High Fat Diet (HFD) fed rats, of the potential therapeutic effect of 3 molecules targeting respectively: a) lipid metabolism (Apolipoprotein A analogue compound -L4F), b) insulin sensitivity (peroxisome proliferator activated receptor delta agonist –PPARd agonist) c) endothelial function (EET Analog). We developed two studies: a clinical observational study and an experimental study. Clinical study: 100 patients with type 2 diabetes were evaluated as far as steatosis is concerned. Among them, 80 had sonographic signs of steatosis. There was no difference in the prevalence between male and female patients. 21 type 2 diabetic patients with liver steatosis were reevaluated after 6 years without any specific treatment. Liver steatosis increases only in less than 1/3 of non-obese diabetic patients and demonstrates that in the majority of them sonographic degree of steatosis improves or recovers concurrently with biohumoral parameters. The presence of increased levels of serum AST and ferritin and lower pulsatility index of haepatic artery seems to be correlated to a worse prognosis and may be used to identify those patients who deserve a higher surveillance. Experimental study: 30 male Wistar rats (4-5 weeks old, 150 grams body weight) were purchased from Charles River Laboratories. 24 rats have been fed with HFD for 8 weeks. After 8 weeks of diet animals have been divided in 4 groups: 7 untreated (HFD); 7 treated with L4F (L4F), 7 treated with PPARd agonist (PPARd) and 3 treated with EET Analog (EET). Treatments lasted 6 weeks. We demonstrate that HFD induced NAFLD reproduces splanchnic haemodynamic alteration of liver steatosis in humans and shows an activation of innate immune system also at early degree of steatosis without hepatic inflammation and fibrosis. The activation of innate immune system can be evaluated by the analysis of lipopolysaccharide (LPS) stimulated/unstimulated CC motif chemockine ligand 2 (CCL2) production in cultured peripheral blood mononuclear cells (PBMCs). PPARd agonist and L4F improved HFD induced liver steatosis and reduced CCL2 production in PBMCs but preserved the ability of PBMCs to react to LPS stimulation EETA administration didn’t improved liver steatosis and further decreased portal vein velocity and reduced the ability of PBMCs to react to LPS stimulation.