Basic Application Research On Nonalcoholic Fatty Liver Disease

Background/aimsNonalcoholic fatty liver disease(NAFLD) refers to a clinicopathological condition characterized by significant lipid deposition in hepatocytes of the liver parenchyma that is unrelated to a history of excessive alcohol consumption.The septum of NAFLD ranges from simple steatosis to steatohepatitis,fibrosis,and cirrhosis.With an increase in the prevalence of obesity,type 2 diabetes mellitus,hypertension and dyslipidemia, the prevalence of NAFLD has rapidly increased in recent years and has become a significant worldwide public health concern.However,the risk factor for NAFLD has not been fully elucidated yet.Serum uric acid level has been suggested to be associated with factors that contribute to the metabolic syndrome.As a hepatic manifestation of the metabolic syndrome,whether NAFLD was associated with serum uric acid level remains unclear.The field of biomarker is another area of fast growing interest in the setting of NAFLD.All of the current diagnostic methods,including imaging methods and liver biopsy,have some significant limitations.Thus,it is necessary to find a novel non-invasive approach for the diagnosis of NAFLD.In addition,hepatic steatosis is a major risk factor for liver surgery and transplantation due to the increased susceptibility to ischemia/reperfusion injury.Ischemic preconditioning could protect the livers from ischemia/reperfusion injury.Nevertheless,it remains unclear whether or not the protective effect was influenced by the grade of steatosis. This dissertation consists of three parts.In the first part,a cross-sectional study was performed to investigate the potential association between serum uric acid levels with NAFLD.In the second part,we combined SELDI-TOF-MS and bioinformatics to discover new biomarkers and establish serum protein fingerprint models for the early diagnosis of NAFLD.In the third part,we analyzed the impact of steatosis on hepatic ischemia/reperfusion injury and protective effect of ischemic preconditioning in mice.MethodsA cross-sectional study was performed and the potential association between serum uric acid and NAFLD was investigated.SELDI-TOF-MS combined with bioinformatic approaches were applied to analyze serum samples from 35 NAFLD patients and 35 controls so as to find new biomarkers and establish serum protein fingerprint models for early diagnosis of NAFLD.The protective effect of ischemic preconditioning against ischemia/reperfusion injury was assessed in graded steatotic mouse livers induced by methionine/choline-deficient diet and potential related mechanisms were studied.ResultsThe cross-sectional study included 8925 subjects(6008 men) with a mean age of 43 years.The prevalence rates of NAFLD and hyperuricemia were 11.78%and 14.71%, respectively.NAFLD patients had significantly higher serum uric acid levels than controls(370.3±86.6 vs.321.1±82.6μmol/L;P＜0.001).The prevalence rate of NAFLD was significantly higher in subjects with hyperuricemia than in those without hyperuricemia(24.75%vs.9.54%;P＜0.001),and the prevalence rate increased with progressively higher serum uric acid levels(P_trend＜0.001).Multiple regression analysis showed that hyperuricemia was associated with an increased risk of NAFLD(odds ratio [OR]:1.291,95%confidence interval[CI]:1.067-1.564;P＜0.001).The first 50 serum samples were analyzed using SELDI-TOF-MS where 135 differentially expressed proteins peaks(59 up- and 76 down-regulated) were observed in NAFLD patients compared to controls(P＜0.01).With the aid of bioinformatic tools, we established a biomarker pattern with a specificity of 88.0%and a sensitivity of 92.0%for NAFLD.The remaining 20 samples were analyzed on the second day and served as an independent experimental set.The analysis of this independent experimental set yielded a specificity of 80.0%and a sensitivity of 70.0%.Hepatic tissues showing lean,mild,moderate,and severe steatosis were successfully induced by methionine/choline-deficient diet.Serum aminotransferases and hepatic histological analysis showed that 75 min ischemia and 2 h reperfusion caused significant hepatic injury,and the severity was correlated with the grade of steatosis. Ischemic preconditioning significantly protected lean and mildly steatotic livers from ischemia/reperfusion injury,while this protective effect was not observed in moderately and severely steatotic livers.Additional experimentation showed that ischemic preconditioning could not preserve the hepatic ATP content that may be a reason for the loss of the protective effect in moderately and severely steatotic livers.ConclusionsOverall,our study indicated that serum uric acid level was significantly associated with NAFLD,and elevated serum uric acid level was an independent risk factor for NAFLD.Serum protein fingerprint analysis using SELDI-TOF-MS technique combined with bioinformatic approaches can help to uncover novel biomarkers for NAFLD.Our results also demonstrated that hepatic steatosis is a significant factor that influenced the protective effect of ischemic preconditioning against ischemia/reperfusion injury.These findings may have important contributions for a better understanding of NAFLD.