Non-alcoholic fatty liver disease (NAFLD) is likely to be associated with elevated plasma branched-chain amino acids (BCAAs) and may precede the development of type 2 diabetes (T2D). We hypothesized that BCAAs may be involved in the pathogenesis of T2D attributable to NAFLD and determined the extent to which plasma BCAAs influence T2D development in NAFLD. We evaluated cross-sectional associations of NAFLD with fasting plasma BCAAs (nuclear magnetic resonance spectroscopy), and prospectively determined the extent to which the influence of NAFLD on incident T2D is attributable to BCAA elevations. In the current study, 5791 Prevention of REnal and Vascular ENd-stage Disease (PREVEND) cohort participants without T2D at baseline were included. Elevated fatty liver index (FLI) ≥60, an algorithm based on triglycerides, gamma-glutamyltransferase, body mass index (BMI) and waist circumference, was used as proxy of NAFLD. Elevated FLI ≥ 60 was present in 1671 (28.9%) participants. Cross-sectionally, BCAAs were positively associated with FLI ≥ 60 (β = 0.208, p < 0.001). During a median follow-up of 7.3 years, 276 participants developed T2D, of which 194 (70.2%) had an FLI ≥ 60 (log-rank test, p < 0.001). Cox regression analyses revealed that both FLI ≥60 (hazard ratio (HR) 3.46, 95% CI 2.45–4.87, p < 0.001) and higher BCAA levels (HR 1.19, 95% CI 1.03–1.37, p = 0.01) were positively associated with incident T2D. Mediation analysis showed that the association of FLI with incident T2D was in part attributable to elevated BCAAs (proportion mediated 19.6%). In conclusion, both elevated FLI and elevated plasma BCAA levels are associated with risk of incident T2D. The association of NAFLD with T2D development seems partly mediated by elevated BCAAs.

Non-alcoholic fatty liver disease is an increasing health issue that develops rather unnoticed with obesity, type 2 diabetes mellitus and metabolic syndrome. We investigated prevalence, determinants and associated metabolic abnormalities of non-alcoholic fatty liver disease in the largest population-based cohort to date. Biochemical characteristics, type 2 diabetes mellitus and metabolic syndrome were determined in the Lifelines Cohort Study (N = 167,729), a population-based cohort in the North of the Netherlands. Non-alcoholic fatty liver disease was defined as Fatty Liver Index (FLI)≥60. Exclusion criteria were age <18 years, immigrants, missing data to assess FLI and metabolic syndrome, excessive alcohol use, previous-diagnosed hepatitis or cirrhosis and non-fasting blood sampling. Out of 37,496 included participants (median age 44 years, 62.1% female), 8,259 (22.0%) had a FLI≥60. Individuals with a FLI≥60 were more often male, older, obese, had higher levels of hemoglobinA1c, fasting glucose, liver enzymes, total cholesterol, low-density lipoprotein cholesterol, triglycerides, c-reactive protein and leucocytes and lower high-density lipoprotein cholesterol (all P<0.0001). Participants with a FLI≥60 showed higher prevalence of type 2 diabetes mellitus (9.3% vs. 1.4%), metabolic syndrome (54.2% vs. 6.2%), impaired renal function (20.1% vs. 8.7%) and cardiovascular disease (4.6% vs. 1.6%) (all P<0.0001). Multivariable logistic analysis showed that smoking, hemoglobin, leucocytes, c-reactive protein, platelets, alanine aminotransferase, alkaline phosphatase, albumin, impaired renal function (OR 1.27, 95%CI 1.15–1.41), metabolic syndrome (OR 11.89, 95%CI 11.03–12.82) and its individual components hyperglycemia (OR 2.53, 95%CI 2.34–2.72), hypertension (OR 1.89, 95%CI 1.77–2.01) and reduced high-density lipoprotein cholesterol (OR 3.44, 95%CI 3.22–3.68) were independently associated with suspected non-alcoholic fatty liver disease (all P<0.0001). Twenty-two percent (22.0%) of the population in the North of the Netherlands is suspected to suffer from non-alcoholic fatty liver disease, coinciding with a significant increased risk of type 2 diabetes mellitus, metabolic syndrome, cardiovascular disease and impaired renal function.

Non-alcoholic fatty liver disease (NAFLD) is featured by increased plasma very low density lipoproteins (VLDL). The extent to which plasma apolipoprotein E (ApoE) levels are elevated in NAFLD is unclear. We determined whether plasma ApoE is elevated in subjects with suspected NAFLD. Plasma ApoE and genotypes were determined in 6,762 participants of the Prevention of Renal and Vascular End-Stage Disease (PREVEND) cohort. A Fatty Liver Index (FLI) ≥ 60 was used as a proxy of NAFLD. A total of 1,834 participants had a FLI ≥ 60, which coincided with increased triglycerides, non-HDL cholesterol, ApoB and ApoE (all P<0.001). In multivariable linear regression analysis, plasma ApoE levels were positively associated with an elevated FLI when taking account of ApoE genotypes and other clinical and laboratory covariates (fully adjusted model: β = 0.201, P<0.001). Stratified analysis for ApoE genotypes (ApoE ε3ε3 homozygotes, ApoE ε2 carriers, and ApoE ε3ε4 and ε4ε4 carriers combined), also showed positive associations of plasma ApoE levels with an elevated FLI in each group (all P<0.001). In conclusion, it is suggested that NAFLD is characterized by increased plasma ApoE levels, even when taking account of the various ApoE genotypes. Increased plasma ApoE may contribute to altered VLDL metabolism and to increased atherosclerosis susceptibility in NAFLD.