Biomarker evaluation

The biomarker determinations were performed using commercially available laboratory kits (XT-1800i, Sysmex for platelets; Vitros 5600, Ortho-Clinical Diagnostics, Johnson & Johnson for biochemical parameters). The noninvasive serum biomarker analysis included APRI and FIB-4, which were calculated according to the published analytic recommendations [6, 7], as follows:

Additionally, modified APRI (M-APRI) and modified FIB-4 (M-FIB-4) were calculated by including BMI z-scores in both formulas, as follows:

A novel simple biomarker, B-AST = BMI z-score × AST (IU/L), was also proposed and calculated.

Liver biopsy was performed percutaneously using a Menghini needle (Hepafix kit 1.4 or 1.6 mm, Braun). An experienced pathologist who was unaware of the clinical data performed the histopathological evaluation. Fibrosis staging was evaluated using the METAVIR scoring system on a 5-point scale (F0 – no fibrosis; F1 – portal fibrosis without septa; F2 – portal fibrosis with few septa; F3 – numerous septa without cirrhosis; and F4 – cirrhosis) [11]. Fibrosis was considered significant if the METAVIR F score was ≥2. Liver steatosis was determined semi-quantitatively according to the percentage of hepatocytes containing fat droplets and was staged as follows: 0 – no steatosis; 1 – minimal (≤ 5% hepatocytes affected); 2 – mild (6–33%); 3 – moderate (34–66%); and 4 – severe (> 66%). Steatosis was considered significant if more than 33% of the hepatocytes were affected (steatosis score > 2).