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Nonalcoholic steatohepatitis (NASH) is a chronic liver disease that can lead to cirrhosis, liver transplant, and even hepatocellular carcinoma. While liver biopsy remains the reference standard for disease diagnosis, analytical and clinical development of non-invasive soluble biomarkers of NASH are of great importance to advance the field. To this end, we performed analytical and clinical validation on a series of pro-inflammatory cytokines and chemokines implicated hepatic inflammation; IL-6, CRP, TNFα, MCP-1, MIP-1β, eotaxin, VCAM-1. Biomarker assays were validated for accuracy and precision. Clinical performance was evaluated in a random sample of 52 patients with biopsy-proven NAFLD/NASH. Patients were categorized into three groups according to their fibrosis stage; advanced (F3-F4), mild (F1-2) and no (F0) fibrosis. Serum IL-6 was increased in patients with advanced fibrosis (2.71 pg/mL; 1.26 pg/mL; 1.39 pg/mL p<0.01) compared to patients with mild or no fibrosis respectively. While, there was no significant difference noted in CRP, TNFα, MCP-1, MIP-1β, eotaxin among the three groups, VCAM-1 levels were increased by 55% (p<0.01) and 40% (p<0.05) in the advanced cohort compared to the mild and no fibrosis groups respectively. VCAM-1 also displayed good clinical performance as a biomarker of advanced fibrosis with an area under the receiver operating curve of 0.87. The VCAM-1 assay demonstrated robust accuracy and precision, and VCAM-1 outperformed IL-6, CRP, TNFα, and the chemokines MCP-1, MIP-1β, and eotaxin as a biomarker of advanced fibrosis in NASH. Addition of biomarkers such as IL-6 and VCAM-1 to panels may yield increased sensitivity and specificity for staging of NASH.

Assay validation and clinical performance of chronic inflammatory and chemokine biomarkers of NASH fibrosis