Novel biomarker candidates to predict hepatic fibrosis in hepatitis C identified by serum proteomics

Background: Liver biopsy remains the gold standard to assess hepatic fibrosis. It is desirable to predict hepatic fibrosis without the need for invasive liver biopsy. Proteomic techniques allow unbiased assessment of proteins and might be useful to identify proteins related to hepatic fibrosis. Aims: We utilized two different proteomic methods to identify serum proteins as candidate biomarkers to predict hepatic fibrosis stage in patients with chronic hepatitis C virus (HCV) infection. Methods: Serum was obtained from 24 people with chronic HCV at time of liver biopsy and from 6 normals. Liver biopsy fibrosis was staged 1-4 (Batts-Ludwig). Pooled serum samples (six in each of four fibrosis groups and controls) were analyzed with 4- and 8-plex isobaric tags for relative and absolute quantitation (iTRAQ), determining protein identification (ID) and ratios of relative protein abundance. Nonpooled samples were analyzed with two-dimensional (2-D) gels and difference in gel electrophoresis (DIGE) comparing different samples on the same gel and across gels. Spots varying among groups were measured with densitometry, excised, digested, and submitted for tandem mass spectrometry (MS/MS) protein ID. Results: iTRAQ identified 305 proteins (minimum 99% ID confidence); 66 were increased or decreased compared with controls. Some proteins were increased or decreased for specific fibrosis scores. From 704 DIGE protein spots, 66 were chosen, 41 excised, and 135 proteins identified, since one gel spot often identified more than one protein. Conclusions: Both proteomic methods identified two proteins as biomarker candidates for predicting hepatic fibrosis: complement C4-A and inter-alpha-trypsin inhibitor heavy chain H4.