OBJECTIVETo study the mechanism of liver fibrogenesis and to find new non-invasive biomarkers.

METHODIn this study, we used subcellular proteomic technology to study the plasma membrane proteins related to immune or alcohol induced liver fibrosis. Rat liver fibrosis models were induced by pig serum or alcohol injection. The liver fibrogenesis were detected by James's staining in the rat models after 2, 4, 6 and 8 weeks of treatment. The liver plasma membrane (PM) of the 2- and 8-week treatment model rats were enriched by two-step sucrose density gradient centrifugation. The purity of PM was verified by western blotting, and the plasma membrane proteins were extracted and analyzed by 2 DE. The differentially expressed proteins were identified by LC-MS/MS. Cellular location and function of these identified differential protein were classified.

RESULTSImmune or alcohol induced liver fibrosis rat models were successfully established. Liver plasma membrane was significantly enriched after sucrose density ultracentrifugation treatment. 87 differential protein spots were find out by 2DE combined with LC-MS/MS from the liver plasma membrane proteins of the 2- and 8-week treatment rat models, which corresponded to 30 non-redundant proteins including annexin A2, keratin 8 and keratin 18.

CONCLUSIONSA list of differentially expressed proteins relate to liver fibrosis were successfully identified. Differential proteins such as annexin A2, keratin 8 and keratin 18 could be new biomarkers for liver fibrosis diagnosis.

Alcohols ; adverse effects ; Animals ; Female ; Keratin-18 ; metabolism ; Keratin-8 ; metabolism ; Liver ; metabolism ; pathology ; Liver Cirrhosis ; chemically induced ; immunology ; metabolism ; pathology ; Male ; Proteome ; metabolism ; Rats ; Rats, Sprague-Dawley

To evaluate the clinical significance of autoantibodies to three major epithelial cytokeratins (CK) -- CK8, CK18, and CK19 -- we compared 66 patients with toluene diisocyanate (TDI)-induced asthma (group I) with three control groups: 169 asymptomatic exposed subjects (group II), 64 patients with allergic asthma (group III), and 123 unexposed healthy subjects (group IV). Serum IgG, specific for human recombinant CKs, were measured by ELISA (enzyme linked immunosorbent assay), and ELISA inhibition tests were performed. The existence of these antibodies was confirmed by IgG immunoblot analysis. Anti-TDI-HSA (human serum albumin) IgE and IgG antibodies were measured by ELISA in the same set of the patients. The prevalence of CK8, CK18, and CK19 auotantibodies in group I was significantly higher than in the other three groups. Results of the ELISA inhibition test showed significant inhibition with the addition of three CKs in a dose-dependent manner. No significant association was found between CK autoantibodies and the prevalence of anti- TDI-HSA IgG and IgE antibodies. These results suggest that autoantibodies to CK18 and CK19 can be used as serologic markers for identifying patients with TDI-induced asthma among exposed workers.
Toluene 2,4-Diisocyanate/*toxicity ; Sensitivity and Specificity ; Occupational Diseases/chemically induced/*diagnosis ; Middle Aged ; Male ; Keratins/*immunology ; Keratin-8/immunology ; Keratin-19/immunology ; Keratin-18/immunology ; Immunoblotting ; Humans ; Female ; Enzyme-Linked Immunosorbent Assay ; Biological Markers/blood ; Autoantibodies/*blood ; Asthma/chemically induced/*diagnosis ; Adult