Antimitochondrial Antibody Positive And Negative Of Primary Biliary Cirrhosis Patients Serum Differences Between Proteomics Research

Objective:DIGE combining with MALDI-TOF/TOF MS was used to explore serological markers that differentially expressed between AMA positive and negative PBC patients.Method:Thirty PBC patients diagnosed according to the criteria of AASLD were recruited in this research. Twenty of them were AMA+PBC. The other ten were AMA--PBC. Both negative of AMA tested by indirect immuno fluorescence and AMA-M2tested by enzyme-linked immuno sorbent assay was defined as AMA-. Firstly, DIGE were applied to the screen cohort (8sex-, age-and pathological staging matched AMA+and AMA-PBC) and achieved differentially expressed spots of proteins in serum. Then these spots were identified by MALDI-TOF/TOF MS. Finally, Western blot were used to verify two of the differentially expressed proteins, SAP and VN, in all the sera of enrolled patients.Results:There were1840proteins spots found in patients with PBC. Twenty-eight of them were differentially expressed between AMA and AMA-PBC (>or<1.5times, P<0.05). And nine of them had a higher level in AMA PBC than AMA-PBC, while the others were lower. Finally, nine proteins were identified, SAP, Serum amyloid A, Platelet basic protein, vitamin D-binding protein isoform3, VN, AMBP, Inter-alpha (Globulin) inhibitor H2, Apolipoprotein A-IV and Complement factor H-related protein2. Results of western blot showed that expression level of VN were indeed higher in AMA+PBC (P<0.01) while no significant difference were found between AMA+and AMA-PBC (P=0.175).Conclusion:This is the first study that using proteomic technology to explore differentially expressed serological markers between AMA PBC and AMA-PBC, and DIGE combining with MALDI-TOF/TOF MS were confirmed as the reliable technology in screening serological markers. Higher level of VN were found in AMA-PBC, and that might be the reason that destruction of bile duct were more severe in AMA-PBC than AMA+PBC. Verification in lager cohort, especially AMA-PBC will help further confirm this results, meanwhile, analyzing the association between VN and the other biochemical/immunological indices will help clarify its clinical significance.