The Analysis And Research Of Antibodies Against Melanocytes In Serum Samples From Patients With Vitiligo And Its Associated Melanocyte Antigens

To detect autoantibodies against melanocytes in serum samples of patients with vitiligo and to identify the associated membrane antigens. To explore the role of immune mechanism on the pathogenesis of vitiligo, and the cause of HBV infection complicated with vitiligo by detecting the levels of heat shock protein 70 and anti-tyrosinase related protein 1antibodies in serum samples of vitiligo patients and further analyzing their correlations.Normal human melanocytes were cultured in vitro. Indirect immunofluorescent assay was used to locate associated antigens. Western blot assay was performed to detect autoantibodies against membrane and cytoplasmic proteins of melanocytes in 50 cases of patients with vitiligo( including 4 cases with HBV infection) and 20 cases normal controls.Membrane antigens with higher frequencies were identified with protein mass spectrometry analysis. ELISA assay was performed to detect the heat shock protein 70 and anti-tyrosinase related protein 1 antibodies in serum samples from 70 patients with vitiligo( including 10 cases with HBV infection) and 26 normal cases.The specific antigens were detected in the cytoplasm and on the membrane of melanocytes. The membrane antigens were detected in 40 out of 50( 80%) randomly selected serum samples of vitiligo patients by western blot assay. The autoantibodies could react to several membrane antigens with approximate molecular weights of 86×103, 75×103, 60×103,52×103 and 44×103 Dalton, showing strip positive rates of 36%, 58%, 22%,2% and 2% respectively. The cytoplasmic antigens with molecularweights of 110×103, 90×103, 75×103, 50×103 and 40×103 Dalton were account for 30% of the 50 tested serum samples of vitiligo patients,showing strip positive rates of 12%, 4%, 12%, 10% and 2% respectively.The positive rates of membrane and cytoplasmic antigens were 15% and5% respectively for healthy subjects.Protein mass spectrometry analysis indicated that membrane proteins with the molecular weights of 86×103 and 75×103 Dalton might be lamin A/C and the membrane protein at a molecular weight of 60×103 Dalton was predicted as vimentin X1. High levels of anti-tyrosinase related protein 1 antibodies in serum samples were detected in patients with vitiligo in combination with HBV infection, which was confirmed to have a positive correlation with heat shock protein 70( r = 0. 927, P < 0. 01).A membrane antigen associated with vitiligo was newly identified which might provide evidence for the investigation on the autoimmune pathogenesis of vitiligo and the formation of autoantibodies. A preliminary study on the relationship between HBV infection and the pathogenesis of vitiligo, and the role of immune mechanisms in the pathogenesis of vitiligo was conducted through detecting the levels of heat shock protein 70 and anti-tyrosinase related protein 1 antibodies in serum samples.