The objective of the study was to identify the keratinocyte antigens recognized by circulating antibodies from patients with pemphigus vulgaris (PV) and pemphigus foliaceus (PF) and to further identify the appropriate substrate(s) for immunoblot detection of these auto-antibodies. Antigenic specificity of 29 PV and 18 PF sera was determined by immunoblot assay on different protein extracts obtaining from human epidermis (HE), bovine tongue epithelium (BT), confluence (CK) and stratified (SK) keratinocyte cell cultures as the source of desmosomal antigens including desmoplakins I/II, desmoglein 1 (Dsg1), desmoglein 3 (Dsg3) and plakoglobin. The 130 kD-Dsg3 was identified by PV sera on all 4 extracts with the highest reactivity on the SK extract (n = 26), 23 cases on HE and 21 sera on CK, and BT. While the 160 kD- Dsg1 was detectable on all 4 extracts by a specific monoclonal antibody, the majority of PF sera (12 out of 18) labeled Dsg1 on the BT extract; 8 on HE and only 4 on CK and SK extracts. Thirteen of the 29 PV sera also labeled the 160 kD-Dsg1, in addition to Dsg3, on the BT or HE extract. PF sera labeled the 130 kD-Dsg3 in only one case. Affinity purified IgG antibodies from the 130 kD antigen further showed an intercellular staining pattern of basal human keratinocytes by immunofluorescence microscopy while the whole PV serum, reacting with the both Dgs1 and Dsg3, stained all the epidermis. A subset of PV (n = 18) and PF (n = 10) sera also contained IgG auto-antibodies against a 190 kD desmosomal plaque antigen. This study provides evidence for the presence of a multiple, auto-immune response against desmosomal antigens in pemphigus and furthermore demonstrates the variable presence of desmosomal antibody reacting sites between the CK, SK, BT and HE protein extracts. Accordingly, the protein extracts obtained from SK and BT are the appropriate substrates for the serological diagnosis of PV and PF, respectively.
Identification of desmosomal antigens recognized by pemphigus vulgaris and pemphigus foliaceus IgG autoantibodies using protein lysates derived from various epithelial cells and tissues
COZZANI, EMANUELE CLAUDIO;
1997-01-01
Abstract
The objective of the study was to identify the keratinocyte antigens recognized by circulating antibodies from patients with pemphigus vulgaris (PV) and pemphigus foliaceus (PF) and to further identify the appropriate substrate(s) for immunoblot detection of these auto-antibodies. Antigenic specificity of 29 PV and 18 PF sera was determined by immunoblot assay on different protein extracts obtaining from human epidermis (HE), bovine tongue epithelium (BT), confluence (CK) and stratified (SK) keratinocyte cell cultures as the source of desmosomal antigens including desmoplakins I/II, desmoglein 1 (Dsg1), desmoglein 3 (Dsg3) and plakoglobin. The 130 kD-Dsg3 was identified by PV sera on all 4 extracts with the highest reactivity on the SK extract (n = 26), 23 cases on HE and 21 sera on CK, and BT. While the 160 kD- Dsg1 was detectable on all 4 extracts by a specific monoclonal antibody, the majority of PF sera (12 out of 18) labeled Dsg1 on the BT extract; 8 on HE and only 4 on CK and SK extracts. Thirteen of the 29 PV sera also labeled the 160 kD-Dsg1, in addition to Dsg3, on the BT or HE extract. PF sera labeled the 130 kD-Dsg3 in only one case. Affinity purified IgG antibodies from the 130 kD antigen further showed an intercellular staining pattern of basal human keratinocytes by immunofluorescence microscopy while the whole PV serum, reacting with the both Dgs1 and Dsg3, stained all the epidermis. A subset of PV (n = 18) and PF (n = 10) sera also contained IgG auto-antibodies against a 190 kD desmosomal plaque antigen. This study provides evidence for the presence of a multiple, auto-immune response against desmosomal antigens in pemphigus and furthermore demonstrates the variable presence of desmosomal antibody reacting sites between the CK, SK, BT and HE protein extracts. Accordingly, the protein extracts obtained from SK and BT are the appropriate substrates for the serological diagnosis of PV and PF, respectively.
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