SSA/Ro antigen expressed on membrane of UVB-induced apoptotic keratinocytes is pathogenic but not detectable in supernatant of cell culture

Abstract

Objectives: To better understand the potential effect of ultraviolet light on the photosensitivity of patients with lupus erythematosus (LE), to elucidate the mechanisms of SSA/Ro antibody formation after UV exposure, and to investigate the role of this autoantibody in the pathogenesis of skin lesions.

Methods: Primary human keratinocytes were cultured in Medium-154. After ultraviolet-B light (UVB) irradiation, the keratinocytes were treated with affinity-purified anti-SSA/Ro sera and stained with FITC-labeled goat-anti-human IgG and propidium iodide (PI), followed by enzyme digestion with RNase, RNase-free DNase or RNase plus DNase. As target cells, the irradiated keratinocytes were incubated with affinity-purified anti-SSA/Ro sera, with or without fresh human sera as complement. The supernatants of irradiated keratinocytes were analyzed with ELISA method for SSA/Ro antigens.

Results: UVB irradiation induced apoptotic blebs on the cell surface. The blebs were composed of ribonucleoproteins and contained SSA/Ro antigens. SSA/Ro antigens expressed on UVB irradiated keratinocytes bound to affinity-purified anti-SSA/Ro sera, leading to complement-dependent cytotoxicity. However, no SSA/Ro antigens were detected in the supernatants.

Conclusions: SSA/Ro, a ribonucleoprotein antigen expressed on UVB irradiated keratinocytes, may be recognized and presented to immune cells by a direct cell-cell contact other than be eliminated into the circulation.