The effects of bullous pemphigoid autoantibodies on apoptosis of keratinocyte and expression of BP180

Abstract

Bullous pemphigoid (BP) is an autoimmune blistering disorder characterized by tense inflammatory subepidermal bullae. BP is caused by binding of IgG autoantibodies to hemidesmosomal proteins BP180 and BP230. Independently from the autoantibodies involved, no study to date has examined direct effects of BP IgG on apoptosis of keratinocytes.

The aim of the present study was to investigate whether apoptosis is triggered by BP IgG binding to HaCaT keratinocytes. Also, this study examined whether BP IgG treatment can deplete BP180 of HaCaT keratinocytes. In MTT assay, treatment with BP IgG decreased the total number of HaCaT keratinocytes comparison with treatment with normal human (NH) IgG. HaCaT keratinocytes treated with 1 mg/ml of BP IgG for 72 hours showed 30 % survival rate whereas the cells treated with NH IgG showed 70 % survival rate. Furthermore, in Annexin V-FITC and PI staining, apoptosis was enhanced in cells treated with BP IgG compared to those treated with NH IgG. Up to 75.1 % apoptosis was observed in HaCaT keratinocytes treated with 1mg/ml of BP IgG for 72 hours. TUNEL staining assay also confirmed the apoptosis were induced in HaCaT keratinocytes after treatment of BP IgG. In immunoblotting, BP180 depletion was observed in BP IgG treated HaCaT keratinocytes.

To the best of our knowledge, the present report first demonstrates that BP IgG autoantibodies promote apoptosis in HaCaT keratinocytes. Apoptosis of HaCaT keratinocytes and depletion of BP180 may weaken the cell-cell adhesion and finally provoke detachment. This study suggests that BP IgG autoantibodies could contribute to the BP pathogenesis by triggering apoptosis of keratinocytes and depletion of BP180. Because apoptotic cell death usually does not provoke inflammatory response, our study provides additional support for an alternative non-inflammatory mechanism of autoantibody-induced tissue damage that could contribute to blister formation. Further studies will be needed to elucidate the detailed pathomechanism of keratinocyte apoptosis and depletion of BP180 induced by BP IgG.