Alteration of YAP in the pathogenesis of Pemphigus Vulgaris

Abstract

Abstract: Background: Compelling evidence indicates that the desmosomal cadherin Desmoglein-3 (Dsg3) is the primary autoantigen in autoimmune blistering disease Pemphigus Vulgaris (PV) that affects both skin and mucous membrane. Many signal pathways have been shown to be involved in the disease pathogenesis in addition to the steric hindrance caused by autoantibodiestargeting Dsg3. However, despite many studies, the pathological mechanisms of blistering remain not fully elucidated and is still a subject under extensive and intensive research in the field. Interestingly, a recent report has shown that Dsg3 is also involved in the regulation of YAP (Yesassociated protein), the downstream effector of the Hippo pathway that controls tissue differentiation and organ size. This finding prompted to ask a question of whether alteration of YAP also occurs in pemphigus which might attribute to the disease process. Aim of study: This study aims to investigate the above mentioned question and evaluate YAP in the expression and distribution in keratinocytes exposed to PV sera and the pathogenic monoclonal antibody specifically binding to Dsg3. Methodology: The alteration in YAP expression and distribution will be examined in oral tissue samples of PV patients in vivo by immunohistochemistry. At least 30 PV cases with the matched controls of healthy individuals will be recruited for this study. In addition, 10 PV sera samples will be collected for the in vitro evaluation. Furthermore, a well-characterised pathogenic monoclonal antibody (AK23) against Dsg3 will be used to confirm the specific effect of antibody on YAP changes. Keratinocyte cultures will be subjected to the treatment of PV sera and AK23, respectively, alongside control sera from healthy individuals or isotype IgG for various time periods (e.g. 6&24 hours) and concentrations (e.g. 40% of PV sera and 1~100µg/ml of AK23) prior to immunofluorescence for YAP staining. All samples will be subjected to microscopic examination and the images acquired from both in vivo and in vitro studies will be analysed using software ImageJ (NIH, available online). The differences between PV and control groups will be subjected by Student’s t-test and for multiple groups’ comparisons obtained from in vitro experiments by one-way of ANOVA. Finally, the results will be presented as mean±SD in the figures along with the representative images. Expected outcomes and significance of the study: we anticipate the positive results obtained from this study that will reveal changes in YAP expression and distribution in PV as well as in keratinocyte cultures treated with PV sera and AK23. This potential novel finding may lead to further investigation of this pathway for a better understanding of the pathogenesis of this devastating disease. Keywords: pemphigus vulgaris; keratinocyte; desmoglein; YAP; Immunofluorescence.