Pemphigus Autoantibodies Induce Blistering in Human Conjunctiva

Abstract

PURPOSE. The autoimmune blistering skin disease pemphigus vulgaris (PV) is caused by autoantibodies against desmosomal adhesion molecules. Patients may suffer conjunctival involvement, yet the underlying mechanisms are largely unclear. We characterized human and murine conjunctiva with respect to the PV autoantigens, and evaluated the effects and mechanisms of PV autoantibodies applied to human conjunctiva ex vivo. METHODS.We obtained human conjunctiva specimens from surgical explants and established a short-term culture model to study the alterations induced by antibody fractions of PV patients (PV-IgG). Furthermore, we applied a mouse model depleted of the desmosomal cadherin desmoglein 3 (Dsg3), the primary autoantigen in PV. Murine and human conjunctiva also was used to analyze the expression pattern of desmosomal proteins by immunostaining and Western blotting.RESULTS. Human and murine conjunctiva samples expressed the majority of desmosomal molecules with an expression pattern similar to the epidermis. Interestingly, Dsg3 knock out animals frequently suffer eye lesions, histologically evident as microblisters in the eyelid epidermis and conjunctiva. Incubation of human specimens with PV-IgG for 12 hours caused blistering in the suprabasal layers of the conjunctiva as well as reduction of Dsg1 and Dsg3 protein levels. Furthermore, PV-IgG prompted activation of p38MAPK in the conjunctiva, which is a central pathomechanism leading to blistering in the epidermis.CONCLUSIONS. PV-IgG leads to blister formation and p38MAPK activation in the conjunctiva and, thus, resembles the effects found in the epidermis. Our data indicate that the ocular involvement observed in PV patients is mainly based on conjunctival blistering.

Mechanisms Causing Loss of Keratinocyte Cohesion in Pemphigus

“…Immunoadsorption using recombinant Dsg1 and Dsg3 abolished pathogenic effects in different models in vitro and in vivo (Amagai et al, 1992;Heupel et al, 2008;Langenhan et al, 2014;Mahoney et al, 1999). Dsg3-specific knockout mice show lesions in the mucosa, conjunctiva, and hair follicles, which, on a histological level, resemble lesions in patients with PV (Koch et al, 1997(Koch et al, , 1998Rotzer et al, 2016;Vielmuth et al, 2016), suggesting that anti-Dsg3 antibodies cause the loss of Dsg3 function. Similarly, antibodies targeting Dsg1 and Dsg3 reduce keratinocyte cohesion of murine and human keratinocytes, at least under mechanical stress as in dissociation assays.…”

Desmoglein 1 Deficiency Causes Lethal Skin Blistering

“…However, antibodies against Dsg1 in contrast to those against Dsg3 where not directly interfering with Dsg binding (Heupel et al, 2008;Waschke et al, 2005). Deletion of Dsg3 without loss of Dsg1 function causes a mild pemphigus vulgaris-like phenotype in the skin and conjunctiva (Koch et al, 1997;Vielmuth et al, 2016), whereas epidermis-specific deletion of desmocollin 3, a desmosomal cadherin also expressed in basal epidermis, but rarely targeted by pemphigus autoantibodies, induces a severe pemphigus vulgaris phenotype (Chen et al, 2008). On the other hand, inactivation of Dsg3 with a specific autoantibody derived from a pemphigus vulgaris mouse model has been shown to cause skin blistering in vivo (Spindler et al, 2013;Tsunoda et al, 2003) and a monoclonal antibody against Dsg1 also induced superficial epidermal splitting in ex vivo human skin (Yoshida et al, 2017).…”

E-Cadherin Is Important for Meibomian Gland Function as Revealed by a New Human ex Vivo Slice Culture Model

“…17,18 Eyes together with eyelids of adult homozygous Dsg3 þ/þ (wild-type) or Dsg3 À/À (knockout) animals were enucleated, and 7-mm cryosections were prepared as described previously. 19 All animal experiments were approved by the Regierung von Oberbayern (number 55.2-1-54-2532-139-2014) and complied with the Association for Research in Vision and Ophthalmology Animal Statement for the Use of Animals in Ophthalmic and Vision Research.…”

“…To further analyze the correlation between differentiation and the formation of strong adhesive contacts in HMGECs, dispase-based dissociation assays were performed after blocking the interactions of certain adhesion molecules by inhibitory antibodies (Figure 3A). Because Dsg3-deficient (Dsg3 knockout) mice not only display a PV phenotype with lesions in the skin 16,28 but also develop conjunctional blistering, 19 the effect of AK23, a monoclonal pemphigus mouse model antibody against Dsg3, 29 and PV-IgG was first studied with autoantibodies against Dsg1 and Dsg3 from a patient with PV. Surprisingly, and in contrast to previous observations in human and mouse keratinocytes, 13,14,17 neither AK23 nor PV-IgG significantly affected intercellular cohesion in HMGECs.…”

“…Given the correlation between DED and pemphigus, our data suggest that ocular involvement in pemphigus is not caused primarily via MGD but rather via direct effects of autoantibodies on human conjunctiva as previously reported. 19 Nevertheless, besides direct impairment of desmosomal adhesion, it is well established that binding of pathogenic PV antibody modulates numerous intercellular signaling pathways (eg, p38 mitogen-activated protein kinase and protein kinase C). 42,43 Therefore, it cannot currently be ruled out that autoantibodies in pemphigus interfere with signaling pathways required for physiologic processes of meibocytes besides cell adhesion and lipid production.…”