Soluble Fas Ligand Is Essential for Blister Formation in Pemphigus.

Roberta Lotti,Marika Quadri,Carlo Pincelli,En Shu,Ann Lin,Lorraine A O’Reilly,Tiziana Petrachi,Alessandra Marconi,Elisabetta Palazzo

doi:10.3389/fimmu.2018.00370

Abstract

Pemphigus is a blistering disease characterized by pemphigus autoantibodies (PVIgG) directed mostly against desmogleins (Dsgs), resulting in the loss of keratinocyte adhesion (acantholysis). Yet, the mechanisms underlying blister formation remain to be clarified. We have shown previously that anti-Fas ligand (FasL) antibody (Ab) prevents PVIgG-induced caspase-8 activation and Dsg cleavage in human keratinocytes, and that sera from pemphigus patients contain abnormally increased levels of FasL. Here, we demonstrate that recombinant FasL induces the activation of caspases prior to Dsg degradation, and anti-FasL Ab prevents acantholysis in cultured keratinocytes. Moreover, the silencing of FasL reduces PVIgG-induced caspase-8 activation and Dsg3 cleavage. Following injection of PVIgG into mice, FasL is upregulated at 1–3 h and is followed by caspase-8-mediated keratinocyte apoptosis, before blister formation. The administration of anti-FasL Ab after PVIgG injection blocks blister formation in mice. Furthermore, we injected PVIgG into two different gene-targeted mutant mice that selectively lack either secreted soluble FasL (sFasL), FasLΔs/Δs mice, or the membrane-bound form of FasL (mFasL), FasLΔm/Δm mice. After PVIgG treatment, blisters are only visible in FasLΔm/Δm animals, lacking mFasL, but still producing sFasL, similar to wild-type (C57BL/6) animals. By contrast, a significant decrease in the relative acantholytic area is observed in the FasLΔs/Δs animals. These results demonstrate that soluble FasL plays a crucial role in the mechanisms of blister formation, and blockade of FasL could be an effective therapeutic approach for pemphigus.

Highlights

Pemphigus is a rare and potentially lethal autoimmune skin disease, characterized by the loss of keratinocyte adhesion at the level of desmosomes, a phenomenon known as acantholysis
We demonstrate that the inhibition of Fas ligand (FasL) prevents apoptosis and acantholysis in vitro, and anti-FasL Ab blocks blister formation in vivo
Since PVIgG have been shown to induce the co-aggregation of FasL and Fas receptor with caspase-8 in death–inducingsignaling complex [7], we first analyzed the expression of Fas in pemphigus skin

Summary

Introduction

Pemphigus is a rare and potentially lethal autoimmune skin disease, characterized by the loss of keratinocyte adhesion at the level of desmosomes, a phenomenon known as acantholysis. This results in the formation of flaccid blisters and/or erosions in both skin and mucous membranes [1]. Autoantibodies play an essential role in the pathogenesis of pemphigus [3], the mechanisms leading to the formation of the blisters remain largely unknown [4, 5]. Several lines of evidence indicate that apoptosis is involved in the pathological mechanisms of pemphigus [reviewed by Grando et al, [6]]. Cleaved caspase-8 and -3 are detected in pemphigus lesions, and caspase-8-positive cells express Fas ligand (FasL)/Fas binding [7, 8]

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Results

Conclusion