Stephanie Goletz

Autoantibodies targeting α6β4 integrin have been identified in individual patients with mucous membrane pemphigoid (MMP). Reactivity against α6 integrin has been associated with oral lesions, while anti-β4 integrin reactivity has been linked to ocular involvement. However, the pathogenic effects of these antibodies have not been fully elucidated. Here, we investigated the pathogenic potential of anti-α6 and anti-β4 integrin IgG both in vitro and in vivo. Immune complexes of anti-α6 and anti-β4 integrin induced the release of reactive oxygen species from normal human leukocytes and stimulated CXCL2 secretion in cultured murine C5N keratinocytes. In vivo, repeated injections of IgG against a recombinant fragment of β4 integrin into C57BL/6 mice led to palpebral conjunctival swelling and mild oral lesions. The latter was observed following injection of IgG against a recombinant fragment of α6 integrin. Histopathological analysis revealed subepithelial inflammatory infiltrates without evidence of split formation. Direct immunofluorescence microscopy showed linear deposits of IgG at the basement membrane zone in most tissues, whereas C3 deposition was largely absent. This lack of complement activation was corroborated by a complement fixation assay, which confirmed that IgG against α6 and β4 integrin failed to induce C3 deposition in normal murine conjunctivae, buccal mucosa, or skin. Collectively, these findings indicate that IgG autoantibodies against α6 and β4 integrin exhibit pathogenic activity in vitro and induce mild disease in vivo, possibly due in part to relatively inefficient complement activation in this model.

This multicentre study compares four immunoassays for detecting antilaminin 332 IgG in mucous membrane pemphigoid. The keratinocyte footprint assay showed the highest diagnostic accuracy, while the laminin 332 Biochip® provided excellent specificity as a standardized and widely accessible diagnostic option.

Zusammenfassung: Hintergrund und Zielsetzung: Die Pemphigoid‐Erkrankungen werden primär durch IgG‐ oder IgA‐Autoantikörper gegen die kutane Basalmembranzone (BMZ) vermittelt. Obwohl rezente Arbeiten auf die Existenz einer ausschließlich IgM‐vermittelten Pemphigoid‐Erkrankung hindeuten, fehlt bislang eine umfassendere Untersuchung. Patienten und Methodik: In dieser prospektiven multizentrischen Studie wurden zehn Patienten mit ausschließlich IgM‐Ablagerungen entlang der BMZ in der direkten Immunfluoreszenz (IF) eingeschlossen. Zirkulierende IgM‐Autoantikörper wurden mittels indirekter IF und Immunoblot mit rekombinantem BP180 nachgewiesen. Ergebnisse: Die Kohorte umfasste vier Frauen und sechs Männer mit einem medianen Alter von 77 Jahren, die überwiegend Prurigo‐artige Hautveränderungen ohne Blasenbildung und Schleimhautbeteiligung aufwiesen. In der indirekten IF auf NaCl‐separierter humaner Spalthaut zeigten die Seren von neun Patienten lineare IgM‐Ablagerungen am Dach der artifiziellen Blase. Bei vier Patienten konnte eine IgM‐Reaktivität gegen BP180 nachgewiesen werden, bei keinem jedoch gegen BP230. Bei keinem der Kontrollpatienten (n   =   100) wurden IgM‐Ablagerungen an der BMZ in der direkten IF beobachtet. Eine IgM‐Reaktivität gegen die BMZ im Serum wurde bei 1 von 30 Kontrollpatienten mit chronischem Pruritus beziehungsweise bei 3 von 60 Kontrollpatienten ohne chronischen Pruritus nachgewiesen. Schlussfolgerungen: Das IgM‐Pemphigoid ist durch gewebsgebundene, exklusive IgM‐Autoantikörper gegen die BMZ, serologische IgM‐Reaktivität mit BP180 als primärem Zielantigen, überwiegend nichtbullösen klinischen Phänotyp, fehlende Schleimhautbeteiligung und eher milden Krankheitsverlauf gekennzeichnet. Unsere Ergebnisse deuten darauf hin, dass das IgM‐Pemphigoid eine eigenständige Entität darstellen könnte.

We evaluated a Biochip™-based immunofluorescence assay for improved diagnosis of dermal-binding pemphigoid diseases by detecting autoantibodies against laminin β4, type VII collagen and laminin 332. Results showed high sensitivity, with anti-p200 pemphigoid (laminin β4 reactivity) being more frequent than epidermolysis bullosa acquisita, suggesting underdiagnosis in clinical practice. The Biochip mosaic is a valuable tool for the parallel detection of key autoantibodies, potentially enhancing diagnostic accuracy and patient management.

Summary: Background and Objectives: Pemphigoid diseases are primarily mediated by IgG or IgA autoantibodies against the cutaneous basement membrane zone (BMZ). Although recent observations suggest the existence of exclusively IgM‐mediated pemphigoid disease, a larger study is lacking. Patients and Methods: This prospective multicenter study included ten patients with exclusive IgM deposition along the BMZ by direct immunofluorescence (IF). Circulating IgM was detected by indirect IF and immunoblotting with recombinant BP180. Results: The cohort, four females and six males with a median age of 77 years, presented predominantly with prurigo‐like lesions without blisters and mucosal involvement. Sera from nine patients demonstrated linear IgM labeling the blister roof of human salt‐split skin by indirect IF. Four patients yielded IgM reactivity against BP180, but none against BP230. In none of the controls (n = 100), anti‐BMZ IgM was seen by direct IF. 1/30 and 3/60 controls with or without pruritic non‐pemphigoid diseases revealed serum IgM reactivity against the BMZ, respectively. Conclusions: IgM pemphigoid is characterized by exclusive tissue‐bound anti‐BMZ IgM, serum anti‐BMZ IgM reactivity with BP180 as main target antigen, a predominantly non‐bullous clinical phenotype, absence of mucosal involvement, and a rather mild disease course. Our findings indicate that IgM pemphigoid may represent a distinct entity.

The 16th non‐collagenous domain (NC16A) of BP180 is the main antigenic target of autoantibodies in bullous pemphigoid (BP) and mucous membrane pemphigoid (MMP). Commercially available assays detect serum autoantibodies against NC16A in the majority of BP (80%–90%) and in approximately 50% of MMP patients. However, a standardized test system for detecting antibodies against other regions of BP180 is still lacking. Moreover, anti‐BP180 autoantibodies have been found in neurological conditions such as multiple sclerosis and Parkinson disease. This study aimed at identifying primary epitopes recognized by BP autoantibodies on the BP180 ectodomain. Serum samples of 51 BP and 30 MMP patients both without anti‐NC16A reactivity were included along with 44 multiple sclerosis and 75 Parkinson disease sera. Four overlapping His‐tagged proteins covering the entire BP180 ectodomain (BP180(ec)1–4) were cloned, expressed, purified and tested for reactivity by immunoblot. IgG antibodies to BP180(ec)3 were detected in 98% of BP, 77% of MMP and 2% of normal human sera. Only weak reactivity was detected for neurological diseases against BP180(ec)1, BP180(ec)2 and BP180(ec)4, in 3%, 11% and 7% of tested multiple sclerosis sera, respectively. 8% of Parkinson disease sera reacted with BP180(ec)2 and 9% with BP180(ec)4. In conclusion, this study successfully identified epitopes recognized by BP autoantibodies outside the NC16A domain in pemphigoid diseases. These findings contribute to a better understanding of the immune response in BP and MMP with potential implications for a future diagnostic assay for NC16A‐negative pemphigoid patients.

Anti-p200 pemphigoid is an autoimmune blistering disease that often resembles other pemphigoid disorders. We developed a test for the detection of autoantibodies against laminin β4, an autoantigen in anti-p200 pemphigoid, to facilitate its diagnosis. The test demonstrated excellent sensitivity and specificity, making it a valuable tool for diagnosing this disease and distinguishing it from similar skin disorders.