Extrusion of pilosebaceous units into blisters of patients affected by a new variant of endemic pemphigus in El Bagre, Colombia, South America.

BACKGROUND: A new variant of endemic pemphigus foliaceus is present in El Bagre, Colombia, and surrounding municipalities (El Bagre-EPF) that affects the skin and in some presentations affects other organs with autoantibodies directed against cell junctions. METHODS: We studied 200 El Bagre-EPF patient perilesional skin biopsies, as well as 200 skin biopsies from normal controls in the endemic area. RESULTS: We observed blister extrusions of sebaceous glands or entire pilosebaceous units via the isthmus in 23% of the patients and not in the controls. CONCLUSIONS: The extrusion of hair follicular unit contents is consistent with our previous pathologic findings of autoreactivity to these units, and their observed clinical decrease in patients affected by El Bagre-EPF.

Membrane attack complex (C5b-9 complex or Mac), is strongly present in lesional skin from patients with endemic pemphigus foliaceus in El Bagre, Colombia.

BACKGROUND: El Bagre endemic pemphigus foliaceus (El Bagre-EPF) is a new variant of endemic pemphigus foliaceus present in the El Bagre area of Colombia, South America. Here, we investigate the presence of complement/C5-b9 in lesional skin of patients and matched controls from the endemic area. We also aim to compare the patient's autoantibody levels using indirect immunofluorescent titers (IIF) and correlate with the lesional presence of complement/C5b-9. METHODS: A case-control study was carried out by testing for the presence of complement/C5b-9 in lesional skin in 43 patients affected by El Bagre-EPF, as well as 43 matched, healthy controls from the endemic area. Skin biopsies were obtained and evaluated via hematoxylin and eosin staining, and immunohistochemistry. RESULTS: The presence of complement/C5b-9 was observed in all cases of the patients affected by El Bagre-EPF and was not observed in the controls from the endemic area (P < 0.001). The patients' autoantibody titers utilizing IIF for IgG and IgM showed correlation between higher autoantibody titers and stronger intensity of staining with complement/C5-b9 staining (P < 0.001). CONCLUSION: Patients affected by El Bagre-EPF have lesional deposition of complement/C5b, which correlates with disease severity and previously established serologies.

Patterns of Antinuclear Antibodies in a New Variant of Endemic Pemphigus in El Bagre, Colombia, Colocalizing with Antigens against MIZAP, ARVCF, p0071, and Desmoplakins I and II.

BACKGROUND: A new variant of endemic pemphigus foliaceus (EPF) has been documented, El Bagre-EPF. We aimed to study antinuclear antibodies (ANAs) in these patients. METHODS: We performed a case-control study, testing 57 patients affected by this disease and 57 controls from the endemic area matched by work activity and demographics. The participants were evaluated clinically as well as by detection of ANAs utilizing HEp-2 cells. We utilized Triton-induced partial permeabilization of the cell membranes, allowing for the visualization of intracellular and intranuclear antigens. We also immunoadsorbed the ANAs using synthetic peptides to elucidate the nature of the ANA. RESULTS: We detected the presence of a new pattern of ANAs. The new pattern of ANAs was seen in 24% of the El Bagre-EPF patients, compared to our controls (P < 0.001). The new ANA pattern consisted of a thin nuclear and nucleolar rim, finely speckled nucleolar, nuclear membrane pores stains, and a positive intranuclear stain directed against small nuclear components, as well as cytoplasmic deposits of autoantibodies were also observed. The new ANAs pattern perfectly colocalized with commercial antibodies to miocardium-enriched zonula occlusans-1 associated protein (MIZAP), armadillo repeat gene deleted in velo-cardio-facial syndrome (ARVCF), p0071 and desmoplakins I-II (all from Progen Biotechnik). Additionally in 14% of patients with El Bagre-EPF forme fruste and hyperpigmented clinical presentations, a classic homogeneous ANA pattern was observed with autoantibodies specific for Ro, La, Sm, and double-stranded DNA antigens. Immunoadsorption with peptide-based sequences from MIZAP, ARVCF, p0071 and desmoplakins I-II removed the new ANA pattern. CONCLUSIONS: We describe a new pattern of ANAs in El Bagre-EPF, colocalizing with autoantibodies directed against MIZAP, ARVCF, p0071, and desmoplakins I-II.

Neural system antigens are recognized by autoantibodies from patients affected by a new variant of endemic pemphigus foliaceus in Colombia.

BACKGROUND: Endemic pemphigus foliaceus (EPF), is also known as "fogo selvagem" or "wild fire," reflecting the intense burning sensation of the skin reported by patients with this disease. Based on this finding, we tested for neural autoreactivity in patients affected by a new variant of EPF (El Bagre-EPF). METHODS: We tested 20 El Bagre-EPF patients, 20 normal controls from the endemic area, and 20 age- and sex-matched normal controls from outside the endemic area. We tested for autoreactivity to several immunoglobulins and complement. Both human skin and bovine tail were used as antigens. RESULTS: We detected autoreactivity to neural structures, mechanoreceptors, nerves, perineural cell layers of the arachnoid envelope around the optic nerve, brain structures, and to neuromuscular spindles; these structures colocalized with several neural markers. The patient antibodies also colocalized with desmoplakins 1 and 2, with the armadillo repeat protein deleted in velo-cardio-facial syndrome and with p0071 antibodies. Autoreactivity was also found associated with neurovascular bundles innervating the skin, and immunoelectron microscopy using protein A gold against patient antibodies was positive against the nerve axons. Paucicellularity of the intraepidermal nerve endings and defragmentation of the neural plexus were seen in 70% of the cases and not in the controls from the endemic area (p<0.005). Neuropsychological and/or behavioral symptoms were detected in individuals from the endemic area, including sensorimotor axonal neuropathy. CONCLUSIONS: Our findings may explain for the first time the "pose of pemphigus," representing a dorsiflexural posture seen in EPF patients vis-a-vis the weakness of the extensor nerves, and furthermore, the autoreactivity to nerves in EPF could explain the "burning sensation" encountered in EPF disease.

Cardiac autoantibodies from patients affected by a new variant of endemic pemphigus foliaceus in Colombia, South America.

Several patients affected by a new variant of endemic pemphigus foliaceus in El Bagre, Colombia (El Bagre-EPF) have experienced a sudden death syndrome, including persons below the age of 50. El Bagre-EPF patients share several autoantigens with paraneoplastic pemphigus patients, such as reactivity to plakins. Further, paraneoplastic pemphigus patients have autoantibodies to the heart. Therefore, we tested 15 El Bagre-EPF patients and 15 controls from the endemic area for autoreactivity to heart tissue using direct and indirect immunofluorescence, confocal microscopy, immunohistochemistry, immunoblotting, and immunoelectron microscopy utilizing heart extracts as antigens. We found that 7 of 15 El Bagre patients exhibited a polyclonal immune response to several cell junctions of the heart, often colocalizing with known markers. These colocalizing markers included those for the area composita of the heart, such as anti-desmoplakins I and II; markers for gap junctions, such as connexin 43; markers for tight junctions, such as ezrin and junctional adhesion molecule A; and adherens junctions, such pan-cadherin. We also detected colocalization of the patient antibodies within blood vessels, Purkinje fibers, and cardiac sarcomeres. We conclude that El Bagre-EPF patients display autoreactivity to multiple cardiac epitopes, that this disease may resemble what is found in patients with rheumatic carditis, and further, that the cardiac pathophysiology of this disorder warrants further evaluation.

Autoantibodies to melanocytes and characterization of melanophages in patients affected by a new variant of endemic pemphigus foliaceus.

BACKGROUND: Melanin and melanophages are commonly seen under the basement membrane zone of the skin in patients affected by a new variant of endemic pemphigus foliaceus in El Bagre, Colombia (El Bagre-EPF). OBJECTIVE: Our study was conducted to determine the nature of these pigmentary alterations. METHODS: We utilized clinical, histopathologic and immunologic techniques including direct and indirect immunofluorescence, immunohistochemistry, Bielschowsky staining and immunoelectron microscopy studies. RESULTS: In the El Bagre-EPF patients, we detected dermal melanin in melanophages and antigen-presenting cells, in close proximity to neural and vascular markers. The melanophages consisted of a mixed population expressing CD68, myeloid/histoid antigen and S-100 protein. By immunoelectron microscopy, the presence of autoantibodies in proximity to melanin granules was confirmed within the melanocytes utilizing 10-nm gold particles. CONCLUSION: Dermal antigen-presenting cells, including melanophages, seem to contain a diverse combination of molecules, representative of an immunologic process where these cells are engulfing both autoantigens and/or cellular debris in El Bagre-EPF. Autoantibodies to discrete components of melanocytes were also identified; the clinical and immunologic significance of these findings remains unknown. Our work may provide a possible explanation of a darkened complexion in patients affected by endemic pemphigus foliaceus.

Antibodies to pilosebaceous units along their neurovascular supply routes in a new variant of endemic pemphigus foliaceus in Colombia, South America.

Senear Usher syndrome is a variant of pemphigus foliaceus, confined to seborrheic sites and considered to be a clinical overlap syndrome, with features of both pemphigus foliaceus and lupus erythematosus. We recently described autoantibodies to skin eyelid meibomian glands in patients with a new variant of endemic pemphigus foliaceus (El Bagre EPF) in South America. We tested for El Bagre EPF patient sera autoreactivity to pilosebaceous units utilizing direct and indirect immunofluorescence, confocal microscopy, immunohistochemistry and immunoelectron microscopy. Hematoxylin and eosin staining of skin biopsies revealed that one third of the patients affected by El Bagre-EPF demonstrated some histologic alteration of the pilosebaceous units. By immunohistochemistry, most El Bagre EPF biopsies demonstrated evidence of an autoimmune response along the neural and vascular supply routes of the pilosebaceous units. An active immune response was seen with antibodies such as anti-human mast cell tryptase, myeloid/histoid antigen, CD8, CD20, CD68, CD117/c-kit, ZAP-70 and vimentin. Immunoelectron microscopy demonstrated autoantibodies within the hair follicle and at the basement membrane area of the sebaceous glands. El Bagre-EPF patients have autoantibodies to pilosebaceous units and to their surrounding neurovascular packages. Our results warrant further characterization and may explain the loss of hair described in severe endemic pemphigus foliaceus before the therapeutic steroid era.

Human eyelid meibomian glands and tarsal muscle are recognized by autoantibodies from patients affected by a new variant of endemic pemphigus foliaceus in El-Bagre, Colombia, South America.

BACKGROUND: Previously, we described a new variant of endemic pemphigus foliaceus (EPF) in Colombia, South America (El Bagre-EPF). OBJECTIVE: Continuing our characterization of this variant of EPF, we now focus on one of our previously reported clinical findings: the presence of ocular lesions. These ocular lesions are seen in patients having extensive skin involvement, as measured by the Lund and Browder scale, which is generally used for patients with skin burns. METHODS: We specifically searched for evidence of autoreactivity to various eyelid structures in these patients and correlated our immunologic data with the clinical findings. We performed indirect immunofluorescence studies using normal-appearing human eyelid skin from routine blepharoplasties as substrate tissue. We tested sera from 12 patients with El Bagre-EPF and ocular lesions, 5 patients with sporadic (nonendemic) pemphigus foliaceus, and 20 healthy control subjects (10 from the El Bagre-EPF endemic area and 10 from nonendemic areas). We used fluorescein isothiocyanate conjugated goat antiserum to human total IgG/IgA/IgM as a secondary antibody. In addition, we used fluorescein isothiocyanate conjugated antibodies to human fibrinogen, albumin, IgG, IgE, C1q, and C3, Texas Red (Rockland Immunochemicals, Inc, Gilbertsville, PA), Alexa Fluor 555, or Alexa Fluor 594 (Invitrogen, Carlsbad, CA). Ki-67 (a cell proliferation marker) was used to determine the cell proliferation rate, and nuclear counterstaining was performed with either 4', 6-diamidino-2-phenylindole or Topro III (Invitrogen, Carlsbad, CA). RESULTS: We observed autoreactivity to multiple eyelid structures, including meibomian glands and tarsal muscle bundles at different levels, and some areas of the epidermis and the dermis close to the isthmus of the eyelids. Tarsal plate autoreactivity was seen in 10 of 12 of the El Bagre-EPF sera and in one control with pemphigus erythematosus. Furthermore, immunoprecipitation using an eyelid sample as a substrate with 1 mmol/L of sodium orthovanodate showed autoreactivity to several antigens, including some of possible lipid origin. LIMITATIONS: The main limitation of this study is the fact that the antigen or antigens remain unknown. CONCLUSION: We identified for the first time to our knowledge autoantibodies to meibomian glands and tarsal muscle in El Bagre-EPF. Our findings suggest that the autoantibodies to the ocular structures cause the clinical and histopathological findings in the ocular lesions in El Bagre-EPF.

Formalin deposition as artifact in biopsies from patients affected by a new variant of endemic pemphigus foliaceus in El Bagre, Colombia, South America.

BACKGROUND: Most autoimmune diseases occur sporadically; however, endemic pemphigus foliaceus (EPF) is present in specific locales restricted to some geographic rural regions mostly in South America, Central America and in Tunisia (Africa). Its geographic restriction makes it an invaluable natural model for studying how the environment, genetic background and host response contribute to the development of autoimmunity. We described a new variant of EPF in El Bagre, Colombia, (El Bagre-EPF). When we examined the skin biopsies from 10 patients and controls from the endemic area, we detected in a systematic manner several types of pigmentation, sometimes intracellular, and sometimes in the extracellular matrix in most biopsies. AIM: We aim to determine the nature of this pigment in these skin biopsies. METHODS: We studied 10 patients and 10 controls matched by sex, age and work activity living in the endemic area by routine hematoxylin and eosin (H&E). RESULTS: We were unable to find any bacteriological or parasitic organism. Specifically, we searched for several tropical disease agents as possible causative agents of this pigment. Iron stains and melanin pigment bleaching techniques failed to determine the etiology of this pigment. We then tried the removal of formalin pigment using picric acid. The pigment was removed after very strong treatment with different acids including picric acid. CONCLUSIONS: Formalin pigment shares many properties with hemozoin. In this case, the authors recommend the use of neutral buffered formalin to prevent the formation of formalin pigment especially after long periods of fixation when taking biopsies under extreme temperature and environmental humidity.

Diagnosis and treatment of cutaneous paraneoplastic disorders.

The skin plays a critical role in the detection of internal malignances. Cutaneous signs of these disorders afford clinicians opportunities for early diagnosis and treatment. We aim to succinctly review the recognition, diagnosis, and treatment of selected cutaneous paraneoplastic diseases. Skin disorders that may be associated with paraneoplastic syndromes include: cutaneous metastases, tripe palms, Sweet's syndrome, glucagonoma, Paget's disease and extramammary Paget's disease, acanthosis nigricans, Birt-Hogg-Dube syndrome, basal cell nevus syndrome, Bazex syndrome (acrokeratosis paraneoplastica), carcinoid syndrome, Cowden's disease(multiple hamartoma syndrome), dermatomyositis, erythema gyratum repens, ichthyosis aquisita, von Recklinghausen's disease, pityriasis rotunda, pyoderma gangrenosum, Quincke's edema (angioedema and paraneoplastic uricaria), paraneoplastic pemphigus, Degos' disease, superior vena cava syndrome, Werner's syndrome, diffuse normolipemic plane xanthomas, and yellow nail syndrome. Treatment for these disorders depends on the nature and anatomic distribution of the primary neoplastic process.

Palm tissue displaying a polyclonal autoimmune response in patients affected by a new variant of endemic pemphigus foliaceus in Colombia, South America.

We previously described a new variant of endemic pemphigus foliaceus in El Bagre, Colombia, South America (El Bagre-EPF). On physical examination, the palms and soles of El Bagre-EPF patients reveal an edematous texture and mild hyperkeratosis, in comparison with the non-glabrous skin of the patients where blisters, pustules or other lesions are commonly found. Based on the preceding observation, we tested the palms of 20 El Bagre-EPF cases and 20 controls from the endemic area for any pathological alterations in the samples by direct immunofluorescence (DIF). Our DIF demonstrated pathological deposits of fibrinogen and albumin, as well as IgG, IgA, IgM, IgD and C3c, at 1) the epidermal basement membrane zone; 2) around isolated areas in the epidermis, 3) within the dermal vessels and nerves, and 4) in areas surrounding dermal neurovascular structures and sweat glands. Specific markers for blood vessels, including 1) anti-intercellular adhesion molecule 1 (ICAM-1)/CD54, and 2) anti-junctional adhesion molecule (JAM-A); as well as specific markers for nerves, including 1) anti-glial fibrillary acidic protein (GFAP), and 2) anti-human neuron specific enolase (NSE) co-localized with the patients' autoantibodies. Although no blisters, ulcerations, pustules or erosions are clinically observed on the palms of El Bagre-EPF patients, our DIF detected distinct immunoreactivity in palm tissue. These alterations may contribute to the clinically edematous texture of the palms and the mild clinical hyperkeratosis found in most of these patients. We propose that normal glabrous skin and non-glabrous skin may be different with regard to the expression of selected molecules, which may vary in number, size or structural organization depending on their anatomical site. Our findings may also partially explain the hyperkeratotic palms that have been clinically well documented in the chronic phase of fogo selvagem i.e., endemic pemphigus foliaceus, in Brazil.

Involvement of the Areae Compositae of the Heart in Endemic Pemphigus Foliaceus.

BACKGROUND: A new variant of endemic pemphigus foliaceus in El Bagre (El Bagre-EPF), Colombia, South America, shares features with Senear-Usher syndrome and occurs in an endemic fashion. Patients affected by El Bagre-EPF have heterogeneous antigenic reactivity not only to the skin but to other organs, including the heart. Here we test for autoantibodies to the areae compositae of the heart (structure consisting of typical desmosomal amalgamated fascia adherens molecules) and evaluate any possible clinical correlation. METHODS: A case-control study comparing 45 patients and 45 controls from the endemic area, matched by demographics including age, gender, weight, work activities, and comorbidities, was performed. Direct and indirect immunofluorescence, immunohistochemistry, confocal microscopic studies, and echocardiogram studies were completed. RESULTS: The main clinical abnormally seen in the El Bagre-EPF patients was left ventricular hypertrophy in 15/45 patients, compared with no such findings in the control population (P < 0.1). Seventy percent of El Bagre-EPF patients and none of the controls displayed polyclonal autoreactivity using different immunoglobulins and complement to the areae compositae of the heart using different methods and antibodies (P < 0.1). CONCLUSIONS: Patients affected by El Bagre-EPF demonstrated autoantibodies to the areae compositae of the heart. This finding was associated with left ventricular hypertrophic cardiomyopathy. The areae compositae may play a role in cell junction tension and the El Bagre-EPF patients' autoantibodies possibly disrupting these junctions and thereby contributing to the left ventricular hypertrophy.

Patients affected by endemic pemphigus foliaceus in Colombia, South America exhibit autoantibodies to optic nerve sheath envelope cell junctions.

BACKGROUND: The majority of the patients affected by a new variant of endemic pemphigus foliaceus in El Bagre, Colombia (El Bagre EPF or pemphigus Abreu-Manu), have experienced vision problems; we have previously reported several ocular abnormalities. METHODS: Here, we aimed to investigate reactivity to optic nerves in these patients. We utilized bovine, rat and mouse optic nerves, and performed immunofluorescence and confocal microscopy to test for optical nerve autoreactivity. We tested 45 patients affected by this disease and 45 controls from the endemic area matched by age, sex and work activity. RESULTS: Overall, 37 of the 45 patient sera reacted to the optic nerve envelope that is composed of leptomeninges; the reactivity was polyclonal and present mostly at the cell junctions (P < 0.001). The immune response was directed against optic nerve sheath cell junctions and the vessels inside it, as well as other molecules inside the nerve. No control cases were positive. Of interest, all the patient autoantibodies co-localized with commercial antibodies to desmoplakins I-II, myocardium-enriched zonula occludens-1- associated protein (MYZAP), armadillo repeat gene deleted in velo-cardio-facial syndrome (ARVCF), and plakophilin-4 (p0071) from Progen Biotechnik (P < 0.001). CONCLUSION: We conclude that the majority of the patients affected by pemphigus Abreu-Manu have autoantibodies to optic nerve sheath envelope cell junctions. These antibodies also co-localize with armadillo repeat gene deleted in velo-cardio-facial syndrome, p0071 and desmoplakins I-II. The clinical significance of our findings remains unknown.

Subclinical oral involvement in patients with endemic pemphigus foliaceus.

BACKGROUND: We have described a variant of endemic pemphigus foliaceus (EPF) in El Bagre area known as pemphigus Abreu-Manu. Our previous study suggested that Colombian EPF seemed to react with various plakin family proteins, such as desmoplakins, envoplakin, periplakin BP230, MYZAP, ARVCF, p0071 as well as desmoglein 1. OBJECTIVES: To explore whether patients affected by a new variant of endemic pemphigus foliaceus (El Bagre-EPF) demonstrated oral involvement. MATERIALS AND METHODS: A case-control study was done by searching for oral changes in 45 patients affected by El Bagre-EPF, as well as 45 epidemiologically matched controls from the endemic area matched by demographics, oral hygiene habits, comorbidities, smoking habits, place of residence, age, sex, and work activity. Oral biopsies were taken and evaluated via hematoxylin and eosin staining, direct immunofluorescence, indirect immunofluorescence, confocal microscopy, and immunohistochemistry. RESULTS: Radicular pieces and loss of teeth were seen in in 43 of the 45 El Bagre-EPF patients and 20 of the 45 controls (P < 0.001) (confidence interval [CI] 98%). Hematoxylin and eosin staining showed 23 of 45 El Bagre-EPF patients had corneal/subcorneal blistering and lymphohistiocytic infiltrates under the basement membrane zone and around the salivary glands, the periodontal ligament, and the neurovascular bundles in all cell junction structures in the oral cavity; these findings were not seen in the controls (P < 0.001) (CI 98%). The direct immunofluorescence, indirect immunofluorescence, confocal microscopy, and microarray staining displayed autoantibodies to the salivary glands, including their serous acini and the excretory duct cell junctions, the periodontal ligament, the neurovascular bundles and their cell junctions, striated muscle and their cell junctions, neuroreceptors, and connective tissue cell junctions. The autoantibodies were polyclonal. IgA autoantibodies were found in neuroreceptors in the glands and were positive in 41 of 45 patients and 3 of 45 controls. CONCLUSIONS: Patients affected by El Bagre-EPF have some oral anomalies and an immune response, primarily to cell junctions. The intrinsic oral mucosal immune system, including IgA and secretory IgA, play an important role in this autoimmunity. Our data contradict the hypothesis that pemphigus foliaceus does not affect the oral mucosa due to the desmoglein 1-desmoglein 3 compensation.

Cardiac rhythm and pacemaking abnormalities in patients affected by endemic pemphigus in Colombia may be the result of deposition of autoantibodies, complement, fibrinogen, and other molecules.

BACKGROUND: We previously showed that one-third of patients affected by endemic pemphigus foliaceus in El Bagre, Colombia (El Bagre-EPF), display autoreactivity to the heart. OBJECTIVE: The purpose of this study was to investigate rhythm disturbances with the presence of autoantibodies and correlate them with ECG changes in these patients. METHODS: We performed a study comparing 30 patients and 30 controls from the endemic area, matched by demographics, including age, sex, weight, work activities, and comorbidities. ECG as well as direct and indirect immunofluorescence, immunohistochemistry, and confocal microscopic studies focusing on cardiac node abnormalities were performed. Autopsies of 7 patients also were reviewed. RESULTS: The main ECG abnormalities seen in the El Bagre-EPF patients were sinus bradycardia (in one-half), followed by left bundle branch block, left posterior fascicular block, and left anterior fascicular block compared with the controls. One-third of the patients displayed polyclonal autoantibodies against the sinoatrial and/or AV nodes and the His bundle correlating with rhythm anomalies and delays in the cardiac conduction system (P <.01). The patient antibodies colocalized with commercial antibodies to desmoplakins I and II, p0071, armadillo repeat gene deleted in velo-cardio-facial syndrome (ARVCF), and myocardium-enriched zonula occludens-1-associated protein (MYZAP; Progen Biotechnik) (P <.01). CONCLUSION: One-third of the patients affected by El Bagre-EPF have rhythm abnormalities that slow the conduction of impulses in cardiac nodes and the cardiac conduction system. These abnormalities likely occur as a result of deposition of autoantibodies, complement, and other inflammatory molecules. We show for the first time that MYZAP is present in cardiac nodes.

Cell junction protein armadillo repeat gene deleted in velo-cardio-facial syndrome is expressed in the skin and colocalizes with autoantibodies of patients affected by a new variant of endemic pemphigus foliaceus in Colombia.

BACKGROUND: We previously described a new variant of endemic pemphigus foliaceus in El Bagre, Colombia, South America (El Bagre-EPF, or pemphigus Abreu-Manu). El Bagre-EPF differs from other types of EPF clinically, epidemiologically, immunologically and in its target antigens. We reported the presence of patient autoantibodies colocalizing with armadillo repeat gene deleted in velo-cardio-facial syndrome (ARVCF), a catenin cell junction protein colocalizing with El Bagre-EPF autoantibodies in the heart and within pilosebaceous units along their neurovascular supply routes. Here we investigate the presence of ARVCF in skin and its possibility as a cutaneous El Bagre-EPF antigen. METHODS: We used a case-control study, testing sera of 45 patients and 45 controls via direct and indirect immunofluorescence (DIF/IIF), confocal microscopy, immunoelectron microscopy and immunoblotting for the presence of ARVCF and its relationship with El Bagre-EPF autoantibodies in the skin. We also immunoadsorbed samples with desmoglein 1 (Dsg1) ectodomain (El Bagre-EPF antigen) by incubating with the positive ARVCF samples from DIF and IIF. RESULTS: ARVCF was expressed in all the samples from the cases and controls. Immunoadsorption with Dsg1 on positive ARVCF immunofluorescence DIF/IIF cases showed that the immune response was present against non-desmoglein 1 antigen(s). Overall, 40/45 patients showed colocalization of their autoantibodies with ARVCF in the epidermis; no controls from the endemic area displayed colocalization. CONCLUSIONS: We demonstrate that ARVCF is expressed in many areas of human skin, and colocalizes with the majority of El Bagre-EPF autoantibodies as a putative antigen.

Periodic Acid-Schiff Staining Parallels the Immunoreactivity Seen By Direct Immunofluorescence in Autoimmune Skin Diseases.

BACKGROUND: In many countries and laboratories, techniques such as direct immunofluorescence (DIF) are not available for the diagnosis of skin diseases. Thus, these laboratories are limited in the full diagnoses of autoimmune skin diseases, vasculitis, and rheumatologic diseases. In our experience with these diseases and the patient's skin biopsies, we have noted a positive correlation between periodic acid-Schiff (PAS) staining and immunofluorescence patterns; however, these were just empiric observations. In the current study, we aim to confirm these observations, given the concept that the majority of autoantibodies are glycoproteins and should thus be recognized by PAS staining. AIMS: To compare direct immunofluorescent and PAS staining, in multiple autoimmune diseases that are known to exhibit specific direct immunofluorescent patterns. MATERIALS AND METHODS: We studied multiple autoimmune skin diseases: Five cases of bullous pemphigoid, five cases of pemphigus vulgaris, ten cases of cutaneous lupus, ten cases of autoimmune vasculitis, ten cases of lichen planus (LP), and five cases of cutaneous drug reactions (including one case of erythema multiforme). In addition, we utilized 45 normal skin control specimens from plastic surgery reductions. RESULTS: We found a 98% positive correlation between DIF and PAS staining patterns over all the disease samples. CONCLUSION: We recommend that laboratories without access to DIF always perform PAS staining in addition to hematoxylin and eosin (H&E) staining, for a review of the reactivity pattern.

Immunofluorescence Patterns in Selected Dermatoses, Including Blistering Skin Diseases Utilizing Multiple Fluorochromes.

BACKGROUND: Autoimmune vesiculobullous disorders represent a heterogeneous group of dermatoses whose diagnosis is made based on clinical history, histologic features, and immunopathologic features. The most commonly used techniques for the diagnosis of these diseases are direct and indirect immunofluorescence (DIF and IIF), including salt-split processing. NaCl split skin is used to determine the level of blister formation, and the localization of autoantibodies relative to the split. Classically, immunofluorescence has been performed with one fluorochrome in the diagnosis of autoimmune bullous skin diseases. AIMS: To compare DIF and IIF of the skin, using a single fluorochrome versus multiple fluorochromes. MATERIALS AND METHODS: We studied 20 autoimmune skin disease cases using fluorescein isothiocyanate (FITC) alone, in comparison to multiple fluorochromes (with or without DNA counterstaining). RESULTS: The use of multiple fluorochromes helped to simultaneously visualize reactivity in multiple skin areas, in contrast to using FITC alone. CONCLUSIONS: Using multiple fluorochromes allows simultaneous labeling of two or more antigens within the same cell/or tissue section, assists in colocalization of unknown antigens with known molecules, and helps in ruling out "background" staining.