Presence of interleukin 10 in the serum and blister fluid of patients with pemphigus vulgaris and pemphigoid

Interleukin 10 (IL-10) is an immunoregulatory cytokine produced by T lymphocytes and macrophages. Recently, it has been suggested that IL-10 may be involved in the pathogenesis of various inflammatory and autoimmune diseases. Using an ELISA we investigated the presence of IL-10 in the serum and blister fluid of pemphigus vulgaris (PV) patients with active disease and those in prolonged clinical remission compared with normal controls. Sera from patients with bullous pemphigoid (BP), ocular cicatricial pemphigoid (OCP), oral pemphigoid (OP) and blister fluid from five patients with BP were also studied. Increased levels of IL-10 were detected in the sera of 87.5% of patients with active PV and were statistically significant (P=0.0003) when compared with levels in normal human serum. Lower levels of IL-10 were detected in 12.5% PV patients in remission and were statistically significant (P=0.0001) when compared with levels in patients with active disease. Levels of IL-10 were detected in sera of 4.6% (1 of 24) of the normal controls. The levels of IL-10 were approximately four times higher in blister fluids than levels in the serum in the same PV patients. This difference was highly statistically significant (P=0.0008). A correlation was observed between serum levels of IL-10 and titres of pemphigus autoantibodies and with disease severity. Elevated level of IL-10 was detected in the blister fluid from five BP patients. Levels of IL-10 in the sera of patients with BP, OCP and OP were not significantly increased. These preliminary data suggest that IL-10 in concert with other cytokines may play an important role in the pathogenesis of PV and BP.     

Desmosomes and disease: pemphigus and bullous impetigo

Desmosomal cadherins are the pathophysiologic targets of autoimmune or toxin-mediated disruption in the human diseases pemphigus and bullous impetigo (including its generalized form, called staphylococcal scalded skin syndrome). Experiments exploiting the production of both pathogenic and nonpathogenic antidesmoglein antibodies in pemphigus patients' sera have afforded data that make an invaluable contribution towards identifying the functional domains of the desmogleins involved in intercellular adhesion. Conformational epitopes of antidesmoglein autoantibodies in pemphigus patients' sera and the specific cleavage site of desmoglein 1 by exfoliative toxin have been identified, implicating the N-terminal extracellular domains of the desmogleins as critical regions for controlling intercellular adhesion. Furthermore, the development of active autoimmune mouse models for pemphigus allows in vivo characterization of the disease and its pathogenesis. These studies offer new insight into the potential mechanisms of acantholysis in pemphigus and staphylococcal-associated blistering disease, with implications for the role of desmogleins in desmosomal structure and function.     

Dominant autoimmune epitopes recognized by pemphigus antibodies map to the N-terminal adhesive region of desmogleins.

Desmoglein (Dsg) is a cadherin-type adhesion molecule found in desmosomes. Dsg1 and Dsg3 are the target Ags in the autoimmune blistering diseases pemphigus foliaceus (PF) and pemphigus vulgaris (PV), respectively. To map conformational epitopes of Dsg1 and Dsg3 in PF and PV, we generated Dsg1- and Dsg3-domain-swapped molecules and point-mutated Dsg3 molecules with Dsg1-specific residues by baculovirus expression. The swapped domains were portions of the N-terminal extracellular domains of Dsg1 (1-496) and Dsg3 (1-566), which have similar structures but distinct epitopes. The binding of autoantibodies to the mutant molecules was assessed by competition ELISAs. Domain-swapped molecules containing the N-terminal 161 residues of Dsg1 and Dsg3 yielded >50% competition in 30/43 (69.8%) PF sera and 31/40 (77.5%) PV sera, respectively. Furthermore, removal of Abs against the 161 N-terminal residues of Dsg1 by immunoadsorption eliminated the ability of PF sera to induce cutaneous blisters in neonatal mice. Within these N-terminal regions, most of the epitopes were mapped to residues 26-87 of Dsg1 and 25-88 of Dsg3. Furthermore, a point-mutated Dsg3 molecule containing Dsg1-specific amino acid substitutions (His(25), Cys(28), Ala(29)) reacted with anti-Dsg1 IgG, thus defining one of the epitopes of Dsg1. Using the predicted three-dimensional structure of classic cadherins as a model, these findings suggest that the dominant autoimmune epitopes in both PF and PV are found in the N-terminal adhesive surfaces of Dsgs.     

A Pathophysiologic Role for Epidermal Growth Factor Receptor in Pemphigus Acantholysis

The pemphigus family of autoimmune bullous disorders is characterized by autoantibody binding to desmoglein 1 and/or 3 (dsg1/dsg3). In this study we show that EGF receptor (EGFR) is activated following pemphigus vulgaris (PV) IgG treatment of primary human keratinocytes and that EGFR activation is downstream of p38 mitogen-activated protein kinase (p38). Inhibition of EGFR blocked PV IgG-triggered dsg3 endocytosis, keratin intermediate filament retraction, and loss of cell-cell adhesion in vitro. Significantly, inhibiting EGFR prevented PV IgG-induced blister formation in the passive transfer mouse model of pemphigus. These data demonstrate cross-talk between dsg3 and EGFR, that this cross-talk is regulated by p38, and that EGFR is a potential therapeutic target for pemphigus. Small-molecule inhibitors and monoclonal antibodies directed against EGFR are currently used to treat several types of solid tumors. This study provides the experimental rationale for investigating the use of EGFR inhibitors in pemphigus.     

Changes in desmoglein 1 expression and subcellular localization in cultured keratinocytes subjected to anti-desmoglein 1 pemphigus autoimmunity

The complexity of pemphigus acantholysis together with the weak expression of desmoglein 1 (Dsg1) in cultured keratinocytes have made the study on the pathogenic action of anti-Dsg1 antibodies quite difficult. The pathophysiology of the acantholytic phenomenon could depend on the reduction of Dsg1 adhesion function occurring after its massive internalization or decrease of its synthesis. Here, we have investigated this hypothesis by using sera of patients having antibodies against Dsg1 or monoclonal anti-Dsg1 antibodies to simulate pemphigus autoimmunity in Dsg1-rich keratinocytes. Similar to pemphigus foliaceus (PF) and vulgaris (PV) sera, monoclonal anti-Dsg1 antibodies induced transient internalization of Dsg1 and reduced the adhesion strength among keratinocytes. However, binding of IgG to Dsg1 did not determine its early depletion from the adhesion complexes but reduced the amount of Dsg1 found in the Triton X-100 soluble pool of proteins. Taken together, our results represent the first demonstration that anti-Dsg1 antibodies induce similar alterations on the subcellular distribution of Dsg1 irrespective of the disease where they come from. Furthermore, the present study provides insight into the mechanisms underlying epithelial blistering observed in the skin type of pemphigus.     

Inhibition of Rho A activity causes pemphigus skin blistering

The autoimmune blistering skin diseases pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are mainly caused by autoantibodies against desmosomal cadherins. In this study, we provide evidence that PV-immunoglobulin G (IgG) and PF-IgG induce skin blistering by interference with Rho A signaling. In vitro, pemphigus IgG caused typical hallmarks of pemphigus pathogenesis such as epidermal blistering in human skin, cell dissociation, and loss of desmoglein 1 (Dsg 1)-mediated binding probed by laser tweezers. These changes were accompanied by interference with Rho A activation and reduction of Rho A activity. Pemphigus IgG-triggered keratinocyte dissociation and Rho A inactivation were p38 mitogen-activated protein kinase dependent. Specific activation of Rho A by cytotoxic necrotizing factor-y abolished all pemphigus-triggered effects, including keratin retraction and release of Dsg 3 from the cytoskeleton. These data demonstrate that Rho A is involved in the regulation of desmosomal adhesion, at least in part by maintaining the cytoskeletal anchorage of desmosomal proteins. This may open the possibility of pemphigus treatment with the epidermal application of Rho A agonists.     

Molecular heterogeneity of the bullous pemphigoid antigens as detected by immunoblotting

Sera from 28 patients with bullous pemphigoid (BP), four patients with cicatricial pemphigoid (CP), and 24 controls (normal volunteers and patients with pemphigus, systemic lupus erythematosus, or other skin diseases) were tested against extracts of human epidermis by immunoblotting techniques. The extraction buffer included 1% SDS, 5% beta-mercaptoethanol, and six protease inhibitors with various specificities. BP sera from individual patients showed different patterns of reactivity with the same epidermal extract, and each pattern consisted of one or more bands. A total of five bands of 240 kD, 200 kD, 180 kD, 97 kD, and 77 kD reacted with BP sera; the 240-kD band reacted with one CP sera, and none of these bands was detected by the control sera. The 240-kD and 180-kD bands reacted very strongly with some sera and were most frequently observed (43% and 29%, respectively). The 200-kD, 97-kD, and 77-kD bands were less frequently observed (25%, 7%, and 7%, respectively), but when present, their reactions were usually strong. Eleven percent of the BP sera did not react with any bands. Contrary to previous reports, this study shows that BP autoantibodies react with several protein bands, as detected by immunoblotting. We have recently shown by immunoelectron microscopy that BP autoantibodies bind to the basal cell hemidesmosomes. It remains to be determined which of these protein bands represent specific hemidesmosomal proteins and which antibody-antigen interactions are relevant to the pathogenesis of this disease.     

Pemphigus vulgaris IgG directly inhibit desmoglein 3-mediated transinteraction

The autoimmune blistering skin disease pemphigus is caused by autoantibodies against keratinocyte surface Ags. In pemphigus vulgaris (PV), autoantibodies are primarily directed against desmosomal cadherins desmoglein (Dsg) 3 and Dsg 1, whereas pemphigus foliaceus (PF) patients only have Abs against Dsg 1. At present, it is unclear whether Dsg autoantibodies contribute to pemphigus pathogenesis by direct inhibition of Dsg transinteraction. Using atomic force microscopy, we provide evidence that PV-IgG directly interfere with homophilic Dsg 3 but, similar to PF-IgG, not with homophilic Dsg 1 transinteraction, indicating that the molecular mechanisms in PV and PF pathogenesis substantially differ. PV-IgG (containing Dsg 3 or Dsg 1 and Dsg 3 autoantibodies) as well as PV-IgG Fab reduced binding activity of Dsg 3 by approximately 60%, comparable to Ca(2+) depletion. Similarly, the mouse monoclonal PV Ab AK 23 targeting the N-terminal Dsg 3 domain and AK 23 Fab reduced Dsg 3 transinteraction. In contrast, neither PV-IgG nor PF-IgG blocked Dsg 1 transinteraction. In HaCaT monolayers, however, both PV- and PF-IgG caused keratinocyte dissociation as well as loss of Dsg 1 and Dsg 3 transinteraction as revealed by laser tweezer assay. These data demonstrate that PV-IgG and PF-IgG reduce Dsg transinteraction by cell-dependent mechanisms and suggest that in addition, Abs to Dsg 3 contribute to PV by direct inhibition of Dsg transinteraction.     

Biphasic Activation of p38MAPK Suggests That Apoptosis Is a Downstream Event in Pemphigus Acantholysis

In pemphigus vulgaris and pemphigus foliaceus (PF), autoantibodies against desmoglein-3 and desmoglein-1 induce epidermal cell detachment (acantholysis) and blistering. Activation of keratinocyte intracellular signaling pathways is emerging as an important component of pemphigus IgG-mediated acantholysis. We previously reported activation of p38 mitogen-activated protein kinase (MAPK) in response to pathogenic pemphigus vulgaris and PF IgG. Inhibition of p38MAPK blocked pemphigus IgG-induced cytoskeletal reorganization in tissue culture and blistering in pemphigus mouse models. We now extend these observations by demonstrating two peaks of p38MAPK activation in pemphigus tissue culture and mouse models. Administration of the p38MAPK inhibitor SB202190 before PF IgG injection blocked both peaks of p38MAPK phosphorylation and blister formation, consistent with our previous findings; however, administration of the inhibitor 4 h after PF IgG injection blocked only the later peak of p38MAPK activation but failed to block blistering. Examination of the temporal relationship of p38MAPK phosphorylation and apoptosis showed that apoptosis occurs at or after the second peak of p38MAPK activation. The time course of p38MAPK activation and apoptotic markers, as well as the ability of inhibitors of p38MAPK to block activation of the proapoptotic proteinase caspase-3, suggest that activation of apoptosis is downstream to, and a consequence of, p38MAPK activation in pemphigus acantholysis. Furthermore, these observations suggest that the earlier peak of p38MAPK activation is part of the mechanism leading to acantholysis, whereas the later peak of p38MAPK and apoptosis may not be essential for acantholysis.Pemphigus is a group of related autoimmune diseases characterized by blistering in the skin. The histologic hallmark of these disorders is termed acantholysis, which describes the loss of adhesion between adjacent epithelial cells. The two major variants are pemphigus foliaceus (PF)² and pemphigus vulgaris (PV). In PF, acantholysis is observed beneath the stratum corneum and within the granular layer of epidermal epithelia, whereas in PV, blister formation occurs above the basal layer of epidermal epithelia and mucosal epithelium. Passive transfer of IgG purified from both PV and PF patient sera reproduces the clinical, histological, and immunologic features of the human diseases, demonstrating that these autoantibodies are pathogenic (1, 2). In PF, autoantibodies target the desmosomal cadherin desmoglein (dsg) 1, whereas in PV, autoantibodies initially target dsg3 (3, 4) in mucosal PV and then subsequently target both dsg1 and dsg3 in mucocutaneous PV (5-7).The mechanism by which pemphigus autoantibodies induce blistering has been under investigation. Work from a number of laboratories has suggested that activation of intracellular events is induced by binding of PF or PV IgG to dsg1 and dsg3, respectively (8-14). Previously, we have reported that PV IgG activate p38MAPK and heat shock protein (HSP) 27 in human keratinocyte tissue cultures (15). Significantly, p38MAPK inhibitors blocked PV IgG-induced keratin filament retraction and actin reorganization in human keratinocyte tissue cultures. Furthermore, we have demonstrated that both PV and PF IgG induce phosphorylation of p38MAPK and HSP25, the murine HSP27 homologue, in mouse models and that inhibitors of p38MAPK block blistering in both the PV (16) and the PF (17) passive transfer mouse models. Additionally, in human skin biopsies from both PV and PF patients, phosphorylation of p38MAPK and HSP27 has been observed (18). Collectively, these observations suggest that activation of p38MAPK within the target keratinocyte contributes directly to loss of cell-cell adhesion induced by pemphigus autoantibodies.Both p38MAPK and HSP27 have been implicated in the regulation of the intermediate filament and actin cytoskeletons (19-25); the ability of p38MAPK inhibitors to block both pemphigus IgG-activated cytoskeletal reorganization and pemphigus IgG-activated blistering suggests that p38MAPK may be acting upstream of the cytoskeleton in the mechanism of acantholysis; however, p38MAPK signaling has been implicated in other cellular responses (reviewed in Ref. 26). For example, there is abundant evidence for p38MAPK involvement in apoptosis (27-29); however, the role of p38MAPK in apoptosis seems to be cell type- and stimulus-dependent. Although p38MAPK signaling promotes cell death in some cell lines, it also functions to enhance survival, growth, and differentiation in other cell lines (30). Several reports describe increased apoptosis of keratinocytes in pemphigus (31-35); however, the relationship between PV IgG-mediated p38MAPK signaling, the induction of apoptosis, and the relationship of apoptosis to blistering has not been defined. This study was undertaken to investigate the relationship between p38MAPK activation, apoptosis, and acantholysis.     

Evidence for a role of autoantibodies to heat shock protein 60, 70, and 90 in patients with dermatitis herpetiformis

Heat shock proteins (Hsp) are highly conserved immunomodulatory molecules upregulated when cells are exposed to stressful stimuli, such as inflammation. Their involvement in various autoimmune diseases, including autoimmune bullous diseases and celiac disease, has been increasingly recognized. To further study the role of Hsp in autoimmune bullous diseases, we have investigated for the first time the humoral autoimmune response to Hsp40, Hsp60, Hsp70, and Hsp90 in patients with dermatitis herpetiformis (DH; n = 26), bullous pemphigoid (BP; n = 23), and pemphigus vulgaris (PV; n = 16), the first representing a cutaneous manifestation of celiac disease. While in patients with active BP and PV, serum levels of autoantibodies against these Hsp did not differ from the corresponding age- and gender-matched healthy controls (n = 9–14); circulating autoantibodies against Hsp60, Hsp70, and Hsp90 were found to be increased at the active disease stage of DH. Further analysis of this latter patient subgroup showed that these anti-Hsp autoantibodies decreased in parallel with serum autoantibodies against epidermal and tissue transglutaminase during remission of skin lesions following a gluten-free diet, revealing significantly positive correlations. Although further studies on larger groups of patients will be needed to confirm the present data, our results support the notion that autoantibodies against Hsp60, Hsp70, and Hsp90 deserve attention in the study of the mechanisms that promote the development and maintenance of DH and possibly also the underlying celiac disease as well as potential novel disease biomarkers.     

Serum from pemphigus vulgaris reduces desmoglein 3 half-life and perturbs its de novo assembly to desmosomal sites in cultured keratinocytes

Defects of cell-cell adhesion underlie disruption of epithelial integrity observed in patients with pemphigus vulgaris (PV), an autoimmune disease characterized by severe mucosal erosions and skin blisters. Pathogenic PV autoantibodies found in patients' sera target desmoglein 3 (Dsg3), a major component of the desmosome, but how does this phenomenon affect Dsg-dependent adhesion and lead to acantholysis still remains controversial. Here, we show that PV serum determines a reduction of Dsg3 half-life in HaCaT keratinocytes, although the total amount of Dsg3 remains unchanged. Immunofluorescence studies suggest that PV IgG exert their effect prevalently by binding non-desmosomal Dsg3 without causing its massive internalization. Furthermore, PV IgG targeting desmosome-assembled Dsg3 do not induce depletion of Dsg3 from the adhesion sites. Conversely, incorporation of PV IgG-Dsg3 complexes into new forming desmosomes appears perturbed. With our study, the basic biochemical changes of Dsg3 in an in vitro model of PV have been defined.     

Recent Advances in the Diagnosis and Treatment of Pemphigus

Immunofluorescent techniques offer considerable promise in facilitating the diagnosis of pemphigus and other bullous diseases and also have provided a means for further investigative studies of these diseases. The immunofluorescent findings have in part stimulated the use of immunosuppressive agents in the management of pemphigus. Their exact status is still undetermined but early experience appears favorable.     

Mechanisms of blister formation by staphylococcal toxins

Many children suffer from the bacterial skin diseases bullous impetigo and staphylococcal scalded skin syndrome (SSSS). Staphylococcus aureus, which produces exfoliative toxins (ETs), causes these diseases. Recently, it was proven that ETs cleave the cell adhesion molecule desmoglein (Dsg) 1, which plays an important role in maintaining the proper structure and barrier function of the epidermis. Surprisingly, Dsg1 is also the antibody target in the autoimmune disease pemphigus foliaceus. Skin biopsies from pemphigus foliaceus patients show the same pathology as those from bullous impetigo and SSSS patients. The crystal structure of ET suggests that it is a serine protease with an inactive catalytic site, which may become activated when ET binds a specific receptor. This receptor binding is thought to cause a change in conformation that exposes the catalytic site. It has recently been shown that Dsg1 specifically binds and activates ET, which in turn cleaves the bound Dsg1 at only one peptide bond. This process is absolutely dependent on the calcium-dependent conformation of Dsg1. These data suggest that ETs have a very high specificity for human Dsg1, and that S. aureus uses ETs to disrupt the barrier of the human epidermis in order to survive and proliferate on the human body.     

Interleukin 4 inhibition as a potential therapeutic in pemphigus

Pemphigus is an autoimmune bullous skin disease that results from desmosomal protein desmoglein 3 and 1 loss in pemphigus vulgaris and foliaceus, respectively. It can be considered as a Th2-dominant disease over-expressed by Th2 cell cytokines. Interleukin (IL)-4 is a key cytokine which can exacerbate Th2 over-expression in addition to isotype switching to immunoglobin (Ig)G1 and IgG4 that are responsible for desmoglein loss. Elevation of IL-4 level has also been reported in various studies. Considering the important role of IL-4 in severe phase of pemphigus and lack of effective and safeness therapy for this potentially fatal disease, anti-IL-4 therapy was introduced as a potential curative for pemphigus disease. This study reviewed all studies about any roles of IL-4 that can directly and indirectly be played in the development of pemphigus and IL-4 inhibition with interferons and dupilumab therapy were introduced as a novel pemphigus treatment for patients who are in relapse phase of the disease. Dupilumab was also introduced as a possible treatment for patients with severe pemphigus. It can directly inhibit IL-4 by targeting IL-4 α-chain receptor. IL-4 inhibition can lead to the creation of Th1:Th2 balance by various pathways, discussed in this study.     

Pemphigus Vulgaris Autoantibody Profiling by Proteomic Technique

Pemphigus vulgaris (PV) is a mucocutaneous blistering disease characterized by IgG autoantibodies against the stratified squamous epithelium. Current understanding of PV pathophysiology does not explain the mechanism of acantholysis in patients lacking desmoglein antibodies, which justifies a search for novel targets of pemphigus autoimmunity. We tested 264 pemphigus and 138 normal control sera on the multiplexed protein array platform containing 701 human genes encompassing many known keratinocyte cell-surface molecules and members of protein families targeted by organ-non-specific PV antibodies. The top 10 antigens recognized by the majority of test patients’ sera were proteins encoded by the DSC1, DSC3, ATP2C1, PKP3, CHRM3, COL21A1, ANXA8L1, CD88 and CHRNE genes. The most common combinations of target antigens included at least one of the adhesion molecules DSC1, DSC3 or PKP3 and/or the acetylcholine receptor CHRM3 or CHRNE with or without the MHC class II antigen DRA. To identify the PV antibodies most specific to the disease process, we sorted the data based on the ratio of patient to control frequencies of antigen recognition. The frequency of antigen recognition by patients that exceeded that of control by 10 and more times were the molecules encoded by the CD33, GP1BA, CHRND, SLC36A4, CD1B, CD32, CDH8, CDH9, PMP22 and HLA-E genes as well as mitochondrial proteins encoded by the NDUFS1, CYB5B, SOD2, PDHA1 and FH genes. The highest specificity to PV showed combinations of autoantibodies to the calcium pump encoded by ATP2C1 with C5a receptor plus DSC1 or DSC3 or HLA-DRA. The results identified new targets of pemphigus autoimmunity. Novel autoantibody signatures may help explain individual variations in disease severity and treatment response, and serve as sensitive and specific biomarkers for new diagnostic assays in PV patients.     

Targeted Disruption of the Pemphigus Vulgaris Antigen (Desmoglein 3) Gene in Mice Causes Loss of Keratinocyte Cell Adhesion with a Phenotype Similar to Pemphigus Vulgaris

In patients with pemphigus vulgaris (PV), autoantibodies against desmoglein 3 (Dsg3) cause loss of cell–cell adhesion of keratinocytes in the basal and immediate suprabasal layers of stratified squamous epithelia. The pathology, at least partially, may depend on protease release from keratinocytes, but might also result from antibodies interfering with an adhesion function of Dsg3. However, a direct role of desmogleins in cell adhesion has not been shown. To test whether Dsg3 mediates adhesion, we genetically engineered mice with a targeted disruption of the DSG3 gene. DSG3 −/− mice had no DSG3 mRNA by RNase protection assay and no Dsg3 protein by immunofluorescence (IF) and immunoblots. These mice were normal at birth, but by 8–10 d weighed less than DSG3 +/− or +/+ littermates, and at around day 18 were grossly runted. We speculated that oral lesions (typical in PV patients) might be inhibiting food intake, causing this runting. Indeed, oropharyngeal biopsies showed erosions with histology typical of PV, including suprabasilar acantholysis and “tombstoning” of basal cells. EM showed separation of desmosomes. Traumatized skin also had crusting and suprabasilar acantholysis. Runted mice showed hair loss at weaning. The runting and hair loss phenotype of DSG3 −/− mice is identical to that of a previously reported mouse mutant, balding (bal). Breeding indicated that bal is coallelic with the targeted mutation. We also showed that bal mice lack Dsg3 by IF, have typical PV oral lesions, and have a DSG3 gene mutation. These results demonstrate the critical importance of Dsg3 for adhesion in deep stratified squamous epithelia and suggest that pemphigus autoantibodies might interfere directly with such a function.     

Imaging and Force Spectroscopy on Desmoglein 1 Using Atomic Force Microscopy Reveal Multivalent Ca2+-Dependent, Low-Affinity Trans-Interaction

Desmoglein 1 is a desmosomal member of the cadherin family expressed in stratified epithelia. Desmoglein 1 is the target adhesion molecule of severe blistering skin diseases such as pemphigus or bullous impetigo. However, despite this enormous pathological relevance, the molecular binding properties of desmoglein 1 are largely unknown. Using atomic force microscopic imaging, we found that desmoglein 1 molecules displayed Ca²⁺-dependent conformational changes of the extracellular domains. By single-molecule force-distance cycles, we provide evidence that desmoglein 1 undergoes Ca²⁺-dependent (K _d = 0.8 mm Ca²⁺) homophilic trans-interaction, which is highly relevant for the contribution of desmoglein 1 homophilic binding to keratinocyte cohesion in distinct epidermal layers. Moreover, while the single-unit unbinding force is comparable to other cadherins (∼40 pN at retrace velocity of 300 nm/s), apparent differences with respect to multivalency of interaction and lifetime of single bonds (0.17 s) were observed. Thus, besides the biophysical characterization of desmoglein 1, a main outcome of the study is that desmoglein 1 differs from other members of the cadherin family in terms of some molecular binding properties. Jens Waschke, Carlos Menendez-Castro, and Paola Bruggeman contributed equally to this study.     

Cytokine profiles in pemphigus vulgaris patients treated with intravenous immunoglobulins as compared to conventional immunosuppressive therapy

Pemphigus vulgaris (PV) is a potentially fatal blistering disease of the skin and mucous membranes, characterized by the presence of autoantibodies against adhesion molecules (desmoglein, Dsg3) present on the surface of keratinocytes, which lead to the loss of cellular adhesion or acantholysis. The mainstay of treatment is conventional immunosuppressive therapy (CIST), i.e. high dose, long-term systemic corticosteroids with or without immunosuppressive drugs. Intravenous immunoglobulin (IVIg) has been used in patients refractory to CIST, and its use has resulted in long-term clinical remission. Since cytokines play an important role in the immunopathogenesis of PV, it would be useful to compare how both IVIg and CIST therapies affect cytokine levels in the serum of PV patients. Thus, the goal of this study was to conduct a comparative analysis of levels of various cytokines, during an 18 month consecutive period, after the initiation of CIST or IVIg treatment in PV patients, with similar extent and severity of disease in the two study groups, with 11 patients in each group. The cytokines measured were IL-1β, IL-6, IL-8, IFN-γ, IL-4 and IL-10. The levels of most of these cytokines were higher in the sera of untreated patients in both groups, compared to normal controls. The cumulative data collected over an 18 month period of treatment demonstrates that there is a gradual reduction in the levels of these cytokines, until they are at levels observed in normal individuals. The conclusions from this limited number of patients, prospectively studied, would suggest that both CIST and IVIg therapies are similar in their ability to influence a panel of cytokines in patients with pemphigus vulgaris.     

Bullous pemphigoid and pemphigus vulgaris—incidence and mortality in the UK: population based cohort study

Objective To determine the incidence of and mortality from bullous pemphigoid and pemphigus vulgaris in the United Kingdom.Design Retrospective historical cohort study.Setting Computerised medical records from the health improvement network, a large population based UK general practice database.Participants Patients with pemphigus vulgaris and bullous pemphigoid diagnostic codes and age, sex, and practice matched controls.Main outcome measures Incidence and mortality compared with the control population by calendar period, age group, sex, geographical region, and degree of social deprivation.Results 869 people with bullous pemphigoid and 138 people with pemphigus vulgaris were identified. The median age at presentation for bullous pemphigoid was 80 (range 23-102) years, and 534 (61%) patients were female. The median age at presentation for pemphigus vulgaris was 71 (21-102) years, and 91 (66%) patients were female. Incidences of bullous pemphigoid and pemphigus vulgaris were 4.3 (95% confidence interval 4.0 to 4.6) and 0.7 (0.6 to 0.8) per 100 000 person years. The incidence of bullous pemphigoid increased over time; the average yearly increase was 17% (incidence rate ratio=1.2, 95% confidence interval 1.1 to 1.2). An average yearly increase in incidence of pemphigus vulgaris of 11% (incidence rate ratio=1.1, 1.0 to 1.2) occurred. The risk of death for patients with bullous pemphigoid was twice as great as for controls (adjusted hazard ratio=2.3, 95% confidence interval 2.0 to 2.7). For pemphigus vulgaris, the risk of death was three times greater than for controls (adjusted hazard ratio=3.3, 2.2 to 5.2).Conclusions Incidences of bullous pemphigoid and pemphigus vulgaris are increasing. The reasons for the changes in incidence are not clearly understood but have implications for identifying causative factors. Both disorders are associated with a high risk of death. Previous estimates may have underestimated the risk of death associated with these diseases.     

Catalase and Lipid Peroxidation Values in Serum of Tunisian Patients with Pemphigus Vulgaris and Foliaceus

Pemphigus is an autoimmune disorder resulting from the interaction between autoantibodies and desmoglein. Oxidative stress seems to be responsible for the onset/aggravation of many human diseases. Actually, it is considered as one of the several factors for the etiopathogenesis of pemphigus. The present study aims to evaluate the oxidative state in the sera of pemphigus vulgaris and pemphigus foliaceus patients by assessing lipid peroxidation, proteins oxidation, and antioxidant enzyme activity. This study included 36 pemphigus vulgaris and 42 pemphigus foliaceus patients as well as a group of controls consisting of 78 healthy volunteers. Malondialdehyde levels (p?<?0.001) and catalase activity (p?<?0.001) are higher in both groups of patients than in the control group. The two groups of patients showed a nonsignificant decrease in the thiol groups compared with the healthy one. A nonsignificant difference was shown between pemphigus vulgaris and pemphigus foliaceus patients, except for the catalase which shows an increase in the pemphigus vulgaris group. We have also found significant correlations between serum oxidative stress marker levels and serum anti-desmoglein antibody levels in the two pemphigus groups. These findings underline the implication of oxidative stress in the physiopathology of pemphigus by the increase in the autoantibodies?? reactivity.