Differential effects of caffeine on hair shaft elongation, matrix and outer root sheath keratinocyte proliferation, and TGF-beta2-/IGF-1-mediated regulation of hair cycle in male and female human hair follicles in vitro.

BACKGROUND: Caffeine reportedly counteracts the suppression of hair shaft production by testosterone in organ-cultured male human hair follicles (HFs). OBJECTIVES: We aimed at investigating the impact of caffeine a) on additional key hair growth parameters, b) on major hair growth-regulatory growth factors and c) on male versus female HFs in the presence of testosterone. METHODS: Microdissected male and female human scalp HFs were treated in serum-free organ culture for 120 h with testosterone alone (0,5 mug/ml) or in combination with caffeine (0.005-0.0005%), and effects on hair shaft elongation, HF cycling (i.e. anagen-catagen transition), hair matrix keratinocyte proliferation and expression of a key catagen inducer, transforming growth factor beta2 (TGF-beta2), and anagen-prolonging insulin-like growth factor 1 (IGF-1) were evaluated by quantitative (immuno-) histomorphometry. Caffeine effects were further investigated in human outer root sheath keratinocytes (ORSK). RESULTS: Caffeine enhanced hair shaft elongation, prolonged anagen duration and stimulated hair matrix keratinocyte proliferation. Female HFs showed higher sensitivity to caffeine compared to male HFs. Caffeine counteracted testosterone-enhanced TGF-beta2 protein expression in male HFs. In female HFs, testosterone failed to induce TGF-beta2 expression, while caffeine reduced it. In male and female HFs, caffeine enhanced IGF-1 protein expression. In ORSK, caffeine stimulated cell proliferation, inhibited apoptosis/necrosis, up-regulated IGF-1 gene expression and protein secretion, while TGF-beta2 protein secretion was down-regulated. CONCLUSIONS: This study reveals new growth-promoting effects of caffeine on human hair follicles of both genders at different (molecular, cellular and organ) levels. This article is protected by copyright. All rights reserved

Genome-wide mapping of gene–microbiota interactions in susceptibility to autoimmune skin blistering

Susceptibility to chronic inflammatory diseases is determined by immunogenetic and environmental risk factors. Resident microbial communities often differ between healthy and diseased states, but whether these differences are of primary aetiological importance or secondary to the altered inflammatory environment remains largely unknown. Here we provide evidence for host gene–microbiota interactions contributing to disease risk in a mouse model of epidermolysis bullosa acquisita, an autoantibody-induced inflammatory skin disease. Using an advanced intercross, we identify genetic loci contributing to skin microbiota variability, susceptibility to skin blistering and their overlap. Furthermore, by treating bacterial species abundances as covariates with disease we reveal a novel disease locus. The majority of the identified covariate taxa are characterized by reduced abundance being associated with increased disease risk, providing evidence of a primary role in protection from disease. Further characterization of these putative probiotic species or species assemblages offers promising potential for preventative and therapeutic treatment development

Tight Clustering of Extracellular BP180 Epitopes Recognized by Bullous Pemphigoid Autoantibodies

Bullous pemphigoid is a blistering skin disease associated with autoantibodies against the BP180 antigen, a transmembrane component of the hemidesmosome. Anti-BP180 antibodies have been demonstrated to be pathogenic in a passive transfer mouse model. One extracellular site on human BP180 (MCW-1) was previously shown to be recognized by 50–60% of bullous pemphigoid sera. To facilitate the identification of additional autoantibody-reactive epitopes, recombinant forms of the BP180 ectodomain were generated using both bacterial and mammalian expression systems. One recombinant protein, sec180e, that was expressed in COS-1 cells and that contained the entire BP180 ectodomain, provided us with a tool to detect conformational epitopes. Bullous pemphigoid sera immuno-adsorbed against the major noncollagenous NC16A domain no longer reacted with sec180e, indicating that autoantibody reactivity to the BP180 ectodomain is restricted to the NC16A region. Immunoblot analysis of bullous pemphigoid sera immunoadsorbed with a series of recombinant NC16A peptides revealed the presence of three novel autoantigenic sites that, along with the MCW-1 epitope, are clustered within the N-terminal 45 amino acid stretch of NC16A. All 15 bullous pemphigoid sera tested reacted with a recombinant protein containing this BP180 segment. No disease-associated epitopes were detectable within the remaining 28 amino acids of NC16A. Thus, bullous pemphigoid patient autoantibodies react with a set of epitopes on the BP180 ectodomain that are highly clustered. This autoantibody-reactive region on human BP180 shows overlap with the corresponding murine BP180 site that is targeted by antibodies that are pathogenic in the mouse model of bullous pemphigoid. These findings suggest new directions for the development of diagnostic and therapeutic tools for this disease

Identification of Quantitative Trait Loci in Experimental Epidermolysis Bullosa Acquisita

Epidermolysis bullosa acquisita (EBA) is a chronic mucocutaneous autoimmune skin blistering disease. Several lines of evidence underscore the contribution of autoantibodies against type VII collagen (COL7) to the pathogenesis of EBA. Furthermore, EBA susceptibility is associated with the MHC haplotype in patients (HLA-DR2) and in immunization-induced EBA in mice (H2s). The latter study indicated an additional contribution of non-MHC genes to disease susceptibility. To identify non-MHC genes controlling EBA susceptibility, we intercrossed EBA-susceptible MRL/MpJ with EBA-resistant NZM2410/J and BXD2/TyJ as well as Cast mice. Mice of the fourth generation of this four-way autoimmune-prone advanced intercross line were immunized with a fragment of murine COL7 to induce EBA. Anti-COL7 autoantibodies were detected in 84% of mice, whereas deposition of complement at the dermal–epidermal junction (DEJ) was observed in 50% of the animals; 33% of immunized mice presented with overt clinical EBA. Onset of clinical disease was associated with several quantitative trait loci (QTLs) located on chromosomes 9, 12, 14, and 19, whereas maximum disease severity was linked to QTLs on chromosomes 1, 15, and 19. This more detailed insight into the pathogenesis of EBA may eventually lead to new treatment strategies for EBA and other autoantibody-mediated diseases

Macrophage Migration Inhibitory Factor (MIF) Drives Murine Psoriasiform Dermatitis

The immunomodulator Macrophage Migration Inhibitory Factor (MIF) exerts pleiotropic immunomodulatory activities and has been implicated in the pathogenesis of diverse inflammatory diseases. Expression levels of MIF are also significantly elevated in the skin and serum of psoriasis patients, but the pathogenic significance of MIF in psoriasis is unknown. We have therefore addressed the role of MIF in two mouse models of psoriasis, namely in the imiquimod-induced psoriasiform dermatitis (IIPD) and the IL-23-induced dermatitis model. Daily treatment with Aldara™ cream, containing imiquimod, markedly increased the abundance of MIF in the skin and generated a cellular skin expression pattern of MIF closely resembling that in human plaque psoriasis. Deficiency in MIF significantly alleviated IIPD. On the clinical level, all hallmarks of psoriasiform dermatitis, including erythema, skin infiltration, and desquamation were reduced in Mif−/− mice. On the histopathological level, MIF deficiency decreased keratinocyte hyperproliferation, inflammatory cell infiltration, specifically with respect to monocyte-derived cells, and dermal angiogenesis, suggesting that MIF may be involved in the pathogenesis of psoriasiform dermatitis through several mechanisms. Similarly, MIF deficiency also significantly reduced disease in the IL-23-induced dermatitis model, suggesting that MIF is involved in the pathogenic pathways activated by IL-23 and required to achieve full-blown psoriasiform dermatitis. Collectively, our results lend support to a possible disease-promoting role of MIF in psoriasis, which should be further investigated

Oral administration of the selective GPR120/FFA4 agonist compound A is not effective in alleviating tissue inflammation in mouse models of prototypical autoimmune diseases

ω3-polyunsaturated free fatty acids (ω3-PUFAs), particularly docosahexaenoic (DHA) and eicosapentaenoic acid (EPA), are thought to exert health promoting effects in metabolic and in inflammatory diseases. The molecular mechanisms of these beneficial effects are only partially understood. DHA and EPA activate Free Fatty Acid receptor 4 (GPR120/FFA4). Recently, the first orally available, synthetic ligand of FFA4, 3-[2-chloro-5-(trifluoromethoxy)phenyl]-3-azaspiro[5.5]undecane-9-acetic acid ("compound A"; cpd A) has been developed. Cpd A exhibits distinctly higher potency, efficiency, and selectivity at FFA4 than ω3-PUFAs and ameliorates insulin resistance and adipose tissue inflammation in the mouse. With GPR120/FFA4 activation believed to also attenuate tissue inflammation in autoimmune diseases, cpd A may also have a beneficial effect in these diseases. We have therefore addressed the therapeutic potential of cpd A in mouse models of three prototypical autoimmune diseases, specifically psoriasis, rheumatoid arthritis, and bullous pemphigoid. The effect of cpd A on the course of Aldara™-induced psoriasis-like dermatitis, K/BxN serum transfer arthritis, and antibody transfer pemphigoid disease-like dermatitis was scrutinized. Cpd A did not alter the course of Aldara-induced psoriasis-like dermatitis, K/BxN serum transfer arthritis, or antibody transfer pemphigoid disease-like dermatitis. Our results suggest that therapeutic regimens solely relying on FFA4 activation do not bear the potential to treat inflammatory diseases. With cpd A distinctly more potent in activating GPR120/FFA4 than ω3-PUFAs, this also suggests that GPR120/FFA4 activation by ω3-PUFAs does not significantly contribute to the health-promoting effects of ω3-PUFAs in autoimmune diseases

Evaluation of Nomacopan for Treatment of Bullous Pemphigoid:A Phase 2a Nonrandomized Controlled Trial

Importance: Bullous pemphigoid is a difficult-to-treat autoimmune blistering skin disease that predominantly affects older adults and is associated with an increased mortality rate. Objective: To examine the safety and therapeutic potential of nomacopan, an inhibitor of leukotriene B4and complement C5, in patients with bullous pemphigoid. Design, Setting, and Participants: This multicenter, single-group, phase 2a nonrandomized controlled trial was conducted in the dermatology departments of universities in the Netherlands and Germany. Participants were enrolled between September 2018 and April 2020. Older adult patients (aged ≥55 years) with mild to moderate, new-onset or relapsing bullous pemphigoid were recruited into the study. Interventions: Patients received nomacopan, 90 mg, subcutaneously on day 1 and 30 mg subcutaneously daily until day 42. Main Outcomes and Measures: The primary end point was the proportion of patients with grade 3 to 5 (severe) adverse events associated or possibly associated with nomacopan. Secondary end points included mean absolute and percentage changes in the Bullous Pemphigoid Disease Area Index (BPDAI) activity score, the BPDAI pruritus score, and the patient-reported outcome measures Dermatology Life Quality Index (DLQI) and Treatment of Autoimmune Bullous Disease Quality of Life (TABQOL). Results: A total of 9 patients (median [range] age, 75 [55-85] years) with bullous pemphigoid were included in the trial, of whom 5 were women (55.6%). No serious adverse events associated with nomacopan were found. The mean (90% CI) BPDAI activity score decreased from 32.0 (8.7) points on day 1 to 19.6 (9.0) points on day 42. Seven of 9 patients (77.8%) responded to nomacopan with a reduction in the BPDAI activity score of at least 8 points between days 1 and 42; in 3 responders, the reduction was 80% or greater. On day 42, the mean (90% CI) BPDAI pruritus score had decreased by 6.8 (4.6) points from 17.6 (4.0) points on day 1. The mean (90% CI) DLQI score decreased from 11.3 (4.2) points at baseline to 6.4 (3.8) points by day 42, and the mean (90% CI) TABQOL score decreased from 14.6 (5.4) points at baseline to 10.3 (5.0) points on day 42. Conclusions and Relevance: Results of this nonrandomized controlled trial suggest that nomacopan can be well tolerated in older patients with bullous pemphigoid and may have therapeutic benefits for suppressing acute flares of this disease. A larger, placebo-controlled randomized clinical trial is warranted to confirm this safety profile and to establish nomacopan as a new therapeutic option for bullous pemphigoid. Trial Registration: ClinicalTrials.gov Identifier: NCT04035733

Inhibition of phosphodiesterase-4 significantly decreases oral mucosa lesions in experimental anti-laminin 332 mucous membrane pemphigoid

Mucous membrane pemphigoid (MMP) is characterized by autoantibodies against the dermal-epidermal-junction and a mucosal disease predominating over skin involvement. As the treatment of MMP patients still relies on high-dose corticosteroids, there is an unmet need for new and more specific therapies. Here, we made use of a recently established experimental model that recapitulated major clinical and immunopathological characteristics of human MMP by the injection of IgG against the murine alpha3 chain of laminin 332 (Lam332) into adult mice. In a prophylactic approach, the specific PDE4 inhibitor roflumilast (ROF) led in 2 independent, blinded experiments, to the reduction of oral lesions compared to vehicle-treated mice as quantified by endoscopy (p¼0.029). In contrast, an increase in skin lesions was observed in ROF-treated mice (p<0.0001). In a quasi-therapeutic approach, i.e. when ROF/vehicle was not used until mice had developed first skin lesions, ROF-treated mice showed significantly less oral lesions compared to vehicle-treated mice, while skin lesions did not differ. To investigate the discrepant effect of ROF on oral and skin lesions, a transcriptome analysis of both tissues in ROF- and vehicle-treated anti-Lam332 MMP mice as well as mice injected with normal rabbit IgG was performed. An up-regulation of IL-6 and an impact of CXCL2 were found by Gene Set Enrichment and STRING analysis, respectively, in both the skin and buccal mucosa of vehicle-treated mice. The subsequent incubation of murine keratinocytes with anti-mLam332 IgG resulted in a dose-dependent release of IL-6 and CXCL2, which was inhibited by the addition of ROF. Our data propose IL-6 and CXCL2 as relevant pathogenic factors in MMP and suggest PDE4 inhibition as potential novel treatment options for MMP patients with severe oral lesions