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Institutional and individual stigmatization represent major barriers that prevent children with disabilities from accessing education. It can be presumed that children with disabilities are labeled as such even in inclusive educational settings and that teachers' attitudes toward inclusive education and children with disabilities play a crucial role in this context. Against this background, the present study aims to (a) apply and conceptualize the mixed-blessings model in the context of stigma-related reactions to children's disability labels in inclusive education and (b) shed light on the causal attributions of teachers that underlie stigma-related attitudes toward children with various disabilities. A 3 × 2 × 2 × 2 × 2 online experiment examined the ways in which disability-specific causes and symptoms, the type of disability in question, the children's sex, and efficacy cues regarding educational efforts affect future teachers' attitudes toward and expectations of inclusive education as well as their social distance toward children with disabilities. The participants in this experiment were N  = 605 German student and trainee teachers representing different types of teaching professions. A multivariate analysis of variance (MANOVA) revealed that, in particular, the cause attributed to the disability, the depicted type of disability and the probability of learning success led to changes in attitudes. Respondents' teaching self-efficacy and their status as students or trainees emerged as moderators of the effect of pupils' type of disability. As a result, teacher education and training as well as communication regarding pupils with disabilities require a high degree of sensitivity to disability-specific and efficacy-related cues to prevent (accidental) professional or institutional stigmatization., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Röhm, Grengel, Möhring, Zensen-Möhring, Nellen and Hastall.)

Background: Forkhead-Box-Protein P3 (FoxP3) is a transcription factor and marker of regulatory T cells, converting naive T cells into Tregs that can downregulate the effector function of other T cells. We previously detected the expression of FoxP3 in retinal pigment epithelial (RPE) cells, forming the outer blood-retina barrier of the immune privileged eye., Methods: We investigated the expression, subcellular localization, and phosphorylation of FoxP3 in RPE cells in vivo and in vitro after treatment with various stressors including age, retinal laser burn, autoimmune inflammation, exposure to cigarette smoke, in addition of IL-1β and mechanical cell monolayer destruction. Eye tissue from humans, mouse models of retinal degeneration and rats, and ARPE-19, a human RPE cell line for in vitro experiments, underwent immunohistochemical, immunofluorescence staining, and PCR or immunoblot analysis to determine the intracellular localization and phosphorylation of FoxP3. Cytokine expression of stressed cultured RPE cells was investigated by multiplex bead analysis. Depletion of the FoxP3 gene was performed with CRISPR/Cas9 editing., Results: RPE in vivo displayed increased nuclear FoxP3-expression with increases in age and inflammation, long-term exposure of mice to cigarette smoke, or after laser burn injury. The human RPE cell line ARPE-19 constitutively expressed nuclear FoxP3 under non-confluent culture conditions, representing a regulatory phenotype under chronic stress. Confluently grown cells expressed cytosolic FoxP3 that was translocated to the nucleus after treatment with IL-1β to imitate activated macrophages or after mechanical destruction of the monolayer. Moreover, with depletion of FoxP3, but not of a control gene, by CRISPR/Cas9 gene editing decreased stress resistance of RPE cells., Conclusion: Our data suggest that FoxP3 is upregulated by age and under cellular stress and might be important for RPE function., (© 2022. The Author(s).)

Objectives: AAV vectors are widely used in gene therapy, but the prevalence of neutralizing antibodies raised against AAV serotypes in the course of a natural infection, as well as innate and adaptive immune responses induced upon vector administration, is still considered an important limitation. In ocular gene therapy, vectors applied subretinally bear the risk of retinal detachment or vascular leakage. Therefore, new AAV vectors that are suitable for intravitreal administration for photoreceptor transduction were developed., Methods: Here, we compared human immune responses from donors with suspected previous AAV2 infections to the new vectors AAV2.GL and AAV2.NN-two capsid peptide display variants with an enhanced tropism for photoreceptors-with the parental serotype AAV2 (AAV2 WT). We investigated total and neutralizing antibodies, adaptive and innate cellular immunogenicity determined by immunofluorescence staining and flow cytometry, and cytokine secretion analyzed with multiplex beads., Results: While we did not observe obvious differences in overall antibody binding, variants-particularly AAV2.GL-were less sensitive to neutralizing antibodies than the AAV2 WT. The novel variants did not differ from AAV2 WT in cellular immune responses and cytokine production in vitro., Conclusion: Due to their enhanced retinal tropism, which allows for dose reduction, the new vector variants are likely to be less immunogenic for gene therapy than the parental AAV2 vector.

Molecular or antigenic mimicry is a term for the similarity of different antigens, which can be confused by the immune system. Antigen recognition by antibodies and T cell receptors is specific, but not restricted to a single antigen. Both types of receptors specifically recognize antigens and are expressed with a very high but still restricted variability compared to the number of different antigens they potentially could bind. T cell receptors only can bind to antigen peptides presented on certain self-MHC-molecules by screening only some amino acid side chains on both the presented peptides and the MHC molecule. The other amino acids of the peptide are not directly perceived by the T cell, offering the opportunity for a single T cell to recognize a variety of different antigens with the same receptor, which significantly increases the immune repertoire. The immune system is usually tolerant to autoantigens, especially to those of immune privileged sites, like the eye. Therefore, autoimmune diseases targeting these organs were hard to explain, unless a T cell is activated by an environmental peptide (e.g. pathogen) that is similar, but not necessarily identical with an autoantigen. Here we describe antigenic mimicry of retinal autoantigens with a variety of non-ocular antigens resulting in the induction of intraocular inflammation. T cells that are activated by mimotopes outside of the eye can pass the blood-retina barrier and enter ocular tissues. When reactivated in the eye by crossreaction with autoantigens they induce uveitis by recruiting inflammatory cells., (Copyright © 2020 Wildner and Diedrichs-Möhring.)

Autoimmune uveitis is a sight-threatening, rare disease, potentially leading to blindness. Uveitis is a synonym for intraocular inflammation, presenting as various clinical phenotypes with different underlying immune responses in patients, whereas different animal models usually represent one certain clinical and immunological type of uveitis due to genetic uniformity and the method of disease induction. T cells recognizing intraocular antigens initiate the disease, recruiting inflammatory cells (granulocytes, monocytes/macrophages) to the eyes, which cause the damage of the tissue. The treatment of uveitis so far aims at downregulation of inflammation to protect the ocular tissues from damage, and at immunosuppression to stop fueling T cell reactivity. Uveitis is usually prevented by specific mechanisms of the ocular immune privilege and the blood-eye-barriers, but once the disease is induced, mechanisms of the immune privilege as well as a variety of novel regulatory features including new Treg cell populations and suppressive cytokines are induced to downregulate the ocular inflammation and T cell responses and to avoid relapses and chronicity. Here we describe mechanisms of regulation observed in experimental animal models as well as detected in studies with peripheral lymphocytes from patients.

Purpose: IFN-α is the only treatment capable of inducing long-term remission in some patients with ocular Behçet's disease. In this review, we focus on immune mechanisms of IFN-α in animal models and patients and compare the outcome of different clinical studies., Methods: Review of literature using PubMed and Google and original data from rat models with monophasic/chronic or relapsing experimental autoimmune uveitis treated with IFN-α., Results: The role of IFN-α and its effect on various cell types were investigated, in some cases with contradictory results. Some patients respond very well to IFN-α treatment, while others are non-responders, which was reflected in the uveitis rat models: relapsing uveitis was ameliorated, the monophasic/chronic disease even aggravated., Conclusions: Despite intensive investigations in patients and animal models, the immune mechanisms explaining the therapeutic effect of IFN-α in ocular Behçet's disease are not yet fully understood and need further investigation.

Autoimmune diseases usually follow a relapsing-remitting or a chronic progressive course. To understand the underlying immunopathogenesis we investigated experimental Lewis rat models displaying both disease types, which were only dependent on the autoantigen peptide used for immunization. Retinal S-Antigen-peptide PDSAg induces chronic, monophasic disease, whilst interphotoreceptor retinoid-binding protein (IRBP)-peptide R14 causes a spontaneously relapsing-remitting course. R14-mediated uveitis can be re-induced by immunization; PDSAg-induced disease is even preventable by prior CFA-injection. T cells with different antigen specificities preferentially infiltrate the eyes from different sites, e.g. choroid or retinal vessels, they remain in the retina after resolution of inflammation for many weeks. The major inflammatory cell populations in the eyes during rat uveitis are CD4 + or CD8 + monocytes/macrophages. Chemokine mutants only suppress PDSAg-mediated EAU, while IFN-α-treatment ameliorated R14-, but worsened PDSAg-induced disease. Comparison of T cells revealed upregulated expression of 26 genes related to various signal transduction pathways upstream and downstream of IFN-γ only in T cells causing relapsing EAU. Intraocular injection of IFN-γ induces synchronized relapses in R14-mediated uveitis, while VEGF-expression of PDSAg-specific T cells causing chronic disease induced chorioretinal neovascularization that is suppressed by anti-CD146 antibody. Intraocular T cells from rat eyes during EAU express IL-17, IFN-γ or IL-10, with dynamic changes of the cell populations during the disease course, differing in both disease types. Immunization of animals with a mixture of both antigens suppressed relapses, indicating a dominance of the monophasic disease. Understanding the exact pathogenesis of both disease courses is key to developing novel therapies for autoimmune diseases., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Background: Ischemic-reperfusion (IR) injury still represents a major concern in clinical transplantation, especially in the era of extreme organ shortage and extended criteria donor organs. In the present study we aimed to investigate the hepatoprotective effects of remote ischemic conditioning (RIC) in a rat model of arterialized orthotopic liver transplantation (OLT)., Methods: Male Lewis rats were used (n = 144 / 72 OLT cases; 240-340g) as donors and recipients. Livers were flushed and stored in 4°C HTK-solution for 8h before implantation. Recipients were randomly allocated into three experimental groups: RIC 1, RIC 2, Control. In RIC 1, RIC 2 groups, RIC was applied in the recipient before hepatectomy or after reperfusion (4x5-5min IR via clamping the infrarenal aorta), respectively. Animals were sacrificed at 1, 3, 24, 168h post-reperfusion (n = 6 recipient/group/time point). Hepatocellular injury, graft circulation, serum cytokines, tissue redox-stress and adenosine-triphosphate (ATP) levels have been assessed. Additional markers were analyzed, using Western blotting and reverse-transcription polymerase chain reaction., Results: RIC 1 group showed significantly (p<0.05) improved portal venous and microcirculation flow as well as velocity. RIC has significantly reduced tissue injury according to the serum levels of transaminases and results of histopathological evaluation. Reduced TUNEL-staining (p<0.01 RIC 1-2 vs. Control) and elevated pBAD/BAD ratio was detected in the RIC groups (p<0.01 RIC 1 vs. Control). Supporting findings were obtained from measurements of serum IL-10 as well as tissue malondialdehyde and ATP levels. Hemoxygenase-1 (HO-1) mRNA-expression was significantly higher in RIC 1 compared to Control (p<0.05 RIC 1 vs. Control)., Conclusion: These results suggest that RIC might confer potent protection against the detrimental effects of IR injury including tissue damage, apoptosis, graft circulation, inflammation, tissue energetic status in OLT. HO-1 overexpression might play an orchestrating role in RIC mediated organ protection. An earlier intervention (RIC 1 protocol) was more effective than remote conditioning after graft reperfusion.

Background: Uveitis is a potentially blinding inflammatory disease of the inner eye with a high unmet need for new therapeutic interventions. Here, we wanted to investigate the suppressive effect of the intraocular application of the small molecule dihydroorotate dehydrogenase (DHODH)-inhibitor PP-001 on experimental relapsing rat uveitis and furthermore determine its effect on proliferation and cytokine secretion of human peripheral blood lymphocytes (PBL) and human retinal pigment epithelial (RPE) cells in vitro., Methods: Spontaneously relapsing uveitis was induced in rats by immunization with interphotoreceptor retinoid-binding protein (IRBP) peptide R14. PP-001 was injected intravitreally after resolution of the primary disease to investigate further relapses. Proliferation and metabolic activity of phytohemagglutinin (PHA)-stimulated human peripheral lymphocytes with and without PP-001 and cytokine secretion were determined by XTT assay and bioplex bead assay. The RPE cell line ARPE-19 as well as primary human RPE cells treated with PP-001 or anti-vascular endothelial growth factor (VEGF) antibody bevacizumab were also investigated for metabolic activity and cytokine/chemokine secretion., Results: Injection of PP-001 into rat eyes reduced the number of relapses by 70%, from 20 relapses (57% of the rats affected) in the control group to 6 relapses (33% of the rats) in the treatment group. In human PBL cultures, PP-001 reduced the proliferation in a dose-dependent manner. The secretion of several cytokines such as IL-17, IFN-γ, and VEGF was suppressed by PP-001, as previously observed with rat T cells in the experimental autoimmune uveitis (EAU) model. In contrast, human RPE cells were not affected by PP-001, while the anti-VEGF antibody bevacizumab severely impaired the secretion of various cytokines including VEGF., Conclusions: For the first time, intravitreal injection of PP-001 demonstrated an effective, but transient reduction of relapses in the rat EAU model. In vitro PP-001 suppressed proliferation and cytokine/chemokine secretion of human lymphocytes, while neither human RPE cell line ARPE-19 nor primary RPE cells were affected.

Purpose: The retinal pigment epithelium (RPE) is a main target for complement activation in age-related macular degeneration (AMD). The anaphylatoxins C3a and C5a have been thought to mostly play a role as chemoattractants for macrophages and immune cells; here, we explore whether they trigger RPE alterations. Specifically, we investigated the RPE as a potential immunoregulatory gate, allowing for active changes in the RPE microenvironment in response to complement., Design: In vitro and in vivo analysis of signaling pathways., Methods: Individual activities of and interaction between the two anaphylatoxin receptors were tested in cultured RPE cells by fluorescence microscopy, western blot, and immunohistochemistry., Main Outcome Measures: Intracellular free calcium, protein phosphorylation, immunostaining of tissues/cells, and multiplex secretion assay., Results: Similar to immune cells, anaphylatoxin exposure resulted in increases in free cytosolic Ca 2+ , PI3-kinase/Akt activation, FoxP3 and FOXO1 phosphorylation, and cytokine/chemokine secretion. Differential responses were elicited depending on whether C3a and C5a were co-administered or applied consecutively, and response amplitudes in co-administration experiments ranged from additive to driven by C5a (C3a + C5a = C5a) or being smaller than those elicited by C3a alone (C3a + C5a < C3a)., Conclusion: We suggest that this combination of integrative signaling between C3aR and C5aR helps the RPE to precisely adopt its immune regulatory function. These data further contribute to our understanding of AMD pathophysiology.

The pro-inflammatory chemokine interleukin-8 (CXCL8) exerts its function by establishing a chemotactic gradient in infected or damaged tissues to guide neutrophil granulocytes to the site of inflammation via its G protein-coupled receptors (GPCRs) CXCR1 and CXCR2 located on neutrophils. Endothelial glycosaminoglycans (GAGs) have been proposed to support the chemotactic gradient formation and thus the inflammatory response by presenting the chemokine to approaching leukocytes. In this study, we show that neutrophil transmigration in vitro can be reduced by adding soluble GAGs and that this process is specific with respect to the nature of the glycan. To further investigate the GAG influence on neutrophil migration, we have used an engineered CXCL8 mutant protein (termed PA401) which exhibits a much higher affinity towards GAGs and an impaired GPCR activity. This dominant-negative mutant chemokine showed anti-inflammatory activity in various animal models of neutrophil-driven inflammation, i.e. in urinary tract infection, bleomycin-induced lung fibrosis, and experimental autoimmune uveitis. In all cases, treatment with PA401 resulted in a strong reduction of transmigrated inflammatory cells which became evident from histology sections and bronchoalveolar lavage. Since our CXCL8-based decoy targets GAGs and not GPCRs, our results show for the first time the crucial involvement of this glycan class in CXCL8/neutrophil-mediated inflammation and will thus pave the way to novel approaches of anti-inflammatory treatment., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Understanding the immunopathogenesis of autoimmune and inflammatory diseases is a prerequisite for specific and effective therapeutical intervention. This review focuses on animal models of two common ocular inflammatory diseases, dry eye disease (DED), affecting the ocular surface, and uveitis with inflammation of the inner eye. In both diseases autoimmunity plays an important role, in idiopathic uveitis immune reactivity to intraocular autoantigens is pivotal, while in dry eye disease autoimmunity seems to play a role in one subtype of disease, Sjögren' syndrome (SjS). Comparing the immune mechanisms underlying both eye diseases reveals similarities, and significant differences. Studies have shown genetic predispositions, T and B cell involvement, cytokine and chemokine signatures and signaling pathways as well as environmental influences in both DED and uveitis. Uveitis and DED are heterogeneous diseases and there is no single animal model, which adequately represents both diseases. However, there is evidence to suggest that certain T cell-targeting therapies can be used to treat both, dry eye disease and uveitis. Animal models are essential to autoimmunity research, from the basic understanding of immune mechanisms to the pre-clinical testing of potential new therapies., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Antigen-specific tolerance induction is a desired therapy for uveitis patients. Our relapsing-remitting rat model of experimental autoimmune uveitis (EAU) induced with IRBP peptide R14 enables us to test the effect of oral tolerance on the prevention of relapsing uveitis. We investigated several peptides overlapping the sequence of R14 for prevention and different doses of R14 for therapy to determine the tolerogenic epitope and the most effective therapeutic regimen for uveitis. Lewis rats were immunized with R14-CFA to induce EAU. Oral tolerance was induced prior to immunization (prevention) or after onset of EAU to prevent relapses (therapy). Therapeutic feeding was performed with high and/or low doses of oral antigen for clonal deletion of effector and induction of regulatory T cells. Uveitis was determined clinically and histologically; mesenteric lymph node (mLN) cells of tolerized rats were tested for surface markers, cytokines and Foxp3 expression. Preventive feeding of R14 and its major epitope R16, but none of the overlapping peptides significantly suppressed EAU and also prevented relapses, irrespective of their pathogenicity. Therapeutic feeding with R14 dramatically reduced relapses, while only the consecutive feeding of high and low-dose R14 had an ameliorating effect on the first course of disease. IL-10-producing T cells from mLN decreased after oral tolerization, and with R14-stimulation in vitro the TCRαβ+/Foxp3+ population increased in the low-dose fed group. No mLN population could be clearly assigned to successful oral tolerance induction during active autoimmune uveitis., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Purpose: We investigated the effect of PP-001, a new small molecule inhibitor of dihydro-orotate dehydrogenase in two experimental rat experimental autoimmune uveitis (EAU) models: a spontaneously relapsing-remitting model and a monophasic/chronic disease model that results in late chorioretinal neovascularization. Both of the diseases are induced by immunization with autoantigen peptides., Methods: Prevention was tested using daily oral applications of PP-001 after immunization with the retinal S-antigen peptide PDSAg (for induction of monophasic uveitis and neovascularization) or the interphotoreceptor retinoid-binding protein peptide R14 (for induction of spontaneously relapsing-remitting EAU). Treatment to inhibit relapses and neovascularization was tested using PP-001 daily after the first attack of R14-induced or after onset of PDSAg-induced EAU. Uveitis was graded clinically and histologically. The effect of PP-001 on cytokine secretion and proliferation was evaluated using rat T-cell lines., Results: Preventive feeding of PP-001 abrogated both types of EAU. Starting treatment after the resolution of the first attack led to a significant reduction of the number and intensity of relapses in R14-induced EAU. PP-001-treatment initiated after onset or after peak of PDSAg-induced EAU significantly reduced neovascularization (as determined by histology). Proliferation of antigen-specific T-cell lines and secretion of IFN-γ, IL-17, IL-10, IP-10, and VEGF were efficiently suppressed by PP-001., Conclusions: We investigated a new dihydroorotate dehydrogenase inhibitor as treatment for primary and recurrent disease in relapsing-remitting and chronic rat models of experimental autoimmune uveitis. The small molecule compound PP-001 suppressed proliferation and cytokine secretion of autoreactive T cells (i.e., IFN-g, IL-17, and VEGF) and chorioretinal neovascularization in chronic EAU., (Copyright 2015 The Association for Research in Vision and Ophthalmology, Inc.)

A major limitation of most animal models of autoimmune diseases is that they do not reproduce the chronic or relapsing-remitting pattern characteristic of many human autoimmune diseases. This problem has been overcome in our rat models of experimentally induced monophasic or relapsing-remitting autoimmune uveitis (EAU), which depend on the inducing antigen peptides from retinal S-Antigen (monophasic EAU) or interphotoreceptor retinoid-binding protein (relapsing EAU). These models enable us to compare autoreactive and regulatory T cell populations. Intraocular, but not peripheral T cells differ in their cytokine profiles (IFN-γ, IL-17 and IL-10) at distinct time points during monophasic or relapsing EAU. Only intraocular T cells concomitantly produced IFN-γ, IL-17 and/or IL-10. Monophasic EAU presented rising numbers of cells expressing IFN-γ and IL-17 (Th1/Th17) and cells expressing IL-10 or Foxp3. During relapsing uveitis an increase of intraocular IFN-γ+ cells and a concomitant decrease of IL-17+ cells was detected, while IL-10+ populations remained stable. Foxp3+ cells and cells expressing IL-10, even in combination with IFN-γ or IL-17, increased during the resolution of monophasic EAU, suggesting a regulatory role for these T cells. In general, cells producing multiple cytokines increased in monophasic and decreased in relapsing EAU. The distinct appearance of certain intraocular populations with characteristics of regulatory cells points to a differential influence of the ocular environment on T cells that induce acute and monophasic or relapsing disease. Here we provide evidence that different autoantigens can elicit distinct and differently regulated immune responses. IFN-γ, but not IL-17 seems to be the key player in relapsing-remitting uveitis, as shown by increased, synchronized relapses after intraocular application of IFN-γ. We demonstrated dynamic changes of the cytokine pattern during monophasic and relapsing-remitting disease with strongly increasing IL-10 expression in intraocular T cells during monophasic uveitis.