Genetic landscape and autoimmunity of monocytes in developing Vogt-Koyanagi-Harada disease.

Vogt-Koyanagi-Harada (VKH) disease is a systemic autoimmune disorder affecting multiple organs, including eyes, skin, and central nervous system. It is known that monocytes significantly contribute to the development of autoimmune disease. However, the subset heterogeneity with unique functions and signatures in human circulating monocytes and the identity of disease-specific monocytic populations remain largely unknown. Here, we employed an advanced single-cell RNA sequencing technology to systematically analyze 11,259 human circulating monocytes and genetically defined their subpopulations. We constructed a precise atlas of human blood monocytes, identified six subpopulations-including S100A12, HLA, CD16, proinflammatory, megakaryocyte-like, and NK-like monocyte subsets-and uncovered two previously unidentified subsets: HLA and megakaryocyte-like monocyte subsets. Relative to healthy individuals, cellular composition, gene expression signatures, and activation states were markedly alternated in VKH patients utilizing cell type-specific programs, especially the CD16 and proinflammatory monocyte subpopulations. Notably, we discovered a disease-relevant subgroup, proinflammatory monocytes, which showed a discriminative gene expression signature indicative of inflammation, antiviral activity, and pathologic activation, and converted into a pathologic activation state implicating the active inflammation during VKH disease. Additionally, we found the cell type-specific transcriptional signature of proinflammatory monocytes, ISG15, whose production might reflect the treatment response. Taken together, in this study, we present discoveries on accurate classification, molecular markers, and signaling pathways for VKH disease-associated monocytes. Therapeutically targeting this proinflammatory monocyte subpopulation would provide an attractive approach for treating VKH, as well as other autoimmune diseases.

Vogt-Koyanagi-Harada patients show higher frequencies of circulating NKG2Dpos NK and NK T cells.

Vogt-Koyanagi-Harada (VKH) is an autoimmune disease characterized by inflammation in tissues that contain melanocytes. We aimed to increase the knowledge regarding immunological pathways deregulated in VKH disease. We compared the percentages of circulating natural killer (NK), NK T and T cells expressing the activatory markers: CD16, CD69, NK group 2D (NKG2D), natural cytotoxicity triggering receptor 3 (Nkp30), natural cytotoxicity triggering receptor 1 (Nkp46) and the inhibitory marker: NK group 2 member A (NKG2A) in 10 active VKH patients, 20 control subjects (CTR) and seven patients with Behçet disease (BD) by flow cytometry. Cytotoxic potential of NK cells was determined through the degranulation marker CD107a expression after contact with K562 cells by flow cytometry. Moreover, plasmatic levels of 27 cytokines were determined with a multiplex bead-based assay. VKH patients showed higher percentages of NKG2Dpos NK and NK T cells versus CTR. The cytotoxic potential of NK cells induced by K562 cells was comparable between VKH patients and CTR. Finally, higher concentrations of interleukin (IL)-4, IL-5, IL-7, IL-17 and platelet-derived growth factor-subunits B (PDGF-bb) were detected in plasma of VKH patients versus CTR. The immune profile of VKH patients was similar to that of BD patients.

Identification of Ribosomal Protein S4, Y-Linked 1 as a cyclosporin A plus corticosteroid resistance gene.

Combination of corticosteroids (CS) with cyclosporin A (CsA) is widely used in the treatment of autoimmune diseases, autoinflammatory diseases and transplantation rejection. However, some patients fail to respond or develop resistance to the combination regimen. In Vogt-Koyanagi-Harada (VKH) disease model, we performed RNA sequencing (RNA-seq) based transcriptomics, isobaric tags for relative and absolute quantification (iTRAQ) based proteomics and assays in vitro to screen and validate potential resistant molecules. We found that a total of 1697 differentially expressed genes (DEGs) and 21 differentially expressed proteins (DEPs) in CD4+ T cells between CsA & CS-resistant and -sensitive VKH patients. Ribosomal Protein S4, Y-Linked 1 (RPS4Y1) was verified to regulate the resistance of CD4+ T cells from male VKH patients to CsA & CS. Importantly, we showed that chlorambucil (CLB) could reverse the resistance by RPS4Y1 suppression. Taken together, we identify RPS4Y1 as an important CsA & CS resistance gene in VKH disease. Researchers should consider validating the resistant effect of RPS4Y1 in other autoimmune diseases or organ transplantation.

Comparison of clinical characteristics in patients with Vogt-Koyanagi-Harada disease with and without anti-retinal antibodies.

PURPOSE: To compare the clinical characteristics of Vogt-Koyanagi-Harada (VKH) disease patients with and without anti-retinal antibodies (ARAs) that are frequently detected in autoimmune retinopathy. METHODS: Using immunoblot analyses, serum autoantibodies for recoverin, carbonic anhydrase II, and α-enolase were examined in 20 treatment-naïve patients with VKH disease. Clinical factors before and after systemic corticosteroid therapy, including best-corrected visual acuity (BCVA) and macular outer retinal morphology, were statistically compared between patients with VKH disease with and without ARAs. RESULTS: Serum ARAs were detected in 50.0% of patients with VKH disease. There were no significant differences in clinical factors between the two groups, including final BCVA, frequency of uveitis recurrence, and recovery of the macular ellipsoid zone after systemic corticosteroid therapy. CONCLUSIONS: Our results suggest that the detected ARAs did not influence visual outcomes, the chronicity of uveitis, or outer retinal morphology in patients with VKH disease.

Interleukin-10 gene polymorphisms are associated with Behcet's disease but not with Vogt-Koyanagi-Harada syndrome in the Chinese Han population.

PURPOSE: This study aimed to investigate the association of interleukin (IL)-10 gene polymorphisms with Behcet's disease (BD) and Vogt-Koyanagi-Harada (VKH) syndrome in the Chinese Han population. METHODS: A two-stage association study was performed on 718 BD patients, 300 VKH patients, and 1,753 controls. Genotyping of the IL-10 gene was performed for six single nucleotide polymorphisms (SNPs), including rs1800871, rs1800872, rs1800896, rs3021094, rs3790622, and rs1554286 using PCR-restricted fragment length polymorphism or TaqMan SNP assays. Real-time PCR was performed to test the IL-10 mRNA expression of the associated polymorphisms. RESULTS: The first-stage result showed significantly increased frequencies of the rs1800871 T allele, rs1800872 A allele, and rs1554286 T allele in BD patients compared with controls (Pcorrected (Pcorr)=1.82×10(-5), OR=1.837; Pcorr=6.1×10(-5), OR=1.780; Pcorr=3.15×10(-5), OR=1.794, respectively). There was no association of the tested six SNPs with VKH syndrome. A second-stage study was therefore performed in BD patients to validate the result of the first stage, showing a significantly increased frequency of the rs1800871 T allele (Second stage, Pcorr=5.59×10(-5), OR=1.493; Combined data, Pcorr=3.65×10(-11), OR=1.632). Compared to the controls, an increased frequency of the rs1800871 T allele was observed in BD patients with extraocular findings, including genital ulcers, skin lesions, and a positive pathergy test. No difference was found among the mRNA expressions of IL-10 in the peripheral blood mononuclear cells (PBMCs) of controls with different genotypes of rs1800871 after stimulation of lipopolysaccharide (LPS) or anti-CD3/CD28 antibodies. CONCLUSIONS: The findings showed that IL-10 is a risk gene for BD but not for VKH syndrome.

Vogt-Koyanagi-Harada disease.

PURPOSE OF REVIEW: The purpose of this article is to review the current literature on Vogt-Koyanagi-Harada (VKH) disease, including current treatment options and new research directions. RECENT FINDINGS: Recent publications on VKH disease show an increased focus on the immunogenetics and immune pathways associated with the development of VKH disease. There have also been advances in imaging modalities and techniques that may help to better elucidate the disease process in eyes with VKH disease. SUMMARY: VKH disease is an autoimmune, multisystem inflammatory disorder, the cause of which is still incompletely understood. Continued research may elucidate the causes and triggers of immune dysregulation in this disease, and in doing so, identify novel therapeutic targets.

Absence of recognition of common melanocytic antigens by T cells isolated from the cerebrospinal fluid of a Vogt-Koyanagi-Harada patient.

PURPOSE: Vogt-Koyanagi-Harada (VKH) syndrome is an autoimmune disease characterized by inaugural uveomeningitidis and hearing loss and at late stages a depigmentation in eyes and skin. Melanocytes are the cells common to the four affected tissues, namely eye, brain, inner ear, and skin. Melanocytes are therefore considered as the source of self-antigens. The melanocytic proteins tyrosinase-related protein-1 (TRP1), TRP2, tyrosinase, and gp100 have been proposed as the proteins targeted by autoreactive T cells from VKH patients bearing human leukocyte antigen (HLA)-DRB1*04:05, the HLA allele classically associated with VKH disease. The objective of this work was to determine the antigens recognized by a large number of potentially autoreactive CD4 T lymphocytes obtained from the cerebrospinal fluid of one VKH patient who did not express HLA-DRB1*04:05. METHODS: T cells were isolated from the cerebrospinal fluid of a newly diagnosed HLA-DRB1*14:01,*15:03;-DPB1*01:01,*04:02 patient in the acute phase of the VKH disease and cloned by limiting dilution. Each of the 107 T cell clones, of which 90% were CD4(+), was tested for its ability to secrete cytokines upon contact with autologous antigen-presenting cells loaded with either of the melanocytic proteins TRP1, TRP2, tyrosinase, gp100, Melan-A and KU-MEL-1. The sensitivity of our recombinant bacteria-based approach was validated with a CD4 T cell clone with known antigen specificity. The ability of each of the 107 clones to secrete cytokines upon nonspecific stimulation was verified. RESULTS: None of the 107 T cell clones was able to secrete tumor necrosis factor-α, interferon-γ, interleukin (IL)-5, or IL-17 upon contact with autologous B cells loaded with any of the six common melanocytic proteins. Nine clones secreted high-level IL-17 upon stimulation with beads coated with antibodies. CONCLUSIONS: The self-antigens that triggered the VKH disease in this patient probably derive from proteins other than the six melanocytic proteins mentioned above. Further study of antigens that are recognized by potential autoreactive T cells from VKH patients is likely to benefit from testing a broader set of melanocytic proteins.

Noninfectious immune-mediated uveitis and ocular inflammation.

Noninfectious uveitis encompasses a diverse group of ocular inflammatory disorders that share an underlying immune etiology and may be associated with systemic disease or confined primarily to the eye. Uveitis is commonly classified by anatomical location of inflammation into anterior, intermediate, posterior, and panuveitis. The treatment of noninfectious uveitis consists of corticosteroids, immunosuppressive agents, and surgically placed steroid implants. We review the epidemiology, immunopathology, and clinical features of several noninfectious immune-mediated uveitides, including HLA-B27 acute anterior uveitis, juvenile idiopathic arthritis, intermediate uveitis, sarcoidosis, Behcet's disease, Vogt-Koyanagi-Harada syndrome, sympathetic ophthalmia, and white dot syndromes. We also discuss the stepwise approach to medical treatment of immune-mediated uveitis as well as the characteristics, safety, and efficacy of immunosuppressive agents used to treat ocular inflammatory disease.

CD40 gene polymorphisms confer risk of Behcet's disease but not of Vogt-Koyanagi-Harada syndrome in a Han Chinese population.

OBJECTIVES: Recent genetic surveys including a genome-wide association study have identified CD40 as a susceptibility gene for several autoimmune diseases. This study was designed to investigate the association of CD40 gene polymorphisms with Behçet's disease (BD) and Vogt-Koyanagi-Harada (VKH) syndrome in a Han Chinese population. METHODS: Two single nucleotide polymorphisms (SNPs), rs4810485 and rs1883832, were genotyped using polymerase chain reaction-restriction fragment length polymorphism in 373 BD patients, 519 VKH patients and 402 controls. A binary logistic regression analysis was applied to assess the influence of gender on the association of CD40 polymorphisms with BD. RESULTS: Our results showed significantly increased frequencies of the homozygous rs4810485 TT and rs1883832 TT in BD patients as compared with the controls [Bonferroni-corrected P-value for gender adjustment (Pac) = 0.006, odds ratio (OR) = 1.98, 95% confidence interval (CI) 1.38, 2.83; = 0.012, OR = 1.73, 95% CI 1.22, 2.46, respectively]. A markedly decreased frequency of the heterozygous rs4810485 GT was observed in BD patients as compared with the controls ( = 0.042, OR = 0.68, 95% CI 0.51, 0.90). The genotype and allele frequencies of rs4810485 and rs1883832 were not different between VKH patients and controls. Stratification analysis did not find any association between the tested SNPs and extra-ocular manifestations of both diseases. CONCLUSIONS: The results suggest that TT genotypes of rs4810485 and rs1883832 may be predisposing genotypes for BD, and that the rs4810485 GT genotype may be a protective genotype for BD. The two tested CD40 gene polymorphisms are not associated with VKH syndrome in the investigated Han Chinese population.

Progranulin Suppressed Autoimmune Uveitis and Autoimmune Neuroinflammation by Inhibiting Th1/Th17 Cells and Promoting Treg Cells and M2 Macrophages.

BACKGROUND AND OBJECTIVES: Progranulin (PGRN) is an important immune regulatory molecule in several immune-mediated diseases. The objective of this study is to investigate the role of PGRN in uveitis and its counterpart, experimental autoimmune uveitis (EAU), and experimental autoimmune encephalomyelitis (EAE). METHODS: Serum PGRN levels in patients with Behcet disease (BD) or Vogt-Koyanagi-Harada (VKH) disease and normal controls were measured by ELISA. EAE and EAU were induced in B10RIII, wild-type, and PGRN-/- mice to evaluate the effect of PGRN on the development of these 2 immune-mediated disease models. The local and systemic immunologic alterations were detected by ELISA, flow cytometry, and real-time PCR. RNA sequencing was performed to identify the hub genes and key signaling pathway. RESULTS: A significantly decreased PGRN expression was observed in patients with active BD and active VKH. Recombinant PGRN significantly reduced EAU severity in association with a decreased frequency of Th17 and Th1 cells. PGRN-/- mice developed an exacerbated EAU and EAE in association with strikingly increased frequency of Th1 and Th17 cells and reduced frequency of regulatory T (Treg) cells. In vitro studies revealed that rPGRN could inhibit IRBP161-180-specific Th1 and Th17 cell response and promote Treg cell expansion. It promoted non-antigen-specific Treg cell polarization from naive CD4+ T cells in association with increased STAT5 phosphorylation. Using RAN sequencing, we identified 5 shared hub genes including Tnf, Il6, Il1b, Cxcl2, and Ccl2 and the most significantly enriched MAPK and tumor necrosis factor signaling pathway in PGRN-/- EAU mice. The aggravated EAE activity in PGRN-/- mice was associated with a skew from M2 to M1 macrophages. DISCUSSION: Our results collectively reveal an important protective role of PGRN in EAU and EAE. These studies suggest that PGRN could serve as an immunoregulatory target in the study of prevention and treatment for the Th1/Th17-mediated diseases.

Cytokine profiles in aqueous humor of patients with different clinical entities of endogenous uveitis.

We assayed aqueous humor (AH) samples from patients with Behçet's disease (BD), Vogt-Koyanagi-Harada (VKH) disease, and HLA-B27-associated uveitis and control patients for the proinflammatory cytokines IL-15, IL-17, interferon-γ and tumor necrosis factor-α and the immunosuppressive cytokine IL-10. Cytokine levels were significantly higher in the three disease groups than in controls. In patients with similar disease activity, levels of IL-15 and IFN-γ were significantly higher in BD patients than in VKH and HLA-B27-associated uveitis groups. Logistic regression identified a significant negative correlation between BD and high levels of IL-10 and a significant positive correlation between VKH disease and high levels of IL-10. The proinflammatory cytokines versus IL-10 ratios were significantly higher in BD compared with other groups. These data suggest that both T helper (Th) 1 and Th17 cells are involved in endogenous uveitis immunopathogenesis. BD is characterized by extensive Th1 polarization, severe proinflammatory conditions and a low immunosuppressive status.

Decreased 1,25-Dihydroxyvitamin D3 level is involved in the pathogenesis of Vogt-Koyanagi-Harada (VKH) disease.

PURPOSE: 1,25-Dihydroxyvitamin D(3) [1,25(OH)(2)D(3)] has recently been found to be involved in the development of autoimmune diseases. This study was to investigate the expression and potential role of 1,25(OH)(2)D(3) in the pathogenesis of Vogt-Koyanagi-Harada (VKH) disease. METHODS: Blood samples were obtained from VKH patients and healthy individuals. Serum 1,25(OH)(2)D(3) levels were measured using ELISA. Peripheral blood mononuclear cells (PBMCs) or cluster of differentiation (CD) 4(+) T cells were cultured with or without 1,25(OH)(2)D(3) in the presence of anti-CD3 and anti-CD28 for the measurement of cell proliferation and cytokines. The cell proliferation was detected using the Cell Counting Kit. The levels of interleukin (IL)-17 and interferon (IFN)-γ levels in the supernatants of PBMCs or CD4(+) T cells were detected by ELISA. RESULTS: 1,25(OH)(2)D(3) was significantly decreased in the serum of active VKH patients as compared with inactive VKH patients and controls. It significantly inhibited PBMCs proliferation and CD4(+) T cell proliferation. It was also able to significantly inhibit the production of IL-17 and IFN-γ by both PBMCs and CD4(+) T cells from VKH patients and controls. CONCLUSIONS: These findings suggest that decreased expression of 1,25(OH)(2)D(3) may be involved in the development of VKH disease. 1,25(OH)(2)D(3) may be potentially used in the treatment of this disease.

Overexpressing Kallistatin Aggravates Experimental Autoimmune Uveitis Through Promoting Th17 Differentiation.

Kallistatin or kallikrein-binding protein (KBP) has been reported to regulate angiogenesis, inflammation and tumor progression. Autoimmune uveitis is a common, sight-threatening inflammatory intraocular disease. However, the roles of kallistatin in autoimmunity and autoreactive T cells are poorly investigated. Compared to non-uveitis controls, we found that plasma levels of kallistatin were significantly upregulated in patients with Vogt-Koyanagi-Harada (VKH) disease, one of the non-infectious uveitis. Using an experimental autoimmune uveitis (EAU) model induced by human interphotoreceptor retinoid-binding protein peptide 651-670 (hIRBP651-670), we examined the effects of kallistatin on the pathogenesis of autoimmune diseases. Compared to wild type (WT) mice, kallistatin transgenic (KS) mice developed severe uveitis with dominant Th17 infiltrates in the eye. In addition, the proliferative antigen-specific T cells isolated from KS EAU mice produced increased levels of IL-17A, but not IFN-γ or IL-10 cytokines. Moreover, splenic CD4+ T cells from naïve KS mice expressed higher levels of Il17a mRNA compared to WT naïve mice. Under Th17 polarization conditions, KS mice exhibited enhanced differentiation of naïve CD4+ T cells into Th17 cells compared to WT controls. Together, our results indicate that kallistatin promotes Th17 differentiation and is a key regulator of aggravating autoinflammation in EAU. Targeting kallistatin might be a potential to treat autoimmune disease.

Analyses of circRNA and mRNA Profiles in Vogt-Koyanagi-Harada Disease.

Recent studies revealed that circular RNAs (circRNAs) are important in numerous biological process and involved in autoimmune diseases. However, their role in Vogt-Koyanagi-Harada (VKH) disease, a classical autoimmune disease, is not yet known. This research aimed to study the expression profile of mRNAs, microRNAs (miRNAs) and circRNAs and investigate the influence of circRNAs on the pathogenesis of VKH disease. We identified circRNAs, miRNAs, and mRNAs expression profiles in CD4+ T cells between 4 VKH patients and 3 healthy controls using the whole-transcriptome sequencing (RNA-seq) technique. We discovered that a total of 5088 mRNAs, 451 circRNAs and 433 miRNAs were differently expressed. The GO and KEGG pathway enrichment analyses were performed for significantly differentially expressed circRNAs and mRNAs. GSEA was conducted for all mRNAs. The functional enrichment suggested that the inflammatory response, the adaptive immune response, NF-kappa B signaling pathway, Th17 cell differentiation, Th1 and Th2 cell differentiation and T cell receptor signaling pathway were associated with VKH disease. In addition, based on the immune-related genes we screened, the circRNA-miRNA-mRNA ceRNA network was analyzed and constructed. Ten differently expressed mRNAs (LAT, ZAP70, ITK, ICOS, RASGRP1, PAG1, PLCG1, PRKCQ, LCK, CARD11) and 5 differently expressed circRNAs (hsa_circ_0033144, hsa_circ_0000233, hsa_circ_0000396, hsa_circ_0001924, hsa_circ_0001320) were selected to be validated by Real-time qPCR (RT-qPCR). The results of RT-qPCR turned out to be consistent with RNA-seq data. Further analysis showed that hsa_circ_0001320 and hsa_circ_0001924 may serve as crucial candidate marker genes of VKH disease. These results reveal that circRNAs may have a crucial immunomodulatory function in the pathophysiological process of VKH disease.

The Rs12569232 SNP Association with Vogt-Koyanagi-Harada Disease and Behcet's Disease is Probably Mediated by Regulation of Linc00467 Expression.

Purpose: To investigate whether the rs12569232 SNP association with Vogt-Koyanagi-Harada disease and Behcet's disease is mediated by regulation of Linc00467 expression.Methods: The expression of linc00467 was detected by real-time PCR. Adenovirus carrying the linc00467 was transduced into CD4+T cells and the effect on cell viability was measured by the CCK-8 test. Human proteome microarray and starBase 2.0 were used to identify the binding proteins of linc00467 and RNA Immunoprecipitation (RIP) was used to confirm the identity of bound proteins.Results: The rs12569232 was associated with the expression of linc00467. The expression of linc00467 was up-regulated in PBMCs and CD4+T cells from VKH disease and BD patients. Over-expression of linc00467 increased cell viability of CD4+T cells. HUR was the common binding protein identified by the two methods and confirmed by RIP.Conclusions: The rs12569232 association with VKH disease and BD may be mediated via regulating the expression of linc00467.

HLA-DRB1*04:05 and HLA-DQB1*04:01: Alleles Potentially Associated with Vogt-Koyanagi-Harada in Northern Thai Patients.

PURPOSE: To determine the frequency and association of alleles at human leukocyte antigen (HLA)-DRB1 and HLA-DQB1 loci in VKH disease patients from Northern Thailand. METHODS: A case-control study was conducted with three subject groups: 23 VKH patients, 20 patients with other uveitis entities, and 40 healthy blood donors. HLA-DRB1 and HLA-DQB1 loci were analyzed and the frequency of HLA-DRB1 and HLA-DQB1 alleles was calculated by direct counting. The measure of association was calculated by odds ratio (OR) and 95% confidence interval. RESULTS: In VKH patients, the most prevalent allele was HLA-DRB1*04:05, found in 35% of patients and with the highest OR (42.13). HLA-DQB1*04:01 was the next most prevalent, found in 23.91% of VKH patients. HLA-DQB1*05:02 was also detected in 23.91% of patients; however, a higher prevalence was observed in non-VKH and healthy controls (30% and 35%, respectively). CONCLUSION: HLA-DRB1*04:05 and HLA-DQB1*04:01 could be potential genetic markers for VKH.

A shared HLA-DRB1 epitope in the DR beta first domain is associated with Vogt-Koyanagi-Harada syndrome in Indian patients.

PURPOSE: Vogt-Koyanagi-Harada (VKH) disease and sympathetic ophthalmia (SO) are two distinct entities that share common clinical and histopathological features; however, it remains unknown whether they have a common genetic susceptibility. Several studies have shown an association of human leukocyte antigen (HLA)-DR4 with VKH disease in patients of different ethnic backgrounds. We present in this paper the HLA-DRB1 genotyping analysis of a large cohort of VKH patients from southern India and compare these patients to patients with SO and to healthy individuals from the same geographic area. METHODS: VKH patients were diagnosed according to the revised criteria of the International Committee on VKH disease. Patients with granulomatous uveitis after ocular trauma or multiple eye surgeries were diagnosed as having SO. Genomic DNA was extracted from all patients and controls. Samples were analyzed for HLA-DRB1 alleles by reverse polymerase chain reaction (PCR) sequence-specific oligonucleotide (SSO) hybridization on microbeads, using the Luminex technology, and by PCR sequence-specific primers (SSP) typing for DRB1*04 allele determination. Strength of associations was estimated by odds ratios (OR) and 95% confidence intervals (CI) and frequencies were compared using the Fisher's exact test. RESULTS: HLA-DRB1 alleles were determined in 94 VKH patients, 39 SO patients, and 112 healthy controls. HLA-DRB1*04 frequency was higher in VKH patients (20.2% versus 10.3% in controls; OR=2.2, p=0.005, pc=0.067). This association was lower than the association of HLA-DRB1*04 frequency in cohorts of patients from different origins. No significant DR4 association with SO was detected. HLA-DRB1*0405 and HLA-DRB1*0410 alleles were significantly increased in VKH patients (8.5% versus 0.9% in controls; OR=10.3, 95% CI=2.34-45.5, p<0.001). These two alleles share the epitope S57-LLEQRRAA (67-74) in the third hypervariable region of the HLA-DR molecule. None of the DRB1 alleles was significantly associated with SO. CONCLUSIONS: Based on the association of HLA-DRB1*0405 and HLA-DRB1*0410 alleles with VKH disease, we propose that the epitope S57-LLEQRRAA (67-74) in the third hypervariable region of the HLA-DRbeta1 molecule is the relevant susceptibility epitope. This genetic component seems specific to VKH disease since no correlation could be identified in SO patients. The weaker association with HLA-DR4 in this VKH patient cohort compared to VKH patients from northern India is probably related to the lower frequency of HLA-DRB1*0405 in our study group. The HLA-DRB1 association with susceptibility to VKH syndrome seems weaker in Indian patients compared to Japanese or Hispanic patients, suggesting a different non-HLA immunogenetic background in Indian VKH patients.

IL-10 and TGF-beta immunoregulatory cytokines rather than natural regulatory T cells are associated with the resolution phase of Vogt-Koyanagi-Harada (VKH) syndrome.

The pro-inflammatory cytokines play a critical role in the initiation and propagation of ocular autoimmune diseases. Regulation of these cytokines is generally mediated by the immunoregulatory cytokine such as IL-10 or TGF-beta. In this study, we investigated the immunoregulatory cytokine profile and frequency of natural regulatory T cells (nTregs) in patients with Vogt-Koyanagi-Harada (VKH). We obtained the peripheral blood mononuclear cells (PBMC) from patients with VKH and healthy controls. The cytokine profile from supernatants of PBMC cultured with or without phytohaemagglutinin (PHA) was measured by ELISA, the percentage of CD4(+) Foxp3(+) and CD25(high)Foxp3(+) T regulatory cells were analysed by flow cytometry, and the transcriptional level of Foxp3 expression was analysed by real-time quantitative PCR. The immunoregulatory cytokines, TGF-beta and IL-10, increased in patients with VKH in the inactive stage of the disease. We observed no significant difference in the CD4(+) Foxp3(+) and CD25(high)Foxp3(+) T cells as well as no reduction in FOXP3 mRNA expression in the patients with VKH when compared to healthy controls. We showed in our work, an increase in IFN-gamma secretion by PBMC of patients with VKH in the active stage of the disease when compared to healthy controls and patients in the inactive stage. Our data suggest that IL-10 and TGF-beta cytokines, rather than nTregs are associated with the resolution phase of the disease and may have a more relevant role in controlling this disease.