The Cytokine Interleukin-6 and the Chemokines CCL20 and CXCL13 Are Novel Biomarkers of Specific Endogenous Uveitic Entities

Abstract

Purpose: The purpose of this study was to determine levels of the cytokines IL-1β, IL-6, IL-21, IL-22, and IL-23 and the chemokines CXCL13, CCL19, CCL20, and CCL21 in aqueous humor (AH) samples from patients with specific uveitic entities.

Methods: Paired serum samples (n = 13) and AH samples (n = 111) from patients with active idiopathic granulomatous uveitis (IGU) or with uveitis associated with HLA-B27-related inflammation, Behçet's disease (BD), Vogt-Koyanagi-Harada (VKH) disease, or sarcoidosis and control patients were analyzed in two different multiplex assays.

Results: Cytokines IL-1β, IL-21, IL-22, and IL-23 were not detected in any AH sample. Chemokine CCL21 concentrations in serum were significantly higher than those in AH. CCL19 levels in AH and serum were not significantly different. Levels of CCL20 and CXCL13 in AH were significantly higher than those in serum. IL-6 was not detected in serum samples. IL-6 AH levels were significantly higher in patients with HLA-B27-associated uveitis and in BD patients than in patients with VKH disease, sarcoidosis, and IGU (P < 0.0001). CCL20 AH levels were significantly higher in HLA-B27-associated uveitis than in BD, VKH, sarcoidosis, and IGU (P = 0.001), whereas CXCL13 AH levels were significantly higher in VKH disease and IGU than in HLA-B27-associated uveitis, BD, and sarcoidosis (P = 0.007).

Conclusions: IL-6-driven immune responses are more potent in HLA-B27-associated uveitis and BD than in VKH disease, sarcoidosis, and IGU. CCL20 appears to be a specific biomarker of HLA-B27-associated uveitis, whereas CXCL13 appears to be a biomarker of VKH disease and IGU. Our findings suggest that IL-6, CCL20, and CXCL13 could serve as drug targets for treatment of specific clinical entities of endogenous uveitis.

Endogenous uveitis is a clinically heterogenous group of potentially blinding intraocular inflammatory diseases. They often occur in conjunction with systemic inflammatory diseases, such as HLA-B27-associated uveitis, Behçet's disease (BD), Vogt-Koyanagi-Harada (VKH) disease, and sarcoidosis. Because endogenous uveitis includes a wide range of diverse inflammatory manifestations in the eye, with different clinical phenotypes, it is possible that different immunopathogenic mechanisms are involved in each clinical subtype. Moreover, causes and outcomes of the disease continue to be largely unpredictable and unexplained. 

Nevertheless, management of uveitis has steadily improved in recent decades, mostly due to enhanced understanding of the pathophysiology of the disease, which has been translated into effective therapies. However, despite the remarkable successes achieved, the clinical complexity and the variable responses to therapy leave multiple challenges in uveitis management. Cytokines and chemokines control inflammation, and they have important roles in the pathology of various endogenous uveitis entities,^1–6 thus representing major targets for clinical intervention. For example, biological therapies that inhibit the proinflammatory cytokine tumor necrosis factor-α (TNF-α) are now widely used in routine clinical practice for the treatment of several autoimmune inflammatory diseases, including endogenous refractory uveitis associated with BD.^7,8 TNF-α levels in the aqueous humor (AH) were significantly higher in patients with Behçet's disease than in patients with other causes of endogenous uveitis,³ These therapies display specific modes of action and have been developed on the basis of a detailed understanding of cytokine involvement in inflammation. 

Dysregulated expression of the proinflammatory cytokine interleukin-6 (IL-6) has been shown to be involved in the development of chronic inflammatory autoimmune diseases. IL-6 exerts a variety of biological activities on responsive cell populations through its binding to transmembrane IL-6 receptor (IL-6R) as well as soluble IL-6R (sIL-6R).^9–11 Transforming growth factor-β (TGF-β) and the inflammatory cytokines, such as IL-1β, IL-6, IL-21, and IL-23 contribute to the generation of T helper (T_h) 17 cells that play a crucial role in the pathogenesis of several autoimmune and inflammatory diseases through production of effector molecules, such as IL-17 and IL-22.¹² 

Chemokines are multifunctional mediators that regulate leukocyte recruitment to the inflamed tissue, promote inflammation, and enhance immune responses. Chemokines are divided into four subgroups (CXC, CC, C, and CX₃C) depending on the arrangement of the conserved cysteine residues. The specific effects of chemokines are mediated by binding to distinct members of a family of G protein-coupled receptors.^13,14 CCL20 is the only chemokine known to interact with CC chemokine receptor 6 (CCR6). The CCL20-CCR6 axis is responsible for the chemoattraction of immature dendritic cells, B lymphocytes, and activated and memory CD4⁺ and CD8⁺ T lymphocytes.¹⁵ CCL20 is implicated in several autoimmune diseases, such as rheumatoid arthritis^16–20 and experimental autoimmune encephalomyelitis.^21,22 

There is increasing evidence that B lymphocytes play an important role in the pathogenesis of autoimmune diseases, such as rheumatoid arthritis,^23–25 Sjogren's syndrome,^26–28 multiple sclerosis,^29–31 myasthenia gravis,^32,33 systemic lupus erythematosus^34,35 and autoimmune thyroiditis.³⁶ A key phenomenon in these diseases is the formation of ectopic lymphoid aggregates with germinal center-like structures in the inflamed tissues, which contain proliferating B lymphocytes, plasma cells, follicular helper CD4⁺ T lymphocytes, and a network of follicular dendritic cells.^26–28 Accumulated evidence has demonstrated that the formation and maintenance of ectopic lymphoid structures in these chronic inflammatory conditions is critically dependent on the ectopic expression of the lymphoid chemokines CXCL13, CCL19, CCL21, and CXCL12 and their specific receptors CXCR5 (for CXCL13), CCR7 (for CCL19 and CCL21), and CXCR4 (for CXCL12).^26–28 

The aim of this study was to identify novel molecules involved in the pathogenic mechanisms of endogenous uveitis that could serve as potential targets for selective therapy or that may act as novel biomarkers for different entities of endogenous uveitis. We, therefore, measured the levels of the inflammatory cytokines IL-1β, IL-6, IL-21, IL-22, and IL-23 and the chemokines CXCL13, CCL19, CCL20, and CCL21 in the AH from patients with active uveitis associated with human leukocyte antigen (HLA)-B27-related intraocular inflammation, BD, VKH disease, and sarcoidosis and from patients with idiopathic granulomatous uveitis (IGU). 

Patients and Methods

Patients with active uveitis seen at the outpatient clinic of King Abdulaziz University Hospital were included in the study. Patients who had undergone cataract extraction with no history of uveitis served as the control group. Conditions were diagnosed using established clinical criteria, with supporting laboratory evidence as needed.^37,38 Uveitis was further classified as nongranulomatous if fine endothelial keratic precipitates were seen in the absence of iris nodules and/or choroidal granulomas, and as granulomatous if larger keratic precipitates, including large (“mutton-fat”) keratic precipitates, or Koeppe and/or Busacca iris nodules, and/or optic disc or choroidal granulomas were seen. In each patient, the uveitis activity was graded according to the criteria of the Standardization of the Uveitis Nomenclature working Group grading scheme.³⁹ None of the patients was taking topical or systemic therapy on presentation. 

Aqueous humor (100–200 μL) was aspirated from each patient by means of limbic paracentesis, using a 27-gauge needle attached to a tuberculin syringe, after the application of the topical local anesthetic oxybuprocaine hydrochloride 0.4% (Benoxinate; Chauvin Pharmaceuticals Ltd., Kingston, UK). The procedure was performed using a surgical microscope. The samples were snap frozen and maintained at −70°C until used. Aqueous humor samples from patients with uveitis were obtained before they underwent therapy. All procedures followed the tenets of the Declaration of Helsinki, and informed consent was obtained from all patients and the control subjects. The study was approved by the Research Center, College of Medicine, King Saud University. 

Two sets of samples were assayed for cytokines and chemokines. In the first set of samples (n = 56), CXCL13, CCL19, CCL20, and CCL21 were quantified using a Bio-Plex Pro human chemokine kit (Bio-Rad Laboratories, Temse, Belgium), whereas in the second set of samples (n = 76), CCL20, IL-1β, IL-6, IL-21, IL-22, and IL-23 were quantified using a Milliplex MAP kit (Merck Millipore, Molsheim, France) according to the protocols provided by the manufacturers. The detection ranges of the assays as provided by the manufacturers are listed in Table 1. 

Table 1

 

Multiplex Assay-Dependent, Specific Detection Range of Chemokines and Cytokines

 

The first set of AH samples was obtained from 13 patients with HLA-B27-associated uveitis, 11 with BD, 12 with VKH disease, seven with sarcoidosis, and five with IGU. Eight patients who had undergone cataract extraction served as the control group. The second set of AH samples was obtained from 17 patients with HLA-B27-associated uveitis, 15 with BD, 15 with VKH disease, six with sarcoidosis, and nine with IGU. Fourteen patients who had undergone cataract extraction served as a control group. 

To address whether cytokines and chemokines detected in the AH could originate from blood circulation, we measured the levels of cytokines and chemokines in 13 serum samples. The serum samples were obtained from three patients with HLA-B27-associated uveitis, four with BD, three with VKH disease, two with sarcoidosis, and one with IGU. 

Statistical Methods

Data management was preliminarily done using Excel 2013 (Microsoft, Redmond, WA, USA), and then all statistics were analyzed using SPSS version 20.0 software (IBM, Armonk, NY, USA). Undetected values were treated as “zero” in terms of analysis due to their trivial decimal values, which were quite close to zero. Descriptive statistics were calculated so that categorical variables were presented as frequency (percent) and continuous variables as means (±standard deviation). The chi-square test was used to compare the detection rates. The Kruskal-Wallis test was used to compare means among different disease categories. Mann-Whitney U-test was then used to compare means from two independent groups. Spearman correlation coefficients were computed to investigate correlations between variables. A P value less than 0.05 indicated statistical significance. 

Results

Analysis of Aqueous Humor From Uveitis Patients by Chemokine Bio-Plex Multiplex Assay

Among the chemokines analyzed in the AH from patients, only CCL20 and CXCL13 levels were significantly higher than those in serum samples (Fig. 1). These findings suggest that local chemokine production is the authentic source of CCL20 and CXCL13 within the ocular microenvironment and that a systemic inflow mechanism is rather unlikely. CCL20 and CXCL13 were not detected in any of the AH samples from the control group. 

Figure 1

 

Comparisons of mean chemokine levels in serum with levels in aqueous humor samples from patients with uveitis, using the Bio-Plex multiplex assay kit.

The incidence rates for CCL20 and CXCL13 detection are shown in Table 2. The incidence rate for detection of CCL20 in AH samples from HLA-B27-associated uveitis patients (100%) was significantly higher than in patients with VKH disease (58.3%) and sarcoidosis (42.9%) (P = 0.036; P = 0.014, respectively; chi-square test). 

Table 2

 

Summary Data for CCL20 and CXCL13 Levels by Bio-Plex Multiplex Assay Kit

 

The chemokine levels in the five disease groups were compared (Kruskal-Wallis test), and the results are shown in Table 2. CCL20 and CXCL13 levels in AH samples differed significantly among HLA-B27-associated uveitis, BD, VKH disease, sarcoidosis, and IGU (P = 0.001 and P = 0.007 for CCL20 and CXCL13, respectively). Pairwise comparisons (Mann-Whitney U-test) indicated that CCL20 levels in patients with HLA-B27-associated uveitis were significantly higher than in patients with BD, VKH disease, sarcoidosis, and IGU (P = 0.014; P < 0.0001; P = 0.002; and P = 0.012, respectively). The CXCL13 levels in VKH disease were significantly higher than the levels in HLA-B27-associated uveitis and BD (P = 0.004; and P = 0.007, respectively). In addition, mean CXCL13 levels in IGU were significantly higher than the mean levels in HLA-B27-associated uveitis (P = 0.022). 

When patients were divided into those with nongranulomatous uveitis (HLA-B27-associated uveitis and BD) (n = 24) and those with granulomatous uveitis (VKH disease, sarcoidosis, and IGU) (n = 24), CCL20 levels in the AH from nongranulomatous uveitis (4.9 ± 7.5 ng/mL) were significantly higher than those in granulomatous uveitis cases (0.1 ± 0.2 ng/mL; P < 0.0001). On the other hand, CXCL13 levels in granulomatous uveitis (2.4 ± 4.1 ng/mL) were significantly higher than those in nongranulomatous uveitis (0.7 ± 2.4 ng/mL; P < 0.0001). 

Correlations Between CCL20 Levels in AH and Disease Activity

The levels of CCL20 in AH samples correlated significantly with the disease activity in all patients (r = 0.6; P < 0.0001) (Fig. 2), in patients with BD (r = 0.88; P < 0.0001) and in patients with nongranulomatous uveitis (r = 0.63; P = 0.001). A significant negative correlation was found between AH levels of CCL20 and CXCL13 in all patients (r = 0.38; P = 0.008). 

Figure 2

 

Correlations between aqueous humor (AH) cytokine and chemokine levels and disease activity. In these analyses, all patients were included. The levels of CCL20 in AH (measured by Bio-Plex multiplex assay kit) (A) and by Milliplex multiplex assay kit (B) correlated significantly with disease activity. In addition, CCL20 levels were significantly higher in patients with anterior chamber reaction >2+ than those in patients with anterior chamber reaction ≤2+. Also, the levels of IL-6 in AH (measured by Milliplex multiplex assay kit) correlated significantly with disease activity (C). However, although IL-6 levels were higher in patients with anterior chamber reaction >2+ than those in patients with anterior chamber reaction, ≤2+ statistical significance was not reached.

Determination of Cytokine and Chemokine Concentrations in Aqueous Humor Samples From Uveitis Patients by Milliplex Multiplex Assay

Among the cytokines and chemokines analyzed in uveitis, only IL-6 and CCL20 were detected in AH samples but not in serum samples. IL-1β, IL-21, IL-22, and IL-23 were not detected in any of the AH samples from uveitis patients and the control group. IL-6 was detected in only five of 14 control AH samples, whereas CCL20 was not detected in any of the control AH samples. IL-6 was detected in all AH samples from patients with uveitis, except in three samples from patients with VKH disease (Table 3). When the whole study group was considered, IL-6 levels were significantly higher in the AH of patients (n = 62) (14.7 ± 15.7 ng/mL) than in that of controls (n = 14; 0.3 ± 0.9 ng/mL; P < 0.0001). In a comparison between individual patient groups and the control group, significantly increased IL-6 levels were observed for HLA-B27-associated uveitis, BD, VKH disease, sarcoidosis, and IGU (P < 0.0001; P < 0.0001; P = 0.002; P = 0.003; and P < 0.0001, respectively). 

Table 3

 

Summary Data for CCL20 and IL-6 Levels by Milliplex Multiplex Assay Kit

 

Similar to the results obtained with the first set of AH samples (Table 2), the incidence rate of CCL20 detection in HLA-B27-associated uveitis in the second set of AH samples (Table 3) was significantly higher than in BD, VKH disease, sarcoidosis, and IGU (P = 0.0004; P = 0.029; P = 0.0007; and P = 0.0034, respectively; chi-square test). To evaluate distribution of the IL-6 and CCL20 levels among the five disease groups, we used the Kruskal-Wallis test, and the results are shown in Table 3. IL-6 and CCL20 levels in AH samples differed significantly among patients with HLA-B27-associated uveitis, BD, VKH disease, sarcoidosis, and IGU (P < 0.001 for both comparisons). Pairwise comparisons indicated that IL-6 levels in HLA-B27-associated uveitis were significantly higher than in VKH disease, sarcoidosis, and IGU (P < 0.0001; P = 0.006; and P = 0.016, respectively) and that IL-6 levels in BD were significantly higher than in VKH disease and sarcoidosis (P = 0.007; and P = 0.002, respectively). As shown in the first set of AH samples analyzed with the Bio-Plex assay (Table 2), the Milliplex assay also confirmed that, in the second set of patients (Table 3), CCL20 levels were significantly higher in HLA-B27-associated uveitis than in BD, VKH disease, sarcoidosis, and IGU (P = 0.001; P < 0.0001; P = 0.001; and P = 0.001, respectively). 

When patients were divided into those with nongranulomatous uveitis (HLA-B27-associated uveitis and BD; n = 32) and those with granulomatous uveitis (VKH disease, sarcoidosis, and IGU) (n = 30), IL-6 levels in the AH from nongranulomatous uveitis (23.1 ± 15.8 ng/mL) were significantly higher than those in granulomatous uveitis (5.7 ± 9.5 ng/mL; P < 0.0001). Similarly, CCL20 levels in AH from nongranulomatous uveitis (0.49 ± 0.77 ng/mL) were significantly higher than that in granulomatous uveitis (0.02 ± 0.05 ng/mL; P < 0.0001). 

Correlations Between IL-6 and CCL20 Aqueous Humor Levels and Disease Activity

The levels of IL-6 (r = 0.55; P < 0.0001) and CCL20 (r = 0.59; P < 0.0001) in AH samples correlated significantly with the disease activity in all patients (Fig. 2). In addition, the levels of IL-6 correlated significantly with disease activity in patients with HLA-B27-associated uveitis (r = 0.58; P = 0.014). Finally, a significant positive correlation was found between aqueous humor levels of IL-6 and CCL20 in all patients (r = 0.69; P < 0.0001). 

Discussion

In the present study, IL-6 levels in the AH of uveitis patients were significantly enhanced compared to those in controls and were higher in patients with HLA-B27-associated uveitis and BD than in patients with VKH disease, sarcoidosis, and IGU. Furthermore, a significantly positive correlation between IL-6 levels in the AH from patients with uveitis and the clinical disease activity was detected in all patients and in patients with HLA-B27-associated uveitis. IL-6 was originally identified as B-cell stimulatory factor 2, inducing differentiation of activated B cells into immunoglobulin-producing plasma cells. However, it is a multipotent cytokine, regulating both B and T cell activation/differentiation.^{10,11,40–42} IL-6 in combination with TGF-β, is essential for T_h17 differentiation from naïve CD4⁺ T cells, whereas IL-6 inhibits TGF-β-induced regulatory T cells (Treg) development.^11,12,43 The resultant predominance of T_h17 cells over Treg caused by IL-6 may be responsible for the disruption of immunological tolerance and the development of inflammatory autoimmune diseases. Numerous studies have confirmed the pathological roles of IL-6 in inflammatory and autoimmune diseases.^9–11 Our findings suggest a role for IL-6 in the development of endogenous uveitis, particularly in HLA-B27-associated uveitis and BD and imply that the IL-6–IL-6R axis might be a target pathway for the treatment of these diseases. Clinical trials have demonstrated the efficacy of tocilizumab, a humanized anti-IL-6R antibody, for patients with rheumatoid arthritis, Castleman's disease, and systemic juvenile idiopathic arthritis, leading to approval of this innovative drug for the treatment of these diseases.^9–11 Small retrospective studies demonstrated the efficacy of tocilizumab in the treatment of refractory uveitic macular edema.^44–46 As an alternative for IL-6, T_h17 polarization might be driven through an IL-1β/IL-1R signaling pathway downstream of Mincle/caspase activation and recruitment domain 9 (CARD 9)-dependent signaling and inflammasome activation. The latter pathway is activated by, for example, mycobacterial peptidoglycan⁴⁷ and also plays a central role in autoimmune diseases of the eye.⁴⁸ 

Several studies identified the CCL20-CCR6 axis as a potential mediator of inflammatory cell recruitment in the rheumatoid arthritis joint. Rheumatoid arthritis synovial fluid contained significantly more CCL20 than that in osteoarthritis,^16–18 and CCL20 and CCR6 expression levels were up-regulated in rheumatoid arthritis synovial tissue.^16,17 The upregulated production of CCL20 induced the recruitment of CCR6-expressing mononuclear cells including T_h17 cells to inflamed joints in rheumatoid arthritis and its animal model.^18,20 In addition to expressing CCR6, T_h17 cells also synthesize CCL20, thereby establishing a positive feedback mechanism, which may perpetuate the chronic inflammatory process in rheumatoid arthritis.¹⁸ Several studies reported that the proinflammatory cytokines IL-1β, TNF-α, and IL-17 stimulate the production of CCL20 by rheumatoid fibroblast-like synovial cells^17–19 and that IL-6 increased IL-1β-, TNF-α-, and IL-17-induced CCL20 production.¹⁹ On the other hand, CCL20 enhanced the production of IL-6 in fibroblast-like synovial cells.^20,49 In agreement with these studies, our analysis identified a significant positive correlation between the AH levels of CCL20 and those of IL-6. Administration of blocking anti-CCR6 monoclonal antibody inhibited inflammation in a mouse model of arthritis, supporting the implication of CCL20-CCR6 axis in the pathogenesis of rheumatoid arthritis.¹⁸ In experimental autoimmune encephalomyelitis, a model of T lymphocyte-dependent inflammation, both CCL20 and its receptor CCR6 were upregulated in the spinal cord and draining lymph nodes. Disease severity and accumulation of mononuclear cells were significantly reduced by administration of specific neutralizing anti-CCL20 and anti-CCR6 antibodies and in gene-targeted CCR6-deficient mice.^21,22 

In this study, the concentrations of CCL20 in the AH were quantified with the use of two multiplex assays. Both of these commercial assays showed that CCL20 AH levels significantly correlated with disease activity. Use of the Milliplex assay showed that the incidence of detection of CCL20 in AH from patients with HLA-B27-associated uveitis was significantly higher than in patients with BD, VKH disease, sarcoidosis, and IGU. In addition, the Bio-Plex assay showed that the incidence of CCL20 detection in HLA-B27-associated uveitis was significantly higher than in patients with VKH disease and sarcoidosis. The detection limit in the Bio-Plex assay was lower than that in the Milliplex assay. Moreover, the Bio-Plex assay yielded higher concentrations than the Milliplex assay. It should be noted that other studies have reported differences in the absolute concentrations of analytes when comparing different multiplex cytokine analysis kits.^50,51 By using both the Bio-Plex and Milliplex assays, we found CCL20 levels were significantly higher in patients with HLA-B27-associated uveitis than in the other uveitic entities. Our findings suggest that CCL20 might serve as a biomarker for HLA-B27-associated uveitis and that the CCL20–CCR6 axis may serve as an excellent drug target for HLA-B27-associated uveitis. 

Among the lymphoid chemokines analyzed, only CXCL13 levels in the AH from patients with uveitis were significantly higher than in sera. These findings suggest that CXCL13 is locally produced within the ocular microenvironment and that a systemic inflow mechanism is rather unlikely. In addition, CXCL13 was not detected in any of the control AH samples. 

A sizeable subset of patients with autoimmune diseases, such as rheumatoid arthritis, Sjogren's syndrome, multiple sclerosis, myasthenia gravis, systemic lupus erythematosus, and autoimmune thyroiditis develop ectopic lymphoid structures at the sites of inflammation. CXCL13 expressed by follicular dendritic cells is central in this process as it attracts CXCR5-expressing B lymphocytes and follicular helper CD4⁺ T lymphocytes to the ectopic follicles.^25–28 In rheumatoid arthritis synovia containing lymphoid aggregates, elevated levels of CXCL13 are observed,²⁵ and several studies identified CXCL13 as a new potential marker for severity of inflammation in rheumatoid arthritis,^23,24 systemic lupus erythematosus,^34,35 multiple sclerosis,^30,31 and myasthenia gravis.³² Supportive for a role for CXCL13 as an important mediator of human autoimmune disorders, CXCL13 neutralizing monoclonal antibody significantly reduced disease severity in animal models of rheumatoid arthritis and experimental autoimmune encephalomyelitis.⁵² 

It must be noted that, although research on other extraocular autoimmune disorders can be very informative, the eye is an immune-privileged site that actively regulates and directs immune responses that take place in its “territory.”⁵³ The immune privilege of the eye limits tissue damage after an episode of immune activation and allows allogenic, unmatched corneal transplantation. It implies strong intraocular tolerance, which is achieved through production of the anti-inflammatory cytokine TGF-β and through the presence of different subsets of Tregs in the ocular surface tissues.⁵⁴ However, when immune privilege is faced with a serious challenge, it fails dismally to prevent severe destruction and permanent structural damage in uncontrolled uveitis.⁵⁵ In those occasions, immune therapy might be beneficial. However, the cytokine patterns in uveitis seem to be nonredundant, and tailored immune therapy might be required. Indeed in the present study, levels of the B-cell chemoattractant CXCL13 were significantly higher in patients with VKH disease and IGU than in patients with BD, HLA-B27-associated uveitis, and sarcoidosis. These findings suggest that B lymphocytes may be pathogenetically important in uveitis associated with VKH disease and IGU and that CXCL13 levels in AH might be a useful biomarker for this pathology. Interestingly, several reports described predominance of B lymphocytes in the uveal inflammatory infiltrate in granulomatous uveitic entities, such as sympathetic ophthalmia,^56–59 multifocal choroiditis⁶⁰ and progressive subretinal fibrosis.⁶¹ Additionally, B lymphocytes were identified in choroidal inflammatory infiltrate in two cases of VKH disease with “sunset glow fundus.”⁶² B lymphocyte aggregates were described in the uvea of one patient with end-stage VKH disease.⁶³ These findings suggest that treatment with rituximab, an antibody against CD20 leading to depletion of B lymphocytes, might be effective in patients with VKH disease and IGU. Several clinical trials have demonstrated the effectiveness of rituximab in treating signs and symptoms of multiple autoimmune diseases.^26,64 Recently, rituximab was reported to be effective in the treatment of 2 cases with diffuse subretinal fibrosis⁶⁵ and one case of resistant VKH disease.⁶⁶ 

In summary, our findings suggest that intraocular levels of cytokines and chemokines differ depending on the cause of uveitis. In the ocular inflammatory microenvironment of patients with endogenous uveitis, CCL20 might serve as a biomarker of inflammation in HLA-B27-associated uveitis, and the B cell chemoattractant CXCL13 might serve as a biomarker of uveitis associated with VKH disease. IL-6-driven immune responses are more potent in HLA-B27-associated uveitis and BD than in VKH disease and sarcoidosis. Our findings also suggest that CCL20, CXCL13, and IL-6 could serve as excellent drug targets for the treatment of specific clinical entities of endogenous uveitis. 

Acknowledgments

The authors thank Connie B. Unisa-Marfil for secretarial work. 

Supported by King Saud University through Vice Deanship of Research Chair, Dr. Nasser Al-Rashid Research Chair in Ophthalmology (AMAE-A), the Fund for Scientific Research of Flanders, the Interuniversity Attraction Poles Program, Belgian Science Policy Office (project P7/40), and the Concerted Research Actions (2013/015) of the Regional Government of Flanders (SS and GO). 

Disclosure: A.M. Abu El-Asrar, None; N. Berghmans, None; S.A. Al-Obeidan, None; A. Mousa, None; G. Opdenakker, None; J. Van Damme, None, S. Struyf, None