Effects of topical erythropoietin on healing experimentally-induced avascular scleral damage in a rabbit model.

The present study was designed to investigate the effect of topical erythropoietin on the healing process of induced necrotizing scleritis and to evaluate the ocular side effects of this treatment modality in a rabbit model. Necrotizing scleritis was induced in 8 New Zealand albino rabbits. The animals were then randomly divided into one of two groups: a treated group administered a topical erythropoietin-containing cellulose-based gel every 8 h or a control group treated with a cellulose-based gel without erythropoietin every 8 h. The sizes of the lesions measured at different time points were compared between the groups. After three months, the rabbits' eyes were enucleated and histologically and immunohistochemically evaluated for angiogenesis and apoptosis. The lesions were completely vascularized in all eyes of the treated group and 50% of eyes of the control group. The mean interval from the induction of scleral necrosis to a complete improvement was 28 days in the treated group and 62.5 days in the control group (P = 0.04). Histological examination revealed that erythropoietin enhanced the improvement of necrotizing scleritis by stimulating angiogenesis and reducing apoptosis. Neovascularization of the cornea, iris, or retina was not observed in the treated group. We observed a significantly faster recovery to complete improvement of necrotizing scleritis in rabbit eyes treated with erythropoietin compared to those of the control group. Treated eyes had a higher rate of complete healing and had no ocular safety concerns. This therapeutic modality represents a promising treatment for scleral necrosis following various types of ocular surgery.

Rituximab in the treatment of refractory scleritis in patients with granulomatosis with polyangiitis (Wegener's).

PURPOSE: The purpose of this descriptive study was to evaluate the clinical response to rituximab (RTX) in patients with scleritis due to granulomatosis with polyangiitis (GPA), in patients who had proved refractory to treatment with systemic glucocorticoids and immunosuppressive agents. METHODS: Retrospective analysis of interventional case series. Single referral center study. Eight patients (12 affected eyes) due to scleritis secondary to GPA, refractory to conventional treatment were included to receive RTX as therapy for remission induction. RTX was administered as a 1-g infusion every 2 weeks, for a total of 2 g. Patient follow-up included clinical evaluation (systemic and ophthalmologic), B-cell subset (CD19, CD20, CD22) counts, proteinase-3 anti-neutrophil cytoplasmic antibody (PR-3 ANCA), and Birmingham Vasculitis Activity Score for Wegener's granulomatosis (BVAS-WG). Outcomes were response to treatment and achievement of remission, as well as number of ocular relapses. RESULTS: The main indication for treatment was refractory necrotising anterior scleritis. Four weeks after completion of treatment with RTX, all patients showed clear clinical improvement, with no further progression. In all patients, an absolute depletion of B cells was confirmed in the first 6 weeks after treatment. Seven patients (87.5 %) achieved remission of inflammatory activity in 7 months or less. However, three patients experienced ocular relapse, which comprised reactivation of the anterior scleritis, uveitis, and posterior scleritis, and two patients required a second dose of RTX, with immediate improvement. CONCLUSIONS: RTX is useful in the treatment of refractory necrotising scleritis in patients with GPA. Of note, in those who relapse after remission, RTX can be successfully used for retreatment.

TNF inhibition for ophthalmic indications: current status and outlook.

BACKGROUND: Tumor necrosis factors (TNF) are a group of cytokines that play a role in systemic inflammation, stimulating the acute phase reaction. They are involved in systemic rheumatologic conditions such as rheumatoid arthritis and juvenile idiopathic arthritis, as well as ocular inflammatory conditions in the uveitis spectrum. Several drugs were developed to inhibit the action of TNF, thereby reducing inflammation. The three most commonly used TNF inhibitors in the US are etanercept, infliximab, and adalimumab. Newer drugs include certolizumab and golimumab. In this review, we discuss the differences in the mechanism of action, route of administration, indication, and efficacy of TNF inhibitors used in the treatment of ocular inflammation. METHODS: A review of the literature in the PubMed, MEDLINE, and Cochrane databases was conducted to identify clinical trials, comparative studies, case series, and case reports describing the use of tumor necrosis factor inhibitors in uveitis therapy. The search was limited to primary reports published in English with human subjects from 1990 to the present, yielding 5,238 manuscripts. In addition, referenced articles from the initial searches were hand searched to identify additional relevant reports. After title and abstract selection, duplicate elimination, and manual search, 69 papers were selected for analysis. Exclusion criteria included review articles and case reports on the efficacy of etanercept, infliximab, and adalimumab. Manuscripts with fewer than 20 study subjects were excluded if other larger studies existed on the use of the same drug for a particular indication. Studies with <6 months of patient follow-up were also excluded, except in the case where no other data were available. Articles meeting these criteria were then reviewed by the three authors for inclusion in this review. RESULTS: Tumor necrosis factor inhibitors have been shown to decrease inflammation associated with a number of rheumatologic conditions. Three of the five commercially available TNF inhibitors-etanercept, infliximab, and adalimumab-have been studied for their efficacy in treatment of ocular inflammation. Etanercept appears to be inadequate in controlling ocular inflammation and is not recommended for the treatment of uveitis. Infliximab and adalimumab, however, have shown encouraging results in multiple trials. Serious potential side effects such as infection, including reactivation of latent tuberculosis, malignancy, and demyelinating disease, may limit the use of TNF inhibitors in uveitis. Proper screening of patients prior to initiating these therapies may decrease these risks. DISCUSSION: Early success with infliximab and adalimumab has paved the way for new TNF inhibitors and other corticosteroid-sparing drugs to emerge in the treatment of ocular inflammation. Future studies are on the horizon to determine the long-term safety and efficacy of newer TNF inhibitors such as certolizumab and golimumab.

Intravitreal bevacizumab (avastin) as an adjuvant for the treatment of posterior scleritis.

We report a case of posterior scleritis effectively managed with intravitreal bevacizumab. A 71-year-old woman was diagnosed with posterior scleritis. Although she was initially treated with systemic steroids, her clinical presentation deteriorated. She was then treated with a single intravitreal injection of bevacizumab and aqueous humor collection. The aqueous level of vascular endothelial growth factor prior to the intravitreal injection was 880.51 pg/mL, greater than that in the healthy control group (p < 0.001). One month later, the scleritis was completely resolved, and the patient remained stable during six months of follow-up. Intravitreal bevacizumab appears to be an effective adjuvant therapy for patients with posterior scleritis.

Association of PDGF-BB-induced thrombomodulin with the regulation of inflammation in the corneal and scleral stroma.

PURPOSE: The responses of corneal and scleral stromal cells to platelet-derived growth factor (PDGF)-BB were assessed and inflammatory reactions in the cornea and sclera were investigated. METHODS: Primary cultures of cells obtained from human subjects and strains derived from human corneal or scleral stromal cells (Cs3 and Sc1, respectively) were used. Changes in gene expression after 24 hours of exposure to 10 ng/mL PDGF-BB were analyzed with an Sc1 DNA microarray. The upregulation of several genes in Cs3 and Sc1 was confirmed by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. The expression of bioactive factors was detected immunohistochemically in nine different clinical specimens. RESULTS: DNA microarray analysis revealed that the gene encoding thrombomodulin (TM) was induced in Sc1 by PDGF-BB. RT-PCR confirmed that TM expression at the mRNA level was increased in both corneal and scleral stromal cells. At the protein level, TM expression was increased in scleral stromal cells, but not in corneal cells, and TM was detected in both the membrane and cytoplasmic compartments. TM was detected immunohistochemically in inflamed scleral and several corneal specimens. After TM stimulation, interleukin (IL)-18 transcription was increased in Sc1. CONCLUSIONS: PDGF-BB induced TM mRNA expression in scleral and corneal stromal cells, but Western blot analysis revealed the increase in TM expression only in the scleral cells. TM induced IL-18 in scleral stromal cells. A cascade involving these biologically active factors may regulate scleral and corneal inflammation. The results also reveal differences in the biological response of scleral and corneal stromal cells.

Ocular surface inflammation, and nerve growth factor level in tears in active thyroid-associated ophthalmopathy.

PURPOSE: To measure tear nerve growth factor (NGF) concentrations in cases of active thyroid-associated ophthalmopathy (TAO) before and after glucocorticoid treatment, and to correlate NGF levels with disease inflammatory activity and thyroid autoantibody concentration. METHODS: The study involved 20 patients with active TAO and 20 age- and gender-matched controls. Tear break-up time (BUT) was obtained, the Schirmer test was performed, and tear NGF/total protein ratio was measured in control subjects and patients with active TAO before, and 2 and 4 weeks after, steroid treatment. RESULTS: Tear BUT and Schirmer values significantly increased after 2 and 4 weeks of steroid treatment (p < 0.001 and p = 0.004 respectively). Baseline tear NGF/total protein ratio was higher in patients with active TAO than in control subjects, and the ratio significantly decreased after 2 and 4 weeks of steroid treatment (p < 0.001). Tear NGF/total protein ratio did not correlate with inflammatory activity score, exophthalmos value and thyroid binding inhibiting immunoglobulin (TBII) level (p > 0.05). CONCLUSIONS: Tear NGF may have a specific role in ocular surface inflammation, which protects against ocular surface damage in patients with active TAO. Anti-inflammatory treatment significantly reduced the level of NGF in tears, increased tear film stability and production, and decreased congestive symptoms.

Infliximab for the treatment of active scleritis.

OBJECTIVE: This study aimed to evaluate the possible safety and effectiveness of infliximab in patients with active scleritis. STUDY DESIGN: Prospective, nonrandomized, open-label pilot study (Protocol No. 04-EI-0065). PARTICIPANTS: Five patients with active anterior scleritis. METHODS: This single-centre, pilot study of infliximab for the treatment of active anterior scleritis was conducted at the National Eye Institute, National Institutes of Health, between 2003 and 2007. Scleritis patients with active disease who had used at least 1 conventional immunosuppressive agent in the past were included. Primary outcome was a 2-step decrease in scleral inflammation within 14 weeks. Patients received infliximab (5 mg/kg) at baseline, at weeks 2 and 6, and every 4 weeks through week 30, after which the infusion interval was increased (week 36, 48). RESULTS: All patients met the primary outcome by achieving quiescence of their active scleritis by week 14 with no additional immunosuppressives. However, after 14 weeks 1 patient developed new-onset intraocular inflammation that did not respond to reinduction and was terminated from the study. Side effects attributable to infliximab included ear infection with transient decreased hearing, urinary tract infection, lower respiratory tract infection, and facial rash in 1 patient and urinary tract infection, diarrhea, upper respiratory tract infection, nasal congestion and headache, mouth sores, head tremor, and occasional numbness and tingling in extremities in another patient, all of which resolved spontaneously or with appropriate treatment. CONCLUSIONS: Infliximab may be considered as a viable option in treating patients with active scleritis; however, patients should be monitored closely for potentially serious side effects.

TH17 cells contribute to uveitis and scleritis and are expanded by IL-2 and inhibited by IL-27/STAT1.

T-helper type 17 cells (T(H)17) are implicated in rodent models of immune-mediated diseases. Here we report their involvement in human uveitis and scleritis, and validate our findings in experimental autoimmune uveoretinitis (EAU), a model of uveitis. T(H)17 cells were present in human peripheral blood mononuclear cells (PBMC), and were expanded by interleukin (IL)-2 and inhibited by interferon (IFN)-gamma. Their numbers increased during active uveitis and scleritis and decreased following treatment. IL-17 was elevated in EAU and upregulated tumor necrosis factor (TNF)-alpha in retinal cells, suggesting a mechanism by which T(H)17 may contribute to ocular pathology. Furthermore, IL-27 was constitutively expressed in retinal ganglion and photoreceptor cells, was upregulated by IFN-gamma and inhibited proliferation of T(H)17. These findings suggest that T(H)1 cells may mitigate uveitis by antagonizing the T(H)17 phenotype through the IFN-gamma-mediated induction of IL-27 in target tissue. The finding that IL-2 promotes T(H)17 expansion provides explanations for the efficacy of IL-2R antibody therapy in uveitis, and suggests that antagonism of T(H)17 by IFN-gamma and/or IL-27 could be used for the treatment of chronic inflammation.

Churg-Strauss syndrome associated with leukotriene receptor antagonists (LTRA).

Churg-Strauss syndrome (CSS) is a rare vasculitic disorder that generally occurs in patients with bronchial asthma. CSS is being increasingly recognized in asthmatic patients treated with leukotriene receptor antagonists. However, the nature of this relationship remains to be elucidated. The present report describes three asthmatic patients who developed clinical manifestations highly suggestive of CSS, although one patient lacked the presence of eosinophilia. The patient, however, exhibited biopsy-proven cutaneous necrotizing vasculitis, which improved after withdrawal of montelukast. The second patient presented with systemic constitutional signs including fever, malaise, arthralgias, clinical jaundice, peripheral blood eosinophilia, and biopsy-proven eosinophilic hepatitis. The third patient also had circulating eosinophilia, scleritis, and arthritis. All patients improved after discontinuation of the leukotriene receptor antagonist (montelukast).

Expression of tumor necrosis factor alpha and matrix metalloproteinase-9 in surgically induced necrotizing scleritis.

BACKGROUND: Surgically induced necrotizing scleritis is primarily managed with immunosuppressive regimens. Such treatments can alter inflammatory cytokines and products such as tumor necrosis factor-alpha (TNF-alpha) and matrix metalloproteinase-9 (MMP-9), which are an important proinflammatory cytokine and a tissue-degrading protease involved in necrosis of some ocular tissues. OBJECT: To evaluate TNF-alpha and MMP-9 in the tear or sclera of surgically induced necrotizing scleritis. METHODS: After tear collection from healthy and diseased eyes of 2 patients, immunoblot analysis using monoclonal antibody specific for human TNF-alpha and MMP-9 was performed. In another patient, scleral tissues were obtained during scleral allograft surgery, and the same immunoblot analysis was done. RESULTS: The level of TNF-alpha and MMP-9 was increased in tear fluid from patients compared to that of control volunteers and returned to the control level after treatment. The diseased sclera showed an increased expression of MMP-9 compared to that of the normal donor sclera. CONCLUSIONS: TNF-alpha and MMP-9 may suggest disease activity of surgically induced necrotizing scleritis and can be altered by proper immunosuppressive treatment.

The value of combining anterior segment fluorescein angiography with indocyanine green angiography in scleral inflammation.

PURPOSE: To determine the value of anterior segment indocyanine green (ICG) angiography combined with anterior segment fluorescein angiography in scleral inflammation. DESIGN: Comparative observational case series. PARTICIPANTS: The study included 18 patients with various forms of scleral and episcleral disease and a single normal subject. METHODS: Anterior segment angiography using both ICG and fluorescein was performed to identify any vascular abnormalities and pathologic changes in the anterior segment. MAIN OUTCOME MEASURE: The pathologic criteria for anterior segment fluorescein angiography described by Watson and Bovey (1995) were used to compare and contrast the results of the angiograms. RESULTS: Fluorescein angiography and ICG angiography provide different and complementary information. Both dyes have different leakage patterns caused by their difference in optical and chemical properties. Areas of slow flow are more readily determined with fluorescein angiography. ICG angiography determines damage to and patency of individual vessels. CONCLUSION: Fluorescein angiography and ICG angiography detect areas of damage not clinically visible and can be useful in the differential diagnosis, the selection of appropriate medication, and monitoring and regulation of treatment in scleritis. To obtain the most information both investigations should be performed sequentially.

Leukocyte adhesion molecule expression in scleritis.

OBJECTIVE: To analyze the expression and cellular distribution of intercellular adhesion molecule 1, E-selectin (endothelial leukocyte adhesion molecule), vascular cell adhesion molecule 1, very late antigen 4, and lymphocyte function associated antigen 1 (LFA-1) in diseased and normal human sclera. METHODS: Monoclonal antibodies to vascular cell adhesion molecule 1, very late antigen 4, intercellular adhesion molecule 1, LFA-1, and E-selectin were used to perform immunohistochemical staining on frozen sections of 16 cryopreserved human sclera specimens and 5 conjunctival specimens. RESULTS: The normal human sclera did not express any of the adhesion molecules. The expression of LFA-1 was dramatic in all the scleral and conjunctival specimens on the inflammatory cells. Intercellular adhesion molecule 1, the ligand for LFA-1, was expressed in 7 of 12 scleral specimens. Furthermore, the expression of LFA-1 and intercellular adhesion molecule 1 were focally present in areas of inflammatory infiltrate. E-selectin expression was detected on the vascular endothelial cells in 8 of 12 patients. There was variable expression of vascular cell adhesion molecule 1 and very late antigen 4 in the inflamed sclera and conjunctiva. CONCLUSIONS: Our results demonstrate the presence of LFA-1 in the sclera and in the conjunctiva of patients with scleritis. Variable expression of other leukocyte adhesion molecules was noted in the sclera and the conjunctiva of these patients.

Expression of matrix metalloproteinases by human plasma cells and B lymphocytes.

Matrix metalloproteinases (MMP) are proteolytic enzymes that play a key role in tissue remodelling during physiological and pathological processes, by initiating the degradation of extracellular matrix. MMP overexpression can lead to tissue destruction which is characteristic of chronic inflammatory diseases such as rheumatoid arthritis and scleritis. Plasma cells are often abundant at such sites of chronic inflammation. In the present study we investigated whether plasma cells could contribute to matrix degradation by their expression of MMP In situ hybridization and immunohistochemical analyses on diseased synovial and scleral tissue demonstrated the expression of stromelysin-1 (MMP-3) and gelatinase B (MMP-9), but little or no tissue inhibitor of matrix metalloproteinase 1 (TIMP-1) mRNA, by IgG-positive plasma cells. Northern blot analysis of RNA extracted from a human plasma cell line (ARH-77), Epstein-Barr virus-transformed B cells, and purified peripheral blood B cells, demonstrated expression of stromelysin mRNA. TIMP-1 mRNA was only detected by the more sensitive reverse transcription PCR method in these cell types. Plasma cells and B lymphocytes cultured in the presence of monensin demonstrated cytoplasmic gelatinase B. Gelatin and casein zymography on conditioned media (CM) derived from cytokine treated plasma cells revealed the induction of secreted gelatinase and stromelysin activity. Western blotting confirmed the presence of stromelysin-1 and TIMP-1 proteins in plasma cell CM. These data suggest that plasma cells are not only capable of modulating an inflammatory response by antibody and cytokine production, but also by their ability to produce MMP. Secretion of MMP from focal aggregates of plasma cells may play a critical role in tissue destructive diseases such as rheumatoid synovitis and scleritis.

Increased expression of matrix metalloproteinases in vivo in scleritis tissue and in vitro in cultured human scleral fibroblasts.

Scleritis is a sight-threatening inflammatory disorder of the eye characterized by the degradation of scleral matrix. Matrix metalloproteinases (MMPs) are ubiquitous proteolytic enzymes important in physiological and pathological processes, the activity of which is stringently controlled by the action of a family of natural antagonists, the tissue inhibitors of matrix metalloproteinases (TIMPs). We hypothesized that enhanced expression of MMPs, without the negative regulatory influence of TIMPs, may be a key feature of tissue destruction in inflammatory eye diseases, such as scleritis. The aim of this study was to localize and characterize cells expressing MMPs and TIMPs in sclera affected by necrotizing scleritis and, in a parallel study, to establish whether cytokines modulate MMP expression in cultured human scleral fibroblasts. In situ hybridization and immunohistochemical analyses indicated that resident scleral fibroblasts as well as inflammatory cells such as macrophages and T lymphocytes express stromelysin, gelatinase B, and TIMP-1 in necrotizing scleritis tissue. In addition, cytoplasmic immunoreactivity for tumor necrosis factor-alpha, an inducer of MMPs, was detected in infiltrating inflammatory cells. Cultured scleral fibroblasts stimulated with the combination of interleukin-1 alpha plus tumor necrosis factor-alpha increased TIMP-1 mRNA twofold above constitutive levels. By contrast, these cytokines induced a sevenfold increase in the steady-state levels of stromelysin mRNA. Using Western blotting, stromelysin and TIMP-1 protein production paralleled mRNA induction in cytokine-stimulated human scleral fibroblasts. Culture supernatants harvested from cytokine-stimulated human scleral fibroblasts were subjected to sodium dodecyl sulfate polyacrylamide gel electrophoresis gelatin substrate zymography. Our results revealed a prominent 92-kd gelatinolytic band corresponding to gelatinase B, which was inducible with interleukin-1 alpha. These data provide evidence for our hypothesis, that an imbalance between enzyme/inhibitor ratios may be the underlying mechanism of the tissue destruction characteristic of scleritis. Our results demonstrate the potential involvement of MMPs and their modulation by cytokines produced by infiltrating inflammatory cells in destructive ocular inflammation.