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To evaluate the treatment of autologous serum eye drops (ASED) on dry eyes in patients with graft-versus-host disease (GVHD).A retrospective chart review of 35 patients with a history of ocular GVHD following hematopoietic stem cell transplantation that used ASED to alleviate dry eye symptoms was performed. Patients were categorized into three different groups. If patients had available ophthalmic data before and after starting treatment was group 1 (n = 14), had available ophthalmic data after starting treatment in group 2 (n = 10) and had available ophthalmic data before treatment or did not have any data after starting treatment in group 3 (n = 11). Data were collected on patient's age, gender, primary diagnosis, visual acuity and fluorescein corneal staining were collected on individual eyes in order to evaluate the efficacy of the ASED on alleviating dry eye-related signs and symptoms.No adverse ocular effect from the ASED was found in our series (except one fungal keratitis). All patients reported either improvement (55%) or stability (45%) in their ocular symptoms upon the use of ASED. In patients with available data before and after starting treatment, the corneal staining score improved by a median of 1 (p = 0.003) and the LogMAR visual acuity had a non-significant improvement.In our study, ASED used by patients with ocular GVHD were both safe and effective. ASED should be considered in patients with GVHD who suffer from dry eyes.

To report the clinical course of 6 patients with refractory neurotrophic corneal ulcers that were treated with topical insulin drops.Retrospective chart review of patients who had neurotrophic corneal ulcers or epithelial defects refractory to standard medical and surgical treatment. Insulin drops, prepared by mixing regular insulin in artificial tears with a polyethylene glycol and propylene glycol base at a concentration of 1 unit per milliliter, were prescribed 2 to 3 times daily.Six patients, aged 2 to 73 years, developed neurotrophic corneal ulcers refractory to a range of medical and surgical treatments, including bandage contact lens, amniotic membrane grafting, and permanent tarsorrhaphy. Each patient was administered topical insulin drops with complete corneal reepithelialization within 7 to 25 days.Topical insulin may be a simple and effective treatment for refractory neurotrophic corneal ulcers. Further study is required to determine the clinical efficacy and side effect profile of insulin drops.

Objective. RA increases vascular disease and angiogenesis, yet a 1964 Lancet report paradoxically linked RA to lower diabetic retinopathy. Our objective was to examine RA as a risk factor for diabetic retinopathy compared with other vascular risk factors. Methods. This cohort study compared the prevalence of diabetic retinopathy in diabetes patients with and without RA in a 5% Medicare sample. We analysed the impact of RA on the prevalence of diabetic retinopathy using multivariate logistic regression calculating adjusted rate ratios (ARRs) controlling for sociodemographics, co-morbidity and health utilization. Sensitivity analysis examined eye exam rates. Results. Among 256 331 Medicare diabetes patients, 5572 (2%) had RA. Diabetic retinopathy was less prevalent in patients with RA compared with those without RA (13.7% vs 16.1%, P ≤ 0.01). Compared with patients without RA, the adjusted model demonstrated that patients with diabetes and RA were 28% less likely to have diabetic retinopathy and 4% more likely to receive an eye exam [ARR 0.72 (95% CI 0.67, 0.77), ARR 1.04 (95% CI 1.02, 1.06)]. Conclusion. Findings support the 1964 paradox observing decreased diabetic retinopathy in patients with RA. These findings pose new questions regarding whether RA physiology or treatments protect against diabetic retinopathy and how intraocular factors vary in contrast to adverse vascular changes elsewhere.

Requirements of hydrogels for drug delivery, wound dressings, and surgical implantation can be extensive, including suitable mechanical properties and tailorable degradation time frames. Herein, an adaptable PEG-based hydrogel, whose mechanical properties and degradation rate can be systematically adjusted to meet these criteria by altering simple variables such as the PEG molecular weight, is described. The performance of these hydrogels in three physical manipulations (pushing, pulling, and folding), representative of manipulations that they may undergo during typical biomedical use, is also assessed. While not all of these formulations can withstand these manipulations, a subset did, and it is intended to further optimize these formulations for specific clinical applications. Additionally, the outcomes of the physical manipulation tests indicate that simply having a high modulus does not correlate with biomedical applicability.

We sought to determine the effect on rat conjunctival mast cells of chronic exposure to a degranulating agent. Compound 48/80 (250 micrograms in 10 microliter) was applied daily to the ocular surface of rats for a total of 13 treatments administered in 17 days. A single application of compound 48/80 was given to rats for comparison. Four groups of animals (6 in each group) were evaluated 1 or 48 h after the last application of compound 48/80 in both chronic exposure and single exposure studies. Although a single exposure to compound 48/80 induced significant degranulation of mast cells in animals observed 1 h after treatment, no significant degranulation of mast cells was observed 1 h after the last dose of compound 48/80 was given to rats whose conjunctiva had been chronically exposed to this degranulating agent. Thus, multiple applications of compound 48/80 attenuated the mast cell response to compound 48/80. This acquired tolerance to compound 48/80 was reflected in the lessened clinical signs in rats receiving repeated applications compared with those given a single application.

Purpose of review This review will focus on recent advances in our understanding of the pathogenesis of allergic eye diseases. Common findings in acute allergic conjunctivitis (seasonal and perennial) and chronic allergic conjunctivitis (vernal keratoconjunctivitis, atopic keratoconjunctivitis, and giant papillary conjunctivitis) include evidence of mast cell activation and eosinophil attraction and activation. Cytokine levels found in tears, conjunctival impression cytology and biopsy specimens, and serum have been evaluated as markers of disease, and as targets of therapeutic intervention. Recent findings Human conjunctival epithelial cells respond to tumor necrosis factor alpha, interleukin-1 beta, and interferon-gamma individually and in combination. Intracellular adhesion molecule-1 expression is upregulated by interleukin-1 beta and tumor necrosis factor alpha. Conjunctival epithelial cells release interleukin-8 in response to interleukin-1 beta and tumor necrosis factor alpha but not interferon-gamma. Supernatants from activated mast cells cause increased adhesion of eosinophils to conjunctival epithelium. Tear levels of tumor necrosis factor alpha were elevated in vernal keratoconjunctivitis patients compared with normal controls. T cell lines from chronic allergic eye disease patients showed inconsistent production of cytokines in atopic and vernal keratoconjunctivitis and low levels in giant papillary conjunctivitis. Vernal keratoconjunctivitis patients have differing levels of eosinophil cationic protein in their serum if they were serum specific immunoglobulin E positive compared to serum specific immunoglobulin E negative patients. Summary Recent findings continue to expand our basic knowledge of mechanisms and differences between seasonal and perennial allergic conjunctivitis and atopic and vernal keratoconjunctivitis. Understanding the complex interactions and cross talk between cells, cytokines and other mediators is relevant for new therapeutic approaches directed at specific disease entities.

Background Allergen-mediated mast cell activation is a key feature of ocular allergic diseases. Evidence of eosinophil-derived mediators in tears and conjunctival biopsy specimens has been associated with chronic ocular allergic inflammation. Objective To examine the role of conjunctival mast cell mediators in eosinophil adhesion to conjunctival epithelial cells and eosinophil degranulation. Methods Conjunctival cells were obtained by enzymatic digestion of cadaveric conjunctival tissues. Eosinophils were obtained from peripheral blood samples using negative magnetic bead selection. The effect of IgE-activated mast cell supernates on eosinophil degranulation and adherence to epithelial cells was compared with supernates obtained from mast cells pretreated with a degranulation inhibitor (olopatadine). Eosinophil adhesion was measured by eosinophil peroxidase assay, and eosinophil degranulation was measured by eosinophil-derived neurotoxin radioimmunoassay. Results IgE-activated conjunctival mast cell supernates stimulated adhesion of eosinophils to epithelial cells (20.4% ± 6.3% vs 3.1% ± 1.0%; P = .048). Degranulation was not required for this process (no effect of olopatadine). IgE-activated mast cell supernates stimulated eosinophil-derived neurotoxin release (108.89 ± 8.27 ng/10 6 cells vs 79.45 ± 5.21 ng/10 6 cells for controls, P = .02), which was significantly inhibited by pretreatment of mast cells with a degranulation inhibitor (79.22 ± 4.33 ng/10 6 cells vs 61.09 ± 5.39 ng/10 6 cells for olopatadine pretreated and untreated, respectively, P = .02). Conclusions Mediators released from conjunctival mast cells promote eosinophil adhesion to conjunctival epithelial cells and eosinophil degranulation. Degranulation inhibition studies suggest that different mast cell mediators are involved in regulation of these events.

Allergic eye disease is associated with IgE-mediated conjunctival inflammation leading to signs of immediate hypersensitivity, including redness, itching, and tearing. Pathologic studies using conjunctival mast cells demonstrate that these cells, when sensitized with IgE antibody and exposed to environmental allergens, release mediators involved with allergic inflammation. The type, release kinetics, and concentration of these mediators in the conjunctiva have not been completely characterized. The ability to isolate and purify mast cells and epithelial cells from human conjunctival tissue has permitted the study of mediator release and cell-to-cell signaling in this tissue. Our laboratory has developed in vitro and in vivo models to better understand how inflammatory cells are recruited to and infiltrate conjunctival tissues. These models demonstrate that mast-cell activation may supply sufficient cytokine signaling to initiate and direct the well-orchestrated trafficking of eosinophils to the ocular surface, facilitate their adhesion, and cause release of potent mediators of ocular inflammation.

A 23-year-old man presented with a 2-year history of a slow growing lesion in the left eye. The lesion consisted of a vascularized, balloon-like mass straddling the limbus nasally (A). Histopathologic examination demonstrates (B) a conjunctival epitheliumwith solar elastosis, consistent with a pterygium (left side), and proliferation of melanocytic cells in the stroma and epithelial-stromal interface, consistent with a compound nevus (right side). It is important to note that, except for a small area on the left (black arrowhead) that demonstrates a normal thickness, the remainder of the conjunctiva shows considerable thickening, which measures as high as 6 times the normal value in some areas. This thickening, which gives the lesion its striking clinical appearance, is due to severe reactive fibrosis and solar elastotic changes. Magnification reveals the nevus component in more detail (C [white arrowheads]). Opinion Viewpoint

Allergic eye disease is a bilateral immune-mediated process of the ocular surface that occurs in up to 25 % of the population of developed countries. Four entities are described as having at least in part some portion of their pathogenesis emanating from mast cell activation: seasonal allergic conjunctivitis (SAC), atopic keratoconjunctivitis (AKC), vernal keratoconjunctivitis (VKC), and giant papillary conjunctivitis (GPC). SAC and GPC are non-vision threatening whereas AKC and VKC may lead to vision loss. The severe forms of allergic disease, AKC and VKC have an immune-mediated pathogenesis as well as mast cell activation. GPC is considered iatrogenic due to the presence of an ocular surface foreign body such as contact lens or an exposed suture. Treatment of allergic eye disease may be as simple as administration of artificial tears or topical antihistamines through to as complex as the need for topical or oral steroids as well as immunomodulatory therapy. Contact dermatitis may occur in the periocular region. The periocular form of contact dermatitis is most commonly associated with topical ophthalmic pharmaceutical agents. Antiviral and glaucoma medications are some of the more common medications implicated. In vivo conjunctival provocation testing is an ocular equivalent of skin testing to determine sensitivity to specific antigens. This may be used experimentally to develop a reproducible human model of allergic eye disease and has been utilized to test new pharmaceutical agents.

Allergic eye disease is a common clinical problem adversely affecting the quality of life for millions of sufferers. This ocular process is associated with IgE-mediated conjunctival inflammation leading to signs of immediate hypersensitivity including redness, itching, and tearing. Pathologic studies have shown that the conjunctiva contains mast cells that when sensitized with IgE antibody and exposed to environmental allergens can release mediators of allergic inflammation. The type, release kinetics, and concentration of these mediators in the conjunctiva have not been completely characterized. The ability to isolate and purify mast cells and epithelial cells from human conjunctival tissue has permitted the study of mediator release and cell-to-cell signaling in this tissue. Our laboratory has developed in vitro and in vivo models to better understand how inflammatory cells are recruited to and infiltrate conjunctival tissues. These models demonstrate that mast cell activation may supply sufficient cytokine signaling to initiate and direct the well-orchestrated trafficking of eosinophils to the ocular surface, facilitate their adhesion, and cause release of potent mediators of ocular inflammation.

Background Tumor necrosis factor-α (TNFα) release likely plays a crucial role in allergic ocular inflammation via increasing ICAM-1 on epithelial cells and triggering other proinflammatory events. The immediate and prolonged release of TNFα from human conjunctival mast cells in response to allergen challenge is potentially an important target for therapeutic intervention, yet the effect of ocular anti-allergic agents on this process has not been examined. Olopatadine (Patanol) is a clinically effective dual-action ophthalmic anti-allergic agent that has been shown to inhibit mast cell histamine, tryptase, and PGD 2 release in vitro and promote decreased H 1 receptor binding activity in vitro and functional H 1 receptor antagonism in vivo. Objective To investigate the effect of olopatadine on TNFα release from anti-IgE antibody challenged purified human conjunctival mast cells. Methods Human conjunctival mast cells were purified (>95%) from cadaveric tissues using a procedure combining enzymatic digestion and Percoll gradient centrifugation. These cells were incubated with olopatadine for 30 minutes then challenged with anti-IgE antibody for 90 minutes. Supernatants were analyzed for TNFα. Results Purified human conjunctival mast cells responded to anti-IgE antibody challenge with TNFα release in a concentration dependent manner (optimum concentration was 10 μg/mL). Olopatadine pre-incubation resulted in a dose-dependent decrease in anti-IgE antibody mediated TNFα release (IC 50 = 13.1 μM). At a concentration of 3 mM olopatadine reduced TNFα release to the level of unchallenged controls. Conclusion Olopatadine inhibited anti-IgE antibody-mediated release of TNFα from human conjunctival mast cells. This effect could contribute to the long duration of anti-allergic activity reported for the drug.

Importance Conjunctivitis is a common problem. Objective To examine the diagnosis, management, and treatment of conjunctivitis, including various antibiotics and alternatives to antibiotic use in infectious conjunctivitis and use of antihistamines and mast cell stabilizers in allergic conjunctivitis. Evidence Review A search of the literature published through March 2013, using PubMed, the ISI Web of Knowledge database, and the Cochrane Library was performed. Eligible articles were selected after review of titles, abstracts, and references. Findings Viral conjunctivitis is the most common overall cause of infectious conjunctivitis and usually does not require treatment; the signs and symptoms at presentation are variable. Bacterial conjunctivitis is the second most common cause of infectious conjunctivitis, with most uncomplicated cases resolving in 1 to 2 weeks. Mattering and adherence of the eyelids on waking, lack of itching, and absence of a history of conjunctivitis are the strongest factors associated with bacterial conjunctivitis. Topical antibiotics decrease the duration of bacterial conjunctivitis and allow earlier return to school or work. Conjunctivitis secondary to sexually transmitted diseases such as chlamydia and gonorrhea requires systemic treatment in addition to topical antibiotic therapy. Allergic conjunctivitis is encountered in up to 40% of the population, but only a small proportion of these individuals seek medical help; itching is the most consistent sign in allergic conjunctivitis, and treatment consists of topical antihistamines and mast cell inhibitors. Conclusions and Relevance The majority of cases in bacterial conjunctivitis are self-limiting and no treatment is necessary in uncomplicated cases. However, conjunctivitis caused by gonorrhea or chlamydia and conjunctivitis in contact lens wearers should be treated with antibiotics. Treatment for viral conjunctivitis is supportive. Treatment with antihistamines and mast cell stabilizers alleviates the symptoms of allergic conjunctivitis.

Background: Giant cell arteritis is a common cause of severe visual loss in older individuals. Patients often present to the ophthalmologist having already lost vision in one eye. Detection of early ophthalmoscopic signs that precede irreversible visual loss in giant cell arteritis would allow preventative treatment in an otherwise frequently blinding disease. Methods: Case presentations. Results: Seven patients with mild visual symptoms and results of an ophthalmologic examination significant for cotton-wool spots were found to have giant cell arteritis. On specific questioning, six of seven patients described constitutional symptoms consistent with giant cell arteritis. Six patients had an abnormally elevated Westergren erythrocyte sedimentation rate. Temporal artery biopsy confirmed giant cell arteritis in six patients. The seventh patient received a diagnosis of polymyalgia rheumatica. Prompt treatment with corticosteroids led to preservation of vision and uneventful resolution of the cottonwool spots in all seven patients. Conclusion: Cotton-wool spots are an early ophthalmoscopic finding in giant cell arteritis and can precede severe visual loss. Recognition of the significance of cottonwool spots, use of laboratory studies, and prompt treatment may preserve vision in an otherwise frequently blinding disease.

Publisher Summary Atopy refers to hypersensitivities in persons with a hereditary background of allergic diseases. The major, most commonly recognized atopic conditions include eczema (atopic dermatitis), asthma, hay fever, and allergic rhinitis. Atopic ocular disease includes Seasonal Allergic Conjunctivitis (SAC), Perennial Allergic Conjunctivitis (PAC), Vernal Keratoconjunctivitis (VKC), Atopic Keratoconjunctivitis (AKC), and Giant Papillary Conjunctivitis (GPC). SAC and PAC, with or without nasal involvement, comprise the majority of ocular allergic conditions. The immunopathology of seasonal and perennial allergic conjunctivitis and rhinitis has been characterized as an IgE-mediated, mast cell dependent, hypersensitivity response. These conditions are classified as Type I immediate hypersensitivity. This chapter discusses the therapies that are currently available to treat SAC and PAC. VKC and AKC might cause significant complications and lead to loss of vision. Type I hypersensitivity reactions of the ocular surface are involved in AKC and VKC, but are not considered the only pathophysiologic mechanisms in these similar, yet distinct entities.

We studied the effect of one application of compound 48/80 to the conjunctiva of the rat on the response to a subsequent challenge. Rats treated once showed conjunctival edema and marked degranulation of conjunctival mast cells. A second exposure to compound 48/80, applied after an interval of 24 h, produced slight clinical effects but had marked effects on conjunctival mast cells. Approximately 90% of the mast cells could no longer be observed by light microscopy. As the interval between initial challenge and rechallenge was increased, the clinical response returned, mast cells were again observable in normal numbers, and the extent of degranulation returned to approximately 50%. After an interval of 7 days, rats responded clinically as they had to the first application of compound 48/80. Although the number of mast cells in the conjunctiva was normal and extensive degranulation occurred, exocytosis was not observed. Thus some effects of the first application of compound 48/80 persisted for at least 1 week. Whether the effects observed depend upon the continued presence of compound 48/80 in the mast cell or granule membrane remains to be determined.

Background Staphylococcus aureus colonization is common in atopic keratoconjunctivitis, potentially activating epithelial cells via toll-like receptor 2 (TLR-2) and the receptor for platelet-activating factor (PAFR). Objectives To examine human conjunctival epithelial cells for the expression of TLR-2 in vitro and in vivo and to evaluate the role of TLR-2 in S aureus -mediated activation of these cells. Methods Conjunctival epithelial cells isolated from cadaveric tissues were stimulated with interferon γ (IFN-γ) or a commercial S aureus cell wall extract ( Staphylococcus aureus -CWE) (with or without anti-TLR-2 blocking antibody or PAFR antagonist) and were analyzed for tumor necrosis factor α (TNF-α) and interleukin 8 (IL-8) release; surface expression of TLR-2, intercellular adhesion molecule-1, HLA, and CD14; and TLR-2 messenger RNA expression. Ocular surface cells collected via impression cytology were examined for TLR-2 expression via flow cytometry. Results Expression of TLR-2 was up-regulated on conjunctival epithelial cells by IFN-γ and Staphylococcus aureus -CWE. Expression of TLR-2 messenger RNA was increased by IFN-γ. Staphylococcus aureus -CWE up-regulated intercellular adhesion molecule 1, HLA, and CD14 expression and increased TNF-α and IL-8 release in a dose-dependent manner. Anti-TLR-2 significantly inhibited TNF-α release, whereas PAFR antagonist significantly inhibited IL-8 release. Toll-like receptor 2 was expressed on conjunctival epithelial cells from 4 of 5 patients with atopic keratoconjunctivitis, 3 of 5 with seasonal allergies, and 0 of 3 without allergies. Conclusions Conjunctival epithelial cells express TLR-2 and may play an active role in the chronic ocular inflammatory response to S aureus through pathways that involve TLR-2 and PAFR.

PURPOSE To gain better understanding of conjunctival epithelial cell responses to proinflammatory cytokines, the individual and combined effects of TNFalpha, IL-1beta, and IFNgamma on chemokine release (IL-8, regulated on activation normal T-cell expressed and secreted [RANTES]) and surface receptor expression (intercellular adhesion molecule [ICAM]-1, and HLA-DR, -DP, and -DQ) were examined. METHODS Conjunctival epithelial cells were isolated from cadaveric conjunctival tissues and cultured in 24-well plates until almost confluent. Recombinant cytokines (0.005-50 ng/mL) were added, alone or in various combinations, 24 hours before harvesting of supernates for ELISAs and cells for flow cytometry. RESULTS TNFalpha, IL-1beta, and IFNgamma had distinctive individual and combined effects on the parameters tested. Although TNFalpha and IL-1beta had similar and synergistic effects on increasing expression of ICAM-1, IL-1beta was a more potent upregulator of the release of IL-8 than was TNFalpha. Upregulation of IL-8 was additive when IL-1beta was combined with TNFalpha. Neither TNFalpha nor IL-1beta increased expression of HLA. In contrast, IFNgamma was a potent upregulator of both surface receptors (ICAM-1 and HLA) but IFNgamma alone had no effect on mediator release (IL-8 and RANTES). Release of RANTES required two cytokine signals, with IFNgamma and TNFalpha being the most potent combination. CONCLUSIONS Knowledge of the differential and combined effects of proinflammatory cytokines on conjunctival epithelial cells allows better understanding of ocular inflammation.

JAMA Hormone Replacement Therapy and Dry Eye Syndrome Debra A. Schaumberg, ScD, MPH; Julie E. Buring, ScD; David A. Sullivan, PhD; M. Reza Dana, MD, MPH Context Postmenopausal hormone replacement therapy (HRT) use is common in the United States. Some research suggests that estrogen may have detrimental effects on the tear film and could influence the development of dry eye syndrome, but few data are available on this relationship. Objective To determine the relationship of HRT and dry eye syndrome. Design, Setting, and Participants The Women's Health Study, a large cohort study in which 25 665 postmenopausal women provided information about use of HRT at baseline (1992), 12, and 36 months and dry eye syndrome at 48 months. Main Outcome Measures (1) Clinically diagnosed dry eye syndrome, as reported by participants;(2) severe symptoms (both ocular dryness and irritation either constantly or often); and (3) either clinically diagnosed dry eye syndrome or severe symptoms, compared between women who used HRT vs those who did not. Results For the combined end point of either clinically diagnosed dry eye syndrome or severe symptoms, the multivariable-adjusted odds ratios were 1.69 (95% confidence interval [CI], 1.49-1.91) for estrogen use alone and 1.29 (95% CI, 1.13-1.48) for estrogen plus progesterone/progestin use compared with no HRT use. Each 3-year increase in the duration of HRT use was associated with a significant 15% (95% CI, 11%-19%) elevation in risk of clinically diagnosed dry eye syndrome or severe symptoms. Results were similar for the combined end point of clinically diagnosed dry eye syndrome and severe symptoms. Conclusions These data suggest that women who use HRT, particularly estrogen alone, are at increased risk of dry eye syndrome. Physicians caring for women who are taking or considering HRT should be apprised of this potential complication. Author Affiliations: Departments of Medicine (Dr Schaumberg), Ambulatory Care and Prevention (Dr Buring), and Ophthalmology (Drs Sullivan and Dana), Division of Preventive Medicine, Brigham and Women's Hospital (Drs Schaumberg and Buring), and Schepens Eye Research Institute (Drs Schaumberg, Sullivan, and Dana), Harvard Medical School, and Department of Epidemiology, Harvard School of Public Health (Dr Buring), Boston, Mass. Corresponding Author and Reprints: Debra A. Schaumberg, ScD, MPH, Division of Preventive Medicine, Brigham and Women's Hospital, Harvard Medical School, 900 Commonwealth Ave E, Boston, MA 02215 (e-mail: dschaumberg@rics.bwh.harvard.edu).

Background Olopatadine is a clinically effective dual-action (antihistamine/mast cell stabilizer) ophthalmic antiallergic agent. We have previously demonstrated that olopatadine inhibits tumor necrosis factor α (TNF-α) release from purified human conjunctival mast cells and that supernates from stimulated mast cells upregulate intercellular adhesion molecule 1 (ICAM-1) expression on epithelial cells via TNF-α. Objective To investigate the effect of olopatadine on the TNF-α–mediated mast cell upregulation of ICAM-1 expression on conjunctival epithelial cells. Methods Human conjunctival mast cells and epithelial cells were purified (>95%) from cadaveric tissue. Conjunctival mast cells were preincubated with three doses (30, 300, or 3,000 μM) of olopatadine or buffer alone for 30 minutes followed by 90-minute challenge with anti-immunoglobulin E (10 μg/mL). The resulting supernates were incubated with conjunctival epithelial cell monolayers for 24 hours along with the following treatments: rTNF-α, mast cell supernate + anti-TNF-α, recombinant (r)TNF-α + anti-TNF-α, the three doses of olopatadine, olopatadine supernates, olopatadine supernates + rTNF-α. ICAM-1 expression was measured using flow cytometry. Results Anti-IgE–stimulated human conjunctival mast cell supernates upregulated human conjunctival epithelial cell ICAM-1 expression to the same extent as rTNF-α. ICAM-1 upregulation could be completely blocked with anti-TNF-α. Preincubation of conjunctival mast cells with olopatadine significantly blocked the ability of supernates to upregulate ICAM-1 on conjunctival epithelial cells. ICAM-1 expression could be restored by adding rTNF-α to the olopatadine-preincubated mast cell supernates. Conclusions Olopatadine is able to significantly decrease the anti-immunoglobulin E mast cell supernate-mediated upregulation of ICAM-1 on human conjunctival epithelial cells in vitro. This seems to be mediated through an effect on a TNF-α–specific mechanism.

Primary ocular posttransplantation lymphoproliferative disorder is rare. Epstein-Barr virus is implicated as the cause as a result of systemic immunosuppression after transplant surgery. We studied a patient who developed ocular posttransplantation lymphoproliferative disorder after orthotopic liver transplantation. Slitlamp and light microscopic photographs confirmed the diagnosis.

Interstitial keratitis is a rarely occurring sign in sarcoidosis. We report a case of childhood sarcoidosis with initial ocular presentation as interstitial keratitis. Erythema nodosum preceded the ocular findings by 4 years and arthritis developed 1 year after ocular findings developed. Although no history of intraocular inflammation was present, there was evidence of inactive chorioretinitis and vitritis found at initial presentation. Systemic steroids were used to treat the patient's skin lesions and topical steroids to treat his corneal inflammation. He developed posterior uveitis and optic nerve edema both of which were responsive to oral steroids. Two years after the presentation of his ocular findings, the onset of pulmonary symptoms and resultant transbronchial biopsy confirmed his diagnosis of sarcoidosis.

A double-masked, randomised, placebo-controlled study was conducted to evaluate the effectiveness of nedocromil 2% eyedrops, a mast cell stabiliser, in 20 symptomatic patients with vernal conjunctivitis. A 1-week baseline period was followed by 6 weeks of treatment. Clinical examination and cytological evaluation of tear fluid were performed weekly, and the patients recorded their subjective assessment on a daily diary card. The nedocromil group showed significantly less hyperaemia in the course of treatment than did the placebo group, and significantly less itching at all visits compared with baseline itching. In the nedocromil-treated group, but not in the placebo group, the number of neutrophils, eosinophils and lymphocytes in tears decreased significantly during some treatment weeks when compared with baseline. The overall assessment of treatment efficacy by both clinician and patient was significantly in favour of nedocromil treatment over placebo.

The proinflammatory cytokine TNFα is believed to play an important role in ocular surface inflammation such as conjunctivitis (allergic, bacterial, and viral) and dry eye disease.1-3 However, the regulation of TNFα on the human conjunctival surface has not been studied. It has been demonstrated in vitro that HCECs respond to recombinant TNFα with proinflammatory responses consistent with those observed in vivo, such as the upregulation of intercellular adhesion molecule (ICAM)-1 expression and increased amounts of the chemokine IL-8. However, regulation of the receptor for TNFα has not been studied in HCECs. In ocular allergic inflammation, conjunctival mast cells are an important source of TNFα. We have previously demonstrated that IgE-activated human conjunctival mast cell supernates upregulate the expression of ICAM-1 on HCECs by a mechanism specifically dependent on TNFα released from the conjunctival mast cell.4 Our work has further suggested that conjunctival mast cell supernates render HCECs more sensitive to TNFα-mediated ICAM-1 upregulation. In these experiments, we demonstrated that TNFα in mast cell supernates could promote ICAM-1 upregulation at log (10−3) lower concentrations than recombinant TNFα alone.4 We hypothesized that IgE-activated conjunctival mast cell supernates upregulate the expression of TNFR1 on HCECs and that the upregulation of TNFR1 expression results in increased sensitivity to TNFα-mediated activation responses (e.g., upregulation of ICAM-1). Our first experiments confirmed the first part of our hypothesis, demonstrating that supernates from IgE-activated conjunctival mast cells upregulate the expression of TNFR1 on primary HCECs. However, because purified conjunctival mast cell supernates require large amounts of cadaveric conjunctival tissues, we sought an alternative mechanism to upregulate TNFR1 expression and to test the biological significance of changes in expression of this receptor. Although TNFR1 expression has not been studied in HCECs, in other types of cells it has been demonstrated that TNFR1 is constitutively expressed and that it can, in some cases, be induced at the transcriptional level by IFNγ.5 TNFR1 is primarily regulated posttranscriptionally by the metalloprotease, TNFα-converting enzyme (TACE; also known as a disintegrin and a metalloprotease [ADAM]-17). TACE-mediated proteolytic cleavage of TNFR1 to its soluble form (sTNFR1) is one important mechanism for the down-regulation of TNFα-mediated inflammatory responses.6 TACE is naturally inhibited in vivo by tissue inhibitor of metalloprotease-3 (TIMP-3) and can be inhibited pharmaceutically by a variety of compounds. For the present study, we examined the mechanisms of TNFR1 expression and shedding in HCECs (primary and IOBA-NHC cell line) using the phorbol ester phorbol myristate acetate (PMA), which activates the protein kinase C pathway (a common pathway of HCEC activation) and stimulates reactive oxygen species (known to activate TACE) and the pharmaceutical TACE inhibitor TNFα protease inhibitor-2 (TAPI-2). With these tools we were able to examine the second part of our hypothesis, which demonstrates how changes in surface expression of TNFR1 affect the threshold of responsiveness to TNFα.

PURPOSE The mechanism by which eosinophils adhere to the ocular surface during allergic inflammation is unknown. This study examined whether the incubation of human conjunctival epithelial cells (HCEs) with tears from allergic subjects promotes eosinophil adhesion, and it examined the effect of treatment with olopatadine on this process. METHODS Allergic subjects (n = 6) and nonallergic subjects (n = 4) were treated in season for 1 week with olopatadine in one eye while the other eye remained untreated. Tears were collected from both eyes with the use of a microcapillary tube. HCEs were acquired by enzymatic digestion of cadaveric conjunctival tissues. Confluent cultures of HCEs were treated with diluted tears for 24 hours before incubation with peripheral blood eosinophils (purified with negative magnetic bead selection). Eosinophil adhesion was measured with an eosinophil peroxidase assay. RESULTS Incubation of HCEs with tears from allergic subjects significantly upregulated eosinophil adhesion compared with eosinophil adhesion to untreated HCEs or with HCEs treated with nonallergic tears and untreated HCEs (P < 0.05). Eosinophil adhesion to HCEs treated with tears from olopatadine-treated allergic subjects was inhibited (P < 0.01) compared with tear-stimulated adhesion observed from untreated eyes. Percentage of inhibition was 43.3% +/- 13.9% (mean +/- SD). Blocking antibodies demonstrated that eosinophil adhesion to HCEs in vitro involved beta2 integrins on eosinophils but not intercellular adhesion molecule-1 on human HCEs. CONCLUSIONS Tears collected from allergic subjects contain bioactivity capable of upregulating eosinophil adhesion to HCEs in vitro. Inhibition of this process by treatment of subjects with olopatadine suggests that some of the cellular targets of this drug may play a role in promoting eosinophil adhesion.

Purpose Increased colonization with Staphylococcus aureus (SA) may play a pro-inflammatory role in atopic keratoconjunctivitis (AKC). We previously demonstrated that human conjunctival epithelial cells are activated by SA peptidoglycan via interaction with Toll-like receptor 2 (TLR2) expressed on human conjunctival epithelial cells in vitro. The purpose of this study was to examine human conjunctival epithelial cells derived from subjects with AKC, seasonal allergic conjunctivitis (SAC), and non-allergic subjects for expression of TLR2. Methods For in vivo studies, human conjunctival epithelial cells were acquired via impression cytology, fixed, and examined for TLR2 expression via flow cytometry. Anti-CD45 was used to gate out contaminating leukocytes. For in vitro studies, human conjunctival epithelial cells and conjunctival mast cells were enzymatically dispersed and purified from cadaveric tissues and cultured on 24-well plates. Conjunctival mast cells were challenged with anti-IgE antibody to obtain activated mast cell supernates. Conjunctival epithelial cells were incubated with the activated mast cell supernates for 24 hours, then harvested using trypsin-EDTA for immunostaining via flow cytometry. Results Staining of conjunctival epithelial cells for TLR2 was positive in 4/5 subjects with AKC, 3/5 subjects with SAC, and 0/3 non-allergic subjects. Intensity of TLR2 staining was significantly greater on conjunctival epithelial cells obtained from AKC subjects compared to conjunctival epithelial cells obtained from non-allergic subjects (p Conclusion While TLR2 may not be constitutively expressed on conjunctival epithelium, ocular allergic inflammation appears to increase TLR2 expression, which is significant in AKC. This increased expression could promote ocular defense against colonizing pathogenic gram-positive bacteria.