A Method for Eliminating Fibroblast Contamination in Mouse and Human Primary Corneal Epithelial Cell Cultures.

PURPOSE: This study was designed to determine if previous approaches to eliminate fibroblast contamination in different cells types would be successful in eliminating fibroblast contamination from human and mouse primary corneal epithelial cell cultures, with the primary goal being to describe a simple, easy, and effective method to culture fibroblast-free primary mouse and human corneal epithelial cell cultures. METHODS: Primary human and mouse corneal stromal cells and epithelial cells were isolated and cultured from human corneal rims and mouse corneas, respectively. Several approaches previously used in other tissue types were evaluated using corneal epithelial cells and mixtures of fibroblasts and epithelial cells to determine the most effective purification method. Methods evaluated included 0.25% trypsin-EDTA, low temperature, mitomycin-C, and dispase. Degree of fibroblast contamination was examined using light microscopy evaluation of cell phenotype, immunofluorescence and western blotting using cell type-specific markers. Anti-pancytokeratin (PanCK) was used as the epithelial immunofluorescence label, and anti-α smooth muscle actin (αSMA) as the fibroblast immunofluorescence label. Epithelial western blot antibodies included PanCK, keratin 12, and E-cadherin, while αSMA, collagen 1A1 and collagen 3A1 were used to identify fibroblasts. RESULTS: Fibroblast contamination of human and mouse primary cornea epithelial cell cultures was best controlled using the 0.25% trypsin-EDTA method. The other methods examined were not effective at eliminating cornea fibroblast contamination. CONCLUSIONS: Trypsin-EDTA digestion is a simple and effective method for controlling fibroblast contamination of cultured primary human and mouse corneal epithelial cells.

Systemic Lupus Erythematosus-associated Retinal Vasculitis Treated with Adalimumab.

PURPOSE: To present a case of refractory systemic lupus erythematosus (SLE)-associated retinal vasculitis that responded to the anti-tumor necrosis factor (TNF)-alpha inhibitor adalimumab as corticosteroid-sparing therapy. METHODS: Descriptive case report of a patient with SLE with retinal vasculitis complicated by an ischemic retinal vein occlusion and cystoid macular edema. RESULTS: A 30-year-old female patient with a history of SLE presented with retinal vasculitis and an ischemic, branch retinal vein occlusion with macular edema in the left eye. Oral corticosteroid was administered along with mycophenolate mofetil (MMF) as a corticosteroid-sparing agent. Despite MMF therapy, the patient developed an exacerbation of her vasculitis with the involvement of both eyes. Adalimumab was initiated with a resultant resolution of retinal vasculitis as a corticosteroid-sparing strategy with over 2 years of follow-up. CONCLUSION: Anti-TNF-alpha therapy with adalimumab may be effective as a corticosteroid-sparing agent in select patients with ocular inflammation associated with SLE.

Update on biologic therapies for juvenile idiopathic arthritis-associated uveitis.

Juvenile idiopathic arthritis (JIA) is the most common rheumatic disease of childhood, and juvenile idiopathic associated uveitis (JIA-U) is the most frequently noted extra-articular manifestation. JIA-U can present asymptomatically and lead to ocular complications, so regular screening and monitoring are needed to prevent potentially sight-threatening sequelae. Topical glucocorticoids such as prednisolone acetate are usually the first line of treatment for anterior uveitis associated with JIA-U, but long-term use may be associated with cataract, ocular hypertension and glaucoma. Disease modifying anti-rheumatic drugs (DMARDs) such as methotrexate allow tapering of the corticosteroids to prevent long-term complications. Biologic therapies have been increasingly used as targeted therapies for JIA-U, particularly monoclonal antibodies targeting the proinflammatory cytokine TNF-α such as adalimumab and infliximab. One recent, multicenter, prospective, randomized clinical trial provided evidence of the efficacy of adalimumab with methotrexate for JIA-U compared to methotrexate alone. Another clinical trial studying the interleukin-6 inhibitor tocilizumab for JIA-U showed promise in tapering topical corticosteroids. Additionally, JAK inhibitors are emerging biologic therapies for JIA-U in patients refractory to TNF-α inhibitors, with a clinical trial assessing the efficacy of baricitinib for JIA-U underway. While clinical trials on these novel biologics are limited, further investigation of these agents may provide additional therapeutic options for JIA-U.

World Health Organization High Priority Pathogens: Ophthalmic Disease Findings and Vision Health Perspectives.

Recent Ebola epidemics, the ongoing COVID-19 pandemic, and emerging infectious disease threats have highlighted the importance of global infectious diseases and responses to public health emergencies. Ophthalmologists are essential health care workers who provide urgent and emergent vision care services during outbreaks and address the ocular consequences of epidemic and pandemic infectious diseases. In 2017, the World Health Organization (WHO) identified high priority pathogens likely to cause a future epidemic with the goal of guiding research and development to improve diagnostic tests, vaccines, and medicines. These measures were necessary to better inform and respond to public health emergencies. Given the ocular complications associated with emerging infectious diseases, there is a need to recognize the ophthalmic sequelae for future vision health preparedness for potential future outbreaks. The WHO High Priority pathogens list provides a roadmap for ophthalmologists and subspecialty providers that will guide strategic areas of research for clinical care and preparedness for future pandemic threats. This review summarizes these key viral pathogens, summarizes major systemic disease findings, and delineates relevant ocular complications of the WHO High Priority pathogens list, including Crimean-Congo hemorrhagic fever, Filovirus diseases (Ebola virus disease and Marburg hemorrhagic fever), human Coronaviruses, Lassa Fever, Nipah virus infection, Zika, and Rift Valley fever.

Clinical management of an outbreak of nutritionally variant streptococcus endophthalmitis following intravitreal bevacizumab injection.

BACKGROUND: The management of an outbreak of endophthalmitis associated with intravitreal bevacizumab represents a challenging real-time process involving identification of cases, treatment and mitigation measures during the outbreak. We summarize the clinical presentation and management of a cluster of endophthalmitis cases from contaminated bevacizumab, in addition to mathematical probabilistic assessment of the number of cases that define an outbreak. METHODS: A retrospective study was conducted to assess the management of an endophthalmitis outbreak after intravitreal bevacizumab (IVB) administration. Demographic data, clinical information, individual patient management and public health reporting measures were reviewed. Outcomes of patients who received prophylactic antibiotics for endophthalmitis prevention were also reviewed. Binomial tail probability calculations were performed to determine the likelihood of clusters of endophthalmitis that could inform when an outbreak was evolving that would warrant more public health notification measures and communication. RESULTS: Forty-five eyes of 42 patients who received IVB from a single batch were reviewed. Four cases of endophthalmitis from Granulicatella adiacens, a nutritionally-variant Streptococcus species, were treated successfully with intravitreal antibiotics ± vitrectomy. Thirty-four of the remaining 41 eyes were treated with prophylactic intravitreal vancomycin with no additional cases of endophthalmitis. Outbreak management also included CDC, ASRS and public health authority notification. Binominal tail probabilities demonstrated the rarity of clusters from a single batch (i.e. ~ 1/10,000 for 2 cases; 1/2 million for 3 cases). However, given the U.S. scale of IVB administration, there is an 87% chance of a cluster ≧ 2 and a 1% chance of a cluster ≧ 3 cases annually, which may guide outbreak management. A process diagram was developed to incorporate patient management and public health measures when an outbreak is suspected. CONCLUSION: Intravitreal antibiotics and vitrectomy were effective in the individual management of cases of endophthalmitis, and no serious adverse events occurred with prophylactic intravitreal vancomycin for at-risk eyes. Best practices for outbreaks should be evaluated, given their likelihood within the U.S. and the sight-threatening consequences of endophthalmitis.

Toxoplasmosis chorioretinitis mimicking acute retinal necrosis associated with local corticosteroid.

BACKGROUND: The cases discussed highlight the atypical presentation and diagnostic dilemmas of toxoplasmosis with fulminant retinal necrosis and the potentially devastating visual outcomes of toxoplasma chorioretinitis following local corticosteroid exposure. CASE PRESENTATION: We report a series of three patients who presented with toxoplasmosis mimicking severe acute retinal necrosis. Patients were between 59 and 77 years old and had been exposed to local corticosteroids preceding our evaluation. All patients demonstrated diffuse retinal whitening with severe vision loss on presentation. Polymerase chain reaction testing (PCR) was diagnostic in two patients, and histopathologic examination of a vitrectomy specimen was diagnostic in one patient. All cases of retinitis resolved with anti-parasitic medication; however, visual acuity failed to improve in all patients due to disease severity and presentation. CONCLUSIONS: Local corticosteroid injection may trigger or exacerbate toxoplasmosis chorioretinitis, leading to fulminant retinal necrosis and severe vision loss. Toxoplasma chorioretinitis should be considered in the differential diagnosis of patients presenting with clinical features of acute retinal necrosis, particularly following local corticosteroid injection regardless of their baseline systemic immune status. Diagnostic vitrectomy may be helpful in patients in whom PCR testing is negative and ocular toxoplasmosis is suspected.

Pathogenesis of Uveitis in Ebola Virus Disease Survivors: Evolving Understanding from Outbreaks to Animal Models.

Ebola virus disease (EVD) and emerging infectious disease threats continue to threaten life, prosperity and global health security. To properly counteract EVD, an improved understanding of the long-term impact of recent EVD outbreaks in West Africa and the Democratic Republic of Congo are needed. In the wake of recent outbreaks, numerous health sequelae were identified in EVD survivors. These findings include joint pains, headaches, myalgias, and uveitis, a vision-threatening inflammatory condition of the eye. Retrospective and more recent prospective studies of EVD survivors from West Africa have demonstrated that uveitis may occur in 13-34% of patients with an increase in prevalence from baseline to 12-month follow-up. The clinical spectrum of disease ranges from mild, anterior uveitis to severe, sight-threatening panuveitis. Untreated inflammation may ultimately lead to secondary complications of cataract and posterior synechiae, with resultant vision impairment. The identification of Ebola virus persistence in immune privileged organs, such as the eye, with subsequent tissue inflammation and edema may lead to vision loss. Non-human primate models of EVD have demonstrated tissue localization to the eye including macrophage reservoirs within the vitreous matter. Moreover, in vitro models of Ebola virus have shown permissiveness in retinal pigment epithelial cells, potentially contributing to viral persistence. Broad perspectives from epidemiologic studies of the outbreak, animal modeling, and immunologic studies of EVD survivors have demonstrated the spectrum of the eye disease, tissue specificity of Ebola virus infection, and antigen-specific immunologic response. Further studies in these areas will elucidate the mechanisms of this highly prevalent disease with the potential for improved therapeutics for Ebola virus in immune-privileged sites.

SARS-CoV-2 and the Eye: Implications for the Retina Specialist from Human Coronavirus Outbreaks and Animal Models.

PURPOSE: The current SARS-CoV-2 pandemic has escalated rapidly since December 2019. Understanding the ophthalmic manifestations in patients and animal models of the novel coronavirus may have implications for disease surveillance. Recognition of the potential for viral transmission through the tear film has ramification for protection of patients, physicians, and the public. METHODS: Information from relevant published journal articles was surveyed using a computerized PubMed search and public health websites. We summarize current knowledge of ophthalmic manifestations of SARS-CoV-2 infection in patients and animal models, risk mitigation measures for patients and their providers, and implications for retina specialists. RESULTS: SARS-CoV-2 is efficiently transmitted among humans, and while the clinical course is mild in the majority of infected patients, severe complications including pneumonia, acute respiratory distress syndrome, and death can ensue, most often in elderly patients and individuals with co-morbidities. Conjunctivitis occurs in a small minority of patients with COVID-19 and SARS-CoV-2 RNA has been identified primarily in association with conjunctivitis. Uveitis has been observed in animal models of coronavirus infection and cotton-wool spots have been reported recently. CONCLUSION: SARS-CoV-2 and other coronaviruses have been rarely associated with conjunctivitis. The identification of SARS-CoV and SARS-CoV-2 RNA in the tear film of patients and its highly efficient transmission via respiratory aerosols supports eye protection, mask and gloves as part of infection prevention and control recommendations for retina providers. Disease surveillance during the COVID-19 pandemic outbreak may also include ongoing evaluation for uveitis and retinal disease given prior findings observed in animal models and a recent report of retinal manifestations.