Effects of ROCK Inhibitors on Apoptosis of Corneal Endothelial Cells in CMV-Positive Posner-Schlossman Syndrome Patients.

Purpose: To examine the role of aqueous tumor necrosis factor α (TNF-α)-RhoA-Rho kinase (ROCK) signaling in cytomegalovirus (CMV)-induced apoptosis and the barrier function of cultured human corneal endothelial cells (hCECs) in CMV-positive Posner-Schlossman syndrome (CMV+/PSS) patients. Methods: Aqueous levels of TNF-α, IL-8, IL-10, and several other cytokines in 19 CMV+/PSS patients and 20 healthy control subjects were quantitated using a multiplex assay. The expression of active RhoA in hCECs post-CMV infection was determined using western blotting (WB). The expression levels of TNF-α and nuclear factor kappa B (NF-κB) in CMV-infected hCECs were examined by immunocytochemistry (ICC) and WB with and without ROCK inhibitors. The apoptotic rate and barrier integrity in CMV-infected hCECs were also examined. Results: The expression levels of TNF-α, monocyte chemoattractant protein-1 (MCP-1), IL-8, and IL-10 were upregulated in the aqueous humor of CMV+/PSS patients, and among these upregulated cytokines aqueous TNF-α was negatively correlated with the number of corneal endothelial cells. In CMV-infected hCECs, upregulation of TNF-α and NF-κB was determined by WB and ICC. In hCECs, CMV infection induced apoptosis and significantly impaired cell-cell contacts, effects that were attenuated by treatment with a ROCK inhibitor. Conclusions: Aqueous TNF-α was upregulated in CMV+/PSS patients, which may have triggered corneal endothelial cell loss. Modulation of TNF-α, including its downstream Rho-ROCK signaling, could serve as a novel treatment modality for corneal endothelial cell loss in CMV+/PSS patients.

Human papillomavirus in ocular malignant tumours: a study from a tertiary eye care centre in North India.

OBJECTIVE: The present study aims to detect the presence of human papillomavirus (HPV) in ocular malignant tumours, including retinoblastoma, eyelid squamous cell carcinoma (SCC), and sebaceous gland carcinoma (SGC), in the North Indian population. DESIGN: Prospective observational non randomized study. PARTICIPANTS: In this study, 142 prospective cases of ocular malignant tumours (retinoblastoma, SGC, and SCC) were included. METHODS: HPV was detected by multiplex PCR using PGMY09/11 primers in 142 patients with ocular malignancies. This was followed by genotyping using linear array (reverse hybridization). RESULTS: Of the 142 tumours studied, 72 were retinoblastoma, 30 SGC, and 40 SCC. The HPV genome was detected in 2.8% (4 of 142) of cases by multiplex PCR; all positive cases (4 of 40) were SCC. Genotyping revealed that all positives belonged to the high-risk HPV16 genotype. HPV-positive SCC patients had better disease-free survival. Retinoblastoma and SGC cases were negative for HPV. CONCLUSIONS: Low prevalence of HPV in ocular malignancies was observed in this study. The HPV genome was detected only in ocular squamous cell carcinoma cases and these patients were associated with better prognosis. HPV may not have a role in retinoblastoma and SGC in the North Indian population.

An eye on hepatitis C: a review / O olho na hepatite C: uma revisão

ABSTRACT This review aims to caution ophthalmologists about the ocular consequences leading to the diagnosis of hepatitis C virus infection. In addition, in this context, the effects of old and new drugs are discussed in the ophthalmological setting. The importance of early diagnosis and the curative treatment of the disease has been reported in the national and international literature, demonstrating that its progression has important implications for daily clinical and surgical ophthalmological practice. Despite the scarcity of studies on new direct-acting antiviral drugs, fewer side effects of these drugs have been shown when compared with conventional interferon treatment with or without ribavirin. The ophthalmologist's risk of becoming infected, as demonstrated by the presence of the virus in ocular structures, and the possibility of contamination, is also discussed.

RESUMO Esta revisão objetiva alertar os oftalmologistas sobre as consequências oculares que levam ao diagnóstico da infecção pelo vírus da Hepatite C. Além disso, neste contexto, os efeitos de drogas antigas e novas são discutidos no cenário oftalmológico. A importância do diagnóstico precoce e do tratamento curativo da doença tem sido relatada na literatura nacional e internacional, demonstrando que sua progressão tem implicações importantes para a prática oftalmológica diária. Apesar da escassez de estudos sobre novos medicamentos antivirais de ação direta, foram demonstrados menos efeitos colaterais desses medicamentos quando comparados ao tratamento convencional com interferon, com ou sem ribavirina associado ou não à rivabirina. O risco do oftalmologista de se infectar, como demonstrado pela presença do vírus nas estruturas oculares, e a possibilidade de contaminação, também é discutido.

Management of herpes simplex virus epithelial keratitis.

PURPOSE OF REVIEW: To review recent advancements in the management of herpes simplex virus (HSV) epithelial keratitis. RECENT FINDINGS: Trifluridine eye drop, acyclovir (ACV) ointment, ganciclovir gel, and oral ACV are still the main therapeutic agents. Cryopreserved amniotic membrane has been recently used as an adjuvant treatment. Resistance to ACV has become a concerning issue. The animal models of HSV vaccine are able to reduce HSV keratitis. New antivirals are under development. SUMMARY: Current cases of HSV epithelial keratitis are manageable with available medications, but new advancements are required to decrease disease burden in the future. HSV vaccine can be revolutionary.

Influence of an immunodominant herpes simplex virus type 1 CD8+ T cell epitope on the target hierarchy and function of subdominant CD8+ T cells.

Herpes simplex virus type 1 (HSV-1) latency in sensory ganglia such as trigeminal ganglia (TG) is associated with a persistent immune infiltrate that includes effector memory CD8+ T cells that can influence HSV-1 reactivation. In C57BL/6 mice, HSV-1 induces a highly skewed CD8+ T cell repertoire, in which half of CD8+ T cells (gB-CD8s) recognize a single epitope on glycoprotein B (gB498-505), while the remainder (non-gB-CD8s) recognize, in varying proportions, 19 subdominant epitopes on 12 viral proteins. The gB-CD8s remain functional in TG throughout latency, while non-gB-CD8s exhibit varying degrees of functional compromise. To understand how dominance hierarchies relate to CD8+ T cell function during latency, we characterized the TG-associated CD8+ T cells following corneal infection with a recombinant HSV-1 lacking the immunodominant gB498-505 epitope (S1L). S1L induced a numerically equivalent CD8+ T cell infiltrate in the TG that was HSV-specific, but lacked specificity for gB498-505. Instead, there was a general increase of non-gB-CD8s with specific subdominant epitopes arising to codominance. In a latent S1L infection, non-gB-CD8s in the TG showed a hierarchy targeting different epitopes at latency compared to at acute times, and these cells retained an increased functionality at latency. In a latent S1L infection, these non-gB-CD8s also display an equivalent ability to block HSV reactivation in ex vivo ganglionic cultures compared to TG infected with wild type HSV-1. These data indicate that loss of the immunodominant gB498-505 epitope alters the dominance hierarchy and reduces functional compromise of CD8+ T cells specific for subdominant HSV-1 epitopes during viral latency.

Feline Infectious Peritonitis: Immunohistochemical Features of Ocular Inflammation and the Distribution of Viral Antigens in Structures of the Eye.

Feline infectious peritonitis (FIP) is a serious, widely distributed systemic disease caused by feline coronavirus (FCoV), in which ocular disease is common. However, questions remain about the patterns of ocular inflammation and the distribution of viral antigen in the eyes of cats with FIP. This study characterized the ocular lesions of FIP including the expression of glial fibrillary acidic protein and proliferating cell nuclear antigen by Müller cells in the retina in cases of FIP and to what extent macrophages are involved in ocular inflammation in FIP. Immunohistochemistry for FCoV, CD3, CD79a, glial fibrillary acidic protein, calprotectin, and proliferating cell nuclear antigen was performed on paraffin sections from 15 naturally occurring cases of FIP and from controls. Glial fibrillary acidic protein expression was increased in the retina in cases of FIP. Müller cell proliferation was present within lesions of retinal detachment. Macrophages were present in FIP-associated ocular lesions, but they were the most numerous inflammatory cells only within granulomas (2/15 cats, 13%). In cases of severe inflammation of the ciliary body with damage to blood vessel walls and ciliary epithelium (3/15, 20%), some macrophages expressed FCoV antigens, and immunolabeling for calprotectin on consecutive sections suggested that these FCoV-positive macrophages were likely to be recently derived from blood. In cases of severe and massive inflammation of most ocular structures (4/15, 26%), B cells and plasma cells predominated over T cells and macrophages. These results indicate that gliosis can be present in FIP-affected retinas and suggest that breakdown of the blood-ocular barrier can allow FCoV-bearing macrophages to access the eye.

Ganciclovir ophthalmic gel treatment shortens the recovery time and prevents complications in the adenoviral eye infection.

The purpose of this study was to determine the effectiveness of ganciclovir ophthalmic gel (GOG) in the treatment of adenoviral eye infection (AEI) by looking at the effect of the drug on shortening recovery time, preventing transmission, reducing sequelae, and on complications such as corneal infiltrates and conjunctival pseudomembranes. 200 patients' examination records were evaluated retrospectively. Patients who were within the first 3 days of AEI were divided into two groups: Group 1 with 100 patients who used artificial tears as treatment, and Group 2 with 100 patients who used GOG plus artificial tears (GAT). All patients underwent an eye examination by the same ophthalmologist on the 1st, 5th, 10th, and 15th day after treatment. Using the examination records, variables were compared using SPSS 22.0. There was a statistically significant difference between Groups 1 and 2. Group 2 showed better and faster response to treatment. There was less transmission to the contralateral eye and environment, and less formation of corneal subepithelial infiltrate and conjunctival pseudomembrane in Group 2. Only three patients in Group 2 had corneal involvement. A comparison of each group pre-treatment and during treatment revealed improved signs and symptoms in Group 2 (p < 0.005). The study showed a trend toward more rapid improvement, less corneal and conjunctival involvement, and less transmission to the contralateral eye and environment in the GAT group. These results need to be confirmed by additional studies.

Resident corneal c-fms(+) macrophages and dendritic cells mediate early cellular infiltration in adenovirus keratitis.

The cornea contains a heterogeneous population of antigen-presenting cells with the capacity to contribute to immune responses. Adenovirus keratitis is a severe corneal infection with acute and chronic phases. The role of resident corneal antigen-presenting cells in adenovirus keratitis has not been studied. We utilized transgenic MaFIA mice in which c-fms expressing macrophages and dendritic cells can be induced to undergo apoptosis, in a mouse model of adenovirus keratitis. Clinical keratitis and recruitment of myeloperoxidase and CD45(+) cells were diminished in c-fms depleted, adenovirus infected mice, as compared to controls, consistent with a role for myeloid-lineage cells in adenovirus keratitis.

Histological features of Cytomegalovirus-related corneal graft infections, its associated features and clinical significance.

AIM: To describe the histological features of ITALIC! Cytomegalovirus (CMV)-related corneal graft infections, its associated features and clinical significance. METHODS: This was a retrospective histological study of 48 consecutive cases of failed repeat penetrating keratoplasty cases with a clinical diagnosis of allograft rejection from 2011 to 2013. CMV infection was confirmed with CMV antibody immunohistochemistry (IHC) and electron microscopy. Additional CD163 and CD68 IHCs for macrophages were also performed. Clinical data and previous graft histology were then reviewed. RESULTS: Mean incidence of CMV infection in corneal graft rejection buttons was 6.3% per year. 3/48 graft buttons were CMV antibody positive. Histological features of CMV graft infection include: (1) stromal keratocytes with cytopathic changes; (2) lack of inflammation, only occasional macrophages present and (3) absence of vascularisation. None of the patients had a history of active CMV infection. CONCLUSION: CMV infection is not limited as endotheliitis, but extends into the corneal stroma, and is a potential reservoir for graft infection, especially in partial thickness endothelial surgery. Clinical features are often non-specific, although glaucoma was present in our patients. CMV-infected grafts showed CD163-positive M2 macrophages in close association with the infected keratocytes, suggesting that the macrophage may be important in CMV graft infection. Histological examination with CMV IHC is a useful method to detect CMV infection postoperatively. Post penetrating keratoplasty, CMV systemic treatment with valganciclovir can prevent graft infection and failure. Boston keratoprosthesis may be a potential alternative surgery in active CMV infections that obviates the need for systemic therapy.

Human Papilloma Virus Infection Does Not Predict Response to Interferon Therapy in Ocular Surface Squamous Neoplasia.

PURPOSE: To identify the frequency of human papilloma virus (HPV) in ocular surface squamous neoplasia (OSSN) and to evaluate differences in clinical features and treatment response of tumors with positive versus negative HPV results. DESIGN: Retrospective case series. PARTICIPANTS: Twenty-seven patients with OSSN. METHODS: Ocular surface squamous neoplasia specimens were analyzed for the presence of HPV. Clinical features and response to interferon were determined retrospectively and linked to the presence (versus absence) of HPV. MAIN OUTCOME MEASURES: Clinical characteristics of OSSN by HPV status. RESULTS: Twenty-one of 27 tumors (78%) demonstrated positive HPV results. The HPV genotypes identified included HPV-16 in 10 tumors (48%), HPV-31 in 5 tumors, HPV-33 in 1 tumor, HPV-35 in 2 tumors, HPV-51 in 2 tumors, and a novel HPV in 3 tumors (total of 23 tumors because 1 tumor had 3 identified genotypes). Tumors found in the superior limbus were more likely to show positive HPV results (48% vs. 0%; P=0.06, Fisher exact test). Tumors with positive HPV-16 results were larger (68 vs. 34 mm2; P=0.08, Mann-Whitney U test) and were more likely to have papillomatous morphologic features (50% vs. 12%; P=0.07, Fisher exact test) compared with tumors showing negative results for HPV-16. Human papilloma virus status was not found to be associated with response to interferon therapy (P=1.0, Fisher exact test). Metrics found to be associated with a nonfavorable response to interferon were male gender and tumors located in the superior conjunctivae. CONCLUSIONS: The presence of HPV in OSSN seems to be more common in lesions located in the nonexposed, superior limbus. Human papilloma virus presence does not seem to be required for a favorable response to interferon therapy.

In Vivo confocal microscopic changes of the corneal epithelium and stroma in patients with herpes zoster ophthalmicus.

PURPOSE: To analyze the density and morphology of corneal epithelial cells and keratocytes by in vivo confocal microscopy (IVCM) in patients with herpes zoster ophthalmicus (HZO) as associated with corneal innervation. DESIGN: Prospective, controlled and masked cross-sectional study. METHODS: setting: Single-center study. PATIENTS: Thirty eyes with the diagnosis HZO and their contralateral clinically unaffected eyes, 15 eyes of 15 normal controls. intervention procedures: In vivo confocal microscopy and corneal esthesiometry of the central cornea. MAIN OUTCOME MEASURES: Changes in morphology and density of the superficial and basal epithelial cells and stromal keratocytes, and correlation with corneal sensation. RESULTS: The density of superficial epithelial cells in HZO eyes with severe sensation loss (766.5 ± 25.2 cells/mm(2)) was significantly lower than both healthy control eyes (1450.23 ± 150.83 cells/mm(2)) and contralateral unaffected eyes (1974.13 ± 298.24 cells/mm(2)) (P = .003). Superficial epithelial cell size (1162.5 µm(2)) was significantly larger in HZO eyes with severe loss of sensation, as compared to contralateral (441.46 ± 298.14) or healthy eyes (407.4 ± 47.2µm(2); all P < .05). The density of basal epithelial cells, anterior keratocytes, and posterior keratocytes did not show statistical significance between patients, controls, and contralateral unaffected eyes. Changes in superficial epithelial cell density and morphology correlated strongly with corneal sensation. CONCLUSIONS: In vivo confocal microscopy reveals profound HZO-induced changes in the superficial epithelium, as demonstrated by increase in cell size, decrease in cell density, and squamous metaplasia. We demonstrate that these changes strongly correlate with changes in corneal innervation in eyes affected by HZO.

Ultrastructure of adenovirus keratitis.

PURPOSE: We determined the ultrastructure of mouse adenovirus keratitis, a model for human adenovirus keratitis. METHODS: Adenovirus keratitis was induced in C57Bl/6j mice by intrastromal injection of human adenovirus species D type 37 (HAdV-D37) with a heat-pulled, glass, micropipette needle under compressed air. At select time points after infection, mice were euthanized and their corneas removed, fixed, and sectioned at 70-nm thickness for electron microscopy. RESULTS: Injection of HAdV-D37 into the mouse corneal stroma placed virus predominantly in the pericellular corneal stromal matrix. Virus was seen bound to and entering stromal cells at 1 and 2 hours after infection, respectively. Cell membrane transit by virus was seen to involve two distinct structures resembling caveolae and macropinosomes. However, later during infection intracellular virus was not seen within membrane-bound organelles. By 8 hours after infection, intracellular virus had accumulated into densely packed, perinuclear arrays. Virus disassembly was not obvious at any time point after infection. Infiltrating neutrophils seen by one day after infection had engulfed degraded stromal cells by 4 days after infection. CONCLUSIONS: By transmission electron microscopy, injected HAdV-D37 readily enters stromal cells in the C57Bl/6j mouse cornea and induces stromal inflammation, as was shown previously by light microscopy. However, electron microscopy also revealed dense, static arrays of intracytoplasmic virus, suggesting a block in viral capsid disassembly and viral DNA nuclear entry. These findings may explain why human adenoviruses do not replicate in the mouse corneal stroma.

Morphological characteristics of conjunctival squamous papillomas in relation to human papillomavirus infection.

OBJECTIVE: To determine the prevalence of a broad spectrum of human papillomavirus (HPV) types in conjunctival papillomas and a possible difference in clinical and histopathological presentation of HPV-positive and HPV-negative papillomas. METHODS: Formalin-fixed, paraffin-embedded papilloma tissue specimens obtained from 25 patients were analysed using six different PCR-based methods targeting 87 HPV types from four different papillomavirus (PV) genera: α-PV, ß-PV, γ-PV and µ-PV, and in situ hybridisation for HPV-6/HPV-11. Slides were reviewed for pedunculated or sessile growth, the presence of goblet cells, keratinising or non-keratinising epithelium, elastosis, atypia and koilocytes. RESULTS: α-PV types HPV-6 and HPV-11 were detected in 19/25 (76%) conjunctival papilloma tissue specimens, 9 (47%) of which were also HPV-6/HPV-11 positive with in situ hybridisation. Six different ß-PV types-HPV-9, HPV-12, HPV-20, HPV-21, HPV-22, HPV-24-were additionally detected in four cases, all of which were also HPV-6/HPV-11 positive. No γ-PVs or µ-PVs were found in any of the tested tissues samples. Extralimbal location (p=0.021), presence of goblet cells (p=0.005), non-keratinising squamous epithelium (p=0.005), and absence of elastosis (p=0.005) were associated with the presence of HPV-6/HPV-11. CONCLUSIONS: We demonstrated that certain clinical and histological features are more frequently associated with HPV infection and that HPV genera other than α-PV are most probably not significant factors in conjunctival papilloma occurrence.

Acute retinal necrosis associated with Epstein-Barr virus: immunohistopathologic confirmation.

IMPORTANCE: Acute retinal necrosis (ARN) is an infectious retinitis primarily caused by the herpesviruses. Although the Epstein-Barr virus (EBV) has been implicated as a cause of ARN, to our knowledge, there has been no histopathologic documentation. We report the clinical history and histopathologic confirmation that EBV can cause ARN. OBSERVATIONS: Clinical course and histopathology of a patient diagnosed with ARN caused by infection with EBV confirmed by molecular pathology. CONCLUSIONS AND RELEVANCE: Epstein-Barr virus is a recognized cause of intraocular inflammation and has been implicated as a possible cause of ARN. However, to our knowledge, tissue demonstration of EBV in a patient with ARN has not previously been reported. We identified the organism in the necrotic retina of a patient receiving immunosuppression because of idiopathic pulmonary fibrosis.

Detecting human papillomavirus in ocular surface diseases.

PURPOSE: Human papillomavirus (HPV) infection has been implicated as a possible inducing factor for benign and neoplastic ocular surface diseases such as pterygia and ocular-surface squamous neoplasia (OSSN). However, the wide range in HPV prevalence previously reported for both diseases adds controversy to, and highlights the limitations of, this field. The aim of this study was to determine the prevalence of HPV in pterygia and OSSN and to devise a standardized approach for detecting viral DNA in ocular tissue samples. METHODS: DNA was extracted from a variety of specimens (n = 160), including formalin-fixed paraffin-embedded tissue shavings, fresh tissue, and cultured cells. Nested PCR for HPV with consensus and subtype-specific primers was used to detect viral DNA. Confirmatory assays, including molecular sequencing, histology, and immunohistochemistry for HPV E6 protein and p16 were also performed. RESULTS: HPV was not detected in pterygia or normal conjunctiva. However, 6.5% (3/46) of OSSN samples were HPV-positive by PCR, sequencing, and immunohistochemistry. Positive cases were all squamous cell carcinoma of the conjunctiva (SCCC), the most severe form of OSSN, representing 12.5% (3/24) of SCCCs in our cohort. HPV-16 was the genotype identified in each case and this correlated with the presence of koilocytes and intense immunoreactivity for p16. Our study found no association between pterygia and OSSN with other oncogenic viruses, such as EBV or CMV, as they were just as prevalent in normal conjunctiva. CONCLUSIONS: The low prevalence of HPV-16 in ocular surface disease suggests infection is not a cause but a cofactor in disease development.

Corneal sensitivity may decrease in adenoviral epidemic keratoconjunctivitis--a confocal microscopic study.

OBJECTIVES: To report a new clinical finding, decreased corneal sensitivity, in epidemic keratoconjunctivitis and to evaluate this sign with corneal confocal microscopy. METHODS: Forty-one eyes of 28 patients who developed corneal infiltrates after an outbreak of epidemic keratoconjunctivitis were included in the study. Clinical and confocal microscopic findings are described. RESULTS: In this outbreak of 72 patients, 28 (38.9%) developed corneal infiltrates. The corneal involvement was unilateral in 15 patients (53.6%) and bilateral in 13 patients (46.4%). Corneal sensitivities were measured in 35 eyes of 24 patients and found to be decreased in 26 eyes (74.3%). Decreased corneal sensation was a feature of mainly stage 2 (7 eyes) and stage 3 (11 eyes) keratitis. Corneal sensitivity returned to normal levels in all eyes in a mean of 8.5 days. The main confocal microscopic features during the period of decreased corneal sensitivity were morphologic changes in the infected epithelial cells, extracellular bright microdeposits, infiltration with round inflammatory cells and dendritic cells, increased brightness in the extracellular matrix and the stroma surrounding the corneal nerves, and increased keratocyte activity. The intensity of the inflammatory reaction in the extracellular space and corneal stroma and the reflectivity of the corneal nerves had subsided by the second confocal measurements. CONCLUSION: There may be a transient decrease in the corneal sensitivity during the course of epidemic keratoconjunctivitis. Confocal microscopy can help to evaluate the changes in the cornea during this period. Future studies are needed to understand the nature of this clinical finding.

Bilateral lineal endotheliitis: case report / Endotelite linear bilateral: relato de caso

To report the case of a patient with bilateral herpetic lineal endotheliitis successfully treated with topic steroids and systemic antiviral. 17 year old female with blurred vision, at evaluation localized edema was observed on both corneas associated to Descemet folds and a line of pigmented precipitates. Topic prednisolone and oral acyclovir are initiated with complete resolution of signs and symptoms. Lineal endotheliitis is produced as an answer of endotelial cells to viral infection; maybe due to an immune reaction against some antigens from herpes virus family. It has the potential of relapses even in the absence of viral replication, with secondary untreatable stromal edema. It responds well to antiviral and steroids treatment, although, on those patients who don't improve, is necesary to make additional tests.

Relatar o caso de uma paciente com endotelite linear herpética bilateral tratado com sucesso por meio de corticoides tópicos e antivirais sistêmicos. Paciente do sexo feminino, 17 anos de idade, com a visão turva, na avaliação foi observado edema localizado em ambas as córneas associadas a dobras de Descemet e uma linha de precipitados ceráticos pigmentados. Prednisolona tópica e aciclovir oral foram utilizados com resolução completa dos sinais e sintomas. A endotelite linear é uma resposta das células endoteliais à infecção viral, talvez devido a uma reação imunológica contra alguns antígenos do vírus da família do herpes. Tem o potencial de recidiva, mesmo na ausência de replicação viral, com edema estromal secundário intratável. Ela responde bem ao tratamento antiviral e esteroides, embora, em pacientes que não melhoram, é necessária a realização de testes adicionais.