Unique molecular signatures of microRNAs in ocular fluids and plasma in diabetic retinopathy.

The main objective of this pilot study was to identify circulatory microRNAs in aqueous or plasma that were reflecting changes in vitreous of diabetic retinopathy patients. Aqueous, vitreous and plasma samples were collected from a total of 27 patients undergoing vitreoretinal surgery: 11 controls (macular pucker or macular hole patients) and 16 with diabetes mellitus(DM): DM-Type I with proliferative diabetic retinopathy(PDR) (DMI-PDR), DM Type II with PDR(DMII-PDR) and DM Type II with nonproliferative DR(DMII-NPDR). MicroRNAs were isolated using Qiagen microRNeasy kit, quantified on BioAnalyzer, and profiled on Affymetrix GeneChip miRNA 3.0 microarrays. Data were analyzed using Expression Console, Transcriptome Analysis Console, and Ingenuity Pathway Analysis. The comparison analysis of circulatory microRNAs showed that out of a total of 847 human microRNA probes on the microarrays, common microRNAs present both in aqueous and vitreous were identified, and a large number of unique microRNA, dependent on the DM type and severity of retinopathy. Most of the dysregulated microRNAs in aqueous and vitreous of DM patients were upregulated, while in plasma, they were downregulated. Dysregulation of miRNAs in aqueous did not appear to be a good representative of the miRNA abundance in vitreous, or plasma, although a few potential candidates for common biomarkers stood out: let-7b, miR-320b, miR-762 and miR-4488. Additionally, each of the DR subtypes showed miRNAs that were uniquely dysregulated in each fluid (i.e. aqueous: for DMII-NPDR was miR-455-3p; for DMII-PDR was miR-296, and for DMI-PDR it was miR-3202). Pathway analysis identified TGF-beta and VEGF pathways affected. The comparative profiling of circulatory miRNAs showed that a small number of them displayed differential presence in diabetic retinopathy vs. controls. A pattern is emerging of unique molecular microRNA signatures in bodily fluids of DR subtypes, offering promise for the use of ocular fluids and plasma for diagnostic and therapeutic purposes.

Bilateral serous retinal detachments associated with burkitt lymphoma.

PURPOSE: To report a case of bilateral serous retinal detachments in a patient found to have burkitt lymphoma. OBSERVATIONS: A patient who presented with bilateral serous retinal detachments and "B" symptoms underwent extensive workup and was found to have burkitt lymphoma. Multiagent chemotherapy was initiated with resolution of the serous retinal detachments and visual recovery occurring in parallel to disease remission. CONCLUSIONS AND IMPORTANCE: Burkitt lymphoma can present with serous retinal detachments and should be included in the differential for a patient with bilateral serous retinal detachments along with Vogt-Koyanagi-Harada syndrome.

Role of vitamin E in preventing arteriohyalinization in kidneys of streptozotocin induced diabetic mice.

OBJECTIVE: To evaluate the role of vitamin E on the arteriolar hyalinisation in kidneys of diabetic mice. METHODS: The laboratory-based randomised control trial was conducted at the Department of Anatomy, Army Medical College, Rawalpindi, in collaboration with National Institute of Health, Islamabad, from November 2009 to November 2010. Adult female BALB/C mice were randomly divided into three groups. Group A served as control group. Group B was made diabetic by the intraperitoneal injection of streptozotocin. Group C received streptozotocin injection and was fed with vitamin E (alphatocopherol) supplemented diet. After 12 weeks,the animals were sacrificed and their kidneys were removed for histomorphological study. SPSS 16 was used for statistical analysis. RESULTS: Diabetes caused significant histomorphological changes in arteriole of kidneys of Experimental Group B compared to Control Group A (p>0.05), but these changes were prevented in Group C. In experimental group B, 2(20%) animals had arteriolar hyalinisation of score 1, while score 2 was revealed in 8(80%) animals. Experimental group C showed no hyalinisation in any arteriole. CONCLUSIONS: Vitamin E prevents the arteriohyalinization in kidneys of mice with STZ induced diabetes.

Effect of alphatocopherol on diameter of proximal convoluted tubules of kidney in diabetic mice.

OBJECTIVE: To evaluate the effects of alphatocopherol supplement on proximal convoluted tubular diameter of kidney in diabetic mice. METHODS: The randomised controlled trials was conducted partly at the National Institute of Health (NIH), Islamabad, and partly in Army Medical College, Rawalpindi, from November 2009 to November 2010. Thirty adult female mice BALB/C were randomly divided into three equal groups. Group A served as the control group. Group B was made diabetic by the intraperitoneal injection of streptozotocin. Group C received injection streptozotocin and was fed with alphatocopherol (vitamin E) supplemented diet. After 12 weeks, the animals were sacrificed and their kidneys were removed for histomorphological study. RESULTS: Diabetes caused significant changes in the diameter of proximal tubule of Experimental Group B (diabetic) compared to the controls in Group A, but these changes were prevented in alphatocopherol treated Group C. Tubular diameter in Group B was significantly reduced compared to theControl Group A (p <0.05), but there was no statistical difference in tubular diameter of Group C and Group A (p > 0.05). CONCLUSION: Significant difference in proximal tubular diameter of kidneys between diabetic and alphatocopherol treated diabetic mice confirm that vitamin E does extend a protective role in improving diabetic nephropathy.

Five-year follow-up of macular morphologic changes after rhegmatogenous retinal detachment repair: Fourier domain OCT findings.

PURPOSE: To evaluate serially long-term macular morphologic changes after successful macula-involving rhegmatogenous retinal detachment repair and correlate changes with macular function. METHODS: Repeat Fourier domain optical coherence tomography (FD OCT) imaging and microperimetry (MP-1) testing of 8 of the initial cohort of 17 eyes studied 5 years earlier. RESULTS: The mean follow-up after rhegmatogenous retinal detachment repair was 3.4 months (range, 1-8.5 months) for the first FD OCT and 5 years (range, 3.75-5.75 years) for the follow-up FD OCT. The final postoperative best-corrected visual acuity mean was 20/201 (range, 20/20 to counting fingers). Six eyes with final best-corrected visual acuity >20/40 had an intact external limiting membrane and progressive resolution of photoreceptor inner segment-outer segment junction disruption and/or subretinal fluid on serial FD OCT, which correlated with improvement in macular function on MP-1. Two eyes with poor or worsening best-corrected visual acuity on follow-up had persistent or worsening inner segment-outer segment disruption on serial FD OCT. External limiting membrane was intact in one eye and persistently disrupted in the other. CONCLUSION: Macular function may progressively improve or worsen long-term after successful rhegmatogenous retinal detachment repair. Progressive resolution of subretinal fluid and/or inner segment-outer segment disruption on FD OCT correlated with improvement in macular function, whereas worsening or persistent inner segment-outer segment disruption correlates with worsening or persistently poor visual outcome.

Corneal epithelial proliferation and thickness in a mouse model of dry eye.

Although several studies have previously focused on the conjunctival epithelial response to surface dryness, little is known about the effect of a dry environment on corneal epithelium, which is the most clinically significant tissue affected in dry eye. The aim of this study was to quantitatively evaluate the effect of desiccating stress on the number of proliferating corneal epithelial cells and corneal epithelial thickness in mice placed in a controlled-environment chamber (CEC) that induces dry eye. Corneal epithelial cell proliferation and thickness were studied in 8- to 12-week-old female BALB/c mice placed in the CEC (temperature: 22.3+/-0.7 degrees C; relative humidity: 22.5+/-4.5%; airflow: 15 L/min) for 7 days and compared to a control group of mice with no dry eye. Actively proliferating cells were identified by immunofluorescence using a FITC-conjugated antibody against the Ki-67 protein, a cell proliferation marker expressed during active phases of the cell cycle. To detect the spatial distribution of proliferative cells, Ki-67(+) cells were counted in three areas of the epithelium: center, periphery, and limbus. Corneal epithelial thickness was evaluated in the central cornea after staining with hematoxylin-eosin. Results from each experimental group were compared using the Mann-Whitney test. The number of Ki-67(+) cells observed in the corneal epithelium of mice exposed to the CEC was significantly higher in each area (center: 32.1+/-1.1; periphery: 94.2+/-5.3; limbus: 4.0+/-1.5) than in the control group (center: 13.2+/-1.0, p=0.02; periphery: 42.9+/-2.3, p=0.02; limbus: 0.0, p=0.01). In mice subjected to desiccating stress, a significant number of Ki-67(+) positive cells were detected in the basal and suprabasal cell layers (central area 46%; periphery 30.8%: limbus 0%), whereas in the control group the cells were exclusively distributed through the basal cell layer. Ki-67(+) cells were not found in the corneal stroma or endothelium in any group. The corneal epithelium was found to be significantly thicker in dry eye mice (54.94+/-6.09 microm) as compared to the controls (43.9+/-6.23 microm, p<0.0001) by a mean of 25%. These results demonstrate that desiccating stress increases corneal epithelial turnover and thickness, similar to what is observed in other chronic inflammatory states of other epithelialized surfaces. The CEC can facilitate the study of the regulation of epithelial cell function and turnover at the molecular and cellular levels under desiccating stress conditions.

Modulation of integrin alpha4beta1 (VLA-4) in dry eye disease.

OBJECTIVE: To study the effect of topical application of very late antigen 4 (VLA-4) small-molecule antagonist (anti-VLA-4 sm) in a mouse model of dry eye disease. METHODS: Anti-VLA-4 sm (or control vehicle) was applied topically to mice placed in a controlled-environment chamber. Corneal fluorescein staining and conjunctival T-cell enumeration were performed in the different treatment groups. Real-time polymerase chain reaction was used to quantify expression of inflammatory cytokines in the cornea and conjunctiva. RESULTS: Dry eye syndrome induced increased corneal fluorescein staining, corneal and conjunctival tumor necrosis factor alpha messenger RNA expression, and T-cell infiltration into the conjunctiva. Very late antigen 4 blockade significantly decreased corneal fluorescein staining compared with the untreated dry eye disease and control vehicle-treated groups (P < .001 and P = .02, respectively). In addition, VLA-4 blockade was associated with a significant decrease in conjunctival T-cell numbers (P < .001 vs control vehicle-treated group) and tumor necrosis factor-alpha transcript levels in the cornea (P = .04 vs control vehicle-treated group) and conjunctiva (P = .048 vs control vehicle-treated group). CONCLUSION: Application of topical anti-VLA-4 sm led to a significant decrease in dry eye signs and suppression of inflammatory changes at the cellular and molecular levels. CLINICAL RELEVANCE: Topical blockade of VLA-4 may be a novel therapeutic approach to treat the clinical signs and inflammatory changes accompanying dry eye disease.

Topical omega-3 and omega-6 fatty acids for treatment of dry eye.

OBJECTIVE: To study the efficacy of topical application of alpha-linolenic acid (ALA) and linoleic acid (LA) for dry eye treatment. METHODS: Formulations containing ALA, LA, combined ALA and LA, or vehicle alone, were applied to dry eyes induced in mice. Corneal fluorescein staining and the number and maturation of corneal CD11b(+) cells were determined by a masked observer in the different treatment groups. Real-time polymerase chain reaction was used to quantify expression of inflammatory cytokines in the cornea and conjunctiva. RESULTS: Dry eye induction significantly increased corneal fluorescein staining; CD11b(+) cell number and major histocompatibility complex Class II expression; corneal IL-1alpha and tumor necrosis factor alpha (TNF-alpha) expression; and conjunctival IL-1alpha, TNF-alpha, interferon gamma, IL-2, IL-6, and IL-10 expression. Treatment with ALA significantly decreased corneal fluorescein staining compared with both vehicle and untreated controls. Additionally, ALA treatment was associated with a significant decrease in CD11b(+) cell number, expression of corneal IL-1alpha and TNF-alpha, and conjunctival TNF-alpha. CONCLUSIONS: Topical ALA treatment led to a significant decrease in dry eye signs and inflammatory changes at both cellular and molecular levels. CLINICAL RELEVANCE: Topical application of ALA omega-3 fatty acid may be a novel therapy to treat the clinical signs and inflammatory changes accompanying dry eye syndrome.

Cicatrizing and autoimmune diseases.

Autoimmune disorders of the ocular surface represent a clinically heterogeneous group of conditions where acute and chronic autoreactive mechanisms can cause significant damage to the eye. When severe and affecting the epithelium and substantia propria of the conjunctiva, cicatrization can ensue, leading to significant mechanical alterations as a result of the fibrosis. These conditions, though generally infrequent, can be the cause of profound pathology and visual disability, and often need systemic immune modulation for therapy.

Nonvascular VEGF receptor 3 expression by corneal epithelium maintains avascularity and vision.

Transparency of the cornea, the window of the eye, is a prerequisite for vision. Angiogenesis into the normally avascular cornea is incompatible with good vision and, therefore, the cornea is one of the few tissues in the human body where avascularity is actively maintained. Here, we provide evidence for a critical mechanism contributing to corneal avascularity. VEGF receptor 3, normally present on lymphatic and proliferating blood vascular endothelium, is strongly constitutively expressed by corneal epithelium and is mechanistically responsible for suppressing inflammatory corneal angiogenesis.

The controlled-environment chamber: a new mouse model of dry eye.

PURPOSE: To develop a controlled-environment chamber (CEC) for mice and verify the effects of a low-humidity setting on ocular surface signs in normal mice. METHODS: Eight- to 12-week-old BALB/c mice were used in a controlled-environment chamber (CEC) where relative humidity (RH), temperature (T), and airflow (AF) are regulated and monitored. Mice were placed into the CEC and exposed to specific environmentally controlled conditions (RH = 18.5% +/- 5.1%, AF = 15 L/min, T = 21-23 degrees C) for 3, 7, 14, and 28 days. Control mice were kept in a normal environment (RH = 50%-80%, no AF, T = 21-23 degrees C) for the same duration. Aqueous tear production by means of the cotton thread test, corneal fluorescein staining (score, 0-15), and goblet cell density in the superior and inferior conjunctiva were measured by a masked observer. RESULTS: No statistically significant differences between the groups were found at baseline. Decreased tear secretion and increased corneal fluorescein staining were significantly present on day 3, 7, 14, and 28 in animals kept in the CEC. Goblet cell density was significantly decreased in the superior conjunctiva on day 7, and on day 3, 7, and 14 in the inferior conjunctiva in the CEC-kept mice compared with control animals. CONCLUSIONS: This study indicates that exposure of normal mice to a low-humidity environment in a CEC can lead to significant alterations in tear secretion, goblet cell density, and acquisition of dry eye-related ocular surface signs.

Systemic soluble Tie2 expression inhibits and regresses corneal neovascularization.

This study was designed to determine if soluble Tie2 (sTie2) expression inhibits and regresses corneal neovascularization, and if VEGF contributes to its effect. The corneas of BALB/c mice were scraped and the mice were injected with either an adenovirus expressing soluble Tie2 (Ad.sTie2) or an empty adenoviral vector. When injected at the inhibition timepoint (one day prior to corneal injury), the mean percentage of neovascularized corneal area two weeks later in Ad.sTie2-treated mice vs. controls was 56.37+/-9.15% vs. 85.79+/-3.55% (p=0.04). At the regression timepoint (4 weeks after corneal scrape), the mean area of corneal neovascularization in Ad.sTie2-treated mice was 42.89+/-4.74% vs. 75.01+/-3.22% in the control group (p=0.007). VEGF expression was significantly higher in Ad.sTie2-treated mice at the inhibition timepoint and there was no significant difference at the regression timepoint. These findings suggest that sTie2 inhibits and regresses corneal neovascularization in a VEGF-independent manner.

Acute unilateral leukemic hypopyon in an adult with relapsing acute lymphoblastic leukemia.

PURPOSE: To report acute unilateral hypopyon uveitis as an initial presenting feature of relapsing acute lymphoblastic leukemia (ALL) in an adult patient. DESIGN: Observational case report. METHODS: Anterior chamber paracentesis was performed in a 56-year-old male presenting with treatment-resistant unilateral hypopyon while in the remission phase of ALL. RESULTS: Examination of the aqueous humor aspirate revealed presence of malignant cells compatible with the previous bone marrow biopsy and subsequent spinal tap results. CONCLUSIONS: Atypical hypopyon uveitis can be an indication of relapsing ALL, even in adults. Prompt anterior chamber aspiration is required for the correct diagnosis and subsequent treatment.

Flt-1 intraceptors inhibit hypoxia-induced VEGF expression in vitro and corneal neovascularization in vivo.

PURPOSE: To determine whether subunits of VEGF receptor-1 coupled with an endoplasmic reticulum retention signal can block hypoxia-induced upregulation of VEGF secretion in corneal epithelial cells and block murine corneal angiogenesis induced by corneal injury. METHODS: Human corneal epithelial cells, transfected with plasmids encoding Flt23K or Flt24K (the VEGF-binding domains of the Flt-1 receptor coupled with the endoplasmic reticulum retention peptide KDEL), were subjected 2 days after transfection to 5% hypoxia for 24 hours. Supernatant was sampled at 24 hours and assayed for VEGF by ELISA. For in vivo models, mouse corneas underwent intrastromal injections of plasmids encoding Flt23K or Flt24K, and 2 days later, sustained injury induced by topical NaOH and mechanical scraping. Corneas were assessed 2 days later for VEGF ELISA and leukocyte counting or 1 week later for quantification of neovascularization. RESULTS: Hypoxia induced VEGF by human corneal epithelial cells was sequestered by both Flt23K and Flt24K; Flt-1 23K suppressed VEGF secretion as well. Intrastromal delivery of plasmid Flt23K suppressed VEGF by 40.4% (P = 0.009), leukocytes by 49.4% (P < 0.001), and neovascularization by 66.8% (P = 0.001). Flt24K suppressed VEGF expression by 30.8% (P = 0.042), leukocytes by 25.8% (P < 0.001), and neovascularization by 49.5% (P = 0.015). CONCLUSIONS: Flt-1 intraceptors, which are endoplasmic reticulum retention signal-coupled VEGF receptors, significantly suppress hypoxia-induced VEGF secretion by corneal epithelial cells in vitro. In vivo, delivery of naked plasmids expressing these intraceptors inhibits injury-induced upregulation of VEGF, leukocyte infiltration, and corneal neovascularization.

Chronic ocular surface disease after allogeneic bone marrow transplantation.

Graft-versus-host disease (GVHD) is a common complication of allogeneic bone marrow transplantation (allo-BMT). Ocular surface disease (OSD) is one of the most common manifestations of chronic ocular GVHD, yet little is known about it. In this article, we review the available literature on this condition and present results from our study of the manifestations of OSD in the chronic phase (>3 months duration) post allo-BMT. Our study consisted of a retrospective chart review of 62 allo-BMT patients with chronic OSD evaluated at our center between 1995 and 2002. The clinical features, systemic associations, treatment, and status of OSD at the last follow-up are presented and discussed in the context of other reports of OSD in GVHD.

Atypical pulmonary giant hydatid cyst as bilaterally symmetrical solitary cysts.

A pulmonary giant hydatid cyst, a special clinical entity, is rare. Our case involves a young patient who presented with a bilaterally symmetrical solitary cyst in each lung, a feature consistent with congenital lung cysts. The radiological and immunological findings were equivocal. A diagnosis of giant hydatid cyst was made intraoperatively and both cysts were removed conservatively. A follow-up showed complete recovery.

Management of coincident cataract and uveitis.

Cataract surgery in a patient with uveitis is more complex than senile cataract extraction, because it involves multiple considerations related to the cause of uveitis, prospects of visual rehabilitation, appropriate surgical timing and technique, and the type and material of the intraocular lens used. Establishing the diagnosis, thorough eye examination, careful patient selection and meticulous control of perioperative inflammation are key elements to a successful visual outcome. Our aims in this article are to review the literature on this subject over the past year and highlight the behavior of intraocular lenses of various biomaterials in the uveitic eye. In addition, we also reemphasize the idea of a model of zero tolerance to intraocular inflammation to minimize the incidence of irreversible damage to ocular structures essential to good vision.