MicroRNA expression profiling of cutaneous squamous cell carcinomas and precursor lesions.

Background: Actinic keratoses (AK) are pre-malignant skin lesions caused by chronic sun exposure. Progression from an AK to intraepidermal carcinoma (IEC) and a cutaneous squamous cell carcinoma (SCC) is well known but the rate of transformation to an invasive SCC is highly variable. Since no definitive biomarkers are available, treatment decisions are made ad hoc. Objectives: To fully characterise our AK to SCC progression series, we performed microRNA (miRNA) microarray expression profiling of normal and photodamaged skin, as well as AKs, IEC, and invasive SCCs. Methods: The study recruited 27 patients who donated fresh biopsies of normal skin, photodamaged skin, AK, IEC, and SCC (n = 67 specimens). All miRbase (v.21) miRNAs were profiled to identify miRNAs related to SCC progression. miRNAs were validated using qRT-PCR and in vitro phenotypic assays. Results: There were 234 robustly expressed miRNAs across the tissue collection, which resulted in 20 miRNA that were differentially expressed ((cor)p ≤ 0.05 and ≥ 10 fold) between normal skin and SCC. Hierarchical clustering all samples illustrated that AKs, IEC, and SCCs were largely indistinguishable, which confirms the premalignant status of an AK. A panel of miRNAs showed significant dysregulation between normal and photodamaged skin and AK. Importantly, we found miR-34a-5p and miR-31-5p had significant differential expression between AKs and IEC and IEC and SCC respectively. Phenotypic assays determined that the miR-31 duplex had opposing effects on SCC cell lines which suggests that dysregulation of this duplex may be related to the dynamic control of progression of transformed keratinocytes. Conclusions: This study confirmed the continuum of AK with IEC and SCC highlighting that miRNA expression plays a role in keratinocyte transformation. Development of our putative miRNA biomarker candidates is warranted to aid in clinical management of patients experiencing high AK load to determine the most appropriate treatment.

Mendelian Randomization Shows a Causal Effect of Low Vitamin D on Non-infectious Uveitis and Scleritis Risk.

PURPOSE: To investigate a causal relationship between Vitamin D levels and non-infectious uveitis and scleritis using Mendelian randomization (MR) techniques. DESIGN: Two-sample Mendelian randomization case-control study. METHODS: The study setting was a biobank of an academic, integrated health care system. The patient population comprised 375 case patients with a non-infectious uveitis and/or scleritis diagnosis and no diagnosis of infectious, trauma-related, or drug-induced uveitis/scleritis. In addition, there were 4167 controls with no uveitis or scleritis diagnosis. Causal effect estimates of low 25-hydroxy Vitamin D (25OHD) on uveitis/scleritis risk were calculated. RESULTS: We found an association of genetically decreased 25OHD with uveitis/scleritis risk (odds ratio [OR] = 2.16, 95% CI = 1.01-4.64, P = .049, per SD decrease in log25OHD). In a first sensitivity MR analysis excluding the genetic variants that are unlikely to have a role in biologically active 25OHD, effect estimates were consistent with those from the primary analysis (OR = 2.38, 95% CI =1.06-5.36, P = 0.035, per SD of log25OHD). Furthermore, in a second sensitivity analysis using only the 6 variants within the CYP2R1 locus (which encodes 25OHD hydroxylase, the liver enzyme responsible for converting Vitamin D to 25OHD), genetically decreased 25OHD was strongly associated with increased uveitis/scleritis risk (OR = 6.42, 95% CI = 3.19-12.89, P = 1.7 × 10-7, per SD of log25OHD). CONCLUSIONS: Our findings suggest a causal relationship between low Vitamin D levels and higher risk of non-infectious uveitis and scleritis. Vitamin D supplementation may be a low-cost, low-risk intervention to mitigate non-infectious uveitis and scleritis risk, and should be explored in a prospective trial.

Genome-Scale DNA Methylation Analysis Identifies Repeat Element Alterations that Modulate the Genomic Stability of Melanocytic Nevi.

Acquired melanocytic nevi grow and persist in a stable form into adulthood. Using genome-wide methylation profiling, we evaluated 32 histopathologically and dermoscopically characterized nevi to identify the key epigenetic regulatory mechanisms involved in nevogenesis. Benign (69% globular and 31% nonspecific dermoscopic pattern) and dysplastic (95% reticular/nonspecific dermoscopic pattern) nevi were dissimilar, with only two shared differentially methylated loci. Benign nevi showed an increase in both genome-scale methylation and methylation of Alu/LINE-1 retrotransposable elements, a marker of genomic stability, as well as global methylation. In contrast, dysplastic nevi showed evidence for genomic instability through the hypomethylation of Alu/LINE-1 (Alu: P = 0.00019; LINE-1: P = 0.000035). Using dermoscopic classifications, reticular/nonspecific patterned nevi had 59,572 5'-C-phosphate-G-3' differentially methylated loci (Q < 0.05), whereas globular nevi had no significant differentially methylated loci. In reticular/nonspecific patterned nevi, the tumor suppressor PTEN had the greatest proportion of hypermethylated 5'-C-phosphate-G-3' loci in its promoter region than all other assayed gene promoters. The relative activity of reticular/nonspecific nevi was evidenced by 50,720 hypomethylated loci being enriched for accessible chromatin and 8,852 hypermethylated loci strongly enriched, for example, marks of active gene promoters, which suggests that gain of DNA methylation observed in these nevus types plays a role in gene regulation.

Open-globe injury caused by barbed fish hook repaired using the back-out method: a report of two cases.

Fish hook open-globe injuries (OGIs) are challenging to repair surgically because of the backward-projecting barb near the hook's point that prevents withdrawal of the hook. The most commonly reported ophthalmic surgical technique for removal of barbed hooks is advance-and-cut, wherein the fish hook is pushed through an iatrogenic wound to the exterior of the globe, the barb is cut off, and the shank is backed out of the entry wound. We report 2 cases of zone I OGIs with retained fish hooks successfully repaired using the back-out technique. This strategy involves enlarging the entry wound to allow the entire hook and barb to be backed out, decreasing iatrogenic injuries and eliminating the need for wire cutters.

Outcomes and Reasons for Late Presentation of Lens Induced Glaucoma: A Prospective Study.

PURPOSE: To determine the clinical outcomes and causes of late presentation in lens induced glaucoma (LIG). DESIGN: Prospective observational study. PARTICIPANTS: One hundred sixty-five patients with LIG who underwent cataract surgery. METHODS: Preoperative data collection included a questionnaire about reasons for late presentation, socioeconomic status, visual acuity, intraocular pressure (IOP), and the lens and angle status of the fellow eye. All patients underwent manual small-incision cataract surgery. Postoperative vision, IOP, the anterior segment, and the fundus were evaluated at days 1, 15, and 30. MAIN OUTCOME MEASURES: Reasons for late presentation, status of the fellow eyes, and surgical outcomes, including visual acuity and IOP at the 1-month postoperative visit. RESULTS: Mean age at presentation was 63.8 years, and the female-to-male ratio was 1.4:1. Of the entire cohort, 70.3% were phacolytic and 29.7% had phacomorphic glaucoma. The main causes for late presentation was nonfinancial (81.2%); of these, good vision in fellow eye and lack of escort to the hospital were the major reasons. Most fellow eyes were pseudophakic (72.1%). After surgery, 75.6% gained best-corrected visual acuity of 6/18 or more. Six percent experienced poor visual recovery (≤6/60) with optic atrophy as the major cause. Only 7.9% required further glaucoma management in the form of topical medications. Delayed presentation (>15 days) was associated with poor visual outcome. CONCLUSIONS: Nonfinancial causes, including good vision in the fellow eye and lack of escort, were the major determinants of late presentation. Most patients were pseudophakic in the fellow eye. After cataract extraction, only a few LIG patients required further glaucoma management.

The Effectiveness of Intracameral Moxifloxacin Endophthalmitis Prophylaxis for Trabeculectomy.

PURPOSE: To analyze the effectiveness of intracameral moxifloxacin prophylaxis in reducing acute postoperative endophthalmitis after trabeculectomy and combined trabeculectomy plus cataract extraction. DESIGN: Retrospective clinical registry analysis. PARTICIPANTS: Patients undergoing either trabeculectomy or trabeculectomy plus cataract extraction at Aravind Eye Hospitals (AEH) between 2009 and 2018 (inclusive). METHODS: Electronic health records data were analyzed before and after implementation of routine intracameral moxifloxacin, and acute postoperative endophthalmitis rates were compared. During 2015, routine intracameral moxifloxacin prophylaxis was added in a step-wise fashion throughout AEH. Date of implementation was used to create group 1 (without intracameral moxifloxacin prophylaxis) and group 2 (with intracameral moxifloxacin prophylaxis). MAIN OUTCOME MEASURES: The primary outcome was the difference in acute (≤6 weeks) postoperative endophthalmitis between groups 1 and 2. Review of culture results, visual acuity, and intraocular pressure also was performed for patients with endophthalmitis. RESULTS: Thirty-eight thousand nine hundred eyes (group 1) did not receive intracameral moxifloxacin, whereas 19 086 eyes (group 2) did. Although the rate of noninfectious postoperative complications was not significantly different (0.81% vs. 0.67%; P = 0.07), a significantly lower rate of acute postoperative endophthalmitis was found in group 2 versus group 1 (0.03% vs. 0.08%; P = 0.03). Patients receiving intracameral moxifloxacin showed approximately 2.5-times lower odds of infection (odds ratio, 0.39 for group 2 vs. group 1; 95% confidence interval, 0.16-0.95) and almost 4-times lower odds after adjustment for covariates (odds ratio, 0.26 for group 2 vs. group 1; 95% confidence interval, 0.09-0.74). The rate of early postoperative infection after intracameral moxifloxacin introduction was lower for patients undergoing both trabeculectomy alone (0.09%-0.03%; P = 0.27) and combined trabeculectomy plus cataract extraction (0.08%-0.03%; P = 0.06). Although most cultures yielded no growth, no Staphylococcus or gram-negative growth was found for patients in group 2, who received intracameral moxifloxacin. CONCLUSIONS: Intracameral moxifloxacin prophylaxis was associated with a nearly 4-fold lower rate of early postoperative endophthalmitis in patients undergoing trabeculectomy or combined trabeculectomy plus cataract extraction.

Association of Low Vitamin D Levels with Noninfectious Uveitis and Scleritis.

Purpose: To determine whether an association between Vitamin D and noninfectious ocular inflammation exists. Methods: Retrospective case-control study with 765 patients (333 uveitis cases, 103 scleritis cases, 329 controls). Logistic regression models examined the relationship between hypovitaminosis D and ocular inflammation. Results: The odds of having uveitis were 1.92 times higher for patients with hypovitaminosis D compared to patients with normal Vitamin D levels in the multivariate analysis [odds ratio (OR) = 1.92, 95% Confidence Interval (CI) = 1.36-2.72, p = 2.32 × 10-4]. A secondary analysis demonstrated that the odds of developing uveitis or scleritis were 5% lower and 4% lower, respectively, for every unit increase in Vitamin D level (uveitis: OR = 0.95, 95% CI = 0.94-0.97, p = 9.87 × 10-6; scleritis: OR = 0.96, 95% CI = 0.93-0.99, p = 0.009). Conclusion: Hypovitaminosis D was associated with increased risk of ocular inflammation in this retrospective study.

Defining the Molecular Genetics of Dermoscopic Naevus Patterns.

Melanocytic naevi are common melanocytic proliferations that may simulate the appearance of cutaneous melanoma. Naevi commonly harbour somatic mutations implicated in melanomagenesis but in most cases lack the necessary genomic alterations required for melanoma development. While the mitogen-activated protein kinase pathway and ultraviolet radiation strongly contribute to naevogenesis, the somatic mutational landscape of dermoscopic naevus subsets distinguishes some of the molecular hallmarks of naevi in relation to melanoma. We herein discuss the classification of naevi and theories of naevogenesis and review the current literature on the somatic alterations in naevi and melanoma. This review focusses on the clinical-dermoscopic-pathological and genomic correlation of naevi that shapes the current understanding of naevi.

Whole-Exome Sequencing of Acquired Nevi Identifies Mechanisms for Development and Maintenance of Benign Neoplasms.

The melanoma transformation rate of an individual nevus is very low despite the detection of oncogenic BRAF or NRAS mutations in 100% of nevi. Acquired melanocytic nevi do, however, mimic melanoma, and approximately 30% of all melanomas arise within pre-existing nevi. Using whole-exome sequencing of 30 matched nevi, adjacent normal skin, and saliva we sought to identify the underlying genetic mechanisms for nevus development. All nevi were clinically, dermoscopically, and histopathologically documented. In addition to identifying somatic mutations, we found mutational signatures relating to UVR mirroring those found in cutaneous melanoma. In nevi we frequently observed the presence of the UVR mutation signature compared with adjacent normal skin (97% vs. 10%, respectively). Copy number aberration analysis showed that for nevi with copy number loss of tumor suppressor genes, this loss was balanced by loss of potent oncogenes. Moreover, reticular and nonspecific patterned nevi showed an increased (P < 0.0001) number of copy number aberrations compared with globular nevi. The mutation signature data generated in this study confirms that UVR strongly contributes to nevogenesis. Copy number changes reflect at a genomic level the dermoscopic differences of acquired melanocytic nevi. Finally, we propose that the balanced loss of tumor suppressor genes and oncogenes is a protective mechanism of acquired melanocytic nevi.

RNA-seq reveals more consistent reference genes for gene expression studies in human non-melanoma skin cancers.

Identification of appropriate reference genes (RGs) is critical to accurate data interpretation in quantitative real-time PCR (qPCR) experiments. In this study, we have utilised next generation RNA sequencing (RNA-seq) to analyse the transcriptome of a panel of non-melanoma skin cancer lesions, identifying genes that are consistently expressed across all samples. Genes encoding ribosomal proteins were amongst the most stable in this dataset. Validation of this RNA-seq data was examined using qPCR to confirm the suitability of a set of highly stable genes for use as qPCR RGs. These genes will provide a valuable resource for the normalisation of qPCR data for the analysis of non-melanoma skin cancer.

Use of Crossed Polarizers to Enhance Images of the Eyelids.

PURPOSE: To describe imaging of the external eye with Crossed Polarizers to enhance clinically important features in digital photographs of the eyelids. METHODS: External photographs with and without crossed polarizing filters were taken of patients with blepharitis and controls with no clinical eye pathology. RESULTS: Photographing eyelid skin through Crossed Polarizers decreased reflections on the skin surface and improved visualization of eyelid telangiectasias and blood vessels in patients with a broad range of skin pigmentation and ethnicities. CONCLUSIONS: The use of Crossed Polarizers in imaging the external eye reduces reflections and glare from the eyelid skin and margins, thereby allowing for a more detailed evaluation of underlying structures and analysis of images. These findings suggest that including Crossed Polarizers in clinical photography has informative applications for assessing eyelid disease.

The "melanoma-enriched" microRNA miR-4731-5p acts as a tumour suppressor.

We previously identified miR-4731-5p (miR-4731) as a melanoma-enriched microRNA following comparison of melanoma with other cell lines from solid malignancies. Additionally, miR-4731 has been found in serum from melanoma patients and expressed less abundantly in metastatic melanoma tissues from stage IV patients relative to stage III patients. As miR-4731 has no known function, we used biotin-labelled miRNA duplex pull-down to identify binding targets of miR-4731 in three melanoma cell lines (HT144, MM96L and MM253). Using the miRanda miRNA binding algorithm, all pulled-down transcripts common to the three cell lines (n=1092) had potential to be targets of miR-4731 and gene-set enrichment analysis of these (via STRING v9.1) highlighted significantly associated genes related to the 'cell cycle' pathway and the 'melanosome'. Following miR-4731 overexpression, a selection (n=81) of pull-down transcripts underwent validation using a custom qRT-PCR array. These data revealed that miR-4731 regulates multiple genes associated with the cell cycle (e.g. CCNA2, ORC5L, and PCNA) and the melanosome (e.g. RAB7A, CTSD, and GNA13). Furthermore, members of the synovial sarcoma X breakpoint family (SSX) (melanoma growth promoters) were also down-regulated (e.g. SSX2, SSX4, and SSX4B) as a result of miR-4731 overexpression. Moreover, this down-regulation of mRNA expression resulted in ablation or reduction of SSX4 protein, which, in keeping with previous studies, resulted in loss of 2D colony formation. We therefore speculate that loss of miR-4731 expression in stage IV patient tumours supports melanoma growth by, in part; reducing its regulatory control of SSX expression levels.

Recurrent Bleeding, Survival, and Longitudinal Pulmonary Function following Bronchial Artery Embolization for Hemoptysis in a U.S. Adult Population.

PURPOSE: To report outcomes of bronchial artery embolization (BAE) for hemoptysis, including recurrent bleeding, survival, and longitudinal pulmonary function. MATERIALS AND METHODS: A prospective database identified 69 patients who underwent 97 BAE procedures (n = 1-7 per patient) at a tertiary academic medical center over a period of 11 years. Technical and clinical success were determined. Recurrent bleeding and survival were compared by etiology of lung disease. Rates of change in pulmonary function (forced expiratory volume in 1 s [FEV1] and forced vital capacity [FVC]) were measured and compared before and after index BAE by linear regression in 17 patients. RESULTS: The technical success rate of BAE was 90%. Clinical success rates at 24 hours and 30 days were 82% and 68%, respectively. Thirty percent of patients had recurrent bleeding that required bronchoscopy (7%) or additional embolization (23%). Median time to recurrent bleeding was 29 days among the 13 patients with sarcoidosis, compared with 293 days among patients without sarcoidosis (P = .0013). The hazard ratio for death in patients with sarcoidosis compared with those without sarcoidosis was 4 (95% confidence interval, 2.6-14.6). Analyzing all instances of pulmonary function tests, slopes of decline in FEV1 and FVC were significantly different (FEV1, P = .0048; FVC, P < .0001) before and after index BAE, with an improvement after BAE (FEV1, 0.8%/y; FVC, 1%/y) and a decrease before BAE (FEV1, -1.6%/y; FVC, -1.4%/y). CONCLUSIONS: BAE is an effective therapy for hemoptysis, but patients with sarcoidosis are at significant risk of recurrent bleeding and death compared with patients with other lung diseases. BAE does not accelerate deterioration in lung function.