Normal Conjunctiva Research Articles

Assessment of conjunctival autograft reperfusion after pterygium surgery by optical coherence tomography angiography (OCT-A)

PurposeTo investigate the healing process of conjunctival autografts (CAG) following pterygium surgery using optical coherence tomography angiography (OCTA). MethodsTwenty-one eyes of 21 patients diagnosed with pterygium underwent pterygium excision with CAG without using Mitomycin-C. Over a 12-week follow-up period, changes in vascular density (VD), vascular density index (VDI), and vascular length density (VLD) were assessed at two distinct depths: superficial (<200 μm) and deep (>200 μm) using OCTA. Additionally, the revascularization rate and pattern were evaluated. ResultsDuring the first week, the CAG was edematous and no sign of neovascularization was observed. In 4th week edema decreased and early signs of vascular formation appeared. In the 12th week, the deep vasculature demonstrated a greater density of interconnectivity compared to the superficial layers. VD and VLD significantly increased during the follow-up period (P < 0.05). The CAG blood flow signals exhibited a chaotic pattern, deviating from the expected centrifugal vascular pattern in the surrounding normal conjunctiva. ConclusionOCTA imaging emerges as a reliable tool for the assessment of CAG vascularization, improving the monitoring of the healing process in the postoperative period. The evaluation of CAG revascularization patterns appears to be promising biomarkers that can predict the potential future recurrence.

Z-shaped incision without epithelial resection in pterygium surgery.

To introduce a novel surgical technique using a Z-shaped incision without epithelial resection in ophthalmic pterygia. This was a prospective study. During pterygium surgery, all proliferative tissues were separated from the cornea and conjunctiva without resection of the tissues. The unaffected conjunctiva was incised in a Z-shape. The upper (or lower) conjunctival flap was sutured to the lower (or upper) normal conjunctiva on the limbal sclera, while the proliferative tissue was sutured to the upper conjunctiva (or lower) near the fornix. Ten patients with pterygia were eligible for this study. Eight patients with primary pterygia and 2 with recurrent pterygia were included. The age of patients at surgery ranged from 47 to 90y (average: 71.9y). Five patients each showed right and left-sided pterygia. The postoperative follow-up periods were from 8 to 78mo (average: 25.0mo). The surgery was successfully conducted and wounds were favorably reconstructed in all patients. The proliferative tissues sutured to the normal conjunctiva showed palor and attenuated neovessles, and never showed re-growth after surgery. Nine patients did not show recerrence. Recerrent pterygium was noted in 1 patient, but additional treatments were not required. The procedure involves the reconstruction of pterygial tissue and normal conjunctiva using a Z-shaped incision. The scleral limbal wound can be covered with non-affected conjunctiva without any excision of conjunctival epithelia in patients with primary or recurrent pterygia.

Elevated KDM4D Expression in Pterygium: Impact and Potential Inhibition by Lycium Barbarum Polysaccharide.

Purpose: This study aimed to explore the effects of elevated KDM4D expression and potential therapeutic effects of Lycium barbarum polysaccharide (LBP) on pterygium. Methods: The expression levels of KDM4D in the primary pterygium (n = 29) and normal conjunctiva (n = 14) were detected by immunohistochemistry. The effects of KDM4D on pterygium fibroblasts were detected by the CCK-8 assay, liquid chromatography-mass spectrometry assay, flow cytometry, and scratch wound healing assay. The relative expression of KDM4D in pterygium fibroblasts stimulated by interleukin (IL)-1β, IL-6, IL-8, and LBP was detected by quantitative real-time PCR and Western blot. The effects of LBP on pterygium fibroblasts were detected using flow cytometry and scratch wound healing assays. Results: The expression level of KDM4D in pterygium was higher than that in normal conjunctiva. KDM4D increased the cell viability of pterygium fibroblasts. The differentially expressed genes identified in the LM-MS assay enriched in "actin filament organization" and "apoptosis." KDM4D promoted migration and inhibited apoptosis of pterygium fibroblasts in vitro. Inflammatory cytokines, including IL-1β, IL-6, and IL-8, enhanced the expression of KDM4D in pterygium fibroblasts. LBP inhibited the expression of KDM4D in pterygium fibroblasts and decreased their cell viability. Moreover, LBP attenuated the KDM4D effects on migration and apoptosis of pterygium fibroblasts. Conclusions: Elevated KDM4D expression is a risk factor for pterygium formation. LBP inhibits the expression of KDM4D in pterygium fibroblasts and may be a potential drug for delaying pterygium development.

Investigation of pathogenesis of the pterygium by transcriptomic and proteomic analysis

Aims/Purpose: Extrinsic factors, including ultraviolet rays, are well‐known risk factors in the pathogenesis of pterygia. However, the biomolecular intrinsic factors that could serve as therapeutic targets have not been fully elucidated. Therefore, this study aimed to investigate comprehensive changes in the proteome and transcriptome of conjunctival pterygium tissues from human patients using multi‐omics analysis.Methods: A prospective cross‐sectional study enrolled 30 patients diagnosed with pterygium who underwent surgery from May 2020 to December 2020. During surgery, normal conjunctiva and conjunctival tissues with pterygium were collected. Nano liquid chromatography with tandem mass labelling spectrometry and next‐generation sequencing were performed on the obtained specimens. Proteins with significantly expressed mRNAs between pterygium and control were identified, and their biological processes were analysed using bioinformatic tools.Results: Through the integration of transcriptomic and proteomic analyses, a total of 1288 proteins were identified, which were also confirmed by mRNA expression. Hierarchical clustering heatmap analysis revealed distinct clusters corresponding to each group, and principle component analysis demonstrated a prominent separation between groups. Among the identified proteins, 185 were determined as differentially expressed proteins (DEPs), with 154 upregulated and 31 downregulated in pterygium compared to the control group. Protein–protein interaction network analysis revealed six distinct interconnected networks of DEPs, and gene ontology enrichment analysis highlighted biological processes associated with extracellular matrix organization, insulin‐like growth factor‐related pathways, and the immune system.Conclusions: This study demonstrates proteomic alterations in conjunctival tissues of patients with pterygium, which are corroborated by RNA expression analysis. The identified significant proteins in pterygium are associated with specific biological pathways and could be potential therapeutic targets. The integrated proteo‐transcriptomic analysis provides essential insights into the pathogenesis of pterygium, contributing to the identification of novel treatment strategies.

Transcriptional profilling to identify the inhibitor effect of trehalose as an alternative natural therapeutic agent on pterygium cells

Aims/Purpose: The aim of this study to investigate transcriptome analysis of the inhibitory effects of trehalose on changing expression of regulator genes in pterygium development.Methods: Human primary pterygium cells and normal conjunctiva cells were isolated from the pterygium and conjunctiva tissues of six individuals, three of whom were men and three of which were females. The MTT test was used to investigate cell proliferation in pterygium treated with different concentrations dosage of trehalose. Differential mRNA expression in pterygium cells following indicative treatment was measured using an mRNA microarray and evaluated using GO and KEGG pathways. In addition, we employed QRT‐PCR to confirm the expression levels of genes linked to pterygium pathogenesis.Results: We have found that pterygium cells are more sensitive to trehalose treatment than conjunctival cell. In total, 419 probe were differentially expressed pterygium cells treated with trehalose compared with paired non‐treated pterygium cells. Gene‐Enrichment and Functional Annotation analysis indicated that differentially expressed RNAs were associated with cellular process including especially fibroblastic cell migration, sterol biosynthetic process (p &lt; 0.00034), extracellular matrix recomposition (p &lt; 0.000740288), cell proliferation (p &lt; 0.004391252), steroid biosynthetic process (p &lt; 0.00029971). Composed of Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway, protein–protein interaction (PPI), and gene set enrichment analysis (GSEA) analysis, bioinformatic analyses showed that four hub genes were upregulated (MSMO, RANBP3L, DHCR7, CCL2) and five were downregulated (NR4A1, TENASTN C, CXCL8, CXCL12, ADAMTS12). Additionally, QRT‐PCR is used to confirm the expression of particular genes. Consistent with these findings, the migration capability of pterygium cells decreased after treatment of trehalose for 24 h.Conclusions: We consider that this study provides a scientific basis for elucidating the effect of trehalose on the pathogenesis of pterygium and exhibited that trehalose affects cellular processes that have essential roles in the progression and development of pterygium.

Metagenomic Analysis of DNA Viruses with Targeted Sequence Capture of Canine Lobular Orbital Adenomas and Normal Conjunctiva.

Our study aims are: (1) to evaluate phenotypically normal canine conjunctival and orbital tissue and tissue from canine lobular orbital adenomas (CLOAs) for the presence of viral genomic material and (2) phylogenetically classify detected DNA viruses to determine if a DNA virus is associated with CLOAs. A total of 31 formalin fixed paraffin embedded CLOA tissue samples, 4 papillomas or sarcoid, and 10 fresh clinically normal conjunctival tissues were included in this study. Genomic DNA was isolated from all samples and sequencing libraries were prepared. The libraries were molecularly indexed and pooled and viral DNA was enriched via targeted sequence capture utilizing ViroCap. The libraries were sequenced on the Illumina HiSeq platform and compared to known viral DNA reference genomes to identify viral DNA. Carnivore parvovirus was identified in 6.4% and 20% of CLOA tissue and normal conjunctival samples, respectively. This study showed that conjunctival tissue from healthy dogs and CLOAs uncommonly harbor DNA viruses, and no DNA virus was associated with these tumors. Further studies are needed to evaluate the etiologic cause of CLOAs.

The use of optical coherence tomography angiography in differential diagnosis of conjunctival melanocytic tumors

BACKGROUND: Optical coherence tomography angiography (OCTA) is a noninvasive method of eye microcirculation evaluation. Few reports are published on the use of OCTA for anterior segment (AS) vessels analysis in healthy eyes and in conjunctival tumors, and their vascular characteristics are still not thoroughly investigated. These questions are of importance, as it is known that tumors vasculature is indicative of the patients vital prognosis.&#x0D; AIM: The aim of our study was to investigate the potential of AS-OCTA in evaluation of normal conjunctival vessels architecture as well as that in melanocytic neoplasms.&#x0D; MATERIALS AND METHODS: 20 healthy volunteers (20 eyes) and 20 patients (20 eyes) with conjunctival nevi and melanomas were examined. AS optical coherence tomography (OCT) and AS-OCTA were performed. Scan analysis included qualitative assessment (vessels pattern, lumen, tortuosity) and quantitative assessment [perfusion density (PD, %) index]. Mean (MPD), maximum (MaxPD) and perifocal PD (PPD) were determined.&#x0D; RESULTS: In normal group, predominantly radial pattern of the vessels was revealed, their caliber remaining the same along their entire length; larger vessels were more often discovered in deep conjunctival layers. The lowest PD value (29.9%) was registered in the inferior conjunctival segment, and the highest (36.7%) in the nasal one. In the conjunctival tumors area tortuosity of the vessels, uneven vessels caliber along their length, and increase in the PD value were observed. Melanomas were characterized by an increase in the lace-like pattern and by presence of confluent pattern zones; MaxPD value was more than 50%. Significant difference was found between MPD values of normal conjunctiva and MPD values in conjunctival melanoma.&#x0D; CONCLUSIONS: AS-OCTA is an informative method for the visualization of vessels in normal conjunctiva and in conjunctival tumors. If the tumors vessels are unevenly distributed, MaxPD should be measured.

Bleach-Associated Uveitis: A Case Report

Introduction: Chemical ocular injuries represent ophthalmologic emergencies with potentially blinding complications. Burns with alkalis tend to be more serious due to deeper tissue penetration as a result of colliquative necrosis. Uveitis is usually observed in higher grades of burns with significant alterations in the anterior segment of the eye. This study aimed to report a case of late-onset unilateral uveitis following a mild alkali burns with sodium hypochlorite (bleach).&#x0D; Case presentation: A 64-year-old Caucasian woman reported that 3 weeks ago, she had an accident with bleach coming in contact with both eyes. At that time, she had emergency ocular irrigation with saline to both eyes, and she had also used artificial tears for about a week. Besides bilateral conjunctival hyperaemia and punctate corneal epithelial erosions, no other complications had been noted. Her symptoms had subsided in several days. Upon presentation to the clinic, reduced best-corrected visual acuity was observed in both eyes to 20/30, which was attributed to age-related cataracts, normal intraocular pressure, normal conjunctiva in both eyes, the clear cornea in the right eye (OD) and several white, round, middle-sized granulomatous keratic precipitates in the left eye (OS), without epithelial and stromal alterations. The anterior chamber of OS demonstrated a 0.5+ cellular reaction. The remaining ocular status was normal.&#x0D; Conclusion: Based on the history of chemical ocular injury, the lack of systemic diseases and the mild form of ocular inflammation, no additional diagnostic tests were offered. The patient was started on topical dexamethasone phosphate for 1 month with an initial dose of one drop four times daily, which led to remission.

Synergic Action of Insulin-like Growth Factor-2 and miRNA-483 in Pterygium Pathogenesis.

Pterygium is a multifactorial disease in which UV-B is speculated to play a key role by inducing oxidative stress and phototoxic DNA damage. In search for candidate molecules that are useful for justifying the intense epithelial proliferation observed in pterygium, our attention has been focused on Insulin-like Growth Factor 2 (IGF-2), mainly detected in embryonic and fetal somatic tissues, which regulate metabolic and mitogenic functions. The binding between IGF-2 and its receptor Insulin-like Growth Factor 1 Receptor (IGF-1R) activates the PI3K-AKT pathway, which leads to the regulation of cell growth, differentiation, and the expression of specific genes. Since IGF2 is regulated by parental imprinting, in different human tumors, the IGF2 Loss of Imprinting (LOI) results in IGF-2- and IGF2-derived intronic miR-483 overexpression. Based on these activities, the purpose of this study was to investigate the overexpression of IGF-2, IGF-1R, and miR-483. Using an immunohistochemical approach, we demonstrated an intense colocalized epithelial overexpression of IGF-2 and IGF-1R in most pterygium samples (Fisher's exact test, p = 0.021). RT-qPCR gene expression analysis confirmed IGF2 upregulation and demonstrated miR-483 expression in pterygium compared to normal conjunctiva (253.2-fold and 12.47-fold, respectively). Therefore, IGF-2/IGF-1R co-expression could suggest their interplay through the two different paracrine/autocrine IGF-2 routes for signaling transfer, which would activate the PI3K/AKT signaling pathway. In this scenario, miR-483 gene family transcription might synergically reinforce IGF-2 oncogenic function through its boosting pro-proliferative and antiapoptotic activity.

Design of functional decellularized matrix for conjunctival epithelial stem cell maintenance and ocular surface reconstruction

Conjunctival defects severely impair ocular surface homeostasis and vision.However, the existing conjunctival reconstruction methods are still unable to achieve satisfactory functional replacement of conjunctiva. Scaffold materials that provide a more stem-cell-friendly microenvironment are in urgent need of improvement. Herein we reported a functional conjunctival reconstruction scaffold constructed from decellularized matrix derived from rabbit subconjunctival fibroblasts (DM-SCF) loaded with conjunctival epithelial stem cells (CjESCs) for ocular surface repair. The DM-SCF had a thickness and mechanical strength close to that of normal conjunctiva and showed excellent biocompatibility. CjESCs inoculated on DM-SCF maintained good stem cell properties, which showed long-term maintenance of low differentiation and good proliferation activity.DM-SCF transplantation with CjESCs showed satisfactory results in rabbit models of a large conjunctival defect.The conjunctiva was reconstructed with abundant goblet cell expression and regular fiber arrangement. Protein mass spectrometry revealed the active extracellular matrix components of DM-SCF, regulating the activation of Wnt/β-catenin and Notch pathways for stem cell maintenance. Overall, our study developed a promising new strategy for ocular surface reconstruction by optimizing bioactive scaffold and providing functional stem cell niche, so as to achieve long-standing and superior functional repair.

The role of cyclophilin A and VEGF in the pathogenesis and recurrence of pterygium.

Pterygium is defined as overgrowth of abnormal conjunctival tissue on the cornea. Many proinflammatory cytokines and various growth factors have been implicated in the pathogenesis of pterygium. Cyclophilin A (CyPA) is a protein that is used by cyclosporin A (CsA) as the intracellular receptor and is secreted in response to inflammatory stimuli. Vascular endothelial growth factor (VEGF) is the most important angiogenic factor. This study aimed to assessment CyPA and VEGF immunoreactivity in pterygium specimens. In this cross-sectional study, 32 primary pterygium samples, 25 recurrent pterygium samples and 25 normal bulbar conjunctiva samples were included. The histopathological features, CyPA and VEGF immunoreactivity of surgically excised pterygium specimens were compared with control conjunctiva specimens obtained from normal bulbar conjunctiva. CyPA immunoreactivity in vascular endothelial cells, epithelial cells, and stromal cells was remarkably higher in pterygium specimens than control conjunctiva specimens (p = 0.004, p = 0.012, p = 0.001, respectively). Morever, VEGF immunoreactivity in endothelial cells was remarkably higher in pterygium specimens than control conjunctiva specimens (p < 0.001). When recurrent and primary pterygium specimens were compared, CyPA and VEGF immunoreactivity was remarkably higher in recurrent pterygium (p = 0.001, p = 0.001, respectively). Pearson correlation showed that CyPA immunoreactivity correlated with stromal vascularity, stromal inflammation, and mast cell count in pterygium specimens. This study aimed to assess CyPA and VEGF may have a important function in the pathogenesis and recurrence of pterygium.

Fibroblast Growth Factor-2 (FGF-2) Expression in Pterygia Using Cell Spot Arrays.

Fibroblast growth factor (FGF) is a main regulator of cell differentiation, cell migration and angiogenesis in normal and abnormal conjunctiva epithelia, but specific mechanisms of its aberrant expression are yet to be investigated. In the present study, we investigated FGF-2 protein expression within several pterygia. Using a liquid-based cytology assay, we obtained cell specimens from pterygia and healthy tissues directly from patients. A combination of immunocytochemistry followed by digital image analysis showed significant overexpression of FGF-2 in all the examined pterygia. In 30/60 (50%) cases there were high levels of staining intensity, whereas in the remaining 30/60 (50%) cases there were moderate levels of expression. FGF-2 levels of the control group were significantly lower in comparison with the pterygia group. There was no significant correlation between FGF-2 levels and either sex or location of the pterygium. FGF-2 levels had a significant correlation with morphological characteristics of the pterygia. More specifically, FGF-2 levels were significantly higher in the pterygia with a fleshy morphology. Interestingly, recurrent lesions demonstrated high expression levels. An overexpression of FGF-2 has been observed frequently in pterygia, where it may play a crucial role in determining the lesion’s progression. FGF-2 upregulation correlates with the morphology of pterygia and its tendency to recur. Cell spot analysis based on liquid-based cytology is a simple, yet effective, method for detecting a broad spectrum of protein markers and could be useful in analyzing potential pterygia patient samples.

Expression profiling suggests the involvement of hormone-related, metabolic, and Wnt signaling pathways in pterygium progression.

BackgroundPterygium is an ocular surface disease that can cause visual impairment if it progressively invades the cornea. Although many pieces of research showed ultraviolet radiation is a trigger of pterygium pathological progress, the underlying mechanism in pterygium remains indistinct.MethodsIn this study, we used microarray to evaluate the changes of transcripts between primary pterygium and adjacent normal conjunctiva samples in China. Then, we performed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analyses. Moreover, we constructed protein-protein interaction (PPI) and miRNA-mRNA regulatory networks to predict possible regulatory relationships. We next performed gene set enrichment analysis (GSEA) to explore the similarities and differences of transcripts between Asian studies from the Gene Expression Omnibus database. Furthermore, we took the intersection of differentially expressed genes (DEGs) with other data and identified hub genes of the development of pterygium. Finally, we utilized real-time quantitative PCR to verify the expression levels of candidate genes.ResultsA total of 49 DEGs were identified. The enrichment analyses of DEGs showed that pathways such as the Wnt-signaling pathway and metabolism-related pathways were upregulated, while pathways such as hormone-related and transcription factor-associated pathways were downregulated. The PPI and miRNA-mRNA regulatory networks provide ideas for future research directions. The GSEA of selecting Asian data revealed that epithelial-mesenchymal transition and myogenesis existed in the pathology of pterygium in the Asian group. Furthermore, five gene sets (interferon-gamma response, Wnt beta-catenin signaling, oxidative phosphorylation, DNA repair, and MYC targets v2) were found only in our Chinese datasets. After taking an intersection between selecting datasets, we identified two upregulated (SPP1 and MYH11) and five downregulated (ATF3, FOS, EGR1, FOSB, and NR4A2) hub genes. We finally chose night genes to verify their expression levels, including the other two genes (SFRP2 and SFRP4) involved in Wnt signaling; Their expression levels were significantly different between pterygium and conjunctiva.ConclusionsWe consider hormone-related, metabolic, and Wnt signaling pathways may be important in the pathology of pterygium development. Nine candidate genes we identified deserve further study and can be potential therapeutic targets.

OVOL1 immunohistochemical expression is a useful tool to diagnose invasion in ocular surface squamous neoplasms

Background and objectivesOVOL1 is a gene that negatively regulates mesenchymal transformation, which allows epithelial cells to invade the stroma. On the other hand, it negatively regulates c-Myc, which has a positive effect on cell proliferation. The aim of this study is to evaluate the expression of OVOL1 and c-Myc in ocular surface squamous neoplasia (OSSN). Patients and methodsCross-sectional cohort study of 36 samples including 6 squamous papillomas, 19 conjunctival intraepithelial neoplasms, 6 squamous carcinomas and 7 normal conjunctivae were evaluated using immunohistochemistry against OVOL1 and c-Myc. The expression of both markers was analysed using the H-score (intensity 1–3 multiplied by the percentage of positive cells). ResultsPercentages of 98 and 100 of the OSSN, and 57 and 71% of the normal conjunctivae expressed OVOL1 and c-Myc respectively, however, the mean H-score of OVOL1 and c-Myc was higher in the OSSN than in normal conjunctivae group (P=0.0001 in both). Within the OSSN, OVOL1 demonstrated a higher H-score in the conjunctival intraepithelial neoplasms and papilloma, compared to the squamous carcinoma (P<0.01) group. c-Myc did not show differences between the OSSN groups. An H-score lower than 35 differentiates a squamous cell carcinoma from other OSSN lesions with a sensitivity of 83.3% and a specificity of 100%. ConclusionsThe expression of OVOL1 is a useful tool to differentiate between a squamous carcinoma of conjunctival intraepithelial neoplasms and papilloma. OVOL1 could play a role in the invasiveness of squamous neoplasms and places it as a potential therapeutic target.

Análisis de la expresión inmunohistoquímica de OVOL1 como herramienta diagnóstica de invasión en las neoplasias escamosas de la superficie ocular

Background and objectivesOVOL1 is a gene that negatively regulates mesenchymal transformation, which allows epithelial cells to invade the stroma. On the other hand, it negatively regulates c-Myc, which has a positive effect on cell proliferation. The aim of this study is to evaluate the expression of OVOL1 and c-Myc in ocular surface squamous neoplasia (OSSN). Patients and methodsCross-sectional cohort study of 36 samples including 6 squamous papillomas, 19 conjunctival intraepithelial neoplasms, 6 squamous carcinomas and 7 normal conjunctivae were evaluated using immunohistochemistry against OVOL1 and c-Myc. The expression of both markers was analysed using the H-score (intensity 1-3 multiplied by the percentage of positive cells). ResultsPercentages of 98 and 100 of the OSSN, and 57 and 71% of the normal conjunctivae expressed OVOL1 and c-Myc respectively, however, the mean H-score of OVOL1 and c-Myc was higher in the OSSN than in normal conjunctivae group (P=.0001 in both). Within the OSSN, OVOL1 demonstrated a higher H-score in the conjunctival intraepithelial neoplasms and papilloma, compared to the squamous carcinoma (P<.01) group. c-Myc did not show differences between the OSSN groups. An H-score lower than 35 differentiates a squamous cell carcinoma from other OSSN lesions with a sensitivity of 83.3% and a specificity of 100%. ConclusionsThe expression of OVOL1 is a useful tool to differentiate between a squamous carcinoma of conjunctival intraepithelial neoplasms and papilloma. OVOL1 could play a role in the invasiveness of squamous neoplasms and places it as a potential therapeutic target.

Vascular Endothelial Growth Factor Expression Patterns in non- Human Papillomavirus – Related Pterygia: An Experimental Study on Cell Spot Arrays Digital Analysis

Purpose The role of angiogenic factors –such as vascular endothelial growth factor (VEGF) – in the development and progression of pterygia lesions remains under investigation. In the current study, we analyzed VEGF protein expression in a series of pterygia and normal conjunctiva epithelia. Methods Using a liquid-based cytology assay, thirty (n = 30) cell specimens were obtained by applying a smooth scraping on conjunctiva epithelia and fixed accordingly. None of them had a history of Human Papillomavirus (HPV) infection. Similarly, the same process was applied also in normal conjunctiva epithelia (n = 10; control group). We constructed five (n = 5) slides each containing eight (n = 8) cell spots. An immunocytochemistry (ICC) assay was implemented. Digital image analysis was also performed for evaluating objectively the corresponding immunostaining intensity levels. Results All the examined pterygia cell samples over-expressed the marker. High staining intensity levels were detected in 15/30 (50%), whereas the rest 15/30 (50%) demonstrated moderate expression. Overall VEGF expression was statistically significantly higher in pterygia compared to normal conjunctiva epithelia (p=.0001). Concerning the other parameters, VEGF protein expression did not associate with the gender of the patients (p = 0.518), the presence of a recurrent lesion (p = 0.311), the anatomical location (p = 0.191) or with their morphology (p = 0.316). Interestingly, the recurrent lesions demonstrated the highest levels of VEGF expression. Conclusions VEGF overexpression is a frequent event in pterygia playing a potentially central molecular role in the progression of the lesion. Cell spot array analysis -based on liquid cytology- seems to be an innovative, easy-to-use technique for analyzing a broad variety of molecules in multiple specimens on the same slide by applying different ICC assays.

NLRP3 and NLRP6 expression in pterygium and normal conjunctiva and their relationship with pterygium formation and recurrence.

This study aimed to explore the pterygium formation and recurrence, by detecting the expression of Nod-like receptor pyrin domain 3 (NLRP3) and Nod-like receptor pyrin domain 6 (NLRP6) in pterygium and evaluate the correlation between NLRP3 and NLRP6 in pterygium. In this prospective study, the expression levels of NLRP3 and NLRP6, with their related effectors, were evaluated in primary pterygium (n = 40) and recurrent pterygium (n = 32) tissue samples and compared with normal conjunctiva (n = 11) tissue samples by immunohistochemistry. Compared to the normal conjunctiva group, the expression levels of NLRP3, caspase-1, IL-18, and IL-1β, were significantly higher, and NLRP6 showed an expression that was significantly lower in pterygium tissue samples (P < 0.05, respectively). Compared to the primary pterygium group, the expression levels of NLRP3, caspase-1, IL-18 and IL-1β were significantly higher, and NLRP6 showed an expression that was significantly lower in recurrent pterygium tissue samples (P < 0.05, respectively).There was a negative correlation between NLRP3 expression and NLRP6 expression in normal conjunctival (r = -0.739, P = 0.009) and pterygium (r = -0.533, P = 0.000). NLRP3 and NLRP6 may be involved in the formation and recurrence of pterygium. NLRP6 may play an anti-inflammatory role in normal conjunctival tissue to maintain conjunctival homeostasis.

Differential expression of TGF-β1 receptors ALK1/ALK5 in pterygium and normal conjunctiva tissues

Differential expression of TGF-β1 receptors ALK1/ALK5 in pterygium and normal conjunctiva tissues

Investigation of Cytotoxic Effects of Recombinant Human Interferon Lambda-1 and Its Pegylated Form on Human Conjunctival Epithelial Cells

Currently, there are no efficacious, all-purpose antiviral medicines for the treatment of ocular surface infections caused by viruses. At the same time, type III interferons demonstrate high potency for histological barriers, such as the conjunctiva. Modification of protein molecules in native products can significantly improve their pharmacodynamic properties. Thus, it seems reasonable to develop antiviral medicines based on interferon lambda (IFN-λ1) and its pegylated form (PEG IFN-λ1).The aim of the studywas to evaluate thein vitrocytotoxic effect of recombinant human IFN-λ1 and its pegylated form onChang conjunctiva clone 1-5c-4human conjunctival cells.Materials and methods:PEG IFN-λ1 was obtained by the electron beam immobilisation method. A normal human conjunctival cell lineChang conjunctiva clone 1-5c-4was used for cell cultivation. The MTT test was used to assess the cytotoxic effect. Cell proliferative activity was studied by measuring microelectrode impedance. Ultrastructural changes were assessed by electron microscopy. Statistical processing was performed using the Statistica 10.0 software package.Results:IFN-λ1 (37 μg/mL) and PEG IFN-λ1 (42 μg/mL) had no significant cytotoxic effect on the human conjunctiva cell culture and the cell proliferative activity. The analysis of ultrastructural changes demonstrated that IFN-λ1 activated metabolic processes in the cells, and PEG IFN-λ1 promoted differentiation and keratinisation of epithelial cells and led to modification of the cell membrane. A ten-fold increase in IFN-λ1 and PEG IFN-λ1 concentration (to 370 μg/mL and 420 μg/mL, respectively) reduced the cell viability by 15–20% as compared to the intact control.Conclusions:the study results demonstrated that IFN-λ1 and PEG IFN-λ1 could be used as active pharmaceutical ingredients in the development of medicines for the treatment of conjunctival viral infections.

What's in an Eye?

What's in an Eye?