Krupple-like factor 4 (KLF4) methylation signature in host cell in active viral keratitis with epithelial manifestation.

HSV1 presents as epithelial or stromal keratitis or keratouveitis and can lead to sight-threatening complications. KLF4, a critical transcription factor, and regulator of cell growth and differentiation, is essential in corneal epithelium stratification and homeostasis. Here, we want to understand the epigenetic modification specifically the methylation status of KLF4 in epithelium samples of HSV1 keratitis patients. After obtaining consent, epithelial scrapes were collected from 7 patients with clinically diagnosed HSV1 keratitis and 7 control samples (patients undergoing photorefractive keratectomy). Genomic DNA was isolated from the collected samples using the Qiagen DNeasy Kit. Subsequently, bisulfite modification was performed. The bisulphite-modified DNA was then subjected to PCR amplification using specific primers designed to target the KLF4, ACTB gene region, allowing for the amplification of methylated and unmethylated DNA sequences. The amplified DNA products were separated and visualized on a 3% agarose gel. KLF4 hypermethylation was found in 6 out of 7 (85.71%) eyes with viral keratitis, while 1 eye showed hypomethylation compared to PRK samples. Out of these 6, there were 2 each of epithelial dendritic keratitis, epithelial geographical keratitis, and neurotrophic keratitis. The patient with hypomethylated KLF4 had a recurrent case of HSV1 keratitis with multiple dendrites and associated vesicular lesions of the lip along with a history of fever. KLF4 hypermethylation in most viral keratitis cases indicated the under functioning of KLF4 and could indicate a potential association between KLF4 hypermethylation and the development or progression of HSV1 keratitis.

Evaluation of stresses and displacement in the craniofacial region as a reaction to bone-anchored maxillary protraction in conjugation with posterior bite plane and rapid maxillary expansion in patients with Class III malocclusion: A finite element analysis study.

INTRODUCTION: Treating a Class III malocclusion is often challenging for orthodontists. Bone-anchored maxillary protraction (BAMP) is known for achieving a significant maxillary protraction. The study aimed to evaluate the stress distribution and displacement of craniofacial bones as a reaction to the forces of BAMP, along with rapid maxillary expander and the posterior bite plane, in growing patients with skeletal Class III malocclusion using a finite element method. METHODS: An finite element model was constructed from the spiral computed tomographic images of a skull from an 11-year-old growing patient with skeletal Class III malocclusion along with BAMP, rapid maxillary expander, and the posterior bite plane. The created model had 105,189 nodes and 481,066 elements. After assigning the appropriate material properties and the boundary condition, 800 g of transverse force per side and a Class III intraoral elastic 250 g of force per side were applied to the model, and after the postprocessing, the results were obtained in the form of color bands. RESULTS: The maxilla and the attached structures were displaced and expanded transversely. The maxilla was displaced anteriorly by 0.692 mm, and the mandible was displaced backward by 0.204 mm in the sagittal direction. The anterior region of the maxilla and mandible, dentition, and nasal bone were rotated counterclockwise. Displacement in an upward direction was greatest at the symphysis region of the mandible. The stresses experienced by most of the bones were tensile, with the maxilla and maxillary dentition experiencing the maximum. CONCLUSIONS: Favorable changes were appreciated with maxillary forward and mandibular backward displacement, with appreciable tensile stresses in all the bones.

An artificial neural network approach for rational decision-making in borderline orthodontic cases: A preliminary analytical observational in silico study.

INTRODUCTION: Artificial intelligence (AI) technology has transformed the way healthcare functions in the present scenario. In orthodontics, expert systems and machine learning have aided clinicians in making complex, multifactorial decisions. One such scenario is an extraction decision in a borderline case. OBJECTIVE: The present in silico study was planned with the intention of building an AI model for extraction decisions in borderline orthodontic cases. DESIGN: An observational analytical study. SETTING: Department of Orthodontics, Hitkarini Dental College and Hospital, Madhya Pradesh Medical University, Jabalpur, India. METHODS: An artificial neural network (ANN) model for extraction or non-extraction decisions in borderline orthodontic cases was constructed based on a supervised learning algorithm using the Python (version 3.9) Sci-Kit Learn library and feed-forward backpropagation method. Based on 40 borderline orthodontic cases, 20 experienced clinicians were asked to recommend extraction or non-extraction treatment. The decision of the orthodontist and the diagnostic records, including the selected extraoral and intra-oral features, model analysis and cephalometric analysis parameters, constituted the training dataset of AI. The built-in model was then tested using a testing dataset of 20 borderline cases. After running the model on the testing dataset, the accuracy, F1 score, precision and recall were calculated. RESULTS: The present AI model showed an accuracy of 97.97% for extraction and non-extraction decision-making. The receiver operating curve (ROC) and cumulative accuracy profile showed a near-perfect model with precision, recall and F1 values of 0.80, 0.84 and 0.82 for non-extraction decisions and 0.90, 0.87 and 0.88 for extraction decisions. LIMITATION: As the present study was preliminary in nature, the dataset included was too small and population-specific. CONCLUSION: The present AI model gave accurate results in decision-making capabilities related to extraction and non-extraction treatment modalities in borderline orthodontic cases of the present population.

The secreted Ly-6/uPAR related protein-1 (SLURP1) stabilizes epithelial cell junctions and suppresses TNF-α-induced cytokine production.

The secreted Ly-6/uPAR related protein-1 (SLURP1) is an anti-angiogenic and anti-inflammatory peptide highly expressed by the mucosal epithelial cells. SLURP1 is abundantly expressed by the corneal epithelial cells and is significantly downregulated when these cells are transformed and adapted for culture in vitro. Here we studied the effect of overexpressing SLURP1 in Human Corneal Limbal Epithelial (HCLE) cells cultured in vitro. The expression of DSP1, DSG1, TJP1 and E-Cadherin was significantly upregulated in two different SLURP1-overexpressing HCLE cell (HCLE-SLURP1) clones. HCLE-SLURP1 cells also displayed a significant decrease in tumor necrosis factor-α (TNF-α)-induced upregulation of (i) IL-8 from 7.4- to 2.9- and 2.1-fold, (ii) IL-1ß from 4.9- to 3.9- and 2.9-fold, (iii) CXCL1 from 9- to 3.3- and 5.5-fold, and (iv) CXCL2 from 4.8- to 2.1- and 2.8-fold. ELISAs revealed a concomitant decrease in IL-8 levels in cell culture supernatants from 789 pg/ml in the control, to 503 and 352 pg/ml in HCLE-SLURP1 cells. Consistently, cytosolic IκB expression was elevated in HCLE-SLURP1 cells with a concurrent suppression of TNF-α-activated nuclear translocation of NF-κB. Collectively, these results elucidate the beneficial effects of SLURP1 in stabilizing the HCLE intercellular junctions and suppressing the TNF-α-induced upregulation of inflammatory cytokines by suppressing NF-κB nuclear translocation.

Cancer medicine: a direction.

OBJECTIVE: The medicine of cancer is directed in this paper. The pie theory is applied for the proposed medicine. The improbability and un-constancy are the major theories, which are used to design this anti-cancer medicine.

Assessment of Changes in Nickel and Chromium Levels in the Gingival Crevicular Fluid during Fixed Orthodontic Treatment.

BACKGROUND: Various components of appliances used in fixed orthodontic treatment are fabricated from materials that are highly resistant in nature and have high strength and biocompatibility. Corrosion of materials occurs inside the oral cavity due to numerous environmental or oral factors that act on them. These factors include temperature, pH variation, salivary conditions, mechanical loads, microbiological and enzymatic activity, and various food components. Gingival crevicular fluid (GCF) is the material obtained from the gingival sulcus and might act as a potential source for various biomarkers in the orthodontic setup because inflammatory-induced response is directly related to orthodontic forces in GCF. In the light of above-mentioned data, we planned this study to assess and evaluate the changes occurring in nickel and chromium levels in the GCF during fixed orthodontic treatment. MATERIALS AND METHODS: This study included assessment of 30 patients who underwent fixed orthodontic treatment. Three samples were taken from the GCF of the patients giving a total of 90 samples. The samples were collected at the following time intervals: At baseline (pretreatment time), 1 month after the start of orthodontic treatment, and at 6 months after the commencement of orthodontic treatment. Cellulose strips were used for isolation of the tooth region. For GCF collection, a standardized cellulose acetate absorbent strip was used. Placement of the strips was done in the sulcus for 60 seconds for the collection of the samples. Refrigeration of the specimen bottles was done for a minimum of 7 days and was then sent to a laboratory where specimens were transferred for atomic absorption spectrophotometry. All the results were analyzed by Statistical Package for the Social Sciences software. RESULTS: At 1 month, the mean value of nickel and chromium in GCF was found to be 4.5 and 4.9 ug/gm of GCF respectively. While comparing the mean nickel levels between 1 and 6 months and between baseline and 6 months, significant results were obtained. Significant results were also obtained while comparing the mean values of chromium in GCF between baseline and 6 months and between 1 and 6 months. Gingival health index of the patients was found to be associated with increased inflammation with the progression of time of orthodontic treatment. CONCLUSION: Levels of nickel and chromium might show considerable elevation in the GCF with time along with an increase in the severity of inflammation in the gingival health in patients undergoing fixed orthodontic treatment. CLINICAL SIGNIFICANCE: Regular oral prophylaxis of the patients undergoing orthodontic treatment should be done to avoid tox-icities caused by the release of nickel and chromium and for maintenance of good oral hygiene and oral health.

Isothiocyanates: a class of bioactive metabolites with chemopreventive potential.

In recent years, growing interest has been focused on the field of chemoprevention using natural therapies. The reason to turn toward "natural" remedies is associated with diverse beneficial pharmacological properties of natural compounds. Isothiocyanates (ITCs), the major pharmacological active constituents of cruciferous vegetables, are derived from the enzymatic hydrolysis of glucosinolates (GSLs). ITCs govern many intracellular targets including cytochrome P 450 (CYP) enzymes, proteins involved in antioxidant response, tumorigenesis, apoptosis, cell cycle, and metastasis. Investigation of the mechanisms of anti-cancer drugs has given important information regarding the use of natural chemopreventive compounds. This extensive review covers various molecular aspects of the interactions of ITCs with their recognized cellular targets involved in cancer treatment in order to enhance anti-tumor outcome with decreased toxicity to patients.

Pathological vitreous causes cell line-derived (but not donor-derived) retinal pigment epithelial cells to display proliferative vitreoretinopathy-like features in culture.

BACKGROUND: It is well understood that epithelial mesenchymal transformation occurs when retinal pigment epithelial cells, sourced from either a cell line or cadaver eye, are cultured in the presence of cadaver-derived vitreous. We sought to study the changes in retinal pigment epithelial cells when cell line-derived retinal pigment epithelial cells are cultured in the presence of pathological vitreous. DESIGN: Prospective study. SAMPLES: 42 patients with rhegmatogenous retinal detachments. METHODS: D407 retinal pigment epithelial cells were cultured in the presence of cadaver-derived vitreous or vitreous/subretinal fluid derived from patients undergoing retinal reattachment surgeries. Besides the changes in phenotypic characteristics, the viability, proliferation, migration, mesenchymal marker expression and changes in the extracellular matrix components were also evaluated. MAIN OUTCOME MEASURES: Fibrotic phenotype in cell culture. RESULTS: Our study clearly demonstrates that cell line-derived retinal pigment epithelial cells (unlike donor-derived retinal pigment epithelial cells) cultured in the presence of patient-derived vitreous/subretinal fluid, exhibit characteristic features of proliferative vitreoretinopathy. CONCLUSIONS: We propose that it is the synergistic effect of the combined use of (i) pathological vitreous, rather than cadaver-derived vitreous (since rhegmatogenous retinal detachment-derived pathological vitreous and subretinal fluid contain exaggerated amounts of growth factors, which could predispose to proliferative vitreoretinopathy development) and (ii) cells from an immortal cell culture (cell line), rather than from primary cell cultures (since cells subjected to continuous serial passaging acquire some mesenchymal characteristics), which together result in not only a unique phenotype, but also prime these cells towards display of features associated with proliferative vitreoretinopathy.

Pulmonary oedema producing toxin from Mesobuthus tamulus venom augments cardio-respiratory reflexes through B2 kinin receptors.

The current study was undertaken to compare the effects of pulmonary oedema producing toxin (PO-Tx) isolated from Mesobuthus tamulus venom on cardio-respiratory reflexes with exogenously administered bradykinin (BK) and to delineate the type of BK receptors mediating these responses. Jugular venous injection of phenyldiguanide (PDG) in anaesthetized rats produced reflex bradycardia, hypotension and apnoea. The PDG-induced reflex was augmented (two folds) by PO-Tx. The pulmonary water content in PO-Tx treated group was also increased. The PO-Tx-induced reflex changes as well as pulmonary oedema were blocked by-Hoe-140 implicating the involvement of B2 kinin receptors. Exogenous BK also produced augmentation (two folds) of the PDG-induced reflexes and increased the pulmonary water content. The BK-induced augmentation was blocked by pre-treatment with des-Arg10 Hoe 140 (a B1 receptor antagonist) and Hoe 140 (B2 receptor antagonist). However, these antagonists did not prevent the development of BK-induced pulmonary oedema. Present results indicate that PO-Tx augmented the PDG-induced reflex responses similar to BK and the PO-Tx induced augmentation of reflexes is mediated through B2 receptors.

Chemical (Alkali) Burn-Induced Neurotrophic Keratitis Model in New Zealand Rabbit Investigated Using Medical Clinical Readouts and In Vivo Confocal Microscopy (IVCM).

PURPOSE: Chemical eye injury is an acute emergency that can result in vision loss. Neurotrophic keratitis (NK) is the most common long-term manifestation of chemical injury. NK due to alkali burn affects ocular surface health and is one of its most common causes. Here, we established a rabbit model of corneal alkali burns to evaluate the severity of NK-associated changes. MATERIAL METHODS: Alkali burns were induced in NZ rabbits by treating the cornea with (i) a 5 mm circular filter paper soaked in 0.75 N NaOH for 10 s (Mild NK) and (ii) trephination using a guarded trephine (5 mm diameter and 150-micron depth), followed by alkali burn, with a 5 mm circular filter paper soaked in 0.75 N NaOH for 10 s (a severe form of NK). Immediately after, the cornea was rinsed with 10 mL of normal saline to remove traces of NaOH. Clinical features were evaluated on Day 0, Day 1, Day 7, Day 15, and Day 21 post-alkali burn using a slit lamp, Pentacam, and anterior segment optical coherence tomography (AS-OCT). NK-like changes in epithelium, sub-basal nerve plexus, and stroma were observed using in vivo confocal microscopy (IVCM), and corneal sensation were measured using an aesthesiometer post alkali injury. After 21 days, pro-inflammatory cytokines were evaluated for inflammation through ELISA. RESULTS: Trephination followed by alkali burn resulted in the loss of epithelial layers (manifested using fluorescein stain), extensive edema, and increased corneal thickness (550 µm compared to 380 µm thickness of control) evaluated through AS-OCT and increased opacity score in alkali-treated rabbit (80 compared to 16 controls). IVCM images showed complete loss of nerve fibers, which failed to regenerate over 30 days, and loss of corneal sensation-conditions associated with NK. Cytokines evaluation of IL6, VEGF, and MMP9 indicated an increased angiogenic and pro-inflammatory milieu compared to the milder form of NK and the control. DISCUSSION: Using clinical parameters, we demonstrated that the alkali-treated rabbit model depicts features of NK. Using IVCM in the NaOH burn animal model, we demonstrated a complete loss of nerve fibers with poor self-healing capability associated with sub-basal nerve degeneration and compromised corneal sensation. This pre-clinical rabbit model has implications for future pre-clinical research in neurotrophic keratitis.

In vitro Assay to Examine the Function of Tears on Corneal Epithelial Cells.

Tears contain numerous secreted factors, enzymes, and proteins that help in maintaining the homeostatic condition of the eye and also protect it from the external environment. However, alterations to these enzymes and/or proteins during pathologies such as mechanical injury and viral or fungal infections can disrupt the normal ocular homeostasis, further contributing to disease development. Several tear film components have a significant role in curbing disease progression and promoting corneal regeneration. Additionally, several factors related to disease progression are secreted into the tear film, thereby serving as a valuable reservoir of biomarkers. Tears are readily available and can be collected via non-invasive techniques or simply from contact lenses. Tears can thus serve as a valuable and easy source for studying disease-specific biomarkers. Significant advancements have been made in recent years in the field of tear film proteomics, lipidomics, and transcriptomics to allow a better understanding of how tears can be utilized to gain insight into the etiology of diseases. These advancements have enabled us to study the pathophysiology of various disease states using tear samples. However, the mechanisms by which tears help to maintain corneal homeostasis and how they are able to form the first line of defense against pathogens remain poorly understood and warrant detailed in vitro studies. Herein, we have developed an in vitro assay to characterize the functional importance of patient isolated tears and their components on corneal epithelial cells. This novel approach closely mimics real physiological conditions and could help the researchers gain insight into the underlying mechanisms of ocular pathologies and develop new treatments. Key features • This method provides a new technique for analyzing the effect of tear components on human corneal epithelial cells. • The components of the tears that are altered in response to diseases can be used as a biomarker for detecting ocular complications. • This procedure can be further employed as an in vitro model for assessing the efficacy of drugs and discover potential therapeutic interventions.

Kuragel: A biomimetic hydrogel scaffold designed to promote corneal regeneration.

Cornea-related injuries are the most common cause of blindness worldwide. Transplantation remains the primary approach for addressing corneal blindness, though the demand for donor corneas outmatches the supply by millions. Tissue adhesives employed to seal corneal wounds have shown inefficient healing and incomplete vision restoration. We have developed a biodegradable hydrogel - Kuragel, with the ability to promote corneal regeneration. Functionalized gelatin and hyaluronic acid form photo-crosslinkable hydrogel with transparency and compressive modulus similar to healthy human cornea. Kuragel composition was tuned to achieve sufficient adhesive strength for sutureless integration to host tissue, with minimal swelling post-administration. Studies in the New Zealand rabbit mechanical injury model affecting corneal epithelium and stroma demonstrate that Kuragel efficiently promotes re-epithelialization within 1 month of administration, while stroma and sub-basal nerve plexus regenerate within 3 months. We propose Kuragel as a regenerative treatment for patients suffering from corneal defects including thinning, by restoration of transparency and thickness.

Design methodology of a new wavelet basis function for fetal phonocardiographic signals.

Fetal phonocardiography (fPCG) based antenatal care system is economical and has a potential to use for long-term monitoring due to noninvasive nature of the system. The main limitation of this technique is that noise gets superimposed on the useful signal during its acquisition and transmission. Conventional filtering may result into loss of valuable diagnostic information from these signals. This calls for a robust, versatile, and adaptable denoising method applicable in different operative circumstances. In this work, a novel algorithm based on wavelet transform has been developed for denoising of fPCG signals. Successful implementation of wavelet theory in denoising is heavily dependent on selection of suitable wavelet basis function. This work introduces a new mother wavelet basis function for denoising of fPCG signals. The performance of newly developed wavelet is found to be better when compared with the existing wavelets. For this purpose, a two-channel filter bank, based on characteristics of fPCG signal, is designed. The resultant denoised fPCG signals retain the important diagnostic information contained in the original fPCG signal.

Lightweight convolution transformer for cross-patient seizure detection in multi-channel EEG signals.

BACKGROUND: Epilepsy is a neurological illness affecting the brain that makes people more likely to experience frequent, spontaneous seizures. There has to be an accurate automated method for measuring seizures frequency and severity to assess the efficacy of pharmacological therapy for epilepsy. The drug quantities are often derived from patient reports which may cause significant issues owing to inadequate or inaccurate descriptions of seizures and their frequencies. METHODS AND MATERIALS: This study proposes a novel deep learning architecture-based Lightweight Convolution Transformer (LCT). The Transformer model is able to learn spatial and temporal correlated information simultaneously from the multi-channel electroencephalogram (EEG) signal to detect seizures at smaller segment lengths. In the proposed work, the lack of translation equivariance and localization of ViT is reduced using convolution tokenization, and rich information from the Transformer encoder is extracted by sequence pooling instead of the learnable class token. RESULTS: Extensive experimental results demonstrate that the proposed model on cross-patient learning can effectively detect seizures from the raw EEG signals. The accuracy and F1-score of seizure detection in the cross-patient case on the CHB-MIT dataset are 96.31% and 96.32%, respectively, at 0.5 sec segment length. In addition, the performance metrics show that the inclusion of inductive biases and attention-based pooling in the model enhances the performance and reduces the number of Transformer encoder layers, which significantly reduces the computational complexity. In this research, we provide a novel approach to enhance efficiency and simplify the architecture for multi-channel automated seizure detection.

Computer-aided diagnostic system for hypertensive retinopathy: A review.

Hypertensive Retinopathy (HR) is a retinal disease caused by elevated blood pressure for a prolonged period. There are no obvious signs in the early stages of high blood pressure, but it affects various body parts over time, including the eyes. HR is a biomarker for several illnesses, including retinal diseases, atherosclerosis, strokes, kidney disease, and cardiovascular risks. Early microcirculation abnormalities in chronic diseases can be diagnosed through retinal examination prior to the onset of major clinical consequences. Computer-aided diagnosis (CAD) plays a vital role in the early identification of HR with improved diagnostic accuracy, which is time-efficient and demands fewer resources. Recently, numerous studies have been reported on the automatic identification of HR. This paper provides a comprehensive review of the automated tasks of Artery-Vein (A/V) classification, Arteriovenous ratio (AVR) computation, HR detection (Binary classification), and HR severity grading. The review is conducted using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) protocol. The paper discusses the clinical features of HR, the availability of datasets, existing methods used for A/V classification, AVR computation, HR detection, and severity grading, and performance evaluation metrics. The reviewed articles are summarized with classifiers details, adoption of different kinds of methodologies, performance comparisons, datasets details, their pros and cons, and computational platform. For each task, a summary and critical in-depth analysis are provided, as well as common research issues and challenges in the existing studies. Finally, the paper proposes future research directions to overcome challenges associated with data set availability, HR detection, and severity grading.

Retinoblastoma: A review of the molecular basis of tumor development and its clinical correlation in shaping future targeted treatment strategies.

Retinoblastoma is a retinal cancer that affects children and is the most prevalent intraocular tumor worldwide. Despite tremendous breakthroughs in our understanding of the fundamental mechanisms that regulate progression of retinoblastoma, the development of targeted therapeutics for retinoblastoma has lagged. Our review highlights the current developments in the genetic, epigenetic, transcriptomic, and proteomic landscapes of retinoblastoma. We also discuss their clinical relevance and potential implications for future therapeutic development, with the aim to create a frontline multimodal therapy for retinoblastoma.

Road Traffic Accidents among Patients Visiting Department of Emergency of a Tertiary Care Centre: A Descriptive Cross-sectional Study.

Introduction: Road Traffic Accidents, are one of the major neglected global health burdens which are predicted to be the 7th leading cause of global deaths by 2030 as per the World Health Organization hence, seem to be one of the major global threats in near future. Most road traffic accidents affect the most vulnerable age groups in developing countries. The aim of this study was to find out the prevalence of road traffic accidents among patients visiting the Department of Emergency of a tertiary care centre. Methods: A descriptive cross-sectional study was conducted among patients visiting the department of emergency of a tertiary care centre from 16 September 2022 to 15 October 2022. Ethical approval was obtained from the Institutional Review Committee (Reference number: IRC-DMCRI: 307/079/080). All the road traffic accidents cases recorded in the Emergency Department from 14 April 2021 to 13 April 2022 were taken. Convenience sampling was used. Point estimate and 95% confidence interval were calculated. Results: Among 29735 patients, the prevalence of road traffic accidents was 1340 (4.50%) (4.26-4.74, 95% Confidence Interval). Among these, 1037 (77.4%) were male and 303 (22.6%) were female. Road traffic accidents among two-wheelers were 1065 (79.48%) followed by pedestrian 703 (52.46%). Mangsir witness the higher number of cases, 137 (13.90%) followed by Kartik, 170 (12.69%). Conclusions: The prevalence of road traffic accidents was similar to other studies done in similar settings. In our study, young people of highly productive and active age groups were the most common victims. Keywords: emergencies; prevalence; traffic accidents.

Prolonged Inflammation and Infectious Changes in the Corneal Epithelium Are Associated with Persistent Epithelial Defect (PED).

Purpose: Failure of rapid re-epithelialization within 10-14 days after corneal injury, even with standard supportive treatment, is referred to as persistent corneal epithelial (CE) defect (PED). Though an array of genes regulates reepithelization, their mechanisms are poorly understood. We sought to understand the network of genes driving the re-epithelialization in PED. Method: After obtaining informed consent, patients underwent an ophthalmic examination. Epithelial scrapes and tears samples of six PED patients and six individuals (control) undergoing photorefractive keratectomy (PRK) were collected. RNA isolation and quantification were performed using either the epithelial scrape taken from PED patients or from HCLE cells treated with control tears or tears of PED patients. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to detect the expression of a few important genes in CE homeostasis, inflammation, and cell-cell communication, viz., Kruppel-like factor 4 (KLF4), GPX4, IL6, TNFα, STING, IL8, desmoglein, and E-cadherin, among others. Their expressions were normalized with their respective housekeeping genes and fold changes were recorded. KLF4 localization and MMPs activity was carried out via immunofluorescence and zymography, respectively. Results: KLF4, a transcription factor important for CE homeostasis, was upregulated in tears-treated HCLE cells and downregulated in PED patients compared to the healthy PRK group. Cell-cell communication genes were also upregulated in tears-treated cells, whereas they were downregulated in the PED tissue group. Genes involved in proinflammation (IL6, 282-fold; TNFα, 43-fold; IL8, 4.2-fold) were highly upregulated in both conditions. MMP9 activity increased upon tears treatment. Conclusions: This study suggests that tears create an acute proinflammatory milieu driving the PED disease pathology, whereas the PED patients scrapes are an indicator of the chronic stage of the disease. Interferons, pro-inflammatory genes, and their pathways are involved in PED, which can be a potential target for inducing epithelialization of the cornea.

Secreted Ly-6/uPAR-related protein-1 (SLURP1) is a pro-differentiation factor that stalls G1-S transition during corneal epithelial cell cycle progression.

PURPOSE: Previously we demonstrated that the secreted Ly-6/uPAR related protein-1 (SLURP1), abundantly expressed in the corneal epithelium (CE) and secreted into the tear fluid, serves as an anti-inflammatory and anti-angiogenic molecule. Here we describe the Slurp1-null (Slurp1X-/-) mouse corneal phenotype for the first time. METHODS: We compared the 10-week-old wild type (WT) and Slurp1X-/- mouse corneal (i) histology by hematoxylin-eosin and periodic acid-Schiff's reagent staining, (ii) cell proliferation by immunostaining for Ki67, (iii) cell adhesion molecules by immunostaining for desmosomal and tight junction proteins, (iv) barrier function by fluorescein staining and (v) wound-healing by epithelial debridement. Effect of SLURP1 on cell cycle was quantified in human corneal limbal epithelial (HCLE) cells engineered to express SLURP1 (HCLE-SLURP1). RESULTS: WT and Slurp1X-/- corneal histology was largely comparable, other than a few loosely attached superficial cells in Slurp1X-/- corneas. Compared with the WT, Slurp1X-/- corneas displayed (i) increase in Ki67+ cells, (ii) altered expression and/or localization of tight junction proteins Tjp1 and Pard3, and desmosomal Dsp, (iii) increased superficial fragility and (iv) slower CE wound healing. HCLE-SLURP1 cells displayed (i) decrease in Ki67+ cells, (ii) increased cell number doubling time, (iii) stalling in G1-S phase transition during cell cycle, and (iv) downregulation of cyclins CCNE and CCND1/D2, cyclin-dependent kinases CDK4 and CDK6, and upregulation of CDK inhibitor p15/CDKN2B. CONCLUSIONS: Collectively, these results elucidate that Slurp1X-/- CE cell homeostasis is altered and suggest that SLURP1 is a pro-differentiation factor that stalls G1-S transition during cell cycle progression by downregulating cyclins and upregulating p15/CDKN2B.