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

Abstract

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.

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

“…We found levels of MMP9 to be decreased in the severe form compared to direct alkali burn. Our observation of decreased MMP9 in the severe form of NK is in line with Fini’s group observation, where MMP9 knockout showed delayed epithelial wound healing [ 6 , 7 ]. Henceforth, clinically relating it to the extent of opacity obtained in the two model systems, with an opaque cornea in trephination with alkali (a severe form of NK) vs. milder opacity in direct alkali burn.…”

“…Laboratory tests, such as a tear film analysis, can be used to measure tear production and evaluate signs of infection. IL6 is a known pro-inflammatory cytokine and plays a very important role in persistent epithelial defect, which is a key feature of NK [ 6 ]. MMP9 is important during tissue remodeling, wound healing, and fibrosis [ 7 ].…”

Ferroptosis: a novel mechanism of cell death in ophthalmic conditions

“…GPX4, an essential player in ferroptosis, is integral to maintaining the oxidative balance, ensuring cell survival, and facilitating the wound healing process in corneal epithelial cells ( 228 ). Research conducted by Sakai et al.…”