Reversed Corneal Fibroblasts Therapy Restores Transparency of Mouse Cornea after Injury.

Cell-based therapies using corneal stromal stem cells (CSSC), corneal keratocytes, or a combination of both suppress corneal scarring. The number of quiescent keratocytes in the cornea is small; it is difficult to expand them in vitro in quantities suitable for transplantation. This study examined the therapeutic effect of corneal fibroblasts reversed into keratocytes (rCF) in a mouse model of mechanical corneal injury. The therapeutic effect of rCF was studied in vivo (slit lamp, optical coherence tomography) and ex vivo (transmission electron microscopy and immunofluorescence staining). Injection of rCF into the injured cornea was accompanied by recovery of corneal thickness, improvement of corneal transparency, reduction of type III collagen in the stroma, absence of myofibroblasts, and the improvement in the structural organization of collagen fibers. TEM results showed that 2 months after intrastromal injection of cells, there was a decrease in the fibril density and an increase in the fibril diameter and the average distance between collagen fibrils. The fibrils were well ordered and maintained the short-range order and the number of nearest-neighbor fibrils, although the averaged distance between them increased. Our results demonstrated that the cell therapy of rCF from ReLEx SMILe lenticules promotes the recovery of transparent corneal stroma after injury.

The effect of lactoferin on the free radical and cytokine status of cornea in the experimental thermal burn.

The free radical and cytokine statuses of the cornea during its thermal burn and the possibility of its correction by lactoferrin have been studied in Soviet Chinchilla rabbits. The development of a corneal thermal burn was accompanied by the development of oxidative stress (increased levels of TBA-reactive substances and carbonyl derivatives of proteins, decreased activity of SOD and GPx enzymes) and a pronounced inflammatory reaction with increased levels of TNF-1α, IL-10, TGF-1ß. The use of lactoferrin had a pronounced therapeutic effect, which was manifested by accelerated healing, prevention of the development of complications (corneal perforations), a decrease in the severity of oxidative stress, an increase in the concentrations of TNF-1α (in the early stages), IL-10 (in the later stages), TGF-1ß (throughout the experiment). At the same time, by the end of regeneration more severe corneal opacification was recognized compared to the control group. This may be associated with an increased level of anti-inflammatory cytokines, especially TGF-1ß.

PAX6/CXCL14 regulatory axis promotes the repair of corneal injury by enhancing corneal epithelial cell proliferation.

BACKGROUND: Corneal injuries, often leading to severe vision loss or blindness, have traditionally been treated with the belief that limbal stem cells (LSCs) are essential for repair and homeostasis, while central corneal epithelial cells (CCECs) were thought incapable of such repair. However, our research reveals that CCECs can fully heal and maintain the homeostasis of injured corneas in rats, even without LSCs. We discovered that CXCL14, under PAX6's influence, significantly boosts the stemness, proliferation, and migration of CCECs, facilitating corneal wound healing and homeostasis. This finding introduces CXCL14 as a promising new drug target for corneal injury treatment. METHODS: To investigate the PAX6/CXCL14 regulatory axis's role in CCECs wound healing, we cultured human corneal epithelial cell lines with either increased or decreased expression of PAX6 and CXCL14 using adenovirus transfection in vitro. Techniques such as coimmunoprecipitation, chromatin immunoprecipitation, immunofluorescence staining, western blot, real-time PCR, cell colony formation, and cell cycle analysis were employed to validate the axis's function. In vivo, a rat corneal epithelial injury model was developed to further confirm the PAX6/CXCL14 axis's mechanism in repairing corneal damage and maintaining corneal homeostasis, as well as to assess the potential of CXCL14 protein as a therapeutic agent for corneal injuries. RESULTS: Our study reveals that CCECs naturally express high levels of CXCL14, which is significantly upregulated by PAX6 following corneal damage. We identified SDC1 as CXCL14's receptor, whose engagement activates the NF-κB pathway to stimulate corneal repair by enhancing the stemness, proliferative, and migratory capacities of CCECs. Moreover, our research underscores CXCL14's therapeutic promise for corneal injuries, showing that recombinant CXCL14 effectively accelerates corneal healing in rat models. CONCLUSION: CCECs play a critical and independent role in the repair of corneal injuries and the maintenance of corneal homeostasis, distinct from that of LSCs. The PAX6/CXCL14 regulatory axis is pivotal in this process. Additionally, our research demonstrates that the important function of CXCL14 in corneal repair endows it with the potential to be developed into a novel therapeutic agent for treating corneal injuries.

Trimebutine prevents corneal inflammation in a rat alkali burn model.

Alkaline burns to the cornea lead to loss of corneal transparency, which is essential for normal vision. We used a rat corneal alkaline burn model to investigate the effect of ophthalmic trimebutine solution on healing wounds caused by alkaline burns. Trimebutine, an inhibitor of the high-mobility group box 1-receptor for advanced glycation end products, when topically applied to the burned cornea, suppressed macrophage infiltration in the early phase and neutrophil infiltration in the late phase at the wound site. It also inhibited neovascularization and myofibroblast development in the late phase. Furthermore, trimebutine effectively inhibited interleukin-1ß expression in the injured cornea. It reduced scar formation by decreasing the expression of type III collagen. These findings suggest that trimebutine may represent a novel therapeutic strategy for corneal wounds, not only through its anti-inflammatory effects but also by preventing neovascularization.

Cell spray printing combined with Lycium barbarum glycopeptide promotes repair of corneal epithelial injury.

The corneal epithelium, located as the outermost layer of the cornea, is inherently susceptible to injuries that may lead to corneal opacities and compromise visual acuity. Rapid restoration of corneal epithelial injury is crucial for maintaining the transparency and integrity of the cornea. Cell spray treatment emerges as an innovative and effective approach in the field of regenerative medicine. In our study, a cell spray printing platform was established, and the optimal printing parameters were determined to be a printing air pressure of 5 PSI (34.47 kPa) and a liquid flow rate of 30 ml/h. Under these conditions, the viability and phenotype of spray-printed corneal epithelial cells were preserved. Moreover, Lycium barbarum glycopeptide (LBGP), a glycoprotein purified from wolfberry, enhanced proliferation while simultaneously inhibiting apoptosis of the spray-printed corneal epithelial cells. We found that the combination of cell spray printing and LBGP facilitated the rapid construction of multilayered cell sheets on flat and curved collagen membranes in vitro. Furthermore, the combined cell spray printing and LBGP accelerated the recovery of the rat corneal epithelium in the mechanical injury model. Our findings offer a therapeutic avenue for addressing corneal epithelial injuries and regeneration.

Adaptor protein 14-3-3zeta promotes corneal wound healing via regulating cell homeostasis, a potential novel therapy for corneal injury.

Severe corneal injury can lead to blindness even after prompt treatment. 14-3-3zeta, a member of an adaptor protein family, contributes to tissue repair by enhancing cellular viability and inhibiting fibrosis and inflammation in renal disease or arthritis. However, its role in corneal regeneration is less studied. In this study, filter disc of 2-mm diameter soaked in sodium hydroxide with a concentration of 0.5 N was placed at the center of the cornea for 30 s to establish a mouse model of corneal alkali injury. We found that 14-3-3zeta, which is mainly expressed in the epithelial layer, was upregulated following injury. Overexpression of 14-3-3zeta in ocular tissues via adeno-associated virus-mediated subconjunctival delivery promoted corneal wound healing, showing improved corneal structure and transparency. In vitro studies on human corneal epithelial cells showed that 14-3-3zeta was critical for cell proliferation and migration. mRNA-sequencing in conjunction with KEGG analysis and validation experiments revealed that 14-3-3zeta regulated the mRNA levels of ITGB1, PIK3R1, FGF5, PRKAA1 and the phosphorylation level of Akt, suggesting the involvement of the PI3K-Akt pathway in 14-3-3zeta-mediated tissue repair. 14-3-3zeta is a potential novel therapeutic candidate for treating severe corneal injury.

INOS ablation promotes corneal wound healing via activation of Akt signaling.

Corneal injury leads to impaired normal structure of the cornea. Improving the wound healing process in epithelial cells significantly contributes to ocular damage treatments. Here, we aimed to investigate the potential mechanisms of nitric oxide (NO) and its mediator, inducible nitric oxide synthase (iNOS), in the process of corneal wound healing. We established a corneal injury model of iNOS-/- mice, and treated human corneal epithelial cell lines (HCE-2) with the iNOS inhibitor L-INL, with or without NO replenishment by supplying sodium nitroferricyanide dihydrate (SNP). Our findings showed that inhibition of NO/iNOS accelerated corneal repair, enhanced uPAR (a receptor protein indicating the migration ability), and improved epithelial cell migration. Furthermore, NO/iNOS ablation activated Akt phosphorylation, reduced neutrophil marker protein MPO expression, and downregulated the transcription of inflammation cytokines CXCL-1, CXCL-2, IL-1ß, IL-6, and TNF-α. However, the protective effects of NO/iNOS inhibition are significantly reduced by NO replenishment when treated with SNP. Therefore, we confirmed that inhibiting NO/iNOS improved the corneal wound healing by facilitating epithelial cell migration and reducing inflammatory reactions, which might be related to the activation of the Akt signaling pathway.

Dexamethasone sodium phosphate loaded nanoparticles for prevention of nitrogen mustard induced corneal injury.

Nitrogen mustard (NM) is a potent vesicating chemical warfare agent that is primarily absorbed through skin, inhalation, or ocular surface. Ocular exposure of NM can cause acute to chronic keratopathy which can eventually lead to blindness. There is a current lack of effective countermeasures against ocular exposure of NM despite their imperative need. Herein, we aim to explore the sustained effect of Dexamethasone sodium phosphate (DSP)-loaded polymeric nanoparticles (PLGA-DSP-NP) following a single subconjunctival injection in the management and prevention of corneal injury progression upon exposure to NM. DSP is an FDA approved corticosteroid with proven anti-inflammatory properties. We formulated PLGA-DSP-NP with zinc chelation ion bridging method using PLGA polymer, with particles of approximately 250 nm and a drug loading of 6.5 wt%. Under in vitro sink conditions, PLGA-DSP-NP exhibited a sustained drug release for two weeks. Notably, in NM injured cornea, a single subconjunctival (SCT) injection of PLGA-DSP-NP outperformed DSP eyedrops (0.1%), DSP solution, placebo NP, and saline, significantly mitigating corneal neovascularization, ulceration, and opacity for the two weeks study period. Through PLGA-DSP-NP injection, sustained DSP release hindered inflammatory cytokine recruitment, angiogenic factors, and endothelial cell proliferation in the cornea. This strategy presents a promising localized corticosteroid delivery system to effectively combat NM-induced corneal injury, offering insights into managing vesicant exposure.

Bone marrow mesenchymal stem cell-derived extracellular vesicles promote corneal epithelial repair and suppress apoptosis via modulation of Caspase-3 in vitro.

Corneal injuries are the major cause of blindness and visual impairment. Available treatments are limited by their efficacy and side effects. Mesenchymal stem cell-derived extracellular vesicles are presumed as functional equivalents and potential candidates for cell-free therapy. This study reports isolation and characterization of extracellular vesicles from human bone marrow mesenchymal stem cells and evaluates their role in mediating epithelial repair and apoptosis in cultured corneal epithelial cells through scratch assay, PCR, immunofluorescence, and flow cytometry in vitro. The isolated extracellular vesicles were spherical, < 150 nm in diameter, and characterized as CD9+, CD63+, CD81+, TSG101+, and Calnexin-. Further, these vesicles promoted corneal epithelial repair by enhancing proliferation and suppressed apoptosis by regulating the expression of BAD, P53, BCL-2, and cleaved CASPASE-3. Thus, our results suggest that BM-MSC-EVs might have the potential to be used for the treatment of injury-induced corneal epithelial defects. Clinical translation of this work would require further investigations.

Blast injury: Impact to the cornea.

Visual disorders are common even after mild traumatic brain injury (mTBI) or blast exposure. The cost of blast-induced vision loss in civilians, military personnel, and veterans is significant. The visual consequences of blasts associated with TBI are elusive. Active military personnel and veterans report various ocular pathologies including corneal disorders post-combat blasts. The wars and conflicts in Afghanistan, Iraq, Syria, and Ukraine have significantly increased the number of corneal and other ocular disorders among military personnel and veterans. Binocular vision, visual fields, and other visual functions could be impaired following blast-mediated TBI. Blast-associated injuries can cause visual disturbances, binocular system problems, and visual loss. About 25% of veterans exposed to blasts report corneal injury. Blast exposure induces corneal edema, corneal opacity, increased corneal thickness, damage of corneal epithelium, corneal abrasions, and stromal and endothelial abnormality including altered endothelial density, immune cell infiltration, corneal neovascularization, Descemet membrane rupture, and increased pain mediators in animal models and the blast-exposed military personnel including veterans. Immune response exacerbates blast-induced ocular injury. TBI is associated with dry eyes and pain in veterans. Subjects exposed to blasts that cause TBI should undergo immediate clinical visual and ocular examinations. Delayed visual care may lead to progressive vision loss, lengthening/impairing rehabilitation and ultimately may lead to permanent vision problems and blindness. Open-field blast exposure could induce corneal injuries and immune responses in the cornea. Further studies are warranted to understand corneal pathology after blast exposure. A review of current advancements in blast-induced corneal injury will help elucidate novel targets for potential therapeutic options. This review discusses the impact of blast exposure-associated corneal disorders.

Transient Receptor Potential Vanilloid-1 Channels Facilitate Axonal Degeneration of Corneal Sensory Nerves in Dry Eye.

Corneal nerve impairment contributes significantly to dry eye disease (DED) symptoms and is thought to be secondary to corneal epithelial damage. Transient receptor potential vanilloid-1 (TRPV1) channels abound in corneal nerve fibers and respond to inflammation-derived ligands, which increase in DED. TRPV1 overactivation promotes axonal degeneration in vitro, but whether it participates in DED-associated corneal nerve dysfunction is unknown. To explore this, DED was surgically induced in wild-type and TRPV1-knockout mice, which developed comparable corneal epithelial damage and reduced tear secretion. However, corneal mechanosensitivity decreased progressively only in wild-type DED mice. Sensitivity to capsaicin (TRPV1 agonist) increased in wild-type DED mice, and consistently, only this strain displayed DED-induced pain signs. Wild-type DED mice exhibited nerve degeneration throughout the corneal epithelium, whereas TRPV1-knockout DED mice only developed a reduction in the most superficial nerve endings that failed to propagate to the deeper subbasal corneal nerves. Pharmacologic TRPV1 blockade reproduced these findings in wild-type DED mice, whereas CD4+ T cells from both strains were equally pathogenic when transferred, ruling out a T-cell-mediated effect of TRPV1 deficiency. These data show that ocular desiccation triggers superficial corneal nerve damage in DED, but proximal propagation of axonal degeneration requires TRPV1 expression. Local inflammation sensitized TRPV1 channels, which increased ocular pain. Thus, ocular TRPV1 overactivation drives DED-associated corneal nerve impairment.

Dynamic changes of endothelial and stromal cilia are required for the maintenance of corneal homeostasis.

Primary cilia are distributed extensively within the corneal epithelium and endothelium. However, the presence of cilia in the corneal stroma and the dynamic changes and roles of endothelial and stromal cilia in corneal homeostasis remain largely unknown. Here, we present compelling evidence for the presence of primary cilia in the corneal stroma, both in vivo and in vitro. We also demonstrate dynamic changes of both endothelial and stromal cilia during corneal development. In addition, our data show that cryoinjury triggers dramatic cilium formation in the corneal endothelium and stroma. Furthermore, depletion of cilia in mutant mice lacking intraflagellar transport protein 88 compromises the corneal endothelial capacity to establish the effective tissue barrier, leading to an upregulation of α-smooth muscle actin within the corneal stroma in response to cryoinjury. These observations underscore the essential involvement of corneal endothelial and stromal cilia in maintaining corneal homeostasis and provide an innovative strategy for the treatment of corneal injuries and diseases.

The proliferative effects of stem cells from apical papilla-conditioned medium on rat corneal endothelial cells.

The cornea, positioned at the forefront of the eye, refracts the light for focusing images on the retina. Damage to this transparent structure can lead to various visual disorders. The corneal endothelial cells (CECs) are crucial for transparency and homeostasis, but lack the ability to reproduce. Significant damage results in structure destruction and vision impairment. While extensive research has aimed at the restoring the corneal endothelial layer, including endothelial proliferation for functional monolayers remains challenging. Our previous studies confirmed the proliferative activity of stem cells from apical papilla-conditioned medium (SCAP-CM) on the retinal pigmented epithelium as a single cell layer. This study investigates how SCAP-CM influences the proliferation and migration of CECs. Our results introduced Matrigel, as a new matrix component for in vitro culture of CECs. Moreover, 60% of SCAP-CM was able to stimulate CEC proliferation as well as migrate to repair wound healing during 24 h. Confluent CECs also expressed specific markers, ATP1a1, ZO-1 and CD56, indicative of CEC characteristics, aligning with the recapitulation of differentiation when forming a homogenous monolayer at the same level of isolated CECs without in vitro culture. These findings suggested that SCAP-CM administration could be useful for future preclinical and clinical applications.

Quantitative evaluation of corneal irregularity and scarring after infectious keratitis using anterior segment optical coherence tomography.

PURPOSE: Corneal scars after infectious keratitis lead to insufficient transparency and irregular astigmatism, affecting visual acuity; therefore, they should be accurately evaluated to estimate visual function. This study aimed to quantitatively evaluate corneal irregularity and scarring after infectious keratitis using anterior segment optical coherence tomography (AS-OCT). METHODS: This was an observational clinical study. We included patients who had corneal scarring after treatment of infectious keratitis between 2014 and 2021 at University of Tokyo Hospital. We retrospectively examined best spectacle-corrected visual acuity (BSCVA), average keratometric power, central corneal thickness (CCT), and four components of the Fourier harmonic analysis including spherical and asymmetry components, as well as regular astigmatism and higher-order irregularity. We included anterior and posterior corneal data and compared results with those of contralateral healthy eyes. Additionally, we quantitatively evaluated the densitometry of the cornea obtained using AS-OCT. RESULTS: A total of 122 eyes of 61 patients were examined; male predominance was observed (n = 37), and the mean patient age was 55.3 ± 19.4 years. Comparisons with contralateral healthy eyes showed that BSCVA worsened (0.30 ± 0.83 and 0.93 ± 1.36 logMAR, respectively, P = 0.003), and CCT (531.1 ± 46.2 and 591.8 ± 132.4 µm, respectively, P < 0.001) and corneal densitometry (84.4 ± 11.8 and 111.9 ± 19.2 grayscale units, respectively, P < 0.001) increased significantly in affected eyes. The asymmetry component and higher-order irregularities that were not corrected with spectacles significantly increased (both P < 0.001), and there were no significant differences in the changes among the bacterial, fungal, herpetic, and acanthamoeba types of keratitis. CONCLUSION: Corneal scarring persisted after treatment for infectious keratitis, and the asymmetry and irregularities of corneal astigmatism increased as visual acuity deteriorated. AS-OCT with the Fourier harmonic analysis was useful for evaluating corneal topographic changes in patients with corneal scarring after keratitis.

Establishing a Dexamethasone Treatment Regimen To Alleviate Sulfur Mustard-Induced Corneal Injuries in a Rabbit Model.

Sulfur mustard (SM) is an ominous chemical warfare agent. Eyes are extremely susceptible to SM toxicity; injuries include inflammation, fibrosis, neovascularization (NV), and vision impairment/blindness, depending on the exposure dosage. Effective countermeasures against ocular SM toxicity remain elusive and are warranted during conflicts/terrorist activities and accidental exposures. We previously determined that dexamethasone (DEX) effectively counters corneal nitrogen mustard toxicity and that the 2-hour postexposure therapeutic window is most beneficial. Here, the efficacy of two DEX dosing frequencies [i.e., every 8 or 12 hours (initiated, as previously established, 2 hours after exposure)] until 28 days after SM exposure was assessed. Furthermore, sustained effects of DEX treatments were observed up to day 56 after SM exposure. Corneal clinical assessments (thickness, opacity, ulceration, and NV) were performed at the day 14, 28, 42, and 56 post-SM exposure time points. Histopathological assessments of corneal injuries (corneal thickness, epithelial degradation, epithelial-stromal separation, inflammatory cell, and blood vessel counts) using H&E staining and molecular assessments (COX-2, MMP-9, VEGF, and SPARC expressions) were performed at days 28, 42, and 56 after SM exposure. Statistical significance was assessed using two-way ANOVA, with Holm-Sidak post hoc pairwise multiple comparisons; significance was established if P < 0.05 (data represented as the mean ± S.E.M.). DEX administration every 8 hours was more potent than every 12 hours in reversing ocular SM injury, with the most pronounced effects observed at days 28 and 42 after SM exposure. These comprehensive results are novel and provide a comprehensive DEX treatment regimen (therapeutic-window and dosing-frequency) for counteracting SM-induced corneal injuries. SIGNIFICANCE STATEMENT: The study aims to establish a dexamethasone (DEX) treatment regimen by comparing the efficacy of DEX administration at 12 versus 8 hours initiated 2 hours after exposure. DEX administration every 8 hours was more effective in reversing sulfur mustard (SM)-induced corneal injuries. SM injury reversal during DEX administration (initial 28 days after exposure) and sustained [further 28 days after cessation of DEX administration (i.e., up to 56 days after exposure)] effects were assessed using clinical, pathophysiological, and molecular biomarkers.

A. castellanii and P. aeruginosa mutually exacerbate damage to corneal cells during coinfection.

IMPORTANCE: At the National Cheng Kung University Hospital, numerous cases of amoebic keratitis had been identified with concurrent bacterial infections. Among these bacterial coinfections, Pseudomonas aeruginosa accounted for 50% of the reported cases. However, the impact of pathogenic bacteria on amoeba-induced corneal damage remains unclear. In our study, we successfully demonstrated that P. aeruginosa accumulated on the Acanthamoeba castellanii surface and caused more severe corneal damage. We also indicated that the exposure of P. aeruginosa to amoeba-soluble antigens enhanced its adhesion ability, promoted biofilm formation, and led to more severe corneal cell damage. These findings significantly contributed to our understanding of the risk associated with P. aeruginosa coinfection in the progression of amoeba keratitis.

Biomimetic Corneal Stroma for Scarless Corneal Wound Healing via Structural Restoration and Microenvironment Modulation.

Corneal injury-induced stromal scarring causes the most common subtype of corneal blindness, and there is an unmet need to promote scarless corneal wound healing. Herein, a biomimetic corneal stroma with immunomodulatory properties is bioengineered for scarless corneal defect repair. First, a fully defined serum-free system is established to derive stromal keratocytes (hAESC-SKs) from a current Good Manufacturing Practice (cGMP)-grade human amniotic epithelial stem cells (hAESCs), and RNA-seq is used to validate the phenotypic transition. Moreover, hAESC-SKs are shown to possess robust immunomodulatory properties in addition to the keratocyte phenotype. Inspired by the corneal stromal extracellular matrix (ECM), a photocurable gelatin-based hydrogel is fabricated to serve as a scaffold for hAESC-SKs for bioengineering of a biomimetic corneal stroma. The rabbit corneal defect model is used to confirm that this biomimetic corneal stroma rapidly restores the corneal structure, and effectively reshapes the tissue microenvironment via proteoglycan secretion to promote transparency and inhibition of the inflammatory cascade to alleviate fibrosis, which synergistically reduces scar formation by ≈75% in addition to promoting wound healing. Overall, the strategy proposed here provides a promising solution for scarless corneal defect repair.

Endogenous TSG-6 modulates corneal inflammation following chemical injury.

PURPOSE: Tumor necrosis factor (TNF)-stimulated gene-6 (TSG-6) is upregulated in various pathophysiological contexts, where it has a diverse repertoire of immunoregulatory functions. Herein, we investigated the expression and function of TSG-6 during corneal homeostasis and after injury. METHODS: Human corneas, eyeballs from BALB/c (TSG-6+/+), TSG-6+/- and TSG-6-/- mice, human immortalized corneal epithelial cells and murine corneal epithelial progenitor cells were prepared for immunostaining and real time PCR analysis of endogenous expression of TSG-6. Mice were subjected to unilateral corneal debridement or alkali burn (AB) injuries and wound healing assessed over time using fluorescein stain, in vivo confocal microscopy and histology. RESULTS: TSG-6 is endogenously expressed in the human and mouse cornea and established corneal epithelial cell lines and is upregulated after injury. A loss of TSG-6 has no structural and functional effect in the cornea during homeostasis. No differences were noted in the rate of corneal epithelial wound closure between BALB/c, TSG-6+/- and TSG-6-/- mice. TSG-6-/- mice presented decreased inflammatory response within the first 24 h of injury and accelerated corneal wound healing following AB when compared to control mice. CONCLUSION: TSG-6 is endogenously expressed in the cornea and upregulated after injury where it propagates the inflammatory response following chemical injury.

The Anti-Inflammatory Effect of Multi-Wavelength Light-Emitting Diode Irradiation Attenuates Dry Eye Symptoms in a Scopolamine-Induced Mouse Model of Dry Eye.

Dry eye disease is a common condition in patients of all ages, causing discomfort and potential visual problems. Current treatments, including artificial tears and anti-inflammatory drugs, have certain limitations, encouraging research into alternative therapies. We investigated the therapeutic potential of multi-wavelength light-emitting diode (LED) irradiation of mice with dry eye. First, we showed that multi-wavelength LED irradiation was non-toxic to human corneal epithelial cells and improved cell viability. We then used a scopolamine-induced mouse model of dry eye to assess the effects of multi-wavelength LED irradiation on various clinical parameters. This treatment increased the tear volume and reduced corneal irregularity, thus improving dry eye. Histological analysis revealed that multi-wavelength LED irradiation protected against corneal epithelial damage and the associated reduction in epithelial thickness and would thus improve the corneal health of dry eye patients. Multi-wavelength LED irradiation significantly reduced the corneal levels of pro-inflammatory cytokines IL-6, IL-1ß, and TNF-α; the treatment was thus anti-inflammatory. Our results suggest that multi-wavelength LED irradiation may serve as a safe and effective treatment for dry eye, alleviating symptoms, reducing inflammation, and promoting corneal health.

NBL1 Reduces Corneal Fibrosis and Scar Formation after Wounding.

Corneal scarring is a leading cause of blindness. Currently, there is no treatment to prevent and/or reduce corneal scar formation under pathological conditions. Our previous data showed that the NBL1 protein, also termed the DAN Family BMP (Bone morphogenetic protein) Antagonist, was highly expressed in corneal stromal cells upon wounding. Here, we examined the function of NBL1 in corneal wound healing. Mouse corneas were mechanically wounded, followed by a 2-week treatment using NBL1. Wounded corneas treated with vehicle or an Fc tag served as controls. Compared with the controls, NBL1 treatment facilitated wound re-epithelialization, partially restored the stromal thickness, and significantly reduced corneal scar formation. NBL1 treatment did not decrease immune cell infiltration, indicating that the anti-scarring effect was not dependent on immune suppression. We further examined the anti-fibrotic effect of NBL1 on human corneas. Pairs of human corneas were induced to form myofibroblasts (a key player in fibrosis and scarring) upon wounding and incubation in a medium containing TGF-ß1. The OS corneas were treated with Fc as a control, and the OD corneas were treated with NBL1. Compared with the control, human corneas treated with NBL1 had significantly fewer myofibroblasts, which was consistent with these mouse data. A further study revealed that NBL1 treatment inhibited BMP canonical (phospho-Smad1/5) and no-canonical (phospho-p38) pathways in human corneas. Data show that NBL1 reduced corneal fibrosis and scar formation in mice and cultured human corneas. The underlying molecular mechanism is not certain because both anti-fibrotic Smad1/5 and pro-fibrotic p38 pathways were inhibited upon NBL1 treatment. Whether the p38 pathway dominates the Smad1/5 pathway during corneal fibrosis, leading to the anti-fibrotic effect of NBL1, needs further investigation.