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OBJECTIVE: Ocular chemical burns are a leading cause of blindness. The cornea is injured by alkali-induced oxidative disturbances and an inflammatory response. The aim of this study was to evaluate the protective effects of aloin, an antioxidant, and anti-inflammatory compound, on corneal alkali burn. MATERIALS AND METHODS: Mice eyes were injured by NaOH and subsequently treated with aloin eye drop and intraperitoneal injection. Pathological characteristics of the eyes were examined, and corneal samples were collected for further analysis. RESULTS: Aloin diminished neutrophil infiltration and the production of proinflammatory cytokines. Aloin also attenuated apoptosis in human corneal epithelial cells (HCEs) by reducing oxidative stress through the activation of the Nrf2 pathway. Additionally, aloin suppressed the formation of neutrophil extracellular traps (NETs) and inhibited their deposition on the cornea. Moreover, aloin mitigated alkali-induced apoptosis in HCEs caused by NETs. CONCLUSIONS: These findings suggest that aloin has potential as an antioxidant and anti-inflammatory compound for treating corneal alkali burn by inhibiting NETs formation and promoting Nrf2.
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Our objective was to determine the effect of a semi-synthetic sodium alginate hydrogel and its combination with platelet-rich plasma (PRP) on histopathological, biochemical, clinical, and anterior segment optical coherence tomography (AS-OCT) data. Alkali chemical burn of the cornea was induced. Injured rats were randomly divided into five equal groups and topically treated with phosphate-buffered saline (sham), platelet-rich plasma (PRP), 0.5% sodium citrate, a semi-synthetic sodium alginate hydrogel, or a combination of PRP and hydrogel (combined group) three times daily. The degree of corneal opacity (CO), corneal epithelial staining (CES), percentage of corneal epithelial defects (CEDP), degree of ciliary hyperemia (CH), neovascularization size (NVS), and extent of neovascularization (NVE) were evaluated. AS-OCT was performed at nine days, and then rats were sacrificed. Histological examination and enzyme-linked immunosorbent assays were performed to detect the concentrations of IL-1ß and MMP-9 in the cornea. There were no significant differences between the groups regarding CEDP, CO, CES, CH, NVS, or NVE on the first day after corneal alkali burn injury (p > 0,05). At the last examination, CO was significantly lower in the PRP group than in the sham group (p = 0,044), while the CO concentrations were similar in terms of NVS (p > 0,05). Similarly, in terms of tissue MMP-9 levels, there were no significant differences between groups (p > 0,05). However, there was a significant difference in tissue IL-1ß levels between the groups (p < 0,001). In the PRP and combined groups, the level of IL-1ß was significantly lower than that in the sham group (p = 0,043 and p = 0,036, respectively). There was a significant difference in epithelial necrosis between the PRP, and it was the lowest in the combined group (p = 0,003). Epithelial thickness was highest in the combined group (p = 0,002). CEDP was significantly different at the last visit between the groups (p = 0.042). The fastest epithelial closing rate was observed for the combined group (p = 0,026). There was a significant negative correlation between tissue MMP-9 levels and corneal solidity and between tissue MMP-9 levels and the corneal area according to the AS-OCT measurements (p = 0,012 and p = 0,027, respectively). When used alone, topical hydrogel application did not significantly enhance the healing of corneal wounds. However, when combined with PRP, it leads to an increased rate of epithelial closure and neovascularization. This combination did not exacerbate inflammation or corneal opacity compared to PRP alone. The anticoagulant citrate solution in the PRP tube did not prove effective. The synergistic use of PRP and hydrogel could enhance epithelial thickness and reduce epithelial necrosis. The use of new parameters for corneal wound healing assessment was facilitated through AS-OCT image processing.
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Alginatos , Quemaduras Químicas , Modelos Animales de Enfermedad , Quemaduras Oculares , Hidrogeles , Plasma Rico en Plaquetas , Tomografía de Coherencia Óptica , Cicatrización de Heridas , Animales , Quemaduras Químicas/tratamiento farmacológico , Quemaduras Oculares/inducido químicamente , Ratas , Cicatrización de Heridas/efectos de los fármacos , Cicatrización de Heridas/fisiología , Masculino , Metaloproteinasa 9 de la Matriz/metabolismo , Ratas Sprague-Dawley , Ensayo de Inmunoadsorción Enzimática , Interleucina-1beta/metabolismo , Lesiones de la Cornea/inducido químicamente , Segmento Anterior del Ojo/diagnóstico por imagenRESUMEN
Background: Corneal neovascularization (CNV) is a sight-threatening condition that necessitates epigenetic control. The role of lysine-specific demethylase 1 (LSD1) in CNV remains unclear, despite its established significance in tumor angiogenesis regulation. Methods: An alkali burn-induced CNV mouse model was used in vivo. The effects of LSD1 inhibitor tranylcypromine hydrochloride (TCP) were examined through slit lamp, histological staining, and immunofluorescence. The expression of malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione (GSH) levels were assessed in corneal tissues. Oxidative stress and ferrous ion expression during CNV were determined using 4-HNE, GPX4, and FerroOrange staining. In vitro, a hypoxia-reoxygenation (H/R) model was established using human umbilical vein endothelial cells (HUVECs) to study LSD1 or hypoxia-inducible factor (HIF-1α) knockdown and lentiviral overexpression of HIF-1α. The effects on HUVECs migration, invasion, and angiogenesis were evaluated through cell scratching assay, transwell migration assay and tube formation assay. The role of ferroptosis was investigated using ROS staining, FerroOrange staining, and key ferroptosis proteins. Further, The JAK2/STAT3 pathway's involvement in CNV regulation was explored through in vivo experiments with subconjunctival injection of AG490. Results: The results showed a substantial correlation between corneal damage and LSD1 levels. In addition, HIF-1α expression was also elevated after alkali burns, and subconjunctival injection of TCP reduced corneal inflammation and neovascularization. Corneal alkali burns increased ROS levels and reduced antioxidative stress indicators, accompanied by elevated ferrous ion levels, which were reversed by TCP injection. In vitro, TCP or siRNAs inhibited H/R-induced ferroptosis and angiogenesis in HUVECs by affecting specific protein expressions and MDA, SOD, and GSH levels. HIF-1α levels, associated with ROS production, ferroptosis, and angiogenesis, increased during H/R, but were reversed by TCP or siRNA administration. HIF-1α overexpression counteracted the effects of LSD1 inhibition. Additionally, AG490 injection effectively reduced HIF-1α and VEGFA expression in the CNV model. Discussion: These findings suggest that LSD1 inhibition via the HIF-1α-driven pathway prevents angiogenesis, oxidative stress, and ferroptosis in corneal alkali burn-induced CNV, highlighting LSD1 as a potential therapeutic target.
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The cornea serves as an essential shield that protects the underlying eye from external conditions, yet it remains highly vulnerable to injuries that could lead to blindness and scarring if not promptly and effectively treated. Excessive inflammatory response constitute the primary cause of pathological corneal injury. This study aimed to develop effective approaches for enabling the functional repair of corneal injuries by combining nanoparticles loaded with anti-inflammatory agents and an injectable oxidized dextran/gelatin/borax hydrogel. The injectability and self-healing properties of developed hydrogels based on borate ester bonds and dynamic Schiff base bonds were excellent, improving the retention of administered drugs on the ocular surface. In vitro cellular assays and in vivo animal studies collectively substantiated the proficiency of probucol nanoparticle-loaded hydrogels to readily suppress proinflammatory marker expression and to induce the upregulation of anti-inflammatory mediators, thereby supporting rapid repair of rat corneal tissue following alkali burn-induced injury. As such, probucol nanoparticle-loaded hydrogels represent a prospective avenue to developing long-acting and efficacious therapies for ophthalmic diseases.
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Quemaduras Químicas , Lesiones de la Cornea , Dextranos , Gelatina , Hidrogeles , Cicatrización de Heridas , Animales , Dextranos/química , Hidrogeles/química , Hidrogeles/farmacología , Gelatina/química , Ratas , Cicatrización de Heridas/efectos de los fármacos , Lesiones de la Cornea/tratamiento farmacológico , Quemaduras Químicas/tratamiento farmacológico , Quemaduras Químicas/patología , Álcalis/química , Oxidación-Reducción , Nanopartículas/química , Córnea/efectos de los fármacos , Córnea/metabolismo , Córnea/patología , Masculino , Quemaduras Oculares/tratamiento farmacológico , Quemaduras Oculares/inducido químicamente , Quemaduras Oculares/patología , Antiinflamatorios/farmacología , Antiinflamatorios/química , InyeccionesRESUMEN
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.
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Proteínas 14-3-3 , Quemaduras Químicas , Lesiones de la Cornea , Cicatrización de Heridas , Animales , Humanos , Masculino , Ratones , Proteínas 14-3-3/metabolismo , Proteínas 14-3-3/genética , Proteínas 14-3-3/biosíntesis , Western Blotting , Quemaduras Químicas/metabolismo , Quemaduras Químicas/patología , Quemaduras Químicas/tratamiento farmacológico , Movimiento Celular , Proliferación Celular , Células Cultivadas , Lesiones de la Cornea/metabolismo , Lesiones de la Cornea/patología , Lesiones de la Cornea/genética , Modelos Animales de Enfermedad , Epitelio Corneal/metabolismo , Epitelio Corneal/efectos de los fármacos , Epitelio Corneal/lesiones , Quemaduras Oculares/inducido químicamente , Regulación de la Expresión Génica , Homeostasis , Ratones Endogámicos C57BL , Hidróxido de Sodio , Cicatrización de Heridas/efectos de los fármacos , Cicatrización de Heridas/fisiologíaRESUMEN
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.
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Álcalis , Quemaduras Químicas , Córnea , Modelos Animales de Enfermedad , Quemaduras Oculares , Cicatrización de Heridas , Animales , Quemaduras Químicas/tratamiento farmacológico , Quemaduras Químicas/patología , Quemaduras Químicas/metabolismo , Ratas , Quemaduras Oculares/inducido químicamente , Quemaduras Oculares/tratamiento farmacológico , Quemaduras Oculares/patología , Álcalis/efectos adversos , Córnea/metabolismo , Córnea/patología , Córnea/efectos de los fármacos , Cicatrización de Heridas/efectos de los fármacos , Interleucina-1beta/metabolismo , Masculino , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Lesiones de la Cornea/tratamiento farmacológico , Lesiones de la Cornea/metabolismo , Lesiones de la Cornea/patología , Lesiones de la Cornea/inducido químicamente , Inflamación/tratamiento farmacológico , Inflamación/patología , Inflamación/metabolismo , Ratas Sprague-Dawley , Colágeno Tipo III/metabolismo , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Antiinflamatorios/farmacología , Soluciones Oftálmicas , Miofibroblastos/metabolismo , Miofibroblastos/efectos de los fármacosRESUMEN
OBJECTIVE: To investigate the effects of emodin on alkali burn-induced corneal inflammation and neovascularization. METHODS: The ability of emodin to target vascular endothelial growth factor receptor 2 (VEGFR2) was predicted by molecular docking. The effects of emodin on the invasion, migration, and proliferation of human umbilical vein endothelial cells (HUVEC) were determined by cell counting kit-8, Transwell, and tube formation assays. Analysis of apoptosis was performed by flow cytometry. CD31 levels were examined by immunofluorescence. The abundance and phosphorylation state of VEGFR2, protein kinase B (Akt), signal transducer and activator of transcription 3 (STAT3), and P38 were examined by immunoblot analysis. Corneal alkali burn was performed on 40 mice. Animals were divided randomly into two groups, and the alkali-burned eyes were then treated with drops of either 10 µM emodin or phosphate buffered saline (PBS) four times a day. Slit-lamp microscopy was used to evaluate inflammation and corneal neovascularization (CNV) in all eyes on Days 0, 7, 10, and 14. The mice were killed humanely 14 d after the alkali burn, and their corneas were removed and preserved at ï¼80 â until histological study or protein extraction. RESULTS: Molecular docking confirmed that emodin was able to target VEGFR2. The findings revealed that emodin decreased the invasion, migration, angiogenesis, and proliferation of HUVEC in a dose-dependent manner. In mice, emodin suppressed corneal inflammatory cell infiltration and inhibited the development of corneal neovascularization induced by alkali burn. Compared to those of the PBS-treated group, lower VEGFR2 expression and CD31 levels were found in the emodin-treated group. Emodin dramatically decreased the expression of VEGFR2, p-VEGFR2, p-Akt, p-STAT3, and p-P38 in VEGF-treated HUVEC. CONCLUSION: This study provides a new avenue for evaluating the molecular mechanisms underlying corneal inflammation and neovascularization. Emodin might be a promising new therapeutic option for corneal alkali burns.
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Quemaduras Químicas , Neovascularización de la Córnea , Emodina , Humanos , Ratones , Animales , Neovascularización de la Córnea/tratamiento farmacológico , Neovascularización de la Córnea/genética , Neovascularización de la Córnea/metabolismo , Factor A de Crecimiento Endotelial Vascular/genética , Factor A de Crecimiento Endotelial Vascular/metabolismo , Quemaduras Químicas/tratamiento farmacológico , Quemaduras Químicas/metabolismo , Quemaduras Químicas/patología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Receptor 2 de Factores de Crecimiento Endotelial Vascular/genética , Receptor 2 de Factores de Crecimiento Endotelial Vascular/metabolismo , Simulación del Acoplamiento Molecular , Neovascularización Patológica/tratamiento farmacológico , Neovascularización Patológica/genética , Transducción de Señal , Células Endoteliales de la Vena Umbilical Humana , Inflamación/tratamiento farmacológico , Modelos Animales de EnfermedadRESUMEN
Purpose: This study aimed to evaluate the effects of a new treatment-conditioned medium from human orbital adipose-derived stem cells (OASC-CM)-on corneal recovery after alkali burns in a rabbit model. Methods: The corneal alkali burn rabbit model was established and treated with OASC-CM, conditioned medium from human abdominal subcutaneous adipose-derived stem cells (ABASC-CM), and fresh control culture medium (con-CM) three times a day for 7 days, respectively. Subsequently, the treatment effects were evaluated and compared through clinical, histological, immunohistochemical, and cytokine evaluations. Results: Clinically, OASC-CM alleviated corneal opacity and edema and promoted recovery of corneal epithelium defect. Histologically and immunohistochemically, OASC-CM inhibited neovascularization, conjunctivalization, and immuno-inflammatory reaction, while promoting corneal regeneration and rearrangement. Increased secretion of interleukin-10 and inhibited protein levels of cluster of differentiation 45, interferon-γ, and tumor necrosis factor-α were observed in the alkali-burned cornea after OASC-CM treatment, which might be the relevant molecular mechanism. Conclusions: OASC-CM showed significant effects on the recovery of rabbit corneal alkali burns and eliminated immunological and ethical limitations, representing a new option for corneal wound treatment.
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Tejido Adiposo , Quemaduras Químicas , Modelos Animales de Enfermedad , Quemaduras Oculares , Células Madre , Animales , Conejos , Medios de Cultivo Condicionados/farmacología , Quemaduras Químicas/terapia , Quemaduras Químicas/tratamiento farmacológico , Quemaduras Oculares/inducido químicamente , Quemaduras Oculares/terapia , Quemaduras Oculares/tratamiento farmacológico , Humanos , Tejido Adiposo/citología , Álcalis , Masculino , ÓrbitaRESUMEN
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.
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Quemaduras Químicas , Enfermedades de la Córnea , Queratitis , Conejos , Animales , Quemaduras Químicas/tratamiento farmacológico , Álcalis , Hidróxido de Sodio/uso terapéutico , Córnea , Microscopía Confocal/métodos , CitocinasRESUMEN
Corneal alkali burns often occur in industrial production and daily life, combined with infection, and may cause severe eye disease. Oxidative stress and neovascularization (NV) are important factors leading to a poor prognosis. URP20 is an antimicrobial peptide that has been proven to treat bacterial keratitis in rats through antibacterial and anti-NV effects. Therefore, in this study, the protective effect and influence mechanism of URP20 were explored in a rat model of alkali burn together with pathogenic bacteria (Staphylococcus aureus and Escherichia coli) infection. In addition, human umbilical vein endothelial cells (HUVECs) and human corneal epithelial cells (HCECs) were selected to verify the effects of URP20 on vascularization and oxidative stress. The results showed that URP20 treatment could protect corneal tissue, reduce corneal turbidity, and reduce the NV pathological score. Furthermore, URP20 significantly inhibited the expression of the vascularization marker proteins VEGFR2 and CD31. URP20 also reduced the migration ability of HUVECs. In terms of oxidative stress, URP20 significantly upregulated SOD and GSH contents in corneal tissue and HCECs (treated with 200 µM H2O2) and promoted the expression of the antioxidant protein Nrf2/HO-1. At the same time, MDA and ROS levels were also inhibited. In conclusion, URP20 could improve corneal injury combined with bacterial infection in rats caused by alkali burns through antibacterial, anti-NV, and antioxidant activities.
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Infecciones Bacterianas , Quemaduras Químicas , Lesiones de la Cornea , Neovascularización de la Córnea , Quemaduras Oculares , Ratas , Humanos , Animales , Quemaduras Químicas/complicaciones , Quemaduras Químicas/tratamiento farmacológico , Quemaduras Químicas/metabolismo , Neovascularización de la Córnea/metabolismo , Antioxidantes/farmacología , Antioxidantes/uso terapéutico , Peróxido de Hidrógeno/farmacología , Neovascularización Patológica/metabolismo , Lesiones de la Cornea/tratamiento farmacológico , Células Endoteliales de la Vena Umbilical Humana , Antibacterianos/uso terapéutico , Antibacterianos/farmacología , Quemaduras Oculares/inducido químicamente , Quemaduras Oculares/tratamiento farmacológico , Quemaduras Oculares/patología , Modelos Animales de Enfermedad , Álcalis/toxicidadRESUMEN
Ocular alkali burns recruit neutrophils and triggers neutrophil extracellular trap (NET)-neovascularization cascade effects that limit ocular surface reconstruction and functional repair. However, effective inhibition of the release of neutrophil extracellular traps after a corneal chemical injury, coordination of intrinsic immunity with corneal repair, and exploration of more effective and non-invasive drug-delivery modes are still urgently needed. Using an in vitro coculture system, we found that an alkaline environment stimulates neutrophils to release NETs, which can be regulated by deoxyribonuclease I (DNase I). Inspired by this, we loaded DNase I, which effectively regulates NETs, onto chitosan nanoparticles and combined them with silk fibroin to construct a composite hydrogel that can sustainably regulate NETs. The hydrogel reduced neutrophil extracellular trap production by 50% and neovascularization by approximately 70% through sustained DNase I release after a corneal alkali burn. The complex hydrogel promotes ocular surface reconstruction by modulating the intrinsic immune-cascade neovascularization effect, providing a new research basis for the construction of nanobiomaterials that modulate pathological neovascularization.
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This study aims to investigate the efficiency and the underlying mechanism of myeloid-derived suppressor cells (MDSCs) in corneal alkali burns (CAB). In the study, CD11b+ Gr-1+ cells from C57BL/6J mice bone marrow were cultured and induced. Cell activity and immunoregulatory function were assessed by flow cytometry in vitro. The optimal strategy of MDSCs therapy was assessed by slit-lamp microscopy, and flow cytometry in vivo. The therapeutic effects of MDSCs and the critical signaling pathway were investigated by hematoxylin-eosin (HE) staining, slit-lamp microscopy, flow cytometry, and immunofluorescence. The expression level of the NLRP3 inflammasome pathway was examined. The crucial biochemical parameters of MDSCs were examined by RNA-seq and qPCR to screen out the key regulators. The mechanism of MDSCs' therapeutic effects was explored using MDSCs with IL-10 knockout/rescue by slit-lamp microscopy, HE staining, and qPCR evaluation. The cell frequencies of macrophages and neutrophils in the cornea were examined by flow cytometry in vivo. The results demonstrated that the induced MDSCs meet the standard of phenotypic and functional characteristics. The treatment of 5â¯×â¯105 MDSCs conjunctival injection on alternate days significantly ameliorated the disease development, downregulated the NLRP3 inflammasome pathway, and decreased the cell frequencies of macrophages and neutrophils in vivo significantly. IL-10 was screened out to be the critical factor for MDSCs therapy. The therapeutic effects of MDSCs were impaired largely by IL-10 knock-out and saved by the IL-10 supplement. In conclusion, MDSCs therapy is a promising therapeutic solution for CAB. MDSCs fulfilled immunoregulatory roles for CAB by IL-10-dependent anti-inflammatory properties.
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Quemaduras Químicas , Células Supresoras de Origen Mieloide , Animales , Ratones , Células Supresoras de Origen Mieloide/metabolismo , Interleucina-10 , Inflamasomas/metabolismo , Quemaduras Químicas/terapia , Quemaduras Químicas/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Ratones Endogámicos C57BLRESUMEN
Corneal alkali burn is one of the most devastating ophthalmic emergencies correlated with remarkable morbidity resulting in severe visual impairment. Appropriate intervention in the acute phase determines the eventual outcome for later corneal restoration treatment. Since the epithelium plays an essential role in inhibiting inflammation and promoting tissue repair, sustained anti-matrix metalloproteinases (MMPs) and pro-epithelialization are the prior remedies during the first week. In this study, a drug-loaded collagen membrane (Dox-HCM/Col) that could be sutured to overlay the burned cornea was developed to accelerate the early reconstruction. Doxycycline (Dox), a specific inhibitor of MMPs, was encapsulated in collagen membrane (Col) through hydroxypropyl chitosan microspheres (HCM) to develop Dox-HCM/Col, affording a preferable pro-epithelialization microenvironment and an in-situ controlled release. Results showed that loading HCM into Col prolonged the release time to 7 days, and Dox-HCM/Col could significantly suppress the expression of MMP-9 and -13 in vitro and in vivo. Furthermore, the membrane accelerated the corneal complete re-epithelialization and promoted early reconstruction within the first week. Overall, Dox-HCM/Col was a promising biomaterial membrane for treating alkali-burned cornea in the early stage, and our attempt may provide a clinically feasible method for the ocular surface reconstruction.
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Quitosano , Lesiones de la Cornea , Quemaduras Oculares , Humanos , Doxiciclina/farmacología , Quitosano/metabolismo , Álcalis/metabolismo , Microesferas , Colágeno/metabolismo , Lesiones de la Cornea/tratamiento farmacológico , Lesiones de la Cornea/metabolismo , Córnea/metabolismo , Quemaduras Oculares/inducido químicamente , Quemaduras Oculares/tratamiento farmacológico , Quemaduras Oculares/metabolismo , Trastornos de la Visión/metabolismoRESUMEN
Limbal stem cell deficiency (LSCD) is a debilitating ocular surface disease that eventuates from a depleted or dysfunctional limbal epithelial stem cell (LESC) pool, resulting in corneal epithelial failure and blindness. The leading cause of LSCD is a chemical burn, with alkali substances being the most common inciting agents. Characteristic features of alkali-induced LSCD include corneal conjunctivalization, inflammation, neovascularization and fibrosis. Over the past decades, animal models of corneal alkali burn and alkali-induced LSCD have been instrumental in improving our understanding of the pathophysiological mechanisms responsible for disease development. Through these paradigms, important insights have been gained with regards to signaling pathways that drive inflammation, neovascularization and fibrosis, including NF-κB, ERK, p38 MAPK, JNK, STAT3, PI3K/AKT, mTOR and WNT/ß-catenin cascades. Nonetheless, the molecular and cellular events that underpin re-epithelialization and those that govern long-term epithelial behavior are poorly understood. This review provides an overview of the current mechanistic insights into the pathophysiology of alkali-induced LSCD. Moreover, we highlight limitations regarding existing animal models and knowledge gaps which, if addressed, would facilitate development of more efficacious therapeutic strategies for patients with alkali-induced LSCD.
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Deficiencia de Células Madre Limbares , Limbo de la Córnea , Animales , Células Madre Limbares , Fosfatidilinositol 3-Quinasas/metabolismo , Células Madre/metabolismo , Inflamación/patología , Modelos AnimalesRESUMEN
The cornea serves as an important barrier structure to the eyeball and is vulnerable to injuries, which may lead to scarring and blindness if not treated promptly. To explore an effective treatment that could achieve multi-dimensional repair of the injured cornea, the study herein innovatively combined modified mRNA (modRNA) technologies with adipose-derived mesenchymal stem cells (ADSCs) therapy, and applied IGF-1 modRNA (modIGF1)-engineered ADSCs (ADSCmodIGF1) to alkali-burned corneas in mice. The therapeutic results showed that ADSCmodIGF1 treatment could achieve the most extensive recovery of corneal morphology and function when compared not only with simple ADSCs but also IGF-1 protein eyedrops, which was reflected by the healing of corneal epithelium and limbus, the inhibition of corneal stromal fibrosis, angiogenesis and lymphangiogenesis, and also the repair of corneal nerves. In vitro experiments further proved that ADSCmodIGF1 could more significantly promote the activity of trigeminal ganglion cells and maintain the stemness of limbal stem cells than simple ADSCs, which were also essential for reconstructing corneal homeostasis. Through a combinatorial treatment regimen of cell-based therapy with mRNA technology, this study highlighted comprehensive repair in the damaged cornea and showed the outstanding application prospect in the treatment of corneal injury.
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Enfermedades de la Córnea , Lesiones de la Cornea , Células Madre Mesenquimatosas , Ratones , Animales , Factor I del Crecimiento Similar a la Insulina/genética , Factor I del Crecimiento Similar a la Insulina/farmacología , Tejido Adiposo , Córnea , Lesiones de la Cornea/genética , Lesiones de la Cornea/terapia , Lesiones de la Cornea/metabolismo , Células Madre Mesenquimatosas/metabolismo , Cicatrización de Heridas/genéticaRESUMEN
Zinn's zonule is a fragile and thin tissue, and little is known about its pathogenesis. The aim of this study was to develop an experimental setup for a comprehensive analysis of Zinn's zonule. Rats were divided into two groups: a control group (n = 4) and an alkali injury group (n = 4). Seven days after injury, the eyes were enucleated, the anterior eye was dissected and embedded in gelatin, and macroscopic observations were made. The gelatin specimens were then embedded in paraffin and observed in detail by low-vacuum scanning electron microscopy, immunofluorescence, and quantitative reverse transcription polymerase chain reaction (RT-qPCR). The results show qualitative changes in Zinn's zonules in both macroscopic and microscopic observations. In addition, macrophage infiltration and increased matrix metalloproteinase 2 (MMP2) expression were observed in the injured group, consistent with the RT-qPCR results. The experimental system in this study allowed us to capture the morphological and molecular biological changes of Zinn's zonule and to gain insight into its pathogenesis. In conclusion, this study presents a new experimental setup for the comprehensive analysis of the rat Zinn's zonule. The results suggest that this system can be used in the future to study and analyze a variety of paraffin-embedded tissues and specimens.
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Extracción de Catarata , Metaloproteinasa 2 de la Matriz , Animales , Ratas , Metaloproteinasa 2 de la Matriz/genética , Gelatina , OjoRESUMEN
Corneal alkali burn (AB) is a blindness-causing ocular trauma commonly seen in clinics. An excessive inflammatory reaction and stromal collagen degradation contribute to corneal pathological damage. Luteolin (LUT) has been studied for its anti-inflammatory effects. In this study, the effect of LUT on cornea stromal collagen degradation and inflammatory damage in rats with corneal alkali burn was investigated. After corneal alkali burn, rats were randomly assigned to the AB group and AB + LUT group and received an injection of saline and LUT (200 mg/kg) once daily. Subsequently, corneal opacity, epithelial defects, inflammation and neovascularization (NV) were observed and recorded on Days 1, 2, 3, 7 and 14 post-injury. The concentration of LUT in ocular surface tissues and anterior chamber, as well as the levels of collagen degradation, inflammatory cytokines, matrix metalloproteinases (MMPs) and their activity in the cornea were detected. Human corneal fibroblasts (HCFs) were co-cultured with interleukin (IL)-1ß and LUT. Cell proliferation and apoptosis were assessed by CCK-8 assay and flow cytometry respectively. Measurement of hydroxyproline (HYP) in culture supernatants was used to quantify the amount of collagen degradation. Plasmin activity was also assessed. ELISA or real-time PCR was used to detect the production of matrix metalloproteinases (MMPs), IL-8, IL-6 and monocyte chemotactic protein (MCP)-1. Furthermore, the immunoblot method was used to assess the phosphorylation of mitogen-activated protein kinases (MAPKs), transforming growth factor-ß-activated kinase (TAK)-1, activator protein-1 (AP-1) and inhibitory protein IκB-α. At last, immunofluorescence staining helped to develop nuclear factor (NF)-κB. LUT was detectable in ocular tissues and anterior chamber after intraperitoneal injection. An intraperitoneal injection of LUT ameliorated alkali burn-elicited corneal opacity, corneal epithelial defects, collagen degradation, NV, and the infiltration of inflammatory cells. The mRNA expressions of IL-1ß, IL-6, MCP-1, vascular endothelial growth factor (VEGF)-A, and MMPs in corneal tissue were downregulated by LUT intervention. And its administration reduced the protein levels of IL-1ß, collagenases, and MMP activity. Furthermore, in vitro study showed that LUT inhibited IL-1ß-induced type I collagen degradation and the release of inflammatory cytokines and chemokines by corneal stromal fibroblasts. LUT also inhibited the IL-1ß-induced activation of TAK-1, mitogen-activated protein kinase (MAPK), c-Jun, and NF-κB signaling pathways in these cells. Our results demonstrate that LUT inhibited alkali burn-stimulated collagen breakdown and corneal inflammation, most likely by attenuating the IL-1ß signaling pathway. LUT may therefore prove to be of clinical value for treating corneal alkali burns.
Asunto(s)
Quemaduras Químicas , Opacidad de la Córnea , Ratas , Humanos , Animales , Quemaduras Químicas/complicaciones , Quemaduras Químicas/tratamiento farmacológico , Quemaduras Químicas/metabolismo , Factor A de Crecimiento Endotelial Vascular/metabolismo , Luteolina/farmacología , Luteolina/uso terapéutico , Álcalis/toxicidad , Interleucina-6/metabolismo , Córnea/metabolismo , Citocinas/metabolismo , Neovascularización Patológica/metabolismo , Colágeno Tipo I/metabolismo , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Opacidad de la Córnea/metabolismo , Inflamación/metabolismo , Metaloproteinasas de la Matriz/metabolismoRESUMEN
Alkali burns are potentially blinding corneal injuries. Due to the lack of available effective therapies, the prognosis is poor. Thus, effective treatment methods for corneal alkali burns are urgently needed. Codelivery nanoparticles (NPs) with characteristics such as high bioavailability and few side effects have been considered effective therapeutic agents for ocular diseases. In this study, we designed a new combination therapy using liposomes and trimethyl chitosan (TMC) for the codelivery of insulin (INS) and vascular endothelial growth factor small interfering RNA (siVEGF) to treat alkali-burned corneas. We describe the preparation and characterization of siVEGF-TMC-INS-liposome (siVEGF-TIL), drug release characteristics, intraocular tracing, pharmacodynamics, and biosafety. We found that siVEGF-TIL could inhibit oxidative stress, inflammation, and the expression of VEGF in vitro and effectively maintained corneal transparency, accelerated epithelialization, and inhibited corneal neovascularization (CNV) in vivo. Morever, we found that the therapeutic mechanism of siVEGF-TIL is possibly relevant to the inhibition of the ferroptosis signaling pathway by metabolomic analysis. In general, siVEGF-TIL NPs could be a safe and effective therapy for corneal alkali burn.
RESUMEN
FROUNT is an intracellular protein that promotes pseudopodia formation by binding to the chemokine receptors CCR2 and CCR5 on macrophages. Recently, disulfiram (DSF), a drug treatment for alcoholism, was found to have FROUNT inhibitory activity. In this study, we investigated the effect of DSF eye drops in a rat corneal alkali burn model. After alkali burn, 0.5% DSF eye drops (DSF group) and vehicle eye drops (Vehicle group) were administered twice daily. Immunohistochemical observations and real-time reverse transcription-polymerase chain reaction (RT-PCR) analyses were performed at 6 h and 1, 4, and 7 days after alkali burn. Results showed a significant decrease in macrophage accumulation in the cornea in the DSF group, but no difference in neutrophils. RT-PCR showed decreased expression of macrophage-associated cytokines in the DSF group. Corneal scarring and neovascularization were also suppressed in the DSF group. Low-vacuum scanning electron microscopy imaging showed that macrophage length was significantly shorter in the DSF group, reflecting the reduced extension of pseudopodia. These results suggest that DSF inhibited macrophage infiltration by suppressing macrophage pseudopodia formation.