Corneal Alkali Burn Research Articles

S100A4 Silencing Facilitates Corneal Wound Healing After Alkali Burns by Promoting Autophagy via Blocking the PI3K/Akt/mTOR Signaling Pathway.

PurposeThis study investigated the role of S100 calcium binding protein A4 (S100A4) in corneal wound healing and the underlying mechanism of the S100A4-mediated PI3K/Akt/mammalian target of rapamycin (mTOR) pathway.MethodsThe rabbit corneal alkali burn model was established in vivo. S100A4 expression, wound healing, inflammation, and autophagy in rabbit cornea after alkali burn were detected. The NaOH-treated rabbit corneal stromal cells (rCSCs) were transfected with overexpressed S100A4 or silencing S100A4 to examine the effect of S100A4 on corneal wound healing in vitro. The effect of S100A4 on cell viability, proliferation, migration, invasion, fibrosis, and autophagy of rCSCs after alkali burn was analyzed. Then the functional rescue experiments were carried out. The PI3K inhibitor, LY294002, was used to elucidate the PI3K/Akt/mTOR signaling pathway in rCSCs.ResultsS100A4 silencing promoted rabbit corneal wound healing by inhibiting fibrosis and inflammation and promoting autophagy in alkali-burned cornea, corresponding to increased levels of LC3, Beclin 1, and Atg4B but lowered α-smooth muscle actin, TNF-ɑ, and p62 levels. Moreover, silencing S100A4 inhibited proliferation, migration, invasion, and fibrosis of NaOH-treated rCSCs and promoted the differentiation of rCSCs into corneal cells and the autophagy of damaged rCSCs. The inhibitory role of S100A4 in wound healing was achieved via activation of the PI3K/Akt/mTOR pathway.ConclusionsS100A4 silencing confers a promising effect on wound healing of alkali-burned cornea by blocking the PI3K/Akt/mTOR pathway, supporting the advancement of corneal gene therapies for wound healing.

Comparison between the Effect of Hyaluronic Acid and Mesenchymal Stem Cell Therapy on Cornea of Adult Albino Rats after Exposure to Alkali Burn (Light and Electron Microscopic Study)

Background: Corneal epithelial injuries are common in both physician and veterinary ophthalmology that often causes extensive damage and results in permanent visual impairment. Aim of work: to evaluate the beneficial effect of hyaluronic acid and MSCs in treatment of experimental alkali corneal burn. Subjects and methods: thirty adult albino rats of both sexes. Ten rats were used to harvest BM-MSCs while the others were divided into four groups. Group I was the control group. Group II with unilateral alkali-burnt cornea these rats were sacrificed after 1 day and the other sacrificed after 2 weeks. Group III were rats with unilateral hyaluronic acid-treated alkali-burnt corneas for 2 weeks. Group IV were rats with MSCs treated alkali-burnt corneas. Immune histochemical staining for CD44 and vimentin was performed. The corneas were examined using light microscopic, transmission electron microscopic and morphometric studies. Results: Corneal alkali burn resulted in desquamation of corneal epithelium in group II. The epithelial cell layers had vacuolated cytoplasm with pyknotic nuclei. The stroma contained irregularly arranged collagen fibers with wide spacing and congested blood vessels with cellular infiltration. Groups III and IV showed improvement of the histological and electron microscopic changes described in group II. Conclusions: The use of MSCs in the acute stage of corneal chemical trauma was associated with faster recovery of the wounded cornea when compared with the effect of hyaluronic acid sodium salt.

Neutrophil extracellular traps promote corneal neovascularization-induced by alkali burn

ObjectivesTo investigate the effects of neutrophil extracellular traps (NETs) on angiogenesis in vitro and in vivo and the regulatory role of mammalian target of rapamycin (mTOR) activity in it. MethodsThe regulatory role of mTOR in NETs formation was explored. In vitro, human neutrophils were pretreated with rapamycin. NETs formation was measured using immunofluorescence staining of NETs markers, SYTOX Green and PicoGreen after NaOH stimulation. In vivo, mice were treated with rapamycin, and NETs formation in cornea was measured using immunofluorescence staining 7 days after alkali burn. Then, the effects of NETs on angiogenesis were investigated. In vitro, human neutrophils were treated with DNase I or rapamycin. NETs were isolated after NaOH stimulation and the isolated NETs were co-culture with human umbilical vein endothelial cells (HUVECs). HUVECs migration, proliferation, and inflammatory activation were measured. In vivo, mice were injected subconjunctivally with supernatant containing NETs. Corneal neovascularization was visualized by immunofluorescence staining. ResultsNETs structures can be observed in NaOH-stimulated neutrophils and alkali-burned mouse cornea compared with normal group. Treated with rapamycin enhanced NETs formation in response to NaOH management compared with DMSO control in vitro and in vivo. NETs increased the migration, proliferation and inflammatory activation of HUVECs, and subconjunctival injection of NETs promoted inflammatory and angiogenic response in corneal alkali burn model. ConclusionsNETs formation can be triggered by NaOH stimulation. mTOR activity has a negative regulatory effect on NETs formation. NETs promoted angiogenic responses and inflammatory activation of HUVECs and increased corneal neovascularization and inflammatory response.

Hydrogen promotes the activation of Cu, Zn superoxide dismutase in a rat corneal alkali-burn model.

To investigate the effects of hydrogen (H2) on Cu, Zn superoxide dismutase (SOD1) activation in a rat model of corneal alkali burn. In each rat, one cornea was subjected to alkali exposure. Physiological saline (saline group) or H2-dissolved saline (H2 group) was instilled continuously on the cornea for 5min before and after alkali exposure. Inflammatory cells, neovascularization, and cytoplasmic SOD1 levels were evaluated immunohistochemically in enucleated eyes from both groups. Three-dimensional ultrastructural tissue changes in the eyes were analyzed using low-vacuum scanning electron microscopy. The numbers of both inflammatory and vascular endothelial cells were significantly reduced in the corneas of the H2 group (P<0.01). Furthermore, H2 treatment increased both cytoplasmic SOD1 levels (P<0.01) and activity in corneal epithelial cells (P<0.01). Notably, the SOD1 activity level in the H2 group was approximately 2.5-fold greater than that in the saline group. H2 treatment suppresses inflammation and neovascularization in the injured cornea and indirectly suppresses oxidative insult to the cornea by upregulating the SOD1 enzyme protein level and activity.

Peroxisome Proliferator-Activated Receptor Beta/Delta Agonist Suppresses Inflammation and Promotes Neovascularization.

The effects of peroxisome proliferator-activated receptor (PPAR)β/δ ophthalmic solution were investigated in a rat corneal alkali burn model. After alkali injury, GW501516 (PPARβ/δ agonist) or vehicle ophthalmic solution was topically instilled onto the rat’s cornea twice a day until day 7. Pathological findings were evaluated, and real-time reverse transcription polymerase chain reaction was performed. GW501516 strongly suppressed infiltration of neutrophils and pan-macrophages, and reduced the mRNA expression of interleukin-6, interleukin-1β, tumor necrosis factor alpha, and nuclear factor-kappa B. On the other hand, GW501516 promoted infiltration of M2 macrophages, infiltration of vascular endothelial cells associated with neovascularization in the wounded area, and expression of vascular endothelial growth factor A mRNA. However, 7-day administration of GW501516 did not promote neovascularization in uninjured normal corneas. Thus, the PPARβ/δ ligand suppressed inflammation and promoted neovascularization in the corneal wound healing process. These results will help to elucidate the role of PPARβ/δ in the field of ophthalmology.

Combination of Peroxisome Proliferator-Activated Receptor (PPAR) Alpha and Gamma Agonists Prevents Corneal Inflammation and Neovascularization in a Rat Alkali Burn Model

Peroxisome proliferator-activated receptor alpha (PPARα) and gamma (PPARγ) agonists have anti-inflammatory and anti-neovascularization effects, but few reports have tested the combination of PPARα and PPARγ agonists. In this study, we investigated the therapeutic effects of ophthalmic solutions of agonists of PPARα, PPARγ, and the combination in a rat corneal alkali burn model. After alkali injury, an ophthalmic solution of 0.05% fenofibrate (PPARα group), 0.1% pioglitazone (PPARγ group), 0.05% fenofibrate + 0.1% pioglitazone (PPARα+γ group), or vehicle (vehicle group) was topically instilled onto the rat’s cornea twice a day. After instillation, upregulation was seen of PPAR mRNA corresponding to each agonist group. Administration of agonists for PPARα, PPARγ, and PPARα+γ suppressed inflammatory cells, neovascularization, and fibrotic changes. In addition, the PPARγ agonist upregulated M2 macrophages, which contributed to wound healing, whereas the PPARα agonist suppressed immature blood vessels in the early phase. Administration of PPARα+γ agonists showed therapeutic effects in corneal wound healing, combining the characteristics of both PPARα and PPARγ agonists. The results indicate that the combination of PPARα and γ agonists may be a new therapeutic strategy.

Microporous Drug Delivery System for Sustained Anti-VEGF Delivery to the Eye.

PurposeTo describe a novel microporous drug delivery system (DDS) for sustained anti- vascular endothelial growth factor (VEGF) delivery to the eye and to evaluate its efficacy in a corneal injury model.MethodsA macro-porous DDS (1.5 × 1.5 × 4 mm) loaded with 2 mg of bevacizumab was implanted subconjunctivally in three Dutch-belted pigmented rabbits after corneal alkali injury (2N NaOH). Three rabbits received sham DDS. Animals were followed for three months and assessed in vivo and ex vivo for corneal neovascularization (NV), epithelial defect, stromal scarring, endothelial cell loss, and expression of angiogenic and inflammatory markers in the cornea and retina.ResultsAnti-VEGF DDS treatment led to complete inhibition of superior cornea NV and complete corneal re-epithelialization by day 58 whereas sham DDS resulted in severe cornea NV and persistent epithelial defect (9%∼12% of total cornea area) through the end of the study. Histologically, anti-VEGF DDS significantly reduced CD45+ and F4/80 CD11b+ cell accumulation (79%, P < 0.05) in the cornea, ameliorated tumor necrosis factor–α expression (90%, P < 0.05), reduced corneal stromal scarring and prevented corneal endothelial cell loss, as compared to sham DDS. Moreover, anti-VEGF DDS achieved retinal penetration and reduction in retinal VEGF levels at 3 months.ConclusionsUse of subconjunctival anti-VEGF DDS suppresses cornea NV, inflammation, stromal scarring, prevents endothelial cell loss, and abrogates retinal VEGF upregulation in a rabbit corneal alkali burn model. Moreover, it delivers anti-VEGF antibodies to the retina for three months. This delivery platform could enable antibody therapy of other corneal and retinal vascular pathologies.Translational RelevanceWe describe a method for sustained anti-VEGF delivery to the eye for the treatment of ocular injuries.

An Immunohistochemical Study of the Increase in Antioxidant Capacity of Corneal Epithelial Cells by Molecular Hydrogen, Leading to the Suppression of Alkali-Induced Oxidative Stress.

Corneal alkali burns are potentially blinding injuries. Alkali induces oxidative stress in corneas followed by excessive corneal inflammation, neovascularization, and untransparent scar formation. Molecular hydrogen (H2), a potent reactive oxygen species (ROS) scavenger, suppresses oxidative stress and enables corneal healing when applied on the corneal surface. The purpose of this study was to examine whether the H2 pretreatment of healthy corneas evokes a protective effect against corneal alkali-induced oxidative stress. Rabbit eyes were pretreated with a H2 solution or buffer solution, by drops onto the ocular surface, and the corneas were then burned with 0.25 M NaOH. The results obtained with immunohistochemistry and pachymetry showed that in the corneas of H2-pretreated eyes, slight oxidative stress appeared followed by an increased expression of antioxidant enzymes. When these corneas were postburned with alkali, the alkali-induced oxidative stress was suppressed. This was in contrast to postburned buffer-pretreated corneas, where the oxidative stress was strong. These corneas healed with scar formation and neovascularization, whereas corneas of H2-pretreated eyes healed with restoration of transparency in the majority of cases. Corneal neovascularization was strongly suppressed. Our results suggest that the corneal alkali-induced oxidative stress was reduced via the increased antioxidant capacity of corneal cells against reactive oxygen species (ROS). It is further suggested that the ability of H2 to induce the increase in antioxidant cell capacity is important for eye protection against various diseases or external influences associated with ROS production.

AMPK Activation by 5-Amino-4-Imidazole Carboxamide Riboside-1-β-D-Ribofuranoside Attenuates Alkali Injury-Induced Corneal Fibrosis.

PurposeIncreased TGF-β1 synthesis after corneal alkali injury is implicated in corneal fibrosis, as it promotes transdifferentiation of keratocytes into myofibroblasts. The activation of 5′-adenosine monophosphate-activated protein kinase (AMPK) by 5-amino-4-imidazole carboxamide riboside-1-β-d-ribofuranoside (AICAR) inhibits TGF-β1-induced fibrosis in other cell types. We investigated the antifibrotic effect of AICAR in corneal fibroblasts after alkali injury.MethodsMouse models of corneal alkali burn, produced by placing 2-mm-diameter filter paper soaked in 0.1-N NaOH on the right cornea for 30 seconds, were treated with the test drugs 4× daily for 21 days. The central cornea was scanned by optical coherence tomography (OCT). Corneal tissues were obtained and processed for western blotting and immunohistochemistry. For in vitro analysis, primary human corneal fibroblasts were treated directly with TGF-β1 to induce fibrosis, with or without AICAR pretreatment. Myofibroblast activation and extracellular matrix (ECM) protein synthesis were detected by western blotting, real-time PCR, and collagen gel contraction assay. Signaling proteins were analyzed by western blotting.ResultsAlkali injury induced the upregulation of TGF-β1 expression, which led to increased α-smooth muscle actin (α-SMA) and fibronectin synthesis and myofibroblast differentiation. AMPK activation by AICAR significantly suppressed TGF-β1 and ECM protein expression. The antifibrotic effect of AICAR was AMPK dependent, as treatment with the AMPK inhibitor Compound C attenuated the antifibrotic response.ConclusionsAMPK activation by AICAR suppresses the myofibroblast differentiation and ECM synthesis that occur after alkali injury in corneal fibroblasts.

Effect of nintedanib thermo-sensitive hydrogel on neovascularization in alkali burn rat model.

To investigate the effects of nintedanib thermo-sensitive hydrogel (NTH) on neovascularization and related markers in corneal alkali burns of Wistar rats. NTH was prepared by grinding, and its phase-transition temperature was determined. Thirty specific-pathogen-free Wistar rats served as a model of corneal alkali burn in the right eye were randomly divided into 3 groups (n=10, each): model group treated with 0.9% saline once a day, NTH group with 0.2% nintedanib b.i.d, and dexamethasone group with dexamethasone ointment once a day. The left eye of rats served as the controls. The corneal transparency was observed under a slit-lamp microscope, and the area of neovascularization was calculated. On day 7, the rats were sacrificed, and the cornea was removed and embedded with paraffin, then stained with hematoxylin-eosin, and the expression of vascular endothelial growth factor receptor 2 (VEGFR-2) and CD31 in the corneal tissues of each group was detected by immunofluorescence. The phase-transition temperature of nintedanib obtained by grinding was 37°C after adding artificial tears. The results of the alkali burn model indicated that the growth rate of neovascularization in the NTH group was slower than that in the model group, and the neovascularization area was significantly smaller than that in the model group (P<0.05). Moreover, CD31 and VEGFR-2 expression levels in the NTH group were significantly lower than those in the model group. NTH becomes colloidal at body temperature, which is beneficial for releasing the drug slowly and can significantly inhibit the neovascularization of corneal induced by alkali burn in rats.

Observation of Corneal Wound Healing and Angiogenesis Using Low-Vacuum Scanning Electron Microscopy.

PurposeWound healing processes in a rat corneal alkali burn model were observed using low-vacuum scanning electron microscopy (LV-SEM), a new observation method that can use paraffin sections for light microscopic immunostaining.MethodsInjured cornea was observed under immunohistochemistry, LV-SEM, and transmission electron microscopy. In LV-SEM, periodic acid-methenamine silver staining was used to observe collagen and platinum blue staining was used to observe vascular endothelial cells. Analyses of the messenger RNA expression involved in neovascularization processes after wound creation were also performed.ResultsLV-SEM depicted progression of corneal wound healing in a stereoscopic fashion. In neovascularization processes after wound creation, LV-SEM with osmification clearly demonstrated detachment of pericytes from the vascular endothelial cells, in association with up-regulation of angiopoietin-2 messenger RNA expression.ConclusionsLV-SEM enables high magnification observation of paraffin sections used for immunohistochemistry. LV-SEM provides easy, detailed observations and offers a promising new observational modality in the field of ophthalmology.Translational RelevanceHigh magnification analysis was easily available using LV-SEM with conventional paraffin sections for light microscopy.

The Therapeutic Effects and Possible Mechanism of Pranoprofen in Mouse Model of Corneal Alkali Burns.

Objective To investigate therapeutic effects and possible mechanism of pranoprofen in a mouse model of corneal alkali burns and provide new evidence for the clinical treatment of corneal alkali burns. Methods A unilateral alkali burn was created in the central cornea by placing a piece of 2 mm diameter filter paper soaked in 1N NaOH on the right eye for 30 seconds. After the model was performed, C57BL/6J mice received topical treatment with saline eye drops or pranoprofen eye drops and were, respectively, categorized as saline group and pranoprofen group, whereas the remaining normal mice that were not subjected to alkali burns served as control, each group containing 15 mice (n = 45). On the 5th day after model establishment, the corneal fluorescein sodium staining score was evaluated in order to assess corneal epithelial damage. Tissue HE stain was used to observe the pathological changes of corneal tissue in each group. Real-time RT-PCR and western blot were also performed to detect the mRNA and protein expression of NLRP3, IL-1β/p17, and matrix metallopeptidase MMP-13. Results 5 days after burns, microscopic observations of the pranoprofen group showed less corneal opacity and neovascularization development than the saline group. Sodium fluorescein staining showed obvious corneal structure disorders, poor corneal epithelium continuity, and a larger corneal epithelial defect area in the saline group (10.33±+−0.57) as opposed to the pranoprofen group (8.33 ± 0.57） (p < 0.05). HE stain results showed the saline group had obvious corneal structure disorder and the corneal epithelial layer was incomplete as opposed to the pranoprofen group. PCR and western blot results suggested that the pranoprofen group expressed less NLRP3, IL-1β, and MMP-13 mRNA and protein expression in corneal tissue than the saline group (p < 0.05). Conclusion Pranoprofen may alleviate inflammatory response by inhibiting the expression levels of NLRP3 and IL-1β at the early stage of corneal alkali injury, lowering the expression of MMP-13 and ultimately reducing corneal epithelial damage.

LRG1 promotes corneal angiogenesis and lymphangiogenesis in a corneal alkali burn mouse model.

To investigate the potential effect and mechanism of leucine-rich α-2-glycoprotein-1 (LRG1) on corneal angiogenesis and lymphangiogenesis. Corneal neovascularization and lymphatics were induced by establishing alkali burn mouse model. Immunofluorescence staining was performed to detect the location of LRG1 in cornea tissues and to verify the source of LRG1-positive cells. Corneal whole-mount staining for CD31 (a panendothelial cell marker) and lymphatic endothelial hyluronan receptor-1 (LYVE-1; lymphatic marker) was performed to detect the growth of blood and lymphatic vessels after local application of exogenous LRG1 protein or LRG1 siRNA. In addition, expressions of the proangiogenic vascular endothelial growth factor (VEGF) related proteins were detected using Western blot analysis. LRG1 was dramatically increased in alkali burned corneal stroma in both the limbal and central areas. LRG1-positive cells in the corneal stroma were mainly derived from Vimentin-positive cells. Local application of exogenous LRG1 protein not only aggravated angiogenesis but also lymphangiogenesis significantly (P<0.01). LRG1 group upregulated the levels of VEGF and the vascular endothelial growth factor receptor (VEGFR) family when compared with the phosphate-buffered saline (PBS) control group. We also found that LRG1-specific siRNA could suppress corneal angiogenesis and lymphangiogenesis when compared with the scramble siRNA-treated group (P<0.01). LRG1 can facilitate corneal angiogenesis and lymphangiogenesis through heightening the stromal expression of VEGF-A, B, C, D and VEGFR-1, 2, 3; LRG1-specific siRNA can suppress corneal angiogenesis and lymphangiogenesis in corneal alkali burn mice.

Novel ultrasmall nanomicelles based on rebaudioside A: A potential nanoplatform for the ocular delivery of pterostilbene.

Rebaudioside A (RA) self-assembled into ultrasmall nanomicelles can be utilized as ocular drug delivery system; nevertheless, the therapeutic efficacy of RA micelles has not evaluated thus far. In this manuscript, the RA micelles are thought to strengthen the therapeutic effects of pterostilbene (Pt). Results showed that Pt can be highly encapsulated into RA micelles with ultrasmall particle sizes (3.99±0.03nm) with a uniform distribution (polydispersity index, PDI=0.184±0.008). RA-Pt exhibited pronounced improvement for the in vitro antioxidant activity and the in vitro membrane permeation of Pt. RA-Pt exhibited good ocular tolerance. The use of RA-Pt led to the significant improvement in the in vivo intraocular permeation of Pt as well as the improvement in the in vivo anti-inflammatory efficacy. The in vivo evaluation of antioxidant effects revealed that RA-Pt exhibits improvement in suppressing the generation of oxidative stress and inducing the enzymes of reactive oxygen species (ROS) degradation. Furthermore, RA-Pt was confirmed to exhibit a strong treatment efficiency for corneal alkali burns. These findings indicate that RA-based self-assembled ultrasmall nanomicelles demonstrate tremendous potential toward the improvement in the ocular bioavailability as well as a therapeutic effect of poor aqueous soluble drugs such as Pt.

Histological study of the effect of granulocyte colony-stimulating factor on experimentally induced corneal burn in adult male albino rats

ABSTRACT Chemical injuries to the eye represent one of the true ophthalmic emergencies that require immediate and intensive intervention to minimize severe complications and visual loss. Granulocyte colony-stimulating factor (G-CSF) is a potent hematopoietic cytokine that influences the proliferation, survival, maturation, and the functional activation of granulocytes. The present work was performed to evaluate the histological effect of G-CSF in treating rat corneal alkali burn model. Thirty adult male albino rats were divided equally into three main groups: Group I was served as a control group, and in Group II and III, their corneas of the right eyes were injured by applying a piece of filter paper soaked in 1M NaOH. Group II (alkali burn-induced group) was left without any treatment, while Group III (G-CSF-treated group) was injected subcutaneously by 100 µg/kg of G-CSF for 5 consecutive days. All animals were sacrificed after 3 weeks. Cornea specimens were processed for histological and immunohistochemical staining for P63 followed by morphometry. Microscopic examination of Group II revealed marked alterations in the corneal epithelium, inflammatory cellular infiltration, and neovascularization. Treatment with G-CSF showed great improvement of the corneal structure, disappearance of the neovascularization and the inflammatory cells, and decreased p63 reaction of the basal layers.

The Wound Healing Effect of Doxycycline after Corneal Alkali Burn in Rats.

Purpose To evaluate the wound healing effect of doxycycline and its underlying mechanisms in a rat model of corneal alkali burn. Methods Male SD rats were administered 1.0 N NaOH in the right cornea for 25 seconds and randomly divided into the doxycycline group and the control group, with 84 rats in each group. 1.0 g·L−1 doxycycline eye drops (doxycycline group) or vehicle (control group) was topically instilled onto the rat cornea after chemical injury. Three days, 7 days, and 14 days after alkali burn, microscopy was used to observe corneal wound healing by fluorescein staining and the degree of corneal opacity. The expression of transforming growth factor-beta 1 (TGF-β1) and matrix metalloproteinase-9 (MMP-9) was detected by RT-PCR and ELISA, alpha-smooth muscle actin (a-SMA) levels were measured by immunofluorescent staining, and Western blot assays for TGF-β1, a-SMA, and nuclear factor-kappa B (NF-κB) were also performed. Results Corneal wound healing and corneal opacity scores were better in the doxycycline group than in the control group. Three, 7, and 14 days after corneal alkali burn, a significant increase in TGF-β1 was observed in corneas from the control group, compared with the corneas from the doxycycline group (P < 0.05). The corneal levels of MMP-9 in the doxycycline group were lower than those in the control group 3 days and 7 days after alkali burn (P < 0.05). In addition, doxycycline inhibited α-SMA expression and suppressed NF-κB expression. Conclusion Doxycycline treatment promoted corneal healing and reduced corneal opacity in SD rats. Doxycycline protected the cornea from alkali burn injury by reducing TGF-β1, MMP-9, NF-κB, and α-SMA expression.

Experimental study of riboflavin-ultraviolet A collagen crosslinking for the treatment of alkali-induced corneal burns in rabbits

Objective To evaluate the efficacy of riboflavin-ultraviolet A (UVA) collagen crosslinking (CXL) combined with drugs for the treatment of alkali-induced corneal burns in rabbits. Methods Prospective experimental study.The corneas of left eyes of 40 healthy New Zealand rabbits were established alkali burn model. All rabbits were randomly divided into 4 groups: the model group, drugs treatment group, crosslinking group and combined group, with 10 cases in each group. The model goup received no treatments. The drug goup received drug treatment. Riboflavin-UVA collagen crosslinking was appied in crosslinking group and the combined group received CXL combined with drug treatment. Therapeutic efficacy of each group were compared at 1 and 3 months after operation. Results The corneal epithelial healing time in the model group, drug group, cross-linking group and combined group were (32.9±5.8 ), (12.8±2.3), (25.9±3.2 ) and (10.7±2.5 )days, respectively. The difference were statistically significant among the four groups (F=67.647, P=0.000) . The corneal turbidity score of combined group was the lowest among the 4 groups, followed by the drug group, and the score of model group was the heaviest (F=6.446, P=0.001). The number of inflammatory cells were (162.6±33.3), (46.0±21.3 ), (87.9±7.5)and (8.3±6.5)cells respectively under single confocal microscopy field of vision ( 400 μm×400 μm). The difference among the 4 groups was statistically significant (χ2=36.040, P=0.000). The corneal collagen fibers with pathological examination in the combined group were the thickest and densely arranged, followed by crosslinking group. Conclusion The efficacy of drug combined with cross-linking for the treatment of corneal alkali burn in rabbits is better than that of simple drug therapy or simple corneal collagen cross-linking. Key words: Riboflavin-ultraviolet A; Collagen; Crosslinking; Burn, alkali, coreal

Therapeutic effects of three human-derived materials in a mouse corneal alkali burn model*

Purpose: To compare the therapeutic effects of human derivatives in a mouse alkali burn model.Methods: The right eyes of mice were injured using NaOH. After alkali injury, one of the following agents was topically administered for 7 d: human amniotic membrane (hAM) suspension, human umbilical cord serum (hUCS), and human peripheral blood serum (hPBS), or saline. The epithelial defect areas on days 1, 2, and 3 degrees of opacity on days 2, 3, and 7, and corneal neovascularization (NV) areas on day 7 were evaluated. Histologic examination and mRNA expression levels of tumour necrosis factor (TNF)-α, interleukin (IL)-6, vascular endothelial growth factor (VEGF), matrix metalloproteinase (MMP)-2, MMP-8, and MMP-9 were also evaluated on day 7.Results: The epithelial defect areas in the hUCS group were smaller than those in the control and hPBS groups on day 3 (p < .05, respectively). The epithelial defect areas in the hAM suspension group showed smaller than those in the control and hPBS groups on days 1 and 2 (p < .05, respectively). The degrees of opacity were lower in all treatment groups than that of the saline control group on day 7 (p < .05, respectively). Corneal NV areas were not different among groups on day 7 (p = 0.20). The expression levels of TNF-α, IL-6, MMP-8, and MMP-9 mRNA and the infiltration of the inflammatory cells in all treatment groups were lesser than those in the control group on day 7 (p< .05, respectively).Conclusions: All treatments reduced inflammatory reactions and corneal opacity development. Corneal reepithelialization was faster in the hUCS group.

Investigation the effect of Hypericum perforatum on corneal alkali burns

Purpose: To investigate the effect of Hypericum perforatum on corneal alkali burn.Methods: We studied 45 250 g weighing, 4 months old Wistar albino rats. Alkaline burns were performed in the corneas of all experimental animals with 2 mol/L NaOH after general anaesthesia. Rats were divided into five groups according to the subsequent process applied to them: group 1 was the topical Hypericum perforatum group, group 2 was the topical pure olive oil group, group 3 was the oral Hypericum perforatum group, group 4 was the oral pure olive oil group, and group 5 was the control untreated group. Rats were sacrificed under general anaesthesia on the 14 day. The rate of corneal inflammation, neovascularization, fibroblastic activity, and cluster of differentiation 31 (CD31) staining was investigated.Result: There were 45 rats at the beginning of the study. One rat in groups 1, 2, and 3 died during the study; therefore, 42 rats could be evaluated. There were 8 rats in group 1, 8 rats in group 2, 8 rats in group 3, and 9 rats in group 4. We found less corneal neovascularization (CNV), inflammation, and fibroblastic activity in group 1 and group 2 than in the other groups (p ˂ 0.001 for all parameters). CNV, inflammation, fibroblastic activity, and CD31 staining rates were lower in group 1 than in group 2 (p ˂ 0.001 for all parameters). There was no difference between groups 3, 4, and 5 (respectively, p = 0.436, 0.634, and 0.750).Conclusions: We found that both topical Hypericum perforatum oily extract and olive oil have anti-inflammatory, anti-angiogenic, and anti-fibroblastic effects when applied after corneal alkali burns in rat corneas. Further studies should be conducted in this field.

Tetrahedral Framework Nucleic Acids Promote Corneal Epithelial Wound Healing in Vitro and in Vivo.

Poor post-traumatic wound healing can affect the normal function of damaged tissues and organs. For example, poor healing of corneal epithelial injuries may lead to permanent visual impairment. It is of great importance to find a therapeutic way to promote wound closure. Tetrahedral framework nucleic acids (tFNAs) are new promising nanomaterials, which can affect the biological behavior of cells. In the experiment, corneal wound healing is used as an example to explore the effect of tFNAs on wound healing. Results show that the proliferation and migration of human corneal epithelial cells are enhanced by exposure to tFNAs in vitro, possibly relevant to the activation of P38 and ERK1/2 signaling pathway. An animal model of corneal alkali burn is established to further identify the facilitation effect of tFNAs on corneal wound healing in vivo. Clinical evaluations and histological analyses show that tFNAs can improve the corneal transparency and accelerate the re-epithelialization of wounds. Both in vitro and in vivo experiments show that tFNAs can play a positive role in corneal epithelial wound healing.