Emodin suppresses alkali burn-induced corneal inflammation and neovascularization by the vascular endothelial growth factor receptor 2 signaling pathway.

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 －80 ℃ 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.

Sorafenib-loaded nanostructured lipid carriers for topical ocular therapy of corneal neovascularization: development, in-vitro and in vivo study.

Sorafenib (SRB), a multikinase inhibitor, is effective in reducing experimental corneal neovascularization (CNV) after oral administration; however, its therapeutic use in ocular surface disorders is restricted due to poor solubility and limited bioavailability. This study aimed to develop and optimize SRB-loaded nanostructured lipid carriers (SRB-NLCs) for topical ocular delivery by a central composite design response surface methodology (CCD-RSM). It was spherical and uniform in morphology with an average particle size of 111.87 ± 0.93 nm and a narrow size distribution. The in vitro drug release from the released SRB-NLC formulation was well fitted to Korsmeyer Peppas release kinetics. The cell counting kit-8 (CCK-8) cell viability assay demonstrated that SRB-NLC was not obviously cytotoxic to human corneal epithelial cells (HCECs). An in vivo ocular irritation test showed that SRB-NLC was well tolerated by rabbit eyes. Ocular pharmacokinetics revealed 6.79-fold and 1.24-fold increase in the area under concentration-time curves (AUC0-12h) over 12 h in rabbit cornea and conjunctiva, respectively, treated with one dose of SRB-NLC compared with those treated with SRB suspension. Moreover, SRB-NLC (0.05% SRB) and dexamethasone (0.025%) similarly suppressed corneal neovascularization in mice. In conclusion, the optimized SRB-NLC formulation demonstrated excellent physicochemical properties and good tolerance, sustained release, and enhanced ocular bioavailability. It is safe and potentially effective for the treatment of corneal neovascularization.

Cannabis and the Cornea.

Purpose: While cannabis has the potential to reduce corneal pain, cannabinoids might induce side effects. This review article examines the effects of cannabinoids on the cornea. As more states and countries consider the legalization of adult cannabis use, health-care providers will need to identify ocular effects of cannabis consumption.Methods: Studies included in this review examined the connection between cannabis and the cornea, more specifically anti-nociceptive and anti-inflammatory actions of cannabinoids. NCBI Databases from 1781 up to December 2019 were consulted.Results: Five studies examined corneal dysfunctions caused by cannabis consumption (opacification, decreased endothelial cell density). Twelve studies observed a reduction in corneal pain and inflammation (less lymphocytes, decreased corneal neovascularization, increased cell proliferation and migration).Conclusion: More than half of the studies examined the therapeutic effects of cannabinoids on the cornea. As the field is still young, more studies should be conducted to develop safe cannabinoid treatments for corneal diseases.

Comparison of the effect of topical bevacizumab and sorafenib in experimental corneal neovascularization.

PURPOSE: The purpose of this study was to compare the neovascularization inhibiting the effect of topical bevacizumab and sorafenib and to determine the effective dose of sorafenib. MATERIAL AND METHODS: Forty-two healthy Wistar albino rats were randomly divided into six groups. The right corneas of all rats except group 1 were cauterised with silver nitrate. Group 2 received DMSO, group 3 received topical bevacizumab (5 mg/dL, 3 times a day) and group 4, 5 and 6 received topical sorafenib (2.5 mg/dl, 5 mg/dL, 7.5 mg/dL, 2 times a day respectively), between days 1 and 7. Corneal photographs were taken on day 8 and the corneal neovascular area percentage was calculated. Following decapitation, the corneas were removed to determine the levels of VEGF ELISA and corneal immune staining. The Mann-Whitney U-test was used for statistical analysis. RESULTS: The neovascular corneal area percentage was statistically significantly lower in the treatment groups than group 2 (p < 0.05). The intensity of VEGF immune staining was also lower in groups 3, 5 and 6 from the group 2. Group 3, 5 and 6 were no significant differences compared to group 1. The VEGF ELISA levels were statistically significantly lower in group 3, 5 and 6 compared to group 2 (p < 0.05). There was no statistically difference between VEGF ELISA levels of group 2 and 4 (p > 0.05). CONCLUSIONS: Sorafenib was as effective as bevacizumab in the regression of corneal neovascularization. The effect of sorafenib seems to be dose-dependent. The low doses and twice a day administration are important advantages of sorafenib.

Osthole: A Traditional Chinese Medicine for Ocular Anti-Angiogenic Therapy.

PURPOSE: Osthole is an agent isolated from Cnidium monnieri (L.) Cusson and has been used to treat several disorders. Corneal neovascularization is a sight-threatening condition associated with several inflammatory or infectious ocular disorders. In this study, we investigated the anti-angiogenic effects of osthole on corneal neovascularization and the underlying mechanism. METHODS: MTT assay, HE staining, and calcein-AM/propidium iodide staining was conducted to detect the toxicity of osthole in vitro and in vivo. Corneal neovascularization of ICR mice was induced by alkali burn and observed by a slit lamp microscopy on day 7 after alkali injury. EdU assay, Ki67 immunofluorescence assay, Transwell migration assay, and Matrigel assay were conducted to investigate the role of osthole in endothelial angiogenic effects in vitro. Western blots were conducted to investigate the anti-angiogenic mechanism of osthole in corneal neovascularization. RESULTS: Administration of osthole ranging from 0.05 to 25 µM had no detectable cytotoxicity or tissue toxicity in vivo and in vitro. Topical administration of osthole inhibited corneal neovascularization induced by alkali burn. Osthole decreased the proliferation, migration, and tube-formation of endothelial cells induced by VEGF. Osthole inhibited endothelial angiogenic functions through blocking the phosphorylation of ERK1/2, JNK, and p38. CONCLUSION: Our study provides evidence that osthole is a promising drug for the treatment of corneal neovascularization.

Inhibition of COX-2, mPGES-1 and CYP4A by isoliquiritigenin blocks the angiogenic Akt signaling in glioma through ceRNA effect of miR-194-5p and lncRNA NEAT1.

BACKGROUND: Arachidonic acid (AA) metabolic enzymes including cyclooxygenase-2 (COX-2), microsomal prostaglandin E synthase-1 (mPGES-1) and cytochrome P450 (CYP) 4A11 play important roles in glioma angiogenesis. Thus, there is an urgent need to identify the underlying mechanisms and develop strategies to overcome them. METHODS: A homology model of human CYP4A11 was constructed using SYBYL-X 2.0. Structure-based virtual screening against COX-2, mPGES-1 and CYP4A11was performed using the Surflex-Dock of the SYBYL suite. The candidates were further evaluated their antiangiogenic activities in a zebrafish embryo and rabbit corneal angiogenesis model. Laser doppler analysis was used to measure tumor perfusion. The expression of CD31 and α-SMA was measured by immunofluorescence. Western blot was used to measure the expression of HIF-1, Akt and p-Akt. The gene expression of FGF-2, G-CSF, PDGF, TGF-ß, Tie-2, VEGF, lncRNA NEAT1 and miR-194-5p were determined using qPCR. The production of FGF-2, TGF-ß and VEGF were analyzed using ELISA. Bioinformatic analysis and luciferase reporter assays confirmed the interaction between lncRNA NEAT1 and miR-194-5p. RESULTS: The nearly 36,043 compounds from the Traditional Chinese Medicine (TCM) database were screened against COX-2, mPGES-1 and CYP4A11 3D models, and the 17 top flavonoids were identified. In zebrafish screening, isoliquiritigenin (ISL) exhibited the most potent antiangiogenic activities with the EC50 values of 5.9 µM. Conversely, the antiangiogenic effects of ISL in the zebrafish and rabbit corneal models were partly reversed by 20-hydroxyeicosatetraenoic acid (20-HETE) or prostaglandin E2 (PGE2). ISL normalized glioma vasculature and improved the efficacy of temozolomide therapy in the rat C6 glioma model. Inhibition of COX-2, mPGES-1 and CYP4A by ISL decreased FGF-2, TGF-ß and VEGF production in the C6 and U87 glioma cells with p-Akt downregulation, which was reversed by Akt overexpression. Furthermore, ISL downregulated lncRNA NEAT1 but upregulated miR-194-5p in the U87 glioma cell. Importantly, lncRNA NEAT1 overexpression reversed ISL-mediated increase in miR-194-5p expression, and thereby attenuated FGF-2, TGF-ß and VEGF production. CONCLUSIONS: Reprogramming COX-2, mPGES-1 and CYP4A mediated-AA metabolism in glioma by flavonoid ISL inhibits the angiogenic Akt- FGF-2/TGF-ß/VEGF signaling through ceRNA effect of miR-194-5p and lncRNA NEAT1, and may serve as a novel therapeutic strategy for human glioma.

Cucurbita argyrosperma Seed Extracts Attenuate Angiogenesis in a Corneal Chemical Burn Model.

Severe corneal inflammation produces opacity or even perforation, scarring, and angiogenesis, resulting in blindness. In this study, we used the cornea to examine the effect of new anti-angiogenic chemopreventive agents. We researched the anti-angiogenic effect of two extracts, methanol (Met) and hexane (Hex), from the seed of Cucurbita argyrosperma, on inflamed corneas. The corneas of Wistar rats were alkali-injured and treated intragastrically for seven successive days. We evaluated: opacity score, corneal neovascularization (CNV) area, re-epithelialization percentage, and histological changes. Also, we assessed the inflammatory (cyclooxigenase-2, nuclear factor-kappaB, and interleukin-1ß) and angiogenic (vascular endothelial growth factor A, VEGF-A; -receptor 1, VEGFR1; and -receptor 2, VEGFR2) markers. Levels of Cox-2, Il-1ß, and Vegf-a mRNA were also determined. After treatment, we observed a reduction in corneal edema, with lower opacity scores and cell infiltration compared to untreated rats. Treatment also accelerated wound healing and decreased the CNV area. The staining of inflammatory and angiogenic factors was significantly decreased and related to a down-expression of Cox-2, Il-1ß, and Vegf. These results suggest that intake of C. argyrosperma seed has the potential to attenuate the angiogenesis secondary to inflammation in corneal chemical damage.

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.

Comparison of the Effects of Subconjunctival Injections of Bevacizumab and Interferon Alpha-2a on Corneal Angiogenesis in a Rat Model.

BACKGROUND AND OBJECTIVE: Corneal neovascularization (CNV) is a vision-threatening condition arising from various corneal diseases. The aim of this study is to compare the effectiveness of bevacizumab and interferon alpha-2a (IFNα-2a) treatment on corneal neovascularization. MATERIALS AND METHODS: Twenty-four Wistar albino rats were used in this study. After cauterization of the cornea with a silver nitrate applicator stick, the control group received 0.1 mL saline solution, the second group received 0.1 mL IFNα-2a (IFNα-2a, 6 million international units [MIU]/0.5 mL), and the third group received 2.5 mg bevacizumab by subconjunctival injection. An additional injection was administered to each group on the fourth day. After one week, the corneal neovascularization rate and the longest neovascular sprout length were determined. RESULTS: The neovascularization rate (saline 0.65 ± 0.05; IFNα-2a 0.62 ± 0.07; bevacizumab 0.42 ± 0.11) with bevacizumab was significantly lower, more than those with IFNα-2a and saline (p < 0.001 and p < 0.001). The longest neovascular sprout length (saline, 4.00 ± 0.6 mm; IFNα-2a, 3.63 ± 0.52 mm; bevacizumab, 2.81 ± 0.65 mm) with bevacizumab was significantly shorter than those with saline and IFNα-2a (p = 0.001 and p = 0.012). CONCLUSIONS: Subconjunctival IFNα-2a has limited efficacy in the treatment of corneal neovascularization.

Ginsenoside Rh2 inhibits vascular endothelial growth factor-induced corneal neovascularization.

VEGF-induced neovascularization plays a pivotal role in corneal neovascularization (CoNV). The current study investigated the potential effect of ginsenoside Rh2 (GRh2) on neovascularization. In HUVECs, pretreatment with GRh2 largely attenuated VEGF-induced cell proliferation, migration, and vessel-like tube formation in vitro. At the molecular level, GRh2 disrupted VEGF-induced VEGF receptor 2 (VEGFR2)-Grb-2-associated binder 1 (Gab1) association in HUVECs, causing inactivation of downstream AKT and ERK signaling. Gab1 knockdown (by targeted short hairpin RNA) similarly inhibited HUVEC proliferation and migration. Notably, GRh2 was ineffective against VEGF in Gab1-silenced HUVECs. In a mouse cornea alkali burn model, GRh2 eyedrops inhibited alkali-induced neovascularization and inflammatory cell infiltrations in the cornea. Furthermore, alkali-induced corneal expression of mRNAs/long noncoding RNAs in cornea were largely attenuated by GRh2. Overall, GRh2 inhibits VEGF-induced angiogenic effect via inhibiting VEGFR2-Gab1 signaling in vitro. It also alleviates angiogenic and inflammatory responses in alkali burn-treated mouse corneas.-Zhang, X.-P., Li, K.-R., Yu, Q., Yao, M.-D., Ge, H.-M., Li, X.-M., Jiang, Q., Yao, J., Cao, C. Ginsenoside Rh2 inhibits vascular endothelial growth factor-induced corneal neovascularization.

Anti-angiogenic effect of hexahydrocurcumin in rat corneal neovascularization.

AIM: This study was to investigate the anti-angiogenic effect of hexahydrocurcumin (HHC) to evaluate gene (p-basic fibroblast growth factor (bFGF)-SAINT-18 & p-vascular endothelial growth factor (VEGF)-SAINT-18 complex)-induced corneal neovascularization (CorNV) in rats. METHODS: CorNV was induced in 24 eyes of 24 rats. Four groups (Group A: 0 µg, B: 0.01 µg, C: 0.1 µg, and D: 1 µg) of HHC were prepared and implanted into the rat subconjunctival substantia propria 1.5 mm from the limbus at temporal side. The 1 µg of p-bFGF-SAINT-18 & p-VEGF-SAINT-18 complex were prepared and implanted into the rat corneal stroma 1.5 mm from the limbus at the same side. Inhibition of CorNV was observed and quantified from day 1 to day 60. bFGF and VEGF protein expression were analyzed by biomicroscopic examination, western blot analysis, and immunohistochemistry. RESULTS: Subconjunctival injection by 1 µg HHC successfully inhibited gene-induced CorNV in rats. bFGF and VEGF protein expression were reduced after 6 days. Meanwhile, the reduction of HLA-DR expression was detected. CONCLUSIONS: Our study showed that the HHC might provide an important anti-angiogenesis factor to inhibit CorNV development at the corneal experimental angiogenesis model.

Therapeutic effects of zerumbone in an alkali-burned corneal wound healing model.

Cornea is an avascular transparent tissue. Ocular trauma caused by a corneal alkali burn induces corneal neovascularization (CNV), inflammation, and fibrosis, leading to vision loss. The purpose of this study was to examine the effects of Zerumbone (ZER) on corneal wound healing caused by alkali burns in mice. CNV was induced by alkali-burn injury in BALB/C female mice. Topical ZER (three times per day, 3µl each time, at concentrations of 5, 15, and 30µM) was applied to treat alkali-burned mouse corneas for 14 consecutive days. Histopathologically, ZER treatment suppressed alkali burn-induced CNV and decreased corneal epithelial defects induced by alkali burns. Corneal tissue treated with ZER showed reduced mRNA levels of pro-angiogenic genes, including vascular endothelial growth factor, matrix metalloproteinase-2 and 9, and pro-fibrotic factors such as alpha smooth muscle actin and transforming growth factor-1 and 2. Immunohistochemical analysis demonstrated that the infiltration of F4/80 and/or CCR2 positive cells was significantly decreased in ZER-treated corneas. ZER markedly inhibited the mRNA and protein levels of monocyte chemoattractant protein-1 (MCP-1) in human corneal fibroblasts and murine peritoneal macrophages. Immunoblot analysis revealed that ZER decreased the activation of signal transducer and activator of transcription 3 (STAT3), with consequent reduction of MCP-1 production by these cells. In conclusion, topical administration of ZER accelerated corneal wound healing by inhibition of STAT3 and MCP-1 production.

Antiangiogenic activity of a bevacizumab-loaded polyurethane device in animal neovascularization models.

PURPOSE: To evaluate the antiangiogenic activity of bevacizumab-loaded polyurethane using two animal models of neovascularization. METHODS: The percentage of blood vessels was evaluated in a chicken chorioallantoic membrane model (n=42) and in the rabbit cornea (n=24) with neovascularization induced by alkali injury. In each model, the animals were randomly divided into the groups treated with the bevacizumab-loaded polyurethane device, phosphate-buffered-saline (negative control) and bevacizumab commercial solution (positive control). Clinical examination, as well as histopathological and immunohistochemical evaluation, were performed in the rabbit eyes. Microvascular density in hot spot areas was determined in semi-thin sections of corneal tissue by hematoxylin-eosin staining and factor VIII immunohistochemistry. Immunohistochemical analysis was also performed to evaluate VEGF expression. RESULTS: In the evaluated models, the use of bevacizumab (Avastin®) and the bevacizumab-loaded polyurethane device led to similar results with regard to inhibition of neovascularization. In the chorioallantoic membrane model, the bevacizumab-loaded polyurethane device reduced angiogenesis by 50.27% when compared to the negative control group. In the rabbit model of corneal neovascularization, the mean density of vessels/field was reduced by 46.87% on analysis of factor VIII immunohistochemistry photos in the bevacizumab-loaded polyurethane device group as compared to the negative control (PBS) sections. In both models, no significant difference could be identified between the bevacizumab-loaded polyurethane device and the positive control group, leading to similar results with regard to inhibition of neovascularization. CONCLUSIONS: The present study shows that the bevacizumab-loaded polyurethane device may release bevacizumab and inhibit neovascularization similarly to commercial bevacizumab solution in the short-term.

Curcumin inhibits angiogenesis by up-regulation of microRNA-1275 and microRNA-1246: a promising therapy for treatment of corneal neovascularization.

OBJECTIVE: Curcumin (capable of inhibiting angiogenic growth of human umbilical vein endothelial cells [HUVECs]), can be employed in vitro as a model of pathogenesis of corneal neovascularization (CRNV). The aim of this study was to explore regulatory mechanisms of microRNA (miR) levels after curcumin treatment. MATERIALS AND METHODS: Expression profiles of miRs in curcumin-treated HUVECs were investigated by miR microassay. Specific mimics and inhibitors of miR-1275 or miR-1246 were transfected into HUVECs. Then, their target genes, vascular endothelial growth factor B (VEGFB) and nuclear transcription factor kappa B acting protein (NKAP) were detected by quantitative real-time PCR, Western blotting assay or immunofluorescence assay. Cell proliferation and cell cycle parameters were measured with the help of CCK-8 assay and flow cytometry. RESULTS: MiR-1275 and miR-1246 expression levels were up-regulated by curcumin. Administration of the specific mimics and inhibitors of the two miRs led to significant changes in expression of VEGFB and NKAP as well as the indicators related to angiogenesis. Anti-angiogenic effect of curcumin depended on expression patterns of the two miRs in that inhibition of either miR interfered with the effect of curcumin. Furthermore, overexpression of NKAP interrupted effects of curcumin on the cells. CONCLUSION: Collectively, our findings demonstrate that curcumin inhibited HUVEC proliferation by up-regulation of miR-1275 and miR-1246.

Therapeutic effects of a novel PIGF-1 derived peptide, ZY-1, on corneal neovascularization in vitro and in vivo.

Corneal neovascularization (NV) is one of the major sight-threatening pathological changes caused by corneal diseases. Current therapeutics generate various adverse effects. Small peptides derived from endogenous protein display certain advantages. This study aims to evaluate the anti-angiogenic effect and molecular mechanism of a novel peptide ZY-1, derived from placental growth factor-1 (PlGF-1), on corneal NV by topical administration, and to investigate its safety profile after long-term treatment. CCK-8 assay and tube formation assay were used to evaluate the effect of ZY-1 on human umbilical vein endothelial cells (HUVECs). The anti-angiogenic effect of topical ZY-1 was estimated in a rat model of alkali burn induced corneal NV. The safety profile of topical ZY-1 was analyzed by CCK-8 assay, tear film break-up time (BUT), and histological examination. Firstly, we found that ZY-1 co-localized with membrane vascular epithelial growth factor receptor-1 (VEGFR-1) and effectively inhibited VEGF/PlGF-1 induced proliferation and tube formation of HUVECs. The topical ZY-1 administration efficiently inhibited alkali-burn induced corneal NV, while it did not show any significant effect on human corneal epithelial cell (HCEC) proliferation, as well as the functionality and morphology of cornea and conjunctiva. Our findings suggested that topical administration of ZY-1 could effectively and safely inhibit corneal NV partially through competing for VEGFR-1 binding, and it would be a promising alternative for ocular topical anti-angiogenic therapy.

[Neovascularization corneal regression in patients treated with photodynamic therapy with verteporfin]. / Regresión en la neovascularización corneal en pacientes tratados con terapia fotodinámica con verteporfirina.

BACKGROUND: Corneal neovascularization is a vision-threatening condition usually associated with inflammatory or infectious disorders of the ocular surface. One current treatment is photodynamic therapy, which uses a photosensitizer to occlude the vessel, is successfully produced microvascular thrombosis with minimal damage to surrounding normal tissue. The aim of this article is to quantitatively determine the percentage of regression of corneal neovascularization experienced by patients treated with photodynamic therapy with verteporfin. METHODS: A before and after treatment; experimental, analytical, prospective and longitudinal. RESULTS: Of the 25 new vessels analyzed, 8 glasses (32 %) had total occlusion one month after, 15 vessels (60 %) had a partial occlusion in the range of 15.3 to 85.1 %, and 2 vessels (8 %) worsening in corneal vascularization. The mean area of corneal neovascularization decreased significantly a 70 % from 0.147 ± 0.118 mm2 to 0.045 ± 0.046 mm2 (p < 0.0005) after photodynamic therapy. No side efects were reported. CONCLUSIONS: Photodynamic therapy with verteporfin is a safe and effective method of reducing corneal neovascularization and can be used to inhibit angiogenesis in the cornea.

Introducción: la neovascularización corneal es una condición que amenaza la visión, por lo general, se asocia a trastornos inflamatorios o infecciosos de la superficie ocular. Uno de los tratamientos actuales es la terapia fotodinámica, el uso de un fotosensibilizador para ocluir los vasos ha producido con éxito la trombosis microvascular con un daño mínimo al tejido normal circundante. El objetivo de este artículo es determinar cuantitativamente el porcentaje de regresión en la neovascularización corneal que presentan los pacientes tratados con terapia fotodinámica con verteporfirina. Métodos: estudio de tratamiento de antes y después; experimental, analítico, prospectivo y longitudinal. Resultados: de los 25 neovasos analizados, 8 vasos (32 %) presentaron al mes una oclusión total del 100 %, 15 vasos (60 %) una oclusión parcial en el rango de 15.3 al 85.1 %, y 2 vasos (8 %) empeoramiento en la vascularización corneal. El promedio del área de neovascularización corneal disminuyó significativamente en un 70 % de 0.147 ± 0.118 mm2 a 0.045 ± 0.046 mm2, (p < 0.0005) posterior a la terapia fotodinámica. No se reportaron efectos adversos. Conclusiones: la terapia fotodinámica con verteporfina es un procedimiento seguro y efectivo para reducir la neovascularización de la córnea y puede utilizarse para inhibir la angiogénesis en la córnea.

Diospyros kaki Extract Inhibits Alkali Burn-Induced Corneal Neovascularization.

The purpose of this study was to evaluate the effect of ethanol extract of Diospyros kaki (EEDK) leaves on corneal neovascularization (CoNV) in rats. One week after the alkali burns in the corneas, the CoNV area coverage in the CoNV-positive control group, 100 mg/kg EEDK group, and 200 mg/kg EEDK group was 43.3% ± 5.5%, 337.7% ± 2.5%, and 27.2% ± 4.3%, respectively. The areas of CoNV in the EEDK-treated groups were significantly different from those of the CoNV group. EEDK significantly attenuated the upregulation of vascular endothelial growth factor, fibroblast growth factor, interleukin-6, and matrix metalloproteinase-2 (MMP-2) protein levels. Orally administrated D. kaki inhibited CoNV development in rats.

[Vascular morphological and microdensity changes of corneal neovascularization induced by topical bevacizumab and sunitinib in an animal model]. / Cambios en la morfología y la microdensidad neovascular corneal inducidos tras la administración tópica de bevacizumab y sunitinib en un modelo animal.

OBJECTIVE: To evaluate the effects of topical bevacizumab and topical sunitinib on vascular microdensity and morphology of corneal neovascularization (NV). METHODS: A total of 33 rabbits were distributed into 3 groups: group 1 (control; n=11): saline; group 2 (n=11): bevacizumab 5mg/ml; and group 3 (n=11): sunitinib 0.5mg/ml. A corneal NV model was used, based on sutures in the right eye of each rabbit. Each treatment was administered topically 3 times daily for 14 days. Corneas were then processed for the study of vascular microdensity (6 eyes) and vascular morphology analysis (5 eyes) using enzymatic staining histological techniques RESULTS: The vascular response in group 3 was limited to small-sized tree formations with various vascular axes compared with the extensive, lush and directional corneal NV of group 1 and 2. In the histological sections near the limb, there were no differences in vascular microdensity studies between the three groups. However, the mean sectional area of vessels (MSAV) in group 3 was 41.88% lower than in group 1 and 19.19% lower than in group 2. In distal sections, there were no differences between groups 1 and 2. However, group 3 was characterized by absence of vessels. CONCLUSIONS: Bevacizumab produced no changes in the morphology of the vessels or the vascular microdensity. Sunitinib reduced the size of the new vessels and induced changes in the vascular tree.

Celastrol nanoparticles inhibit corneal neovascularization induced by suturing in rats.

PURPOSE: Celastrol, a traditional Chinese medicine, is widely used in anti-inflammation and anti-angiogenesis research. However, the poor water solubility of celastrol restricts its further application. This paper aims to study the effect of celastrol nanoparticles (CNPs) on corneal neovascularization (CNV) and determine the possible mechanism. METHODS: To improve the hydrophilicity of celastrol, celastrol-loaded poly(ethylene glycol)-block-poly(ɛ-caprolactone) nanopolymeric micelles were developed. The characterization of CNPs was measured by dynamic light scattering and transmission electron microscopy analysis. Celastrol loading content and release were assessed by ultraviolet-visible analysis and high performance liquid chromatography, respectively. In vitro, human umbilical vein endothelial cell proliferation and capillary-like tube formation were assayed. In vivo, suture-induced CNV was chosen to evaluate the effect of CNPs on CNV in rats. Immunohistochemistry for CD68 assessed the macrophage infiltration of the cornea on day 6 after surgery. Real-time quantitative reverse transcription-polymerase chain reaction and enzyme-linked immunosorbent assay were used to evaluate the messenger ribonucleic acid and protein levels, respectively, of vascular endothelial growth factor, matrix metalloproteinase 9, and monocyte chemoattractant protein 1 in the cornea. RESULTS: The mean diameter of CNPs with spherical shape was 48 nm. The celastrol loading content was 7.36%. The release behavior of CNPs in buffered solution (pH 7.4) showed a typical two-phase release profile. CNPs inhibited the proliferation of human umbilical vein endothelial cells in a dose-independent manner and suppressed the capillary structure formation. After treatment with CNPs, the length and area of CNV reduced from 1.16 ± 0.18 mm to 0.49 ± 0.12 mm and from 7.71 ± 0.94 mm(2) to 2.29 ± 0.61 mm(2), respectively. Macrophage infiltration decreased significantly in the CNP-treated corneas. CNPs reduced the expression of vascular endothelial growth factor, matrix metalloproteinase 9, and monocyte chemoattractant protein 1 in the cornea on day 6 after suturing. CONCLUSION: CNPs significantly inhibited suture-induced CNV by suppressing macrophage infiltration and the expression of vascular endothelial growth factor and matrix metalloproteinase 9 in the rat cornea.

Inhibition of corneal neovascularization in rats by systemic administration of sorafenib.

PURPOSE: To evaluate the effect of orally administered sorafenib on corneal neovascularization in rat models. METHODS: In male Sprague-Dawley rats, a silver nitrate applicator was placed on the central cornea in both eyes to elicit angiogenesis. Rats were divided into 3 groups, the control group and the 2 sorafenib-treated groups (low dose, 30 mg · kg(-1) · day(-1); high dose, 60 mg · kg(-1) · day(-1)). The area of corneal neovascularization was measured by image analysis. Vascular endothelial growth factor receptor 2 (VEGFR2) messenger RNA expression was measured in rat corneas by reverse transcription-polymerase chain reaction, and the expression of phosphorylated extracellular signal-regulated kinase (ERK) was measured by Western blot analysis 1 week after cauterization. RESULTS: The area of corneal neovascularization was significantly reduced by 44% in the 30 mg · kg(-1) · day(-1) group and by 66% in the 60 mg · kg(-1) · day(-1) group, compared with the control group (P = 0.014 and P < 0.0001). Corneal VEGFR2 messenger RNA expression was higher in the control group than in the sorafenib-treated groups. The expression of phosphorylated ERK in rat corneas was suppressed in the sorafenib-treated groups but not in the control group. CONCLUSIONS: Oral administration of a multikinase inhibitor (sorafenib) significantly reduced the development of experimental corneal neovascularization in a dose-dependent manner. This inhibitory effect is probably related to the suppression of ERK phosphorylation by sorafenib.