Inhibition Of Corneal Neovascularization Research Articles

VEGF-targeted scFv inhibits corneal neovascularization via STAT3 pathway in alkali burn model

We developed a novel anti-vascular endothelial growth factor (VEGF) single-chain variable fragment (scFv) and evaluated its anti-inflammatory and anti-corneal neovascularization (CoNV) effects in the alkali burn model. VEGF-scFv was prepared from the derived sequence of bevacizumab with a glycine and serine repetitive linker. After being evaluated using surface plasmon resonance (SPR), enzyme-linked immunosorbent assay (ELISA), and cellular fluorescent staining, VEGF-scFv showed high specificity and good affinity with VEGF antigen and HUVEC cells. The KD value was 3.07 ± 0.37 nM and the Bmax value was 0.23 ± 0.01 pmol (n = 3). The corneal alkali burn model was built on male Sprague Dawley (SD) rats. ScFv was injected subconjunctivally immediately after the alkali burn. On 1, 3, 7, and 14 d after alkali burn, the area of CoNV in the VEGF-scFv group was similar to that of the dexamethasone group (2 mg/mL) and significantly smaller than the 0.9% NaCl and the bevacizumab control groups (2 mg/mL). Especially on 14 d, the CoNV area was 45.15% ± 8.46% in the VEGF-scFv group. Hematoxylin and eosin staining (HE) confirmed that few new blood vessels in the VEGF-scFv group. Both immunofluorescence staining and western blotting verified that the expression levels of VEGF, vascular endothelial growth factor receptor 2 (VEGFR2), and signal transducer and activator of transcription 3 (STAT3) in the burned corneal tissue in VEGF-scFv group were significantly lower than other groups (P < 0.05), suggesting this scFv inhibited CoNV by regulating the VEGF/VEGFR2/STAT3 pathway. Therefore, our study prepared an anti-VEGF scFv which may provide a potential treatment for corneal alkali burn for future clinical application.

Inhibition of Corneal Neovascularization with the Combination of Aflibercept and Plasmid Pigment Epithelium-Derived Factor-Synthetic Amphiphile INTeraction-18 Vector

Aflibercept, a recent VEGF inhibitor, is a recombinant fusion protein consisting of portions of human VEGF receptors 1 and 2 extracellular domains, fused to the Fc portion of human IgG1 formulated as an iso-osmotic solution for intravitreal administration. In this study, we have estimated that the combination of aflibercept and plasmid pigment epithelium-derived factor-synthetic amphiphile INTeraction-18 (p-PEDF-SAINT-18) has a favorable antiangiogenic effect on corneal neovascularization (NV). Four groups (Group A: 0 μg + 0 μg, B: 0. 1μg + 0.1 μg, C: 1 μg + 1 μg, D: 10 μg + 10 μg) of aflibercept + p-PEDF-SAINT-18 were prepared and implanted into rat subconjunctival substantia propria 1.5 mm from the limbus on the temporal side. The 2 μg of p- bFGF-SAINT-18 were prepared and implanted into the rat corneal stroma 1.5mm from the limbus on the same side. Inhibition of NV was observed and quantified from day 1 to day 90. 18-kDa bFGF and VEGF protein expression was analyzed through biomicroscopic examination, western blot analysis and immunohistochemistry. No inhibition activity for normal limbal vessels was noted. Subconjunctival injection by a combination of aflibercept and p-PEDFSAINT-18 successfully inhibited corneal NV. Successful genes of bFGF and PEDF were expressed through western blot analysis; immunohistochemistry staining showed mild immune response for HLA-DR. We conclude that the combination of aflibercept and p-PEDF-SAINT-18 may have more potent and prolonged antiangiogenic effects, making it possible to reduce the frequency of subconjunctival aflibercept administration, while maintaining relatively high safety and low toxicity.

Effect of Nintedanib Nanothermoreversible Hydrogel on Neovascularization in an Ocular Alkali Burn Rat Model

Purpose To compare the therapeutic effects of different forms of nintedanib ophthalmic preparations on neovascularization corneal alkali burns in rats. Methods Forty rat models of left eye corneal alkali burns were constructed, and the five groups (N = 8) were treated with normal saline, dexamethasone ointment (dexamethasone), 0.2% nintedanib aqueous solution and nintedanib nano thermoreversible hydrogel (NNTH). A slit lamp microscope was used to observe the area of neovascularization. The levels of the inflammatory factors were detected by ELISA. HE staining was performed on the rat corneas. Vascular endothelial growth factor (VEGFA) was detected by immunohistochemistry, and the expression of corneal VEGFA and CD31 was detected by western blotting. An MTT assay was performed to detect the cytotoxicity of nintedanib on human corneal epithelial cells (HCECs) and human umbilical vein vascular endothelial cells (HUVECs). Cell migration was detected by a cell scratch assay, and the proportion of apoptotic cells was detected by Annexin/PI double staining. Immunofluorescence and western blotting were performed to detect the protein expression of VEGFA and CD31. Results NNTH had a stronger inhibitory effect on corneal neovascularization (CNV) in alkali-burned rats while reducing the level of inflammatory factors. NNTH had a longer drug duration of release than nanoformulations in vitro. Nintedanib at low concentrations (<8 μM) had no significant cytotoxicity to HCECs but significantly induced apoptosis and inhibited the expression of VEGFA and CD31 and the migration of HUVECs. Conclusions Nanomorphic thermoreversible hydrogel is superior among the nintedanib ophthalmic preparations, showing better inhibition of CNV in alkali-burned eyeballs and it blocked the migration and proangiogenic ability of HUVECs.

Printable Metal-Polymer Conductors for Local Drug Delivery.

Safe and effective local drug delivery is challenging due to complex physiological barriers that limit the entry of drugs. Here, we report the metal-polymer conductors (MPCs) for local drug delivery via iontophoresis or electroporation. The MPCs are stretchable, conductive, and biocompatible. The flexible MPCs of different geometries are used both on a dry, flat surface (skin) and a moist, curved surface (cornea) with conformability. Conformal integration with the tissues enables good mechanical/electrical properties and realizes application of electrical voltage to the target areas for local drug delivery. By iontophoresis and electroporation, the MPCs achieve efficient delivery of doxorubicin and siRNA, leading to tumor regression and inhibition of corneal neovascularization, respectively. Our work presents an efficient strategy to harness the power of the MPCs to broaden the scope of local drug delivery to dry and wet organs with different surface topography.

Preparation and in vitro and in vivo evaluation of an isoliquiritigenin-loaded ophthalmic nanoemulsion for the treatment of corneal neovascularization

Isoliquiritigenin (ISL), as a natural flavonoid, has been proven to have therapeutic potential for corneal neovascularization (CNV) treatment; however, its therapeutic use is restricted due to its poor aqueous solubility and limited bioavailability. To overcome these limitations, a novel ISL-loaded nanoemulsion (ISL-NE) was designed for inhibiting CNV in this study. ISL-NE formulation was composed of propylene glycol dicaprylate (PGD), Cremophor® EL (EL35), polyethylene glycol 400 (PEG 400) and adding water with sodium hyaluronate, its particle size was 34.56 ± 0.80 nm with a low polydispersity index of less than 0.05, which suggested a narrow size distribution. The results demonstrated that ISL-NE released higher and permeated more drug than ISL suspension (ISL-Susp) in in vitro drug release and ex vivo corneal permeation study. ISL-NE showed no cytotoxicity in human corneal epithelial cells toxicity study, which was consistent with the result of ocular irritation study in rabbit eyes. ISL-NE had bioavailability 5.76-fold, 7.80-fold and 2.13-fold higher than ISL-Sups in tears, cornea and aqueous humor after a single dose of ISL-NE, respectively. Furthermore, the efficacy of ISL-NE treatment (0.2% ISL) was comparable to that of dexamethasone treatment (0.025%) in the inhibition of CNV in mice model. Enzyme-linked immunosorbent assay (ELISA) showed that the expressions of corneal vascular endothelial growth factor (VEGF-A) and matrix metalloproteinase (MMP-2) were decreased. In conclusion, the ISL-NE demonstrated excellent physicochemical properties, good tolerance, and enhanced ocular bioavailability. It could be a promising, safe, and effective treatment for CNV.

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.

The effects of subconjunctival bevacizumab, ranibizumab, and aflibercept on corneal neovascularization.

To investigate the effects of subconjunctival bevacizumab, ranibizumab, and aflibercept in an experimental corneal neovascularization model. The eyes of 24 rats were chemically cauterized and randomly divided into four groups: bevacizumab group (0.05mL/1.25mg bevacizumab), ranibizumab group (0.05mL/0.5mg ranibizumab), aflibercept group (0.05mL/1.25mg aflibercept), and control group (0.05mL saline solution). Plasma vascular endothelial growth factor (VEGF) levels were among the major measurement outcomes to assess corneal neovascularization. The collected plasmas were analyzed using the SIGMA RAB0511 Rat VEGF Elisa kit. The PCR technique and VEGF amplification procedures were used for molecular analysis. Each cornea was removed and histologically examined on day 21. Corneal images were evaluated by image analyzer software. In the post-injection period, the number of major corneal arteries decreased significantly in the injection groups when compared to the control group (p = 0.037), but no statistically significant differences were noted among the injection groups (p > 0.05). The aflibercept group had the lowest area of neovascularization. Immunohistochemical staining revealed substantially lower VEGF percentages in neovascularized arteries of the injection groups than the control group (p = 0.015). In TUNEL staining, the mean TUNEL value (number/1hpf) was substantially greater in the control group than in the injection groups, but the mean TUNEL values were similar between the injection groups (p = 0.019, p > 0.05, respectively). No statistically significant differences were observed between the groups in terms of corneal surface area with increased cellularity, edema, and inflammation (p = 0.263). The mean plasma VEGF concentration in the control group was statistically greater than those in the injection groups (p = 0.001). Subconjunctival bevacizumab, ranibizumab, and aflibercept crossed the blood and seemed to be effective in inhibiting corneal neovascularization without causing epitheliopathy in an experimental rat model compared to the controls. However, no significant results were noted between these three anti-VEGF molecules.

Development of a naringenin microemulsion as a prospective ophthalmic delivery system for the treatment of corneal neovascularization: in vitro and in vivo evaluation

Naringenin, a flavonoid, possesses antiangiogenic potential and inhibits corneal neovascularization (CNV); however, its therapeutic use is restricted due to poor solubility and limited bioavailability. In this study, we developed a naringenin microemulsion (NAR-ME) for inhibiting CNV. NAR-ME formulation was composed of triacetin (oil phase), Cremophor RH40 (CRH40), PEG400, and water, its droplet size was 13.22 ± 0.13 nm with a narrow size distribution (0.112 ± 0.0014). The results demonstrated that NAR-ME released higher and permeated more drug than NAR suspension (NAR-Susp) in in vitro drug release and ex vivo corneal permeation study. Human corneal epithelial cells (HCECs) toxicity study showed no toxicity with NAR-ME, which is consistent with the result of ocular irritation study. NAR-ME had high bioavailability 1.45-fold, 2.15-fold, and 1.35-fold higher than NAR-Susp in the cornea, conjunctiva, and aqueous humor, respectively. Moreover, NAR-ME (0.5% NAR) presented efficacy comparable to that of dexamethasone (0.025%) in the inhibition of CNV in mice CNV model induced by alkali burning, resulting from the attenuation of corneal vascular endothelial growth factor (VEGF) and matrix metalloproteinase (MMP-14) expression. In conclusion, the optimized NAR-ME formulation demonstrated excellent physicochemical properties and good tolerance, enhanced ocular bioavailability and corneal permeability. This formulation is promising, safe, and effective for the treatment of CNV.

Monolith/Hydrogel composites as triamcinolone acetonide carriers for curing corneal neovascularization in mice by inhibiting the fibrinolytic system

Corneal neovascularization is a serious corneal pathological change caused by various factors. The drug delivery system is of great significance for the effective treatment of corneal neovascularization. Herein, we developed and characterized a monolith/hydrogel composite as the triamcinolone acetonide (TA) carrier for curing corneal neovascularization. The composite was prepared by photo-initiated free radical polymerization of multi-methacrylate substituted dodecamine organic molecular cage and post-modified by the sequential photo-initiated free radical polymerization of acrylated gelatin. The globular morphology and structural property of as-prepared composites were evaluated by scanning electron microscopy, Fourier-transform infrared spectroscopy and solid-state cross polarization magic angle spinning carbon-13 nuclear magnetic resonance. Then swelling ratio and the TA loading capacity were investigated then. Compared with gelatin hydrogel, the composites exhibited a decreased swelling ratio and an improved loading capacity. With good biocompatibility, the composite can sustainedly release TA for up to 28 days, and effectively inhibit corneal neovascularization with an alkali burn-induced corneal neovascularization model. Additionally, tandem mass tags-labeled quantitative proteomics were performed to identify differentially expressed proteins between vascularized and devascularized corneas. The Kyoto Encyclopedia of Genes and Genomes enrichment analysis revealed that the inhibition process could be primarily linked to the fibrinolytic system. These results demonstrated the potential of monolith/hydrogel composites as delivery systems in the therapy for biomedical diseases.

Inhibition of Neovascularization and Inflammation in a Mouse Model of Corneal Alkali Burns Using Cationic Liposomal Tacrolimus.

Eye drops account for more than 90% of commercialized ophthalmic drugs. However, eye drops have certain shortcomings, such as short precorneal retention time and weak corneal penetration. The requirement of frequent instillation of eye drops also causes poor patient compliance, which may lead to further aggravation of the disease. We aimed to develop a cationic liposome formulation to increase the bioavailability of the therapeutic agent and solve the aforementioned problems. In the present study, we prepared cationic liposomal tacrolimus (FK506) with a surface potential of approximately +30 mV, which could bind to the negatively charged mucin layer of the ocular surface. Our results showed that the content of FK506 in the cornea was increased by 93.77, 120.30, 14.24, and 20.36 times at 5, 30, 60, and 90 min, respectively, in the FK506 liposome group (0.2 mg/ml) compared with the free drug group (0.2 mg/ml). Moreover, FITC-labeled FK506 liposomes significantly prolonged the ocular surface retention time to 50 min after a single dose. In addition, the results of the Cell Counting Kit-8 assay, live and dead cell assay, sodium fluorescein staining, and hematoxylin and eosin staining all indicated that FK506 liposomes had good biological compatibility in both human corneal epithelial cells and mouse eyeballs. Compared with the free drug at the same concentration, FK506 liposomes effectively inhibited vascular endothelial growth factor-induced green fluorescent protein-transduced human umbilical vein endothelial cell migration and tube formation in vitro. In a mouse corneal neovascularization model induced by alkali burns, FK506 liposomes (0.2 mg/ml) enhanced corneal epithelial recovery, inhibited corneal neovascularization, and reduced corneal inflammation, and its therapeutic effect was better than those of the commercial FK506 eye drops (1 mg/ml) and the free drug (0.2 mg/ml). Collectively, these results indicate that cationic FK506 liposomes could increase the efficacy of FK506 in the corneal neovascularization model. Therefore, cationic FK506 liposomes can be considered as a promising ocular drug delivery system.

Inhibition of corneal neovascularization by topical application of nintedanib in rabbit models.

To evaluate the potential efficacy and mechanisms of nintedanib in corneal neovascularization (NV) in rabbit models. Corneal NV was induced using 1 mol/L NaOH. Rabbits (n=21) were randomized to 3 groups: Group 1 were treated with 0.9% NaCl, Group 2 with Avastin (5 mg/mL), and Group 3 with nintedanib (1 mg/mL). All treatments started 1d after alkaline burns and were topically performed 3 times a day for 2wk. Photographs were taken on a slit lamp microscope on day 7 and 14. The NV area, the length of the vascularization and angiogenesis index (AI) were used to evaluate the corneal NV. On day 14, the immunohistochemical (IHC) studies of the cornea were examined. Western blot was performed to test the expression levels of vascular endothelial growth factor (VEGF), Akt, p-Akt, P38, p-P38, MMP-2 and MMP-9. The corneal NV area, vessel length and AI in Group 3 were significantly lower than Group 2, with both being lower than Group 1. IHC staining showed that VEGF was significantly overexpressed in the epithelium and stroma of cornea following alkaline burns. In contrast, the level of VEGF was significantly suppressed in both Group 2 and Group 3. Western blot results further confirmed that, compared with Group 1, Group 3 had significantly reduced expressions of VEGF, Akt, p-Akt, p-P38, MMP-2, and MMP-9 in corneal tissues. Trends of lower levels of MMP-2, AKT, and p-AKT in Group 3 than Group 2 were identified. Nintedanib and Avastin can effectively inhibit corneal NV, with P38 MAPK and AKT signaling pathways being possibly involved. Nintedanib seems more effective than Avastin and has the potential to be a novel therapy for preventing corneal NV.

The Effect of Limbal Mesenchymal Stem Cell Secretome on Corneal Neovascularization in Experimental Model of Corneal Alkali Injury

Background: To investigate the effect of limbal mesenchymal stem cell (L-MSC) secretome in inhibition of corneal neovascularization in animal model of alkali chemical injury Methods: Secretome was collected from limbal mesenchymal stem cells of corneoscleral tissue of healthy male rabbit. Rabbits were exposed to NaOH. The research subjects consisted of 3 groups, normal control group, Balance salt solution (BSS) group, and (L-MSC) secretome group. Clinical corneal neovascularization level, number of corneal neovascularization histopathologically with hematoxylin-eosin staining and VEGF expression by immunohistochemical staining were analyzed. Results: After seven days of administration of topical instillation of L-MSC secretome, neovascularization level in L-MSC group was lower than BSS group even though the difference was not statistically significant (p>0.05). The mean histopathological neovascular count in the L-MSC secretome group was lower than that BSS group even though the difference was not statistically significant (p>0.05) Conclusions: In conclusion, topical instillation of limbal mesenchymal stem secretome has a potential effect on inhibiton of corneal neovascularization even though not significant at day-7 in experimental alkali injury model.

Efficacious, safe, and stable inhibition of corneal neovascularization by AAV-vectored anti-VEGF therapeutics

Corneal neovascularization (CoNV) leads to visual impairment, affecting over 1.4 million people in the United States per year. It is caused by a variety of pathologies, such as inflammation, hypoxia, and limbal barrier dysfunction. Injection of the anti-vascular endothelial growth factor (VEGF) drug KH902 (conbercept) can inhibit CoNV but requires repeated dosing that produces associated side effects, such as cornea scar. To explore more efficacious and long-lasting treatment of CoNV, we employed recombinant adeno-associated virus (rAAV)2 and rAAV8 vectors to mediate KH902 expression via a single intrastromal injection and investigated its anti-angiogenic effects and safety in both alkali-burn- and suture-induced CoNV mouse models. Our results showed that rAAV-mediated KH902 mRNA expression in the cornea was sustained for at least 3 months after a single intrastromal injection. Moreover, the expression level of rAAV8-KH902 far exceeded that of rAAV2-KH902. A single-dose rAAV8-KH902 treatment at 8 × 108 genome copies (GCs) per cornea dramatically inhibited CoNV for an extended period of time in mouse CoNV models without adverse events, whereas the inhibition of CoNV by a single intrastromal administration of the conbercept drug lasted for only 10−14 days. Overall, our study demonstrated that the treatment of CoNV with a single dose of rAAV8-KH902 via intrastromal administration was safe, effective, and long lasting, representing a novel therapeutic strategy for CoNV.

Ocular lamellar crystalline gels for sustained release and enhanced permeation of resveratrol against corneal neovascularization

Corneal neovascularization (CNV) is the major cause of blindness after eye injury; however, only several drugs can be applied and the invasive administration ways (i.e., intravitreal injection and subconjunctival injection) are used. Resveratrol is a highly effective anti-VEGF agent against CNV. However, its applications are limited due to its strong hydrophobicity and instability. Here, we developed a resveratrol-loaded ocular lamellar crystalline gel (ROLG) for high inhibition of CNV. ROLGs were composed of resveratrol, glyceryl monooleate (GMO), ethanol, and water, and their lamellar crystalline structures were identified by polarizing light microscopy and small-angle X-ray scattering. High drug loading (4.4 mg/g) of ROLGs was achieved due to the hydrogen bonding between GMO and resveratrol. Resveratrol showed sustained release with 67% accumulative release in 7 h, which was attributed to the slow erosion of gels. Resveratrol in ROLGs had a high corneal permeation 3 times higher than resveratrol in hyaluronic acid suspensions (RHSs). ROLGs were administered to rats only once a day because of their strong retention on the cornea surface. ROLGs were safe due to the very little contact of ethanol in ROLGs to the cornea. CNV post-rat corneal alkaline injury was highly inhibited by ROLGs, resulting from the attenuation of corneal VEGF expression and then corneal healing was improved. The ROLG was a promising ocular medicine for the prevention of CNV.

Bioadhesive glycosylated nanoformulations for extended trans-corneal drug delivery to suppress corneal neovascularization.

Eye-drop formulations as conventional regimens to tackle ocular diseases are far from efficient due to the rapid clearance by eye tears and the blockage of the corneal epithelium barrier. Here, we describe a bioadhesive glycosylated nanoplatform with boric acid pendants as a drug carrier for noninvasive trans-corneal delivery of drugs to treat corneal neovascularization (CNV), a serious corneal disease resulting in significant vision impairment. This biocompatible nanoplatform is formulated from a synthetic amphiphilic boric acid-based copolymer self-assembling to form highly stable micelles with a high loading capacity for dexamethasone (DEX). The nanoplatform is demonstrated to be in contact with the corneal epithelium for a long period under the bioadhesive function of boric acid modules and releases the drug over 96 h in a controlled manner. Our results also suggest that the nanoplatform can be efficiently internalized by corneal epithelial cells in vitro and realize transcytosis in vivo to greatly enhance the transcorneal penetration of the loaded drugs into the pathological corneal stroma. On topical application against rat corneal alkali burn, the nanoformulation presents more robust efficacy on neovascularization suppression and inflammation elimination than free DEX with a negligible effect on normal tissues. This bioadhesive strategy which focuses on extending ocular drug retention and improving trans-corneal drug delivery not only highlights an approach for alternative noninvasive therapy of CNV but also provides a versatile paradigm for other biomedical applications by overcoming protective barriers.

The phototherapy toward corneal neovascularization elimination: An efficient, selective and safe strategy

Corneal neovascularization (CNV) can induce severe visual impairment and even blindness. Current treatments have limited efficacy and some undesirable side effects. Phototherapy (PT) is a modern medical technique in CNV treatment and worth further improvement. Nanotechnology has various advantages and nanoparticle-mediated drug delivery also contributes to CNV elimination. Phototherapy combined with functional nanoparticles featuring photoacoustic imaging contrast properties can accomplish accurate and safe inhibition of CNV, thus is promising in clinical application.

Thermoresponsive gel for sustained release of BMP4 to inhibit corneal neovascularization

This work aimed to seek a sustained drug release system based on poloxamer-based thermoresponsive gel for sustained release drugs to inhibit corneal neovascularization (CNV) after eye operations. Thus, we designed and prepared a thermoresponsive gel with a phase transition temperature from 22 °C to 25 °C. When the concentrations of poloxamer (P) was 18% (w/w) and ε-Polylysine (EPL) was 0.5 mg/mL (P-18-EPL-05) in the thermoresponsive gel solution, the obtained thermoresponsive gel showed a suitable viscosity and strength in physiological condition. The viscosity, storage modulus G' and loss modulus G” of P-18-EPL-05 were 8 × 102 mPa.s, 1.17 × 104 Pa and 3.77 × 103 Pa, respectively. In-vitro release studies indicated that the drug release ratio of P-18-EPL-05 gel better than that of the poloxamer solution alone. The animal experiments indicated that the thermoresponsive gel loading bone morphogenetic protein 4 (BMP4) was better to inhibit CNV than the common solvent one. Overall, these results demonstrated that P-18-EPL-05 gel would be a promising platform as drug sustained systems for inhibiting CNV after eye injury in ophthalmic applications.

Multimodal Photoacoustic Imaging-Guided Regression of Corneal Neovascularization: A Non-Invasive and Safe Strategy.

Corneal neovascularization (CNV) is one of the main factors that induce blindness worldwide. However, current medical treatments cannot achieve non‐invasive and safe inhibition of CNV. A noninvasive photoacoustic imaging (PAI)‐guided method is purposed for the regression of CNV. PAI can monitor the oxygen saturation of cornea blood vessels through the endogenous contrast of hemoglobin and trace administrated drugs by themselves as exogenous contrast agents. An indocyanine green (ICG)‐based nanocomposite (R‐s‐ICG) is prepared for CNV treatment via eye drops and subconjunctival injections. It is demonstrated that R‐s‐ICG can enrich corneal tissues and pathological blood vessels rapidly with minor residua in normal eyeball tissues. Anti‐CNV treatment‐driven changes in the blood vessels are assessed by real‐time multimodal PAI in vivo, and then a safe laser irradiation strategy through the canthus is developed for phototherapy and gene therapy synergistic treatment. The treatment leads to the efficient inhibition of CNV with faint damages to normal tissues.

Corneal neovascularization is inhibited with nucleolin-binding aptamer, AS1411

Corneal neovascularization (CNV) is a common sight-threatening pathology that can be induced by a variety of inflammatory and angiogenic stimuli. Current CNV treatments include anti-inflammatory drugs and antibody-based inhibitors of vascular endothelial growth factor (VEGF). However, these are not always effective and novel therapeutic approaches are needed. Previous work has indicated a role for nucleolin (NCL) in VEGF-mediated neoangiogenesis in a suture-induced CNV model. The major goal for this current study is to test the effect of AS1411, a NCL-binding DNA aptamer that has reached human clinical trials, on neovascularization in a murine model of VEGF-mediated CNV. Our results show that topical administration of AS1411 can significantly inhibit corneal neovascularization in this model. Mechanistic studies indicate that AS1411 reduces the VEGF-stimulated proliferation, migration, and tube formation of primary cells obtained from human limbus stroma (HLSC). AS1411 treatment also significantly reduced VEGF-stimulated induction of miR-21 and miR-221 in HLSC, suggesting a role for these pro-angiogenic miRNAs in mediating the effects of AS1411 in this system. In sum, this new research further supports a role for NCL in the molecular etiology of CNV and identifies AS1411 as a potential anti-angiogenic CNV treatment that works by a novel mechanism of action.

Evaluation of pigment epithelium-derived factor concentration in equine amniotic membrane homogenate and its in-vitro vascular endothelial growth factor inhibition effect in tears of dogs with vascularized ulcerative keratitis.

Background:Corneal neovascularization can result from many pathological processes affecting the ocular surface leading to disturbances and opacifications that reduce corneal clarity and may impact vision. In veterinary medicine, the use of topical corticosteroid is contraindicated in the presence of ulcerative keratitis, and there is sparse research regarding safe medical alternatives to inhibit corneal neovascularization in dogs to improve visual outcome.Aim:To investigate the pigment epithelium-derived factor (PEDF) concentration in equine amniotic membrane homogenate (EAMH) and its in-vitro vascular endothelial growth factor (VEGF) inhibition in tears of dogs with vascularized ulcerative keratitis.Methods:Homogenates from 10 equine amniotic membranes (AM) were analyzed by sandwich enzyme-linked immunosorbent assay (ELISA) for quantification of equine PEDF and VEGF. Forty tear samples were collected from both eyes of dogs diagnosed with vascularized ulcerative keratitis, and 50 samples from healthy dogs. Samples from affected eyes were allocated to G1 – affected undiluted tears; G2 – affected tears diluted with phosphate-buffer solution; G3 – affected tears treated with low-concentrated EAMH; and G4 – affected tears treated with high-concentrated EAMH. Tears from the unaffected contralateral eyes were composed in G5, while G6 was composed by tears from healthy dogs (control). The presence and levels of VEGF were evaluated in all groups by Western blot and ELISA.Results:The PEDF:VEGF ratio in EAMH was 110:1. An increase in VEGF levels was observed in tears from eyes with vascularized corneal ulcers (G1) as well as in contralateral tears (G5), compared to normal dogs (G6). High-concentrated EAMH provided a greater decrease in VEGF levels in-vitro compared to low-concentrated EAMH.Conclusion:EAMHs exhibited high concentrations of PEDF in comparison to VEGF and were able to partially decrease VEGF levels in tears of dogs with vascularized ulcers, in-vitro. Our results suggest that VEGF concentration is elevated in tears of dogs with active vascularized ulcerative keratitis in both affected and contralateral eyes compared to that of healthy dogs.