| Background: Choroidal neovascularization (CNV) is a common pathological endpoint in a host of blind oculopathies. Several therapies in clinical status are available to treat CNV including laser photocoagulation, photodynamic therapy (PDT), transpupillary thermotherapy (TTT), macular translocation or surgical removal and radiation therapy. However, above-mentioned treatments on CNV have some insufficient aspects. As the pathogenesis of CNV is better understood, the anti-neovasular agents for CNV-related diseases has been developed the researched hot point for the past few years. Glucocorticoid, for example dexamethasone and triamcinolone, as an exogenous neovascular inhibitors, has been applied to treat the CNV-related diseases. However, it is difficult to deliver effectively doses of drugs to the posterior part of the eyes. Systemic administration necessitates large doses of the drug, resulting in general side effects. Topical eye drops penetrate poorly into the posterior part of the eye, because of lacrimation and the length of the diffusion path. Intravitreal injection must be administered multiple to maintain drug concentrations within a therapeutic range for a long period of time and sometimes causes a series of complications, such as glaucoma, cataract, vitreous hemorrhage, retinal detachment or endophthalmitis. The development of drug delivery systems to facilitate therapeutic efficacy and to minimize side effects may be the ideal method.Controlled delivery of drugs via poly(D,L-lactide-co-glycolide) (PLGA) polymers as implants, microspheres, and nanoparticles has gained wide acceptance. An ideal controlled release formulation should release the entrapped drugs in a continuous manner over desired time periods. Biodegradable nanoparticles formulated from PLGA and polyvinyl alcohol (PVA) polymers are being extensively investigated for various drug delivery applications. They are used to control drug release rates and to target drugs to specific sites in the body, thereby optimizing their therapeutic response, decreasing toxic side effects, and eliminating the inconvenience of repeated injections. PLGA is biocompatible, and more importantly, the degradation rates of PLGA and the accompanying release of encapsulated drugs can be controlled by the polymer's physicochemical properties such as molecular weight, hydrophilicity, and the ratio of lactide to glycolide. Its advantages is not only no surgical procedures for implantation in contrast to large polymer implants and removal but also their degradation rate ranges from months to years.Purpose: To investigate the inhibitory efficacy of intravitreal dexamethasone actate (DA)-loaded by Poly (D,L-lactic-co-glycolic acid) (PLGA) nanoparticles on choroidal neovascularization (CNV) in a laser-induced rat model,and to assess its release mode.Methods: DA-loaded with PLGA nanoparticles containing 50% DA was prepared using an emulsification/solvent evaporation method with slight modifications. CNV was unilaterally induced by laser photocoagulation in male Brown Norway rats. One hundred and eighty rats were randomly divided into seven groups and the 10μL volume of intravitreal drug suspension was performed the same day after photocoagulation. Different dosages of sterilised DA-loaded PLGA nanoparticles suspension were evaluated: i.e. 50μg (n=30), 100μg (n=30), 200μg (n=30), 400μg (n=30). The others groups as control were 100μg DA (n=30), blank PLGA nanoparticles (n=15) and normal saline (n=15). For each animal studied, the experimental eye received one of aforementioned suspensions. The animals were sacrificed on days 1, 3, 7, 14, 21, 28 and 56 after intravitreal injection, then the eyes were enucleated and frozen in -80℃. The cornea, iris, vitreous and choroid-retina of eyes were retrieved and DA levels were determined using a reversed phase high-performance liquid chromatography (RP-HPLC) method. Flash electroretinography (F-ERG) recordings and transmission electron microscope (TEM) were performed to assess retinal toxicity. Fluorescein fundus angiography (FFA) was performed to evaluate the incidence of CNV on days 14 and 56. The animals were sacrificed at 14 and 56 days, then eyecups were processed for histological analysis.Results: At 1 day after photocoagulation, intravitreal DA-loaded PLGA nanoparticles, blank PLGA nanoparticles and DA suspensions caused immediately the obvious vitreous opacitas in clinical examination. While the time extended, the drug suspension was gradually absorbed and deposited in the beneath of vitreous cavity. DA was fast resolved and almost complete absorbtion at 14 days after photocoagulation. However, PLGA nanoparticles suspension was very slowly absorbed so that small amounts of remanent drug clumping could be observed in the beneath of vitreous at 56 days. By RP-HPLC method, the pharmacokinetics of DA-loaded PLGA nanoparticles in vitreous and choroid-retina of the experimental eyes showed a triphasic pattern: i.e. an initial burst, a lag phase with relatively permanent release rate and another burst, and controlled release at least 56 days. In the cornea, DA was below the detection limit during the observed period. Although DA was slightly detected on day 1 after photocoagulation, it hasn't been determined after 3 days in the iris. During the examination of F-ERG, the results showed no different significant comparing with the a- and b-wave's amplitude and implicit time between pro and after laser photocoagulation in various groups.On day 14 and 56 after photocoagulation, the incidence of CNV was 47.4% or 46.2% for 50μg, 28.2% or 25.0% for 100μg, 15.8% or 15.0% for 200μg and 7.9% or 7.7% for 400μg DA-loaded PLGA nanoparticles, 31.6% or 35.0% for DA, 65.8% or 64.1% for blank PLGA nanoparticles and 65.8% or 65.0% for normal saline group, respectively. These results demonstrated that the treated groups (including DA-loaded PLGA nanoparticles and DA groups) compared with the blank PLGA nanoparticles and normal saline groups (as control) showed a statistically significant decrease in the formation of CNV (P<0.05). On day 56, the mean thickness of FVP in the recovered lesions was 84.77±9.79μm (n=10), 69.52±10.52μm (n=9), 53.17±13.83μm (n=8), 38.39±7.87μm (n=7), 70.49±12.39μm (n=9), 103.86±16.36μm (n=10) and 105.11±13.70μm (n=11), respectively, in the 50μg, 100μg, 200μg, 400μg DA-loaded PLGA nanoparticles, DA, blank PLGA nanoparticles and normal saline group. All treated groups inhibited FVP relative to controls in a statistically significant fashion (P<0.01). The inhibition of DA-loaded PLGA nanoparticles on experimental CNV showed a dose-dependent effect (P<0.05).On day 14, TEM revealed the disorder of RPE, indefinite of outer segment membrane disc apparente vacuolization of mitochondria, swelling of endoplasmic reticulum, disorded arrangement of Golg's complex, derangement of microtubule and microfilament in 400μg-treated PLGA nanoparticles and DA groups. The changes of ultrastructure were almost recovered in RPE, outer segment membrane disc, retinal ganglionic and nerve fiber layer in the 400μg DA-loaded PLGA nanoparticles group on day 56, but there were an invisible changes in the DA group. No signs of ultrastructure destruction in the RPE, outer segment membrane disc, inner and outer nuclear layer, and inner and outer plexiform layer were detected in 50μg, 100μg and 200μg DA-loaded PLGA nanoparticles groups.Conclusions: These results suggest that DA-loaded PLGA nanoparticles can dose-dependently inhibit the development of experimental CNV. Compared with DA, DA-loaded PLGA nanoparticles may be a promising drug delivery system (DDS), which possessed the characteristics of low drug toxicity, controlled release and prolonged action and may have potential as a treatment modality for CNV. . |
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