IndexAbstractIntroductionOcular structure, barriers and delivery systemsNPs used in ophthalmologySpecific applications of NPs in ophthalmologyNPs for the diagnosis of ocular diseasesNPs for the therapy of ocular diseasesNPs for ocular gene therapyOcular Anterior Segment Gene TherapyOcular Posterior Segment Gene TherapyNP for Drug Therapy PurposesAnterior Segment Drug TherapyAbstractNowadays, achieving more efficient and convenient ocular drug delivery methods is a highly welcomed achievement. Nanotechnology via nanoparticles as effective and feasible carriers, considered an attractive treatment method to deliver ocular drugs to specific target cells. Due to the presence of numerous barriers and the unique anatomy of the eye, direct access of the drug to particular sites represents a great challenge. The ocular surface epithelium, tear turnover, the presence of the blood-aqueous barrier and the blood-retinal barrier have a significant impediment impact on the administration of drugs at the ocular level. Furthermore, conventional methods of ocular drug treatment are hampered by low bioavailability and serious ocular adverse effects. Nanocarriers and nanotechnology can be enormously useful in treating ocular diseases and delivering drugs to target regions. In this review, we present recent advances in nanotechnology-based systems and specific nanoparticles used for different purposes in ophthalmology. Say no to plagiarism. Get a tailor-made essay on "Why Violent Video Games Shouldn't Be Banned"? Get an original essay IntroductionEye diseases could have a direct negative influence on human vision and quality of life. According to previous research, 285 million people globally have visual impairments, with 39 million being blind, of which 82% are over the age of fifty. Due to the burden of global population growth, it has been estimated that the overall cost of vision problems will increase significantly by 2020. Considering this growing trend and the critical role of ocular dysfunctions in individuals' lives, achieving diagnostic and therapeutic pathways most convenient and advantageous has been investigated. So far, significant discoveries have been made in the mechanism of ophthalmic pathology and the treatment of ocular diseases. However, effective ocular drugs and gene therapy methods still represent a challenge that requires attention. There are some particular characteristics of our eyes that favor the study of therapeutic methods. Nowadays, non-invasive methods for direct ocular visualization and local treatment can be accessed. In addition, the possibility of simultaneous observation of the other eye is another advantage in this matter. On the other hand, due to ocular anatomical barriers, local and systemic administration of ocular drugs suffers from lack of specificity and low efficiency. Thus, overcoming these obstacles is a major challenge. Furthermore, most of the available and current treatments are not used to restore eye diseases or vision loss. Nowadays, the development of both nanotechnology and the application of nanomedicine makes tremendous progress in medicine. Nanotechnology is a part of science related to the design of functional structures based on the nanoscale. Nanomedicine is the application of nanotechnology for medical interventions including the diagnosis, treatment and prevention of human diseases based on molecular studiesof the human body. In the Nanomedicine system, there are several nanocarriers for ocular delivery purposes, such as: nanoparticles, liposomes, emulsions, nanotubes, nanosuspensions and dendrimers. Nanoparticles (NPs) are microscopic materials measured at the nanoscale, which act as a whole unit. They could be used in two categories of nanocapsules and nanospheres and consist predominantly of lipids, proteins and polymers. Based on recent discoveries regarding the exceptional impacts of NPs in ophthalmology, development of intelligent diagnostic and therapeutic solutions is highly beneficial. In this study we discuss the advantages of applying nanomedicine and nanoparticles in the field of ophthalmology. First, we introduce the ocular anatomical characteristics, barriers and delivery routes. Then, samples of nanoparticles used for various ocular purposes are examined. In the end, we come to a conclusion as well as the future prospects in this field. This review aims to provide better knowledge of the captivating roles of NPs in different sections of ophthalmology in the present and future. Ocular structure, barriers and delivery systems The eye with its spherical shape consists of three main layers. The ocular globe is divided into anterior segment (cornea, conjunctiva, aqueous humor, iris, ciliary body and lens) and posterior segment (retina, choroid, sclera, vitreous humor and optic nerve). The primary barrier of the eye is the cornea which comprises five layers. The corneal epithelial layer, through its cellular tight junctions, plays a considerable hindering role in drug penetration. Furthermore, there is constant tear turnover on the ocular surface between 2 and 20 minutes, which significantly decreases the bioavailability and absorption of topically administered drugs. Although drug permeability is apparently greater in the conjunctiva than in the cornea, it is not adequately efficient due to the conjunctival capillaries and lymphatic presence. The blood-ocular barriers consisting of the blood-aqueous barrier (BAB) and the blood-retinal barrier (BRB) limit the systemic diffusion of drugs. The BAB plays a significant preventive role in the anterior segment of the eye. BAB created by the endothelium of the iris capillaries, the non-pigmented ciliary epithelium and the iris epithelium. The blood-retinal barrier (BRB) is associated with the back of the eye, including the inside and outside parts. Retinal vascular endothelium, pericytes, and astrocytes accumulate in the inner part, and retinal pigment epithelium (RPE) cells correlate with the outer section of the BRB. Overall, in developed countries, posterior segment eye diseases such as age-related maculopathy and diabetic retinopathy are more common than anterior segment dysfunctions. However, medications for the anterior segment (e.g. antibiotics, antiglaucoma medications,…) reflect greater dominance, particularly in the form of eye drops. There are several avenues regarding the applications of ocular drugs. Depending on the ocular target site, the modalities for ocular drug therapy are: topical administration, periocular, transseptal, retroorbital, intracameral, subretinal and intravitreal injections. Anterior segment oculopathies (cornea, conjunctiva, sclera, and anterior uvea) are commonly treated with topical eye drops. Unfortunately, due to rapid tear turnover, blinking and drainage of the nasolacrimal system, the bioavailability of the eye drops is less than 5%. It is obvious that topical therapeutic methods may not be adequately effective on disorders of the posterior segment (retina, vitreous, choroid). Therefore, systemic administration of the drug (intravenous or intravitreal)it is more popular when it comes to posterior segment therapies. However, distinct complications of frequent, high-dose intraocular injections (retinal hemorrhage, retinal detachment, cataract, endophthalmitis) are of concern. Subsequently, outstanding research has been conducted to make new alternative methods with more beneficial results in this regard. For example, by injecting nanoparticles via hollow microneedles through the sclera, the drugs showed themselves more quickly with long-lasting effectiveness. NPs used in ophthalmology as novel carriers have numerous advantages based on their good targeting potential and consistent drug release. Furthermore, various studies have shown descriptions of long-term bioavailability, fair biodegradability, and biocompatibility of NPs, which make them promising tools in ophthalmology. Nanoparticles for molecular therapy can be classified into 4 groups: (1) lipid-based NPs (protamine lipid DNA (LPD)), (2) metal-based NPs, (3) polymer-based and gelatin-based NPs . NP-based delivery mechanisms include cell entry, avoiding endosomal degradation, and DNA entry for therapeutic agent delivery. NPs are made of various materials, such as chitosan, polylactic glycolic acid, hyaluronic acid (HA), cerium oxide, silver, gold, and silica. Specific Applications of NPs in Ophthalmology As mentioned, nanotechnology is the science of synthesis and characterization of nanomaterials, which could functionalize such materials with other different molecules for specific purposes. So far, the combination of nanotechnology and biomedicine has produced interesting results in many fields of medicine. The advent of nanotechnology is promising and can substantially accelerate ophthalmic therapeutic and diagnostic approaches using specific NPs through improved penetration and sustained and controlled drug release. NP for the Diagnosis of Ocular Diseases There are various methods used for the diagnosis of ocular diseases, including fluorescein and indocyanine green angiography, electroretinography, ultrasonography, ocular coherence tomography (OCT), computed tomography (CT), and magnetic resonance imaging ( MRI). Although these methods have greatly influenced the healing process of ocular diseases, each of them suffers from restrictions in the diagnosis and monitoring of the disease. To address these limitations, nanotechnology appears to have provided several avenues. The application of GNPs in ocular imaging is one of its myriad roles in ophthalmology (e.g. photothermal therapy, gene delivery, drug delivery). It has been reported that GNPs could be used as a good contrast agent for OCT. Anderson et al. prepared an emulsion of gold-perfluorocarbon NPs conjugated with the monoclonal anti-integrin antibody (àvß3) DM101. They indicated that after injection of the targeted agent, the mean MRI signal intensity was increased to 25% in the rabbit in vivo model. Zagaynova et al. demonstrated that gold-silica NPs improved the OCT signal intensity and brightness of related parts of the OCT image. Similarly, in a mouse model study, there was a nearly 50-fold increase in OCT contrast after GNR injection into the anterior chamber [109]. Interestingly, biosensing cards made with GNP could be used for the diagnosis of infectious diseases such as keratoconjunctivitis. similarly, using GNPs with Raman spectroscopy has significantly improved detection signals, which analyze human tears. This method not only revealed the discrimination between normal and contagious ocular tissue, but also determined thetype of infected eye (viral, bacterial and allergic). Taken together, GNPs have a potential impact on improving the early diagnosis of ocular diseases. Although the application of nanotechnological methods for the diagnosis and treatment of tumors has recently shown increasing improvement, there are few studies related to their use in ophthalmology. However, approaches used for other diseases could represent a viable guide regarding the diagnosis and treatment of ocular diseases. In one study, the Nano hydrogel system is combined with tumor targeting, induced drug delivery, and photo-to-heat processing. In this report, peptide-based ligands with phage particles, heat-sensitive liposomes (HSLs), or mesoporous silica (MSNPs) were assembled into a hydrogel for tumor-targeted drug delivery. Together, the authors arrived at multimodal imaging and monitored the delivery of therapeutic agents in human tumor xenograft. Recently, there have been promising breakthroughs in nanotechnology for early disease detection using engineered monitoring tools such as biosensors, particularly for chronic eye diseases such as glaucoma and retinal degenerative disease. . Lin et al. demonstrated a new light-based technique targeting NPs for the treatment of pigmented cells in ophthalmology. The nanoindentation system is another interesting example in this regard, which determines the efficient hydraulic conductivity and modulus of the human ocular surface. With this method, ideal drug delivery routes via NPs from the ocular surface can be achieved. As a consequence of the development of these methods, there will be a notable increase in therapeutic interventioneffectiveness and notable reduction in patient expenditure.NP for the therapy of ocular diseasesAs stated, ocular anatomical barriers limit the penetration of drugs from front to back. Furthermore, drugs infiltrate the eye in the different direction of distribution of the liquid. Therefore, sharp obstacles impressively disturb the effectiveness of intraocular drug administration. In view of the increasing prevalence of chronic ocular diseases in the elderly, numerous drug administration methods have been studied. Eye-catching results in gene transfer, stem cell and protein therapy have accentuated the need to improve the stability and bioavailability of new therapeutic units [119]. NP-based delivery methods reduce the frequency of ocular injections and improve efficiency, causing fewer side effects as well as patient convenience. The effectiveness of gene- or protein-based drug delivery is hindered by their chemical and physical instability. Therefore, designing sustainable methods of drug/gene therapy (e.g., encapsulation- or nanotechnology-based methods) could be a great approach to address the obstacles and nominated side effects of frequent ocular injections. The effectiveness of nanoparticle drug/gene/protein delivery depends on their charge, polarity, size, shape and structure. Different formulations based on basic salts, surfactants, etc. have been tested. to improve molecular stability during intraocular administration. The main applications of NPs for ocular therapeutic purposes are explained here. NPs for Ocular Gene Therapy The success of gene therapy is determined by two fundamental factors: (1) safe and effective gene delivery to target cells in vivo and in vitro. (2) effective monitoring of modifying agents oredited cells via non-invasive imaging methods, which allow monitoring of gene delivery. There are two types of gene therapy vectors: viral and non-viral. Although viral vectors are more popular due to their greater efficacy in gene expression, their significant restrictions and side effects have been motivating to study other alternative vectors. The use of NPs with less proven side effects, long-term gene expression, higher capacity, better bioavailability, and biodegradability is receiving more and more attention. There has been considerable use of NPs loading gene transcription factors, which facilitate cellular programming via in vivo studies. Ocular anterior segment gene therapy The cornea is simply accessible, transparent and somewhat separated from the blood circulation. Thus, these properties make the cornea a noble target for gene therapy or childbirth. The main purpose of corneal gene therapy is to transfer the gene to the cornea or ocular tissues close to it. More recent studies have productively used corneal gene therapy to avoid corneal disorders such as: corneal neovascularization, corneal rejection, and herpetic stromal keratitis. In Sharma et al. 15 μmole of polyethylenimine conjugated (PEI) to GNP-containing gene (GFP) was administered locally onto the rabbit cornea. Therapeutic dosing of GNPs accumulated in corneal keratocytes and extracellular matrix did not lead to cytotoxicity. Furthermore, after topical use of GNPs encapsulating the BMP7 (bone morphogenic protein 7) gene, there was a notable reduction in surgery-induced fibrosis in the rabbit cornea. Vicente et al. have achieved magnificent results through solid lipid NP gene delivery by transfecting human corneal epithelial cell lines. In another study, we note noteworthy results of the topical use of PLGA (polylactic-co-glycolic acid) NPs that target the anti-VEGF RNA expression cassette plasmid on the cornea. Finally, a conspicuous decrease in corneal neovascularization was observed 4 weeks after the administration of the formulated materials. A similar case was observed in the subconjunctival application of PLGA NPs for Flt23k (anti VEGF intraceptor), which reduced corneal neovascularization and graft rejection fraction. In Iriyama et al. There was successful subconjunctival delivery of VEGF1-expressing PEG-bP nanomicelles (ASP(DET)) upon regression of corneal neovascularization. Interestingly, it was demonstrated that direct application of dendritic polymers rapidly promoted corneal wound repair compared to sutures. In another study chitosan (CS) and thiolated chitosan (TCS) NPs were used on cultured mouse corneal and human corneal fibroblast cells. CS and TCS NPs enhanced anti-angiogenic and anti-fibrotic therapies by downregulating TGFβ1 and PDGF expression. Ocular Posterior Segment Gene Therapy Choroidal neovascularization (CNV) is a major cause of blindness, the treatment of which depends on the underlying intentions. For CNV therapy, in one study PLGA NPs loading the anti-VEGF plasmid were functionalized with transferrin and surface peptide. As a result, rats studied with this method had a smaller CNV area than the group treated with non-functionalized NPs. Furthermore, CNV suppression was achieved by intravitreal gene therapy of PLGA/chitosan NPs enveloping the proteolytic plasminogen kringle 5 (K5) plasmid. Similarly, PLG NPs loading RGD peptide-labeled anti-VEGF interceptor have been successfully studied forgene expression in laser-induced CNV model via intravenous (IV) injection. Retinal gene therapy has achieved exceptional results particularly in animal models of retinal dystrophic disease. It is known that the retina is immunologically privileged and separated from other parts of the body by BRBs. Interestingly, surgical access to the retina is cost-effective. That's why it's the fascinating part of the eye for gene delivery methods. But one of the key requirements in this regard is the selection of effective carriers such as useful and specific nanoparticles that take into account the high sensitivity of the retina. Both viral and non-viral vectors such as NPs have demonstrated robust success in retinal gene therapy. However, considering the advantages of NPs over viral vectors, their contribution in gene delivery is gaining more attention. Application of PEI NPs via oligonucleotide (ODN) encapsulating the anti TGF-ß2 plasmid was evaluated for retinal gene delivery. This in vivo study showed adequate diffusion of the conjugated NPs into retinal muller ganglion (RMG) cells after intravitreal injection. Furthermore, the accumulation of formulated materials in RMG cells inhibited cell growth procedures. Retinal pigment epithelial (RPE) cells have important roles in the visual system. RPE cells are not only the outer part of the BRB, but are also widely involved in ophthalmic pathological conditions, such as AMD and retinitis pigmentosa (RP). RPE cells are capable of taking up various types of NPs. Bejjani et al. evaluated the delivery of internalized GFP plasmid via PLA and PLGA NPs to the human ARPE-19 cell line, with clear results without toxic effects on RPE cells. Bourges et al. concluded the effective and sustained accumulation of PLA NPs (containing Rh-6G and Nile Red Fluorochromes (RNFPs)) in RPE cells by IV injection. Subsequent studies have been conducted on new methods for rescuing photoreceptor function in degenerative retinal conditions. Furthermore, subretinal injection of DNA NPs with CK30-PEG encapsulating rhodopsin gDNA promoted gene expression and therapeutic efficiency for rhodopsin deletion in the mouse model of RP. In this way, remarkable gene expression occurred in addition to the rescue of photoreceptor function. It is worth knowing that another merit of using PLA/PLGA NPs in gene delivery is that the FDA has already approved their application in this field [78, 150]. Naash et al. evaluated the effects of CK30-PEG driving pZEO-GFP by incorporating the CMV promoter. After subretinal injection into wild-type PI-2 mice, GFP expression was observed in retinal ganglion cells, inner and outer retina. Furthermore, GFP expression was present in the lens and cornea in the eye of P5 mice. This could be a good reason for using cell/gene-specific promoters to reduce the possibility of ectopic expression. In another study, CK30-PEG DNA NPs targeting human ABCA4 cDNA and mouse opsin (MOP-ABCA4) were formulated for gene delivery to mice with Stargardt dystrophy. During subretinal injection of Abca4(-/-) mice at P30, there was constant expression of ABCA4 for more than 8 months in addition to a notable improvement in the structural and functional Stargardt phenotype, as well as improved dark adaptation and depletion of lipofuscin granules. Han et. al. have shown that DNA NPs are a promising replacement for viral vectors, particularly for genes that are too large. Intriguingly, subretinal injection of DNA NPs loading the plasmid with S/MAR (scaffold matrix attachment region) led to the remarkableRPE65 expression in the rpe65(-/-) mouse model with LCA. Similarly, some studies have reported the efficacy of ocular gene therapy via subretinal delivery of compacted DNA NPs for nucleic acid expression in ocular disorders, such as retinal degenerative disease. In another study, a side-by-side comparison was made between subretinal delivery of wild-type rds gene loaded via compacted DNA NPs and naked DNA. Gene delivery by NPs promoted gene expression four times more and continued for up to four months compared to the control group. There are other reports regarding the good efficiency of lipid NPS (e.g. LPD) via sustained protein promoters (such as recoverin) and gene expression in animal models of visual impairment, such as blindness. Furthermore, intravitreal delivery of genes by solid lipid NPs (SLNs) via a special murine rhodopsin promoter (mOPS) facilitated structural improvement in the mouse model of retinoschisis (Rs1h-deficient). Sole et al. reported effective and safe subretinal therapy of Leber congenital amaurosis by administration of ECO/pDNA NPs into mouse RPE cells. NPs could be effectively used to deliver genes to stem cells that aid in the diagnosis and treatment of ocular diseases. For example, subretinal application of compact DNA NPs loading the mouse opsin promoter and wild-type Rds gene have shown promise in mouse models of retinitis pigmentosa. Yanai et al. reported superparamagnetic iron oxide NPs (SPIONs) that internalize rat mesenchymal stem cells for targeted delivery into the retina. There was a notable therapeutic outcome regarding this monitored intravitreal release as well as the concentration of anti-inflammatory agents in the retina. Mitra et al. evaluated the efficacy of loading the miR200-p plasmid CK30PEG10K (an antiangiogenic factor). There was a notable decrease in VEGFR-2 protein expression after intravitreal injection of materials into mouse models of diabetic retinopathy. Retinoblastoma (RB), as a rare pediatric ocular tumor, has also benefited from the application of NPs. There are several nanotechnology-based drug and gene delivery systems that illustrate great impacts for RB therapy. In Mitra et al. provided polyethyleneimine (PEI)-loaded gold NPS in conjugation with epithelial cell adhesion molecule (EpCAM) antibodies and siRNA molecules. There was a notable decrease in EpCAM expression in RB Y79 cells after application of combined GNPs via EpCAM antibody. The authors considered this method as a novel gene delivery, which was significantly internalized to cause cytotoxicity in cultured RB cells. There is a developing trend in gene therapy for several ocular diseases, such as primary open angel glaucoma, Stargardt disease, Leber congenital amaurosis, AMD, retinitis pigmentosa, and red-green ocular blindness. Despite these captivating results in this field, there is still much to be done for effective drugs and genes to achieve explicit cures [166, 167]. Considering these delivery methods, as a potential role in the efficacy of gene delivery, nanoparticle delivery systems could achieve promising results. Furthermore, considerable efforts have been made to overcome the problem of transient expression by nonviral gene delivery systems. NP for Pharmacological Therapeutic Purposes As mentioned above, the emergence of nanomedicine with nanotechnology has provided valuable consequences in ophthalmic pharmacotherapy. A.