Eye bandages

Abstract

The present disclosure relates to an eye bandage (100, 100a) comprising a membrane (10) comprising at least silk fibroin obtained by a porogen and a crosslinking agent, and a central portion (20) that blocks light within a preset wavelength range, the central portion (20) being dimensioned relative to the membrane (10) such that there is at least an outer portion (30) of the membrane that is not covered by the central portion. The present disclosure also relates to a kit for implanting the eye bandage (100, 100a) into ocular tissue, and a method for implanting the eye bandage by optical coupling.

Description

[Technical field]

[0001] The present disclosure relates generally to the field of ophthalmology. [Background technology]

[0002] The cornea is the front-most lens of the eye. It has two main functions: it provides refractive power to the eye and it protects the internal structures of the eye. A clear and healthy cornea is essential for proper vision. The transparency and integrity of the cornea can be compromised by accidents involving abrasions, erosions, or chemical burns, which account for 10% of ophthalmic emergencies. The standard of care for treating an injured cornea is with eye drops and / or bandages. Eye drops are easily applied by the patient, but they are lost in tears and must be reapplied multiple times during treatment. Corneal bandages The use of bandages enhances and strengthens the cornea's healing ability. Potential drugs and / or growth factors are retained by the bandage in contact with the cornea, enhancing its effectiveness. Commercially available corneal bandages include bandage contact lenses. Contact lens and amniotic membrane There are two types:

[0003] Bandage contact lenses are a long-lasting method because they require prolonged contact lens wear on the injured cornea to induce corneal healing. Due to the mechanical properties of the bandage contact lens material and design, the bandage contact lens does not fully contact the injured cornea. Finally, the wrapping of the contact lens exposes the regenerating tissue to abrasive shear forces. Oxygen permeability is also a limiting factor for bandage contact lenses due to the fact that they must be worn for long periods and oxygen permeability must be high. Another disadvantage of bandage contact lenses is that manipulation by the patient poses a risk of corneal infection.

[0004] There are some problems with amniotic membrane. First, amniotic membrane is an allogeneic tissue. Amniotic membrane is derived from human donors, which means it can trigger an immune response in the patient and potentially lead to rejection. In addition, the amniotic membrane must be sutured to the cornea. It must also be stored at -80°C. And finally, it is donor-dependent, which makes it scarce and expensive.

[0005] In recent years, several approaches have been investigated with the aim of improving the current therapies available for treating patients. These approaches include the implantation of scaffolds to promote cell migration and proliferation in the cornea. These strategies consist of two main ones: implantation of the endothelium into the endothelium, which acts as a healing promoter and a barrier against external infection, and membrane grafts.

[0006] Scaffold implantation is based on the development of biocompatible materials that promote the migration and proliferation of fibroblasts and epithelial cells during the healing process.

[0007] In situ-formed collagen / polyethylene glycol hydrogels have been developed with good healing results. These hydrogels are formed directly onto the damaged cornea.

[0008] OcuPair® is an adhesive hydrogel for temporary corneal repair (up to 72 hours) and wound stabilization. It is composed of a biocompatible photocrosslinkable polyamidoamine dendrimer and a biodegradable photocrosslinkable hyaluronic acid. It is designed to be applied directly to the corneal wound and to initiate crosslinking under blue light irradiation.

[0009] These hydrogels likely function by forming a scaffold composed of collagen fibers. Corneal transparency requires a high degree of collagen organization, and it is difficult to reproduce the collagen packing, orientation, and weaving patterns of the natural, healthy cornea.

[0010] Membrane grafts are made by placing an ex-situ membrane on top of the cornea. In this study, ex situ membranes were obtained and typically affixed to the corneal surface during a surgical procedure. These ex situ membranes replace the current standard of care, the use of amniotic membrane, as previously mentioned.

[0011] Regarding membrane transplantation, there are several approaches currently under development.

[0012] One approach is to use cyclodextrin cross-linked collagen implants. The implant is a non-degradable preformed implant that requires corneal sutures and is intended to support re-epithelialization. Because the implant is non-degradable and cannot be removed, it is of utmost importance to fine-tune its shape and refractive index to prevent refractive index changes.

[0013] Other membrane approaches are based on collagen-PEG but require suturing to the cornea. The BIOcular® dressing overcomes this issue as it adheres to the cornea via a suture-free, light-activated mechanism. It is made of a thin, flexible chitosan membrane and is photoadhered to the ocular surface. Its suture-free approach and degradability overcome the shortcomings of previous approaches, but the apparent lack of biocompatibility and the complexity of the manufacturing process seem to make it unfeasible. Furthermore, to date, no in vivo experiments have been performed. Summary of the Invention [Problem to be solved by the invention]

[0014] The present disclosure provides a method for the preparation of silk fibroin for treating any type of corneal injury, including corneal abrasions. Fibroin-based eye bandages The present invention is intended to address the shortcomings of the prior art solutions by providing a bandage. [Means for solving the problem]

[0015] A first aspect of the present disclosure is an eye bandage comprising: A film comprising at least silk fibroin obtained by a porogen and a crosslinking agent; a central portion that blocks light within a preset wavelength range; The central portion is an eye bandage that is sized relative to the membrane such that there is at least an outwardly located portion of the membrane that is not covered by the central portion.

[0016] A second aspect of the present disclosure is an eye bandage, comprising: A membrane comprising at least silk fibroin and polyethylene glycol; a central portion that blocks light within a preset wavelength range; The central portion is an eye bandage that is sized relative to the membrane such that there is at least an outwardly located portion of the membrane that is not covered by the central portion.

[0017] In the present disclosure, polyethylene glycol may be used, although any other porogen and crosslinker may be used.

[0018] As a result, in any of the above-mentioned embodiments, the silk fibroin film is crosslinked and has pores. In any case, the central portion acts as a light blocking element at a preset wavelength, which has two functions: the central portion of the eye bandage does not optically couple with any other surface, and when the eye bandage is applied to the eye, the retina is protected from the light.

[0019] In certain embodiments, the same agent is used as the porogen and the crosslinker, as in the case of polyethylene glycol. It is also possible to use two different agents, one acting as the porogen and the other as the crosslinker, for example, polyethylene glycol and ethanol, respectively.

[0020] These outwardly located portions of the membrane not covered by the central portion are adapted to be adhered to ocular tissue, for example by photoadhesion.

[0021] The proposed eye bandage overcomes the shortcomings of existing solutions because:

[0022] -Silk fibroin is an inert material and has low immunogenicity, i.e. it does not induce an immune response, which is an expected property for any biomaterial. Furthermore, since silk fibroin is naturally produced, it is abundant and affordable compared to amniotic membrane.

[0023] -The silk fibroin membrane can be attached to the cornea by photobonding, eliminating the need for sutures.

[0024] -No expensive storage facilities are required.

[0025] The membrane can be cast with a certain amount of silk fibroin, which can range from 2% to 10% of the total volume, and the amount of porogen and crosslinker (such as polyethylene glycol) used can range between the minimum amount required to crosslink the membrane and the maximum amount that should not be exceeded to maintain a certain stiffness in the membrane. When using polyethylene glycol 300MW, the amount should be kept below 15% of the total volume to prevent the membrane from not having sufficient consistency. When using polyethylene glycol 3350MW, the membrane can be cast using 10% to 20% of the volume of polyethylene glycol (3350MW) in the total solution, and the 10% to 20% volume, in turn, has a concentration that can be kept between 7% to 55%, more preferably 10% to 50%.

[0026] In a particular embodiment, the membrane is obtained using about 2.5% to 3% silk fibroin and about 4% to 5% polyethylene glycol, based on the total volume of solution used to cast the membrane, which provides sufficient stiffness to the membrane while accelerating the healing of the wound to which the eye bandage is applied.

[0027] In certain embodiments, the outer portions of the membrane comprise a photoinitiator. For example, the outer portions may be coated with a photoinitiator. The photoinitiator is activatable by light, the light having a preset wavelength range. The photoinitiator may be a solution containing riboflavin or rose bengal.

[0028] In certain embodiments, not only the outer portions of the film comprise the photoinitiator, but other portions of the film or the entire film comprise the photoinitiator, and the central portion is configured to act as a mask and block light within a pre-set wavelength range, such that when light is applied, only the outer portions actually photocouple to other surfaces.

[0029] In certain embodiments, the central portion is made of a hydrophobic material, thus leaving the central portion undyed when the photoinitiator is delivered (e.g., by syringe or dropwise) to the outlying portions.

[0030] Also, because the silk fibroin membrane is porous, these outer portions of the membrane or the entire membrane can be configured to receive the photoinitiator. In such a case, the membrane can be immersed in the photoinitiator, with the central portion acting as a mask as previously described. In certain embodiments, the central portion is made of a flexible and / or semi-rigid material. Making the central portion from a flexible material facilitates the adaptation of the eye bandage to the ocular tissue. Making the central portion from a semi-rigid material (which typically provides increased stiffness relative to the membrane) facilitates the removal of the central portion once the eye bandage is in place and adhered to the ocular tissue.

[0031] In certain embodiments, the eye bandage further comprises a peripheral portion, which is also arranged to block light within a preset wavelength range. This further peripheral portion may be ring-shaped and has dimensions suitable to cover (and thus protect) the limbus. This peripheral portion may partially cover the membrane or may be dimensioned as a separate element that does not overlap the membrane.

[0032] In certain embodiments, the film further comprises growth factors to stimulate the growth of injured tissue and improve healing time. The film may also comprise a therapeutic agent for the prevention, treatment, or alleviation of symptoms of a disease or abnormal condition.

[0033] The disclosed eye bandage can have a thickness comprised between 5 μm and 150 μm for the same cast volume of silk fibroin membrane and the same shape of casting mold, depending on the porogen and crosslinker used while obtaining the silk fibroin membrane.

[0034] When 4%-5% polyethylene glycol (300 MW) is used, the resulting membrane thickness is 50 μm-150 μm. If the membrane is less than 50 μm thick, it becomes difficult to handle. Keeping the membrane thickness below 150 μm has the advantage that the membrane does not wrinkle when implanted into the cornea (especially in the peripheral area) and the implanted membrane does not cause discomfort to the end user.

[0035] However, with alternative porogens, film thicknesses as low as 5 μm can be engineered. For example, films can be cast using a 15% amount of polyethylene glycol (3350 MW) in the total solution, with the 15% amount having a concentration of 10-30% by weight PEG 3350 MW. In this case, the film thickness is preferably kept below 45 μm to prevent unwanted removal, e.g., by blinking, after the eye bandage is placed on the eye and photobonded.

[0036] The central part can be embodied as a sticker that is glued to the membrane. In this way, the central part can be easily removed after dyeing and / or after implantation. The tackiness of the sticker is preferably lower than that of the bond that is created after the eye bandage is implanted in the ocular tissue, facilitating the removal of the central part. It is also possible that the non-sticky surface of the central part is provided with some gripping means to allow the removal of the central part if the adhesion is too strong.

[0037] In some embodiments, the central portion completely covers at least the pupil of the eye.

[0038] In some embodiments, at least the outer portion of the membrane that is not covered by the central portion completely surrounds the central portion, and if the eye bandage is made in a circular shape, the outer portion of the membrane is a single peripheral ring or annular portion around the central portion.

[0039] An eye bandage such as that disclosed above can be used to treat corneal injuries.

[0040] Another aspect of the present disclosure is a kit for implanting an eye bandage into ocular tissue, comprising: Eye bandages; a photoinitiator for at least partially impregnating the corneal bandage; and a light source for providing light having a wavelength tuned to the excitation wavelength of the photoinitiator.

[0041] In certain embodiments, the eye bandage of the kit is as defined above with respect to the first or second aspect.

[0042] The kit includes a peripheral mask. The kit may further comprise a peripheral mask, the peripheral mask configured to block light within a pre-determined range of wavelengths and dimensioned to protect the limbus. In certain embodiments of the kit, the photoinitiator is a solution including riboflavin or rose bengal, and the light source provides light having ultraviolet, blue and / or green wavelengths.

[0043] A further aspect of the present disclosure relates to a method for obtaining an eye bandage, the method comprising: agitating the silk fibroin solution with polyethylene glycol; pouring the resulting stirred solution into a flat mold; and shaking the mold to ensure uniform distribution.

[0044] A further aspect of the present disclosure relates to a method of implanting an ophthalmic bandage onto an eye, the method comprising: dyeing a peripheral portion of an eye bandage with a photoinitiator, the eye bandage being according to the first aspect defined above; placing the eye bandage on the eye with the center portion facing up against the ocular surface; irradiating at least an outer portion of the eye bandage with light to photochemically bond the eye bandage to ocular tissue; and removing a central portion of the eye bandage.

[0045] In certain embodiments, the method further comprises the step of soaking the eye bandage in a growth factor solution.

[0046] In certain embodiments, the light is applied for less than 400 seconds. And, the step of applying light has a duration of 0.075 W / cm 2 These values ​​have been proven to be valid under in vitro conditions.

[0047] In other embodiments, particularly in in vivo conditions where blinking occurs, the optical coupling is about 0.15 W / cm 2 This is achieved with an illumination intensity of 100 nm and the light is applied for approximately 400 seconds.

[0048] In certain embodiments, the photoinitiator solution concentration is 0.01% (w / v) or less.

[0049] The different aspects and embodiments of the present disclosure defined above can be combined with each other, as long as they are compatible with each other.

[0050] Additional advantages and features of the present disclosure will become apparent from the following detailed description and are particularly pointed out in the appended claims.

[0051] To complete the description and to provide a better understanding of the present disclosure, a set of drawings are provided. The drawings form an integral part of this specification, illustrate embodiments thereof, and should not be interpreted as limiting or restricting the scope of the present disclosure, but merely as examples of how the present disclosure can be implemented. The drawings consist of the following figures: [Brief description of the drawings]

[0052] [Figure 1] FIG. 1 illustrates a possible embodiment of a corneal bandage according to the present disclosure. [Figure 2A] 2A and 2B show another possible embodiment of a corneal bandage. [Figure 2B] 2A and 2B show another possible embodiment of a corneal bandage. [Diagram 3] FIG. 3 shows the infrared spectra of silk fibroin and silk fibroin-PEG films. [Figure 4] Figure 4 shows the tensile force after irradiation with 0.075 W / cm2 and 10-2 % RB. [Diagram 5] Figure 5 shows the tensile force after irradiation with 0.075 W / cm2 and 10-3 % RB. [Figure 6] Figure 6 shows the tensile force after irradiation with 0.019 W / cm2 and 10-4 % RB. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0053] The following description is not to be taken in a limiting sense, but is given solely for the purpose of illustrating the broad principles of the present disclosure.Embodiments of the apparatus and method of the present disclosure are described, by way of example only, with reference to the accompanying drawings, in which:

[0054] Silk fibroin-based bandages Silk fibroin-based bandages (SF) have been developed with the aim of overcoming the shortcomings of previous techniques. In the last few years, silk fibroin-based biomaterials have been developed with applications ranging from biomedicine (tissue engineering, implants, drug delivery) to the food industry. Silk fibroin (SF) is extracted from the cocoons of silkworms. Raw silk contains fibroin and sericin.

[0055] It has been proven that silk fibroin, after extraction and purification, can be adapted into membranes, scaffolds, droplets, etc. with high biocompatibility, non-cytotoxicity, and low immunogenicity.

[0056] In one formulation of the present disclosure, polyethylene glycol (PEG) is added to silk fibroin to add porosity to increase the oxygen permeability of the resulting silk fibroin membrane. In this disclosure, the terms SF-based membrane and SF membrane are used interchangeably.

[0057] This SF-based membrane can be attached to ocular tissues, especially corneal tissues, in a suture-free manner. The silk fibroin bandage disclosed herein can be attached to the cornea by photobonding and can make full contact with the cornea. Furthermore, the SF membrane is a ductile material, adapting its shape to the corneal curvature and reducing potential abrasive shear forces. The degradation rate of the SF membrane can be tuned by chemical procedures in the manufacturing process. Control of the degradation rate allows the design of membranes ranging from dissolving in hours to days to high-retention membranes that function as removable corneal bandages.

[0058] Growth factors or drugs may also be administered with the bandage to improve wound healing.

[0059] The versatility of the photoadhesion process also allows for partial adhesion of the membrane (e.g., a peripheral ring can be used to create a central reservoir) and allows for control of adhesion strength to favor membrane peeling. These silk fibroin membranes are affordable compared to amniotic membranes and other manufactured corneas, and do not require expensive storage devices.

[0060] FIG. 1 is a schematic diagram of a corneal bandage according to the present disclosure. 1 shows a possible embodiment of a corneal bandage 100. The corneal bandage 100 comprises a silk fibroin membrane 10 and a mask 20, which in the illustrated embodiment is a circular black mask for blocking light. In this embodiment, the portion of the membrane not covered by the black mask is a peripheral ring 30.

[0061] 2A shows another possible embodiment of a corneal bandage 100a. In this case, the corneal bandage 100a comprises a silk fibroin membrane 10 and a mask 20a, which in the illustrated embodiment is black but oval in shape to block light. In this embodiment, there are two portions 30 of the membrane that are not covered by the black mask.

[0062] FIG. 2B shows yet another possible embodiment of a corneal bandage 100b. In this case, the corneal bandage 100b is made of a silk fibroin membrane 10, the shape of which is essentially rectangular with rounded edges. The corneal bandage 100b also comprises a mask 20b, which in the illustrated embodiment is black and also oval, for blocking light. In this embodiment, there are two portions 30a of the membrane 100b that are not covered by the black mask.

[0063] The bandages are made from a silk fibroin solution and polyethylene glycol, which are mixed together and poured into a flat mold at room temperature and allowed to evaporate.

[0064] The resulting ocular bandage can be photobonded to a wounded cornea, allowing healing of the bond, as demonstrated in a corneal wound healing model with fibroblast cultures. Wound healing occurred after 15 days in the presence of the silk fibroin bandage. Control samples were treated with no external adjuvant. No spontaneous healing of the cornea was observed.

[0065] Bandages can be manufactured in a variety of forms and have different administration and removal strategies.

[0066] In a possible embodiment, the eye bandage can be sealed to integrate with the cornea and allowed to resorb.

[0067] -Alternatively, the bandage may be removed once the cornea has healed.

[0068] In this approach, a silk fibroin eye bandage is photocoupled to the cornea using rose bengal (a photosensitizer) and green light illumination. The parameters of rose bengal and green light are adjusted to obtain optimal film photocoupling to the cornea with minimal photosensitizer concentration and light illuminance.

[0069] The biocompatibility of the proposed ocular bandage has been demonstrated in in vitro corneal fibroblast cultures.

[0070] Experimental Example Example 1 Silk fibroin (SF) was extracted from the silk cocoons of the silkworm, Bombyx mori. The cocoons were first cut into small pieces and then boiled for 0.2 h. The sericin was removed by degumming in 9.3 M Na2CO3 solution for 40 min. After degumming, the fibroin fibers were rinsed repeatedly with distilled water and dried overnight at 60 °C. The dried fibers were then resuspended in 9.3 M Na2CO3 solution for 40 min. The solution was dissolved in an aqueous LiBr solution at 60° C. for 4 hours. The final solution was then passed through a dialysis membrane (MWCO The silk fibroin was dialyzed against water using a 3.5 kDa filter (Spectrum) at 4 °C for 48 h. Finally, the resulting solution was centrifuged to remove impurities. The final concentration of silk fibroin in the aqueous solution was 7-8%.

[0071] To prepare the bandage (SF-PEG), 2.6% silk fibroin solution was mixed with 4.35% polyethylene glycol (300 MW) for 2 min. The resulting solution was poured into a flat mold with controlled temperature and humidity to allow the water to evaporate. A shaker was used to ensure uniform distribution in the membrane. PEG is a porogen, which generates pores in the membrane. Additionally, PEG induces β-sheets and cross-links the fibers, preventing the membrane from dissolving in water.

[0072] The presence of β-sheets is characterized by IR spectroscopy. Figure 3 shows the IR spectrum of a SF-PEG membrane and a control membrane made without PEG (SF). Three bands are characteristic of β-sheets.

[0073] The effects of degumming and crosslinking on membrane stability and degradation were investigated, specifically analyzing SF-PEG membranes in which silk fibroin had been extracted for 30 and 40 min.

[0074] The degradation rate of the SF-PEG membrane was evaluated in PBS, lysozyme (LS) at 500 U / mL, and protease XIV (PT) at 0.05 U / mL.

[0075] The degumming time did not affect the stability of the membrane. Based on this result and the results of in vitro experiments, all SF-PEG membranes were prepared with silk fibroin degummed for 40 min. All membranes degrade within the first 24 h in proteases.

[0076] Rabbit corneal strips were photobonded to membrane SF-PEG strips with different RB concentrations and green light irradiation with the aim of optimizing the concentration and irradiation parameters. Rabbit corneal strips measuring 15x3 mm were photobonded to SF-PEG membrane strips measuring 15x3 mm. Evaluation of adhesion strength was performed using uniaxial stretching. Adhesion strength was measured after irradiation with different doses and RB concentrations. The stretching strength of both RB-dyed and control samples (non-RB-dyed) varied with the irradiation time (Figures 4-6). Irradiated samples dyed with RB showed higher stretching strength than the control sample (C). Because the SF-PEG membrane is flexible, samples with higher illumination deformed or broke before the bond detached, as seen in the dashed bars.

[0077] Irradiation amount (0.019~0.075W / cm 2 ), irradiation time (0.25~6.6 min), RB concentration (10 -1 ~10 -4 The adhesion strength was examined by varying the irradiance and Rb concentration. The parameters of irradiance and Rb concentration were decreased until a threshold was reached at which the cornea and the membrane strip separated before the membrane was deformed. These parameters were maintained at 0.019 W / cm 2 and 10 -4% RB (Figure 5). Control samples for these parameters could not be measured since there was very little adhesion and it was not possible to mount the samples on the stretcher apparatus.

[0078] As part of this study, corneal stromal cells were transfected into silk fibroin-derived membranes (SFMs) after injury. The stroma cell response was evaluated.

[0079] To evaluate the effect of SFM on various processes occurring during stromal repair, a stromal in vitro wound model was used. Briefly, human corneal stromal cells (HCSCs) were seeded and cultured on a transparent SFM placed on the bottom of a Petri dish. When HCSCs reached confluence, a linear wound was created. A similar culture without SFM served as a control. Wound closure time and cellular organization were assessed microscopically. Proliferation and myofibroblast differentiation were observed. The differentiation of hepatocytes was analyzed by immunocytochemistry. Hepatocyte growth factor (HGF) secretion was measured by ELISA.

[0080] The silk fibroin-derived membrane induced significantly faster wound closure than the control, with complete closure by day 3. HCSCs showed greater organization on the substrate during wound closure than the control. Proliferation peaks were observed on day 1 in both groups, but were significantly increased in HCSCs seeded on SFM (p<0.001). On day 3, the rate of proliferation decreased in both groups, with no difference observed.

[0081] HGF secretion was significantly increased by SFM compared to controls on day 3 (p<0.05). No myofibroblast differentiation was observed in HCSCs seeded on SFM at any of the tested times, compared to the low percentage of myofibroblasts observed in controls on day 3.

[0082] This study showed a beneficial effect of SFM on stromal wound closure. SFM improved the corneal repair process and prevented the development of corneal opacity in terms of promoting early wound closure. The early wound closure could be attributed to an earlier peak in proliferation in the presence of SFM, but it also suggests that SFM supported the migration process. The absence of myofibroblasts in the presence of SFM could be due to the higher secretion of HGF, which has been shown to suppress myofibroblast generation in the corneal stroma.

[0083] Example 2 The following examples demonstrate that the disclosed ocular bandages can be secured to ocular tissue without sutures. In this example, a corneal bandage comprising a SF membrane can be secured to ocular tissue using a light-initiated fixation method such as photoconjugation. The disclosed method overcomes the potential graft rejection, opacity, scarcity, and need for suturing of amniotic membrane. The following examples demonstrate the optimization of this method and photoconjugation parameters in an in vitro and in vivo rabbit eye model.

[0084] Silk fibroin was extracted from silkworm cocoons with an aqueous solution. A solution of 2.6% SF and 4.35% polyethylene glycol (PEG) was cast at 25°C and 40% relative humidity. PEG acts as a cross-linking agent and a porosifying agent.

[0085] Fourier transform infrared spectroscopy (FTIR) was used to characterize the obtained SF films. The SF films were cut into 8x10mm strips, the corners were rounded, and immersed in 0.01% Rose Bengal photosensitizer for 10 min.

[0086] The experiment was carried out using a New Zealand rabbit model, in which four enucleated eyes were Irradiation was performed on corneal strips from 1000 mice (1000 eyes) and on seven eyes in vivo. Irradiation was performed with collimated 532 nm green laser light at 0.15 W / cm. 2 The exposure time was approximately 6.6 min. The ex vivo dyed membrane strips were placed on the corneal strips and exposed with the same exposure parameters as the in vivo SF membrane strips.

[0087] After 24 hours, uniaxial stretching was performed and the photoadhesion strength was measured. In vivo, the dyed membrane was placed on a vertical rectangular deep dissection area centered on the pupil of the same size. An opaque mask was used to cover the limbus and pupil, leaving a 2 mm margin on either side of the bandage for irradiation. Animal follow-up (0-15 days) included clinical symptoms, neovascularization, and membrane stability, adhesion, and transparency.

[0088] The membranes had thicknesses of 35-40 μm, Young's modulus of 10.4 ± 1.8 MPa, and were stable for over one month in PBS and 24 hours in protease XIV. On ex vivo corneas, the adhesion strength was 1.37 ± 0.24 N / cm. 2 In vivo, the SF membrane remained attached for more than 1 week in 85% of rabbits, and no signs of corneal edema were observed. The membrane was completely transparent 10 days after attachment and did not show RB in the non-irradiated areas.

[0089] This example also demonstrates that the SF-based corneal bandage is a suitable alternative to AM dressings and a suture-free photoadhesion paradigm that can be firmly fixed to the cornea both in vitro and in vivo, and the method has been proven to be safe and stable over the long term.

[0090] Example 3 This example demonstrates that growth factors can be administered through the disclosed ocular bandages, resulting in a biocompatible SF membrane that replaces current solutions. This example describes the development and characterization of a SF-based corneal bandage, with particular reference to the loading and release of growth factors in the SF membrane.

[0091] Silk fibroin was extracted from silkworm cocoons using standard methods. SF films were obtained by evaporation in a thermostatic chamber and cross-linked using porogen and a cross-linking agent such as polyethylene glycol (PEG).

[0092] Two types of growth factors (GFs) were used: epidermal growth factor (EGF) and hepatic growth factor (HGF).

[0093] GF was added to the SF membrane in two steps.

[0094] -SF membrane, 1 for EGF, 5, 10μg / ml, and 0.4, Immersion in 2 ml of 0.8 μg / ml at room temperature for 24 hours, and -Carbodiimide reaction Covalent binding of GF to SF membranes by (EDC).

[0095] For both procedures, 200 μl aliquots were taken every 24 hours. The aliquots were analyzed using human EGF and HGF enzyme-linked immunosorbent assay (ELISA) kits. Statistical differences (p<0.01) between the two procedures in terms of added GF and GF concentration were evaluated by ANOVA followed by parametric Bonferroni test for multiple comparisons.

[0096] The immersion method showed a more rapid release than the second method (using EDC) in all cases. With the immersion method, EGF was released at a rate of 6% / day during the first 2 days, 18% on the 5th day, and 15-20% on the 32nd day. HGF was released at a higher rate, with no measurable HGF content on the 22nd day. In contrast, with EDC, release was sustained longer in time, with rates of 3% (at 1 μg / ml), 1.5% (at 5 μg / ml), and 0.75% (at 10 μg / ml) for EGF, and 5% (at 0.4 μg / ml) and 3% (at 0.8 μg / ml) for HGF during the first 2 days.

[0097] The difference between the two procedures (immersion and EDC) was statistically significant for both EGF and HGF (p<0.001), and the difference between the concentrations was statistically significant for EFG (p=0.01) and HGF (p=0.01).

[0098] This example shows that SF bandages can be loaded with GFs, and both their release rate and release time can be tuned by the loading mode and GF concentration, aiming to replace amniotic membrane (AM)-based treatment of corneal wounds. As a result, GF-loaded SF bandages can provide an affordable alternative to amniotic membrane treatment.

[0099] All previous examples have demonstrated that ophthalmic bandages made of SF-based membranes are promising alternatives to amniotic membrane treatment in severe corneal injuries due to their proven advantageous properties, such as transparency, strength, possibility of adding and releasing growth factors, and ease of handling.

[0100] In this text, the term "comprise" and its derivatives (such as "comprising") should not be understood in an exclusive sense, i.e., these terms should not be interpreted as excluding the possibility that what is described and defined may include additional elements, steps, etc.

[0101] However, the present disclosure is clearly not limited to the particular embodiments described herein, but encompasses all modifications that may occur to those skilled in the art (e.g., with regard to selection of materials, dimensions, components, configurations, etc.) within the general scope of the present disclosure as defined by the claims.

Claims

1. An eye bandage (100, 100a, 100b), comprising: A film (10, 10a) containing at least silk fibroin obtained by a porogen and a crosslinking agent; a central portion (20, 20a, 20b) that blocks light within a preset wavelength range; An eye bandage, wherein the central portion (20, 20a, 20b) is dimensioned relative to the membrane (10, 10a) such that there is at least an outer portion (30, 30a) of the membrane that is not covered by the central portion.

2. An eye bandage (100, 100a, 100b), comprising: A membrane (10, 10a) comprising at least silk fibroin and polyethylene glycol; a central portion (20, 20a, 20b) that blocks light within a preset wavelength range; An eye bandage, wherein the central portion (20, 20a, 20b) is dimensioned relative to the membrane (10, 10a) such that there is at least an outer portion (30, 30a) of the membrane that is not covered by the central portion.

3. 3. An eye bandage according to any one of claims 1 to 2, wherein the membrane (10, 10a) is provided with holes.

4. An eye bandage (100, 100a, 100b) as described in any of the previous claims, wherein the outer portion (30, 30a) of the membrane (10, 10a) comprises a photoinitiator activatable by light, the light having the preset wavelength range.

5. An eye bandage (100, 100a, 100b) according to any of the previous claims, wherein said central portion (20, 20a, 20b) is made of a hydrophobic material.

6. An eye bandage (100, 100a, 100b) according to any of the previous claims, wherein said central portion (20, 20a, 20b) is made of a flexible and / or semi-rigid material.

7. 10. An eye bandage (100, 100a, 100b) according to any of the previous claims, wherein said film has a thickness, said thickness being dependent on the porogen and / or cross-linking agent used during obtaining or casting of said film.

8. 10. An eye bandage (100, 100a, 100b) according to any of the previous claims, wherein said membrane has a thickness between 5 μm and 150 μm.

9. An eye bandage (100, 100a, 100b) according to any of the previous claims, wherein the membrane (10, 10a) and the central portion (20, 20a, 20b) are adhered to each other.

10. 10. An eye bandage (100, 100a, 100b) according to any of the previous claims for use in treating corneal injuries.

11. An eye bandage (100, 100a, 100b) according to any of the previous claims, wherein said membrane (10, 10a) is loaded with growth factors, preferably by carbodiimide reaction.

12. A kit for implanting an eye bandage (100, 100a, 100b) into ocular tissue, comprising: Eye bandages (100, 100a, 100b) and a photoinitiator for at least partially impregnating the corneal bandage; and a light source for providing light of a wavelength tuned to the excitation wavelength of the photoinitiator.

13. 13. The kit of claim 12, wherein the eye bandage is as defined in any one of claims 1 to 11.

14. 13. The kit of claim 12, comprising a peripheral mask configured to block light within the preset wavelength range and dimensioned to protect the limbus.

15. 1. A method for implanting an eye bandage on an eye, comprising: i) dyeing a peripheral portion of an eye bandage with a photoinitiator, said eye bandage being as defined in any one of claims 1 to 11; ii) placing the eye bandage on the eye with the central portion facing up against the surface of the eye; iii) irradiating at least the outer portion of the eye bandage with light; and iv) removing the central portion from the eye bandage.

16. 16. The method of claim 15, further comprising the step of immersing the membrane (10) of the eye bandage (100, 100a, 100b) in a growth factor solution after i) and before ii).

17. The step of irradiating with light is performed at a light intensity of 0.15 W / cm 2 17. The method of any one of claims 13 to 16, wherein the method is carried out at an illumination intensity of less than 100 nm and / or the light is applied for less than 400 seconds.

18. 18. The method of any one of claims 13 to 17, wherein the photoinitiator solution concentration is 0.01% (w / v) or less.

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