Application of arbutin in inhibition of ocular surface injury

Abstract

The invention discloses application of arbutin in inhibition of ocular surface injury, and relates to the technical field of medicines. The invention aims to provide new application of arbutin. The arbutin is applied to relieving various ocular surface diseases mediated by inflammation and oxidative stress. The arbutin plays an antioxidant role through an SIRT1 / Nrf2 / HO-1 signal channel to relieve BAC-induced ocular surface injury and ocular surface inflammatory response, corneal limbal epithelial stem cells are protected from BAC damage, and corneal epithelial cell apoptosis is reduced at the same time. The arbutin has an obvious effect on treating ocular surface injury diseases caused by inflammation and oxidative stress.

Description

Technical Field

[0001] The invention relates to the technical field of medicines, and in particular to an application of arbutin in inhibiting ocular surface damage. Background Art

[0002] Ocular surface diseases refer to diseases that damage the structure and function of the ocular surface and are the most common ophthalmic diseases. Common ocular surface diseases in clinical practice mainly include infectious inflammation of the ocular surface (cornea and conjunctiva) (such as bacterial, viral, and chlamydia infections), dry eyes, allergic keratoconjunctivitis, and ocular surface trauma, which are the most common diseases in ophthalmology clinics. Ocular surface damage caused by various ocular surface, anterior segment, and posterior segment surgeries has also become a common ocular surface disease, seriously threatening the patient's visual health. Common drugs that can cause corneal and conjunctival toxicity include aminoglycosides, chloramphenicol, idoxuridine (IDU), adenosine, pilocarpine, beta-blockers, adrenaline, topical anesthetics, preservatives benzalkonium chloride (BAC) and thimerosal, etc. Glaucoma patients need to use eye drops containing preservatives for a long time, which leads to a variety of ocular surface problems including dry eyes. Among all the toxic effects of eye drops, the toxicity of preservatives is easily overlooked. Currently, the more commonly used preservatives include BAC and thimerosal. Among them, BAC is a quaternary ammonium cationic detergent with a surface active structure. It can not only act as a surfactant to dissolve ionic components into other incompatible solvents to promote the effective emulsification and stabilization of drugs, but also is superior to other preservatives in inhibiting microbial activity, so it is used as a preservative in eye drops. Some clinical studies in recent years have suggested that long-term use of eye drops containing preservatives may lead to a variety of ocular surface problems, including dry eye syndrome.

[0003] Arbutin (AR), also known as arbutin, scientific name hydroquinone glucoside, is a naturally occurring glycoside compound from green plants. It is a solute rich in many freeze-resistant plants such as wheat, pear, and bearberry of Ericaceae. It is white needle-shaped crystals or powder, easily soluble in hot water and ethanol. It is slightly soluble in cold water. Arbutin is widely used as a cosmetic skin whitening agent due to its strong inhibitory effect on tyrosine hydroxylation in the melanogenesis pathway. With the deepening of research and the further expansion of the research scope, more effects of arbutin have been highlighted, such as antioxidant, anti-inflammatory, antibacterial, antitussive, expectorant and antiasthmatic, anti-cancer and treatment of acute lung injury. Although arbutin has been confirmed to have anti-inflammatory and antioxidant properties in the study of many diseases and has significant therapeutic effects on the treatment of diseases, there are currently no reports on the use of arbutin as eye drops for ocular surface damage caused by inflammation and oxidative stress. Summary of the invention

[0004] The purpose of the present invention is to provide an application of arbutin in inhibiting ocular surface damage, which can be used to alleviate the ocular surface damage induced by BAC.

[0005] The purpose of the present invention can be achieved through the following technical solutions: An application of arbutin in inhibiting ocular surface damage.

[0006] As a further solution of the present invention: the application is the application of arbutin in the preparation of products for inhibiting ocular surface damage.

[0007] As a further embodiment of the present invention: the dosage forms of the product include tablets, eye drops, gels, eye ointments, injections, solutions, aerosols, and sprays.

[0008] As a further embodiment of the present invention: the effective concentration of the product for a single administration is 0-50 mg / mL.

[0009] As a further solution of the present invention: the optimal effective concentration of the product for a single administration is 5 mg / mL Beneficial effects of the present invention: The present application provides a new use of arbutin, which is to apply arbutin to a pharmaceutical product for alleviating a variety of ocular surface diseases mediated by inflammation and oxidative stress. The present application experimentally applies arbutin as eye drops to an experimental disease animal model, and for the first time verifies the therapeutic effect of arbutin in BAC-induced ocular surface damage.

[0010] The mechanism of action of arbutin in alleviating BAC-induced ocular surface damage and ocular surface inflammation by exerting anti-inflammatory and antioxidant effects through the SIRT1 / Nrf2 / HO-1 signaling pathway was further explored. Arbutin can protect limbal epithelial stem cells from BAC damage and alleviate BAC-induced ocular surface damage; at the same time, it reduces corneal epithelial cell apoptosis and alleviates BAC-induced ocular surface inflammation. Arbutin has a significant effect on the treatment of ocular surface damage diseases caused by inflammation and oxidative stress. BRIEF DESCRIPTION OF THE DRAWINGS

[0011] The present invention will be further described below in conjunction with the accompanying drawings.

[0012] Figure 1 This is a picture of corneal sodium fluorescein staining; Figure 2 It is a statistical chart of tear secretion test results; Figure 3 This is the corneal HE staining picture; Figure 4 This is the HE staining picture of the lacrimal gland; Figure 5 This is a PAS staining image of conjunctival goblet cells; Figure 6 This is the picture of meibomian gland oil red O staining; Figure 7 is the corneal P63 and TUNEL immunofluorescence staining; Figure 8 This is a statistical chart of the results of RT-PCR testing of inflammation-related indicators; Fig. 9 is the corneal CD45 / CD68 immunofluorescence co-staining image; Fig.10 This is the DHE immunofluorescence staining of cornea, conjunctiva and meibomian gland; Fig.11 It is a statistical chart of the ROS test results of cornea, conjunctiva and meibomian gland; Fig.12 This is a statistical chart of the results of RT-PCR detection of corneal oxidative stress related indicators; Fig.13 It is the verification of oxidative stress-related signaling pathways by corneal WB and the statistical chart of the results. DETAILED DESCRIPTION

[0013] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0014] (1) Experimental animals The experimental mice were C57BL / 6 female mice, aged 6-8 weeks, weighing 18-22 g (provided by Hunan Slake Jingda Laboratory Animal Co., Ltd., and the license number for use by the University of South China is SYXK (Xiang) 2020-0002).

[0015] (2) Experimental environment The experimental temperature was 25℃±1℃, the relative humidity was 55%±5%, the ventilation was fan-ventilated, the natural light was 12h / day, the cages were 8 per cage, and the cages were cleaned every three days. All studies were conducted in accordance with the Association for Research in Vision and Ophthalmology (ARVO) Statement on the Use of Animals in Ophthalmic and Vision Research.

[0016] (3) Experimental content The experiment was divided into four groups, with eight mice in each group. The ocular surface of all animals was evaluated before grouping to exclude individuals with ocular abnormalities. Four groups were set up, including the normal group (NT), model group (BAC+PBS), medication group (BAC+AR) and positive control group (BAC+CS). The mice in the normal group did not need any treatment, and the other three groups were induced with ocular surface damage by 0.075% benzalkonium chloride (BAC); the route of administration was eye drops; the number of administrations was 2 times / day; the eye drops were continued for 7 days; the administration volume was 5uL / eye, and the mouse ocular surface damage model was constructed.

[0017] The normal group (NT) mice did not need any treatment; the model group (BAC+PBS) was treated with PBS eye drops; the medication group (BAC+AR) was treated with 5mg AR dissolved in 1mL PBS as eye drops; the positive control group (BAC+CS) was treated with the purchased finished product 0.05% cyclosporine eye drops II (Zirun). The administration frequency was 4 times / day, and the eye drops were continued for 7 days; the administration volume was 5uL / eye. And it was performed at the same time as BAC eye drops.

[0018] Example 1 Effect of arbutin on the repair of ocular surface damage in mice See also Figure 1 , Seven days later, the ocular surface of the mice was stained with sodium fluorescein and photographed; See also Figure 2 , the mice were subjected to Schirmer test, and the wetted length of the test strip was recorded; See also Figure 3 , hematoxylin-eosin (HE) staining was used to evaluate the pathological changes of the cornea; See also Figure 4 , hematoxylin-eosin (HE) staining was used to evaluate the pathological changes of the lacrimal gland; See also Figure 5 , the number of conjunctival goblet cells was detected by periodic acid-Schiff staining (PAS staining); See also Figure 6 , Oil red O staining was used to detect lipid accumulation in the meibomian glands; The results showed that the tear secretion level and corneal epithelial barrier function were evaluated. Compared with the model group (BAC+PBS), the corneal fluorescein sodium staining in the drug group (BAC+AR) was significantly reduced, and arbutin could alleviate BAC-induced ocular surface damage. BAC can cause inflammatory cell infiltration in the cornea and lacrimal glands, significantly reduce the number of conjunctival goblet cells, but has no obvious effect on meibomian glands and tear secretion. Arbutin can reduce the infiltration of inflammatory cells and maintain the number of conjunctival goblet cells.

[0019] Example 2 Corneal P63 and TUNEL Immunofluorescence Staining See also Figure 7, indicating that arbutin can protect limbal epithelial stem cells from BAC destruction and reduce corneal epithelial cell apoptosis to maintain normal corneal epithelial cell barrier function.

[0020] Example 3 RT-PCR detection of inflammation-related gene expression and corneal CD45 / CD68 immunofluorescence co-staining See also Figure 8 , RT-PCR technique was used to detect the expression of inflammatory cytokines in the cornea; See also Fig. 9 Immunofluorescence co-staining was used to detect the expression of inflammatory cell surface specific markers CD45 and CD68 in corneal tissue; The results showed that arbutin could significantly reduce the infiltration of inflammatory cells and the expression of inflammatory factors in the cornea.

[0021] Example 4 RT-PCR detection of oxidative stress-related gene expression and ocular surface DHE immunofluorescence staining See also Fig.10 , DHE staining of ocular surface; See also Fig.11 , Expression of reactive oxygen species (ROS) in the cornea, conjunctiva, and meibomian glands; See also Fig.12 , RT-PCR was used to detect corneal oxidative stress related indicators; The results showed that arbutin could significantly reduce the production of reactive oxygen species and the expression of oxidative stress-related indicators in ocular surface tissues.

[0022] Example 5 Verification of signaling pathway See also Fig.13 , WB was used to verify the oxidative stress-related signaling pathways; The results showed that arbutin could alleviate BAC-induced ocular surface damage through its anti-inflammatory and antioxidant effects via the SIRT1 / Nrf2 / HO-1 signaling pathway.

[0023] The data in this experiment were processed using GraphPad Prism version 7.0 software (GraphPad Software, Inc., San Diego, California, USA). All data are presented as mean ± standard error. Statistical comparisons were performed using unpaired t-test or one-way analysis of variance (ANOVA). P values ​​< 0.05 were considered statistically significant.

[0024] The above is a detailed description of an embodiment of the present invention, but the content is only a preferred embodiment of the present invention and cannot be considered to limit the scope of implementation of the present invention. All equivalent changes and improvements made within the scope of the present invention should still fall within the scope of the patent coverage of the present invention.

Claims

1. An application of arbutin in inhibiting ocular surface damage.

2. The use of arbutin in inhibiting ocular surface damage according to claim 1, characterized in that: The application is the application of arbutin in preparing products for inhibiting ocular surface damage.

3. The use of arbutin in inhibiting ocular surface damage according to claim 2, characterized in that: The dosage forms of the product include tablets, eye drops, gels, eye ointments, injections, solutions, aerosols, and sprays.

4. The use of arbutin in inhibiting ocular surface damage according to any one of claims 2 or 3, characterized in that: The single administration concentration of the product is 0-50 mg / mL.

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