Anti-inflammatory composition

Abstract

To provide an anti-inflammatory composition containing cholesterol.SOLUTION: According to one form, provided is an anti-inflammatory composition containing cholesterol as an active ingredient. In some forms, the anti-inflammatory composition inhibits epidermal inflammation. In some forms, the anti-inflammatory composition suppresses inflammation induced by light irradiation or skin stimulation. In another form, provided is a production inhibitor for at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα, containing cholesterol.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to an anti-inflammatory composition containing cholesterol. [Background technology]

[0002] Cholesterol is one of the components that form a lamellar structure in the stratum corneum of the epidermis, and plays an important role in maintaining the barrier function. It is believed that when the cholesterol synthesis ability in the epidermis decreases, the barrier function decreases and the recovery ability from rough skin decreases. Therefore, cholesterol is sometimes incorporated into cosmetics, quasi-drugs, medicines, etc., in the hope of maintaining and / or improving the barrier function of the skin and improving rough skin, etc. However, since cholesterol is a solid substance at room temperature and has extremely low hydrophilicity, the dosage form of the formulation to be incorporated is limited. Various formulations for stably incorporating cholesterol have been investigated (e.g., Patent Document 1). On the other hand, the physiological significance and functionality of external application of cholesterol have not been fully investigated, and the anti-inflammatory effect has not been known until now. [Prior art documents] [Patent documents]

[0003] [Patent Document 1] Patent Publication No. 2013-227293 Summary of the Invention [Problem to be solved by the invention]

[0004] An object of the present invention is to provide an anti-inflammatory composition containing cholesterol. [Means for solving the problem]

[0005] The present invention includes the following embodiments. [1] An anti-inflammatory composition containing cholesterol as an active ingredient. [2] The composition described in [1] that suppresses inflammation in the epidermis. [3] The composition described in [1] or [2], which suppresses inflammation caused by light exposure to the skin or irritation to the skin. [4] The composition according to any one of [1] to [3], which suppresses inflammation of the skin caused by exposure to ultraviolet rays. [5] The composition described in any of [1] to [4], which suppresses the production of at least one inflammatory cytokine selected from interleukin-1α (IL-1α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor (TNFα). [6] The composition described in [5], which suppresses the production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα in the epidermis. [7] The composition according to any one of [1] to [6], wherein the cholesterol content is 0.1 to 3 mass% based on the total amount of the composition. [8] The composition according to any one of [1] to [7], which is in the form of an aqueous composition. [9] The composition described in [8], further comprising a surfactant having a sterol skeleton.

[10] The composition described in [9], wherein the surfactant having a sterol skeleton includes at least one of polyoxyethylene cholesteryl ether and polyoxyethylene phytosteryl ether.

[11] The composition according to any one of [8] to

[10] , further comprising a polyol.

[12] The composition described in

[11] , wherein the polyol comprises a diol.

[13] The composition according to any one of [1] to

[12] , which is a cosmetic or a topical skin preparation.

[14] The composition according to any one of [1] to

[13] , wherein the composition is in the form of a mist, spray or lotion.

[15] The composition according to any one of [1] to

[14] , which is administered transdermally at a concentration of 0.001 to 0.01% by mass in terms of cholesterol.

[16] A cholesterol-containing inhibitor of the production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα. Effect of the Invention

[0006] According to the present invention, there is provided an anti-inflammatory composition containing cholesterol. The anti-inflammatory composition of the present invention can suppress inflammation (particularly inflammation in the epidermis). According to some embodiments, It is possible to suppress inflammation caused by exposure to or irritation of the skin from the environment, such as light irradiation (eg, ultraviolet irradiation) or skin irritation. The anti-inflammatory composition of the present invention contains as an active ingredient cholesterol, a substance found in the body that constitutes intercellular lipid layers and cell membranes, and therefore has high affinity with the body and is also excellent in safety. The anti-inflammatory composition of the present invention can be suitably used as a cosmetic, quasi-drug, pharmaceutical, etc. (for example, a cosmetic or topical skin preparation). [Brief description of the drawings]

[0007] [Figure 1] 1 shows the RNA concentration recovered from cells in Test Example 1. The vertical axis shows the RNA concentration recovered from a human epidermal keratinocyte cell line (HaCaT) under UVB irradiation (UVB(+), UVB(+)+Ch) or without irradiation (Non-treatment), while the numerical value above "UVB(+)+Ch" on the horizontal axis indicates the final concentration of cholesterol in the medium, and "-" above "UVB(+)" indicates the absence of addition of cholesterol aqueous emulsion. [Diagram 2] Figure 2 shows the results of microscopic photographs of cells in Test Example 1. The left photo is a photograph taken without UVB irradiation (UVB(-)) and without addition of cholesterol aqueous emulsion (Non-treatment), the center photo is a photograph taken with UVB irradiation (UVB(+)) and without addition of cholesterol aqueous emulsion (Non-treatment), and the right photo is a photograph taken with UVB irradiation (UVB(+)) and 0.05% concentration of cholesterol aqueous emulsion (Chaq). [Diagram 3]Fig. 3 shows the inhibitory effect of the cholesterol aqueous emulsion on the inflammatory response induced by UVB irradiation in HaCaT in Test Example 1. Fig. 3(a) to (c) show the expression levels (relative expression) of the IL-1α gene, IL-6 gene, and TNFα gene in HaCaT under UVB irradiation (UVB(+), UVB(+)+Ch) or without irradiation (Non-treatment). In Fig. 3(a) to Fig. 3(c), the numbers above "UVB(+)+Ch" on the horizontal axis indicate the final concentration of cholesterol in the medium, and "-" above "UVB(+)" or "Non-treatment" indicates the absence of addition of cholesterol aqueous emulsion. The non-treated group without UVB irradiation (Non-treatment) and the non-treated group with UVB irradiation (UVB(+)) were analyzed by Student's t-test, and the non-treated group with UVB irradiation (UVB(+)) and the group treated with cholesterol aqueous emulsion in addition to UVB irradiation (UVB(+)+Ch) were analyzed by Dunnett's test using the non-treated group with UVB irradiation (UVB(+)) as the control group. The significance level was 5%, *P<0.05, **P<0.01. [Figure 4] Figure 4 shows the inhibitory effect of cholesterol aqueous emulsion on the inflammatory response induced by a stimulant (SDS) in a skin model (three-dimensional epidermal culture model) in Test Example 2. Figures 4(a) and 4(b) show, from the left, the amount of IL-1α gene and IL-1β gene released in the untreated group, the SDS-exposed and cholesterol-free group, the SDS-exposed and cholesterol-added group before SDS exposure (pretreatment (Pre)), and the SDS-exposed and cholesterol-added group after SDS exposure (pretreatment (Post)). Student's t-test was used for the untreated group and the SDS-exposed and cholesterol-free group, and Dunnett's test was used for the SDS-exposed and cholesterol-added group (pretreatment and posttreatment) using the SDS-exposed and cholesterol-free group as the control group. *P<0.05, **P<0.01 when the significance level was 5%. [Diagram 5]FIG. 5 shows the anti-inflammatory effects of the components having a sterol skeleton (cholesterol, phytosterol, dipotassium glycyrrhizinate) in Test Example 3. FIG. 5(a)-(c) show the expression levels (relative expression) of the IL-1α gene, IL-6 gene, and TNFα gene in HaCaT cells without UVB irradiation (UVB-) or with UVB irradiation (UVB+). In FIG. 5(a)-FIG. 5(c), the numbers above "UVB+" on the horizontal axis indicate the final concentration of the component having a sterol skeleton in the medium, "-" above "UVB-" or "UVB+" indicates the absence of addition of the component having a sterol skeleton, "Chaq" indicates the addition of cholesterol aqueous emulsion 3, "Phy" indicates the addition of phytosterol aqueous emulsion, and "GK2" indicates the addition of dipotassium glycyrrhizinate aqueous solution. The groups treated with UVB irradiation plus Phy or GK2 (UVB(+)+Phy, UVB(+)+GK2) were compared with the group treated with UVB irradiation plus cholesterol aqueous emulsion (UVB(+)+Chaq) in a Dunnett's test. The significance level was set at 5%, *P<0.05, **P<0.01. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0008] Hereinafter, the embodiments of the present invention will be described in detail. Note that the present invention is not limited to the following embodiments, and can be modified and carried out as desired without departing from the gist of the present invention. The upper and lower limit values ​​of the numerical ranges described in this specification can be arbitrarily combined. For example, when "X to Y" and "Z to W" are described, the ranges of "X to W" and "Z to Y" are also included in the scope of the present invention as numerical ranges. In addition, the numerical range "lower limit value to upper limit value" described in this specification means not less than the lower limit value and not more than the upper limit value, unless otherwise specified.

[0009] <<Anti-inflammatory composition>> One aspect of the present invention relates to an anti-inflammatory composition containing cholesterol (hereinafter also referred to as "the anti-inflammatory composition of the present invention" or "anti-inflammatory composition"). The anti-inflammatory composition of the present invention contains cholesterol as an active ingredient.

[0010] (Anti-inflammatory effect) The present inventors have found that cholesterol suppresses the production of inflammatory cytokines such as interleukin-1α (IL-1α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor (TNFα). IL-1α, IL-1β, IL-6, and TNFα are major proinflammatory cytokines, and it is known that abnormal or excessive signaling by cytokines causes various inflammatory symptoms in vivo (Reference: MD Turner et al., Biochimica et Biophysica Acta 1843 (2014) 2563-2582). In the present invention, cholesterol can be used as an inhibitor of the production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα, and is therefore useful as an anti-inflammatory agent. In some embodiments, cholesterol can be used as an inhibitor of the production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα in the epidermis. In some embodiments, cholesterol can be used as an inhibitor of the production of at least one inflammatory cytokine selected from IL-6 and TNFα. In one embodiment, cholesterol can be used as an inhibitor of the production of at least one inflammatory cytokine selected from IL-6 and TNFα in the epidermis. Cholesterol is a substance with extremely low hydrophilicity, so the dosage form of the preparation is limited, and although the function of cholesterol has been studied in terms of endogenous cholesterol, the physiological significance and functionality of external application of cholesterol have not been fully studied. The anti-inflammatory effect of cholesterol mentioned above was discovered for the first time by the present invention.

[0011] As used herein, "inflammation" includes local inflammation and systemic inflammation, and symptoms and conditions of "inflammation" include the four major signs of inflammation, namely pain, redness, heat and swelling; as well as various secondary symptoms induced thereby, for example, inflammatory symptoms in inflammatory diseases such as inflammatory dermatitis, gastritis and ulcerative colitis, arthritis such as rheumatoid arthritis and degenerative osteoarthritis, and various allergic inflammatory diseases such as hay fever, allergic rhinitis, allergic conjunctivitis and atopic dermatitis. In some embodiments, the "inflammation" is inflammation involving at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα. As used herein, "anti-inflammatory agent" refers to an agent that is effective in preventing, treating, improving, or alleviating an inflammatory symptom or condition. In some embodiments, the anti-inflammatory agent prevents, treats, improves, or alleviates an inflammatory symptom or condition in the skin. In some embodiments, the anti-inflammatory agent treats, improves, or alleviates an inflammatory symptom or condition in the skin. In some embodiments, the anti-inflammatory agent is effective against inflammation caused by light irradiation of the skin or irritation of the skin. In some embodiments, the anti-inflammatory agent is effective against inflammation caused by light irradiation of the skin. In some embodiments, the anti-inflammatory agent is effective against inflammation caused by ultraviolet light exposure of the skin.

[0012] The term "anti-inflammatory composition" refers to a composition that has an anti-inflammatory effect or can be used as an anti-inflammatory agent. The present invention can suppress inflammation by using an anti-inflammatory composition containing cholesterol. In some embodiments, the anti-inflammatory composition suppresses inflammation in the skin (particularly inflammation in the epidermis). In some embodiments, the anti-inflammatory composition suppresses inflammation caused by light irradiation of the skin (e.g., visible light, ultraviolet light (ultraviolet A rays (UVA), ultraviolet B rays (UVB))), etc.) or irritation to the skin. In some embodiments, the anti-inflammatory composition suppresses inflammation caused by light irradiation of the skin. In one embodiment, the anti-inflammatory composition suppresses inflammation caused by ultraviolet light exposure of the skin. In some embodiments, the anti-inflammatory composition suppresses inflammation caused by skin irritation. Examples of skin irritation include irritation caused by chemical substances (eg, sodium dodecyl sulfate (SDS) etc.), house dust, medicines, and allergens such as bacteria and viruses.

[0013] In some embodiments, the anti-inflammatory agent or anti-inflammatory composition is administered after light irradiation of the skin (e.g., visible light, ultraviolet light (ultraviolet A rays (UVA), ultraviolet B rays (UVB))), etc.) or stimulation of the skin. By administering after light irradiation of the skin or stimulation of the skin, inflammation caused by light irradiation of the skin or stimulation of the skin can be significantly suppressed.

[0014] In some embodiments, the anti-inflammatory agent or anti-inflammatory composition suppresses the production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα in the epidermis (particularly, human epidermal cells). In some embodiments, the anti-inflammatory composition suppresses the production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα in the epidermis (particularly, human epidermal cells). In some embodiments, the anti-inflammatory composition suppresses the production of at least one inflammatory cytokine selected from IL-6 and TNFα. In one embodiment, the anti-inflammatory composition suppresses the production of at least one inflammatory cytokine selected from IL-6 and TNFα in the epidermis (particularly, human epidermal cells).

[0015] In some embodiments, the anti-inflammatory agent or anti-inflammatory composition reduces the gene expression level of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα, compared to the expression level in a control (cholesterol not added). For example, the anti-inflammatory agent or anti-inflammatory composition preferably reduces the gene expression level of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα in human epidermal cells to preferably 90% or less, more preferably 80% or less, even more preferably 70% or less, and particularly preferably 50% or less, compared to the expression level in a control (cholesterol not added). The expression level of the inflammatory cytokine gene in human epidermal cells can be measured according to the method described in the Examples below.

[0016] In some embodiments, the anti-inflammatory agent or anti-inflammatory composition reduces the protein production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα, compared to the production amount in a control (cholesterol not added). For example, the anti-inflammatory agent or anti-inflammatory composition preferably reduces the protein production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα in human epidermal cells to preferably 90% or less, more preferably 80% or less, even more preferably 70% or less, and particularly preferably 50% or less, compared to the production amount in a control (cholesterol not added). The amount of inflammatory cytokine protein in human epidermal cells can be measured according to the method described in the Examples below.

[0017] (Active ingredient) The method for obtaining the cholesterol used in the present invention is not particularly limited, and a commercially available product or one produced according to a known method may be used. The anti-inflammatory composition of the present invention contains as an active ingredient cholesterol, a substance found in the body that constitutes intercellular lipid layers and cell membranes, and therefore has high affinity with the body and is also excellent in safety.

[0018] (formulation) The anti-inflammatory composition of the present invention can be suitably used in cosmetics, quasi-drugs, pharmaceuticals, or foods and beverages.

[0019] The form of the anti-inflammatory composition is not particularly limited as long as it contains cholesterol. The anti-inflammatory composition can be, for example, liquid, solid, or semi-solid. The dosage form of the anti-inflammatory composition is also not particularly limited. When the anti-inflammatory composition is liquid, it can be, for example, a dispersion (for example, in the form of a mist, spray, lotion, liquid, etc.), when it is solid, it can be, for example, a powder, pellet, etc., and when it is semi-solid, it can be, for example, a paste, cream, mousse, etc.

[0020] The formulation of the anti-inflammatory composition can be produced by known production techniques according to the desired dosage form. The composition of the present invention can be blended with known excipients, disintegrants, binders, lubricants, stabilizers, preservatives, colorants, surfactants, pH adjusters, flavorings, flavorings, sweeteners, preservatives, fragrances, aqueous components, alcohols, sugars, water, etc. as additives, if necessary. In addition, anti-aging agents, antioxidants, skin-whitening agents, various herbal medicines, etc. can also be blended within a range that does not weaken the effect of the composition of the present invention.

[0021] The anti-inflammatory composition of the present invention can be used in parenteral or oral dosage forms. Examples of the form of a composition for parenteral administration include ointments, plasters, liquids, lotions, creams, gels, emulsions, aerosols, liniments, tapes, poultices, powders for external use, injections, drops, packs, bath agents, and the like, and these can be administered as medicines, quasi-drugs, cosmetics, and the like. Examples of the form of the orally administered composition include tablets, capsules, powders, fine granules, pills, liquids, syrups, lozenges, jellies, etc., and these can be administered as medicines, quasi-drugs, foods and beverages, etc. The anti-inflammatory composition is preferably a parenteral administration composition. Among them, the anti-inflammatory composition can be used as cosmetics for various applications. Examples of such cosmetics include skin care cosmetics such as lotion, milky lotion, cream, beauty essence, massage agent, pack agent, hand cream, body lotion, body cream, and sunscreen; finishing cosmetics such as foundation, eye shadow, eye liner, mascara, eyebrow, concealer, lipstick, and lip balm; hair mists; and hair cosmetics such as treatments and hair creams. Examples of the method of use include using the composition on hands, fingers, or cotton, or impregnating nonwoven fabrics or the like. The anti-inflammatory composition can be used as a topical composition. Examples of the composition include topical liquids, topical gels, creams, ointments, liniments, lotions, poultices, plasters, sprays, aerosols, etc. The method of use can be the same as that of the cosmetic preparations. In some embodiments, the topical composition can be used as a skin topical drug, skin topical quasi-drug, or skin cosmetic.

[0022] The content of cholesterol in the anti-inflammatory composition is not particularly limited, and may be appropriately determined taking into consideration the user's sex, age, symptoms, administration method, number of administrations, administration time, dosage form, etc. For example, the content of cholesterol in the anti-inflammatory composition is preferably 0.01 to 1 mass% relative to the total amount (total mass) of the composition, more preferably 0.05 to 1 mass%, and even more preferably 0.1 to 1 mass%, and may be 0.01 to 0.05 mass% or 0.05 to 0.1 mass%. In a specific embodiment, the cholesterol content in the anti-inflammatory composition when used as a topical composition is preferably 0.01 to 1 mass%, more preferably 0.05 to 1 mass%, and even more preferably 0.1 to 1 mass%, or may be 0.01 to 0.05 mass%, or may be 0.05 to 0.1 mass%, relative to the total amount (total mass) of the composition.

[0023] The daily amount of the anti-inflammatory composition to be used may be appropriately determined in consideration of the user's sex, age, symptoms, administration method, number of administrations, administration time, dosage form, etc., and may be used within a range that does not cause side effects, and is not particularly limited. For example, in some embodiments, the amount of cholesterol administered (applied) when used as an external composition is 10 to 500 mg per day for an adult, preferably 50 to 500 mg per day, more preferably 100 to 500 mg per day, and is applied 1 to 6 times per day, preferably 1 to 3 times per day. The amount of cholesterol administered (applied) when used as an external composition may be 10 mg to 50 mg or 50 to 100 mg per day for an adult.

[0024] In some embodiments, the anti-inflammatory composition is administered transdermally at a cholesterol concentration of 0.001 to 0.01% by mass as the ratio of cholesterol. That is, it is preferable to administer the composition so that the ratio of cholesterol in the epidermis is 0.001 to 0.01% by mass. If the concentration is within this range, it is possible to effectively obtain the anti-inflammatory effect.

[0025] (aqueous composition) In some embodiments, the anti-inflammatory composition is in the form of an aqueous composition. By formulating cholesterol in the form of an aqueous composition, for example, when used as a topical composition, the permeability of cholesterol into the epidermis can be improved, thereby enhancing the anti-inflammatory effect. As used herein, the term "aqueous composition" refers to a composition that contains at least water, and the external phase of the composition is an aqueous component.

[0026] In some embodiments, the aqueous composition comprises cholesterol, water, and a surfactant having a sterol skeleton. By blending cholesterol with a surfactant having a sterol skeleton, a composition in which cholesterol is stably dispersed in an aqueous medium containing water can be obtained, and the composition has advantages such as better permeability and lower viscosity than an ointment composition, making it possible to apply the composition to a wide range of areas. The form of the aqueous composition will be described below.

[0027] (a) Cholesterol The cholesterol contained in the aqueous composition is preferably 0.1 to 3% by mass based on the total amount (total mass) of the composition. Within the above range, cholesterol can be stably incorporated into the aqueous medium. In addition, when applied externally, it can exhibit an anti-inflammatory effect, and can be suitably used, for example, as a cosmetic or skin external preparation. In general, cholesterol tends to precipitate or become cloudy in an aqueous medium as the cholesterol concentration increases. Therefore, from the viewpoint of the stability of the composition (reduction or suppression of cholesterol precipitates and cloudiness), it is more preferably 0.1 to 2% by mass, even more preferably 0.1 to 1.5% by mass, and particularly preferably 0.1 to 1.0% by mass.

[0028] (b) Surfactants with a sterol skeleton The aqueous composition preferably contains a surfactant having a sterol skeleton. By containing a surfactant having a sterol skeleton together with cholesterol, cholesterol can be stably dispersed in water, and thus a composition having excellent transparency and reduced or inhibited precipitation or clouding of cholesterol can be obtained.

[0029] A surfactant having a sterol skeleton is a surfactant having a sterol skeleton in a lipophilic group (hydrophobic part). The sterol skeleton is an alcohol having a hydroxyl group bonded to the 3-position of a cyclopentanohydrophenanthrene ring shown in the following formula, and a surfactant having a sterol skeleton is a compound having a hydrophilic group bonded to this sterol skeleton. In addition, the symbols A to D in the following formula represent each ring constituting the cyclopentanohydrophenanthrene ring, and the numbers represent the position numbers of the carbons constituting the cyclopentanohydrophenanthrene ring. The sterol skeleton of the lipophilic group may have a double bond in the cyclopentanohydrophenanthrene ring.

[0030] [ka]

[0031] Among them, sterols having a double bond in the B ring or stanols composed of a saturated ring are preferred.Specific examples include phytosterols such as β-sitosterol, campesterol, stigmasterol, brassicasterol, and the like, or stanols thereof, cholesterol, cholestanol, lanosterol, ergosterol, and the like, and are preferably phytosterols, phytostanols, cholesterol, or cholestanol, and more preferably phytosterol or cholesterol.

[0032] The hydrophilic group is not particularly limited, and examples thereof include nonionic hydrophilic groups such as polyoxyalkylene chains (e.g., polyoxyethylene (POE) chains, polyoxypropylene (POP) chains, etc.), polyglycerin, polyglucosides, sorbitan, and sucrose; anionic hydrophilic groups such as carboxylates, sulfonates, sulfates, and phosphates; and cationic hydrophilic groups such as aliphatic amine salts and quaternary ammonium salts. The alkylene in the polyoxyalkylene chain preferably has 1 to 4 carbon atoms, more preferably has 1 or 2 carbon atoms, and particularly preferably is ethylene. Among these, the hydrophilic group is preferably a nonionic hydrophilic group, more preferably a polyoxyalkylene chain, and even more preferably a polyoxyethylene (POE) chain. The number of moles of polyoxyalkylene chains (preferably polyoxyethylene (POE) chains) added is not particularly limited, but is preferably 12 or more, more preferably 15 or more, and even more preferably 18 or more. The number of moles of polyoxyalkylene chains (preferably polyoxyethylene (POE) chains) added is preferably 25 or less, more preferably 24 or less, and even more preferably 22 or less. In some embodiments, the number of moles of polyoxyalkylene chains (preferably polyoxyethylene (POE)) added is preferably 12 to 25, more preferably 15 to 24, and even more preferably 18 to 22.

[0033] The surfactant having a sterol skeleton is, for example, a surfactant in which a polyoxyalkylene is ether-bonded to the sterol skeleton. More specifically, the surfactant may be, but is not limited to, polyoxyalkylene cholesteryl ether (e.g., POE cholesteryl ether, POP cholesteryl ether), polyoxyalkylene phytosteryl ether (e.g., POE phytosteryl ether, POP phytosteryl ether), polyoxyalkylene cholestanol (e.g., POE cholestanol, POP cholestanol), polyoxyalkylene phytostanol (e.g., POE phytostanol, POP phytostanol), etc. Among them, the surfactant having a sterol skeleton is preferably at least one of polyoxyethylene cholesteryl ether and polyoxyethylene phytosteryl ether in terms of transparency and stability. Furthermore, POE (12-25, more preferably 15-24, even more preferably 18-22) cholesterol and POE (12-25, more preferably 15-24, even more preferably 18-22) phytosteryl ether are preferred.

[0034] The HLB (Hydrophilic-Lipophilic Balance) of the surfactant having a sterol skeleton is not particularly limited, but is preferably 11 or more, more preferably 12 or more, from the viewpoint of improving transparency. The HLB of the surfactant having a sterol skeleton is not particularly limited, but is preferably 16 or less, more preferably 15 or less. The HLB of the surfactant having a sterol skeleton is preferably in the range of 11 to 16, more preferably 12 to 15. In this specification, unless otherwise specified, the HLB value is experimentally determined by comparing the emulsifying power with that of a standard sample.

[0035] The surfactant having a sterol skeleton may be any surfactant having the above-mentioned characteristics, and may be a commercially available product. Examples of commercially available products include, but are not limited to, EMALEX CS series (EMALEX CS-15, EMALEX CS-20, EMALEX CS-24, etc.) manufactured by Nippon Emulsion Co., Ltd. and NIKKOL BPS series (NIKKOL BPS-20, etc.) manufactured by Nikko Chemicals Co., Ltd.

[0036] These surfactants having a sterol skeleton can be used alone or in combination of two or more.

[0037] The content of the surfactant having a sterol skeleton in the aqueous composition is not particularly limited, but from the viewpoint of the feeling upon use, it is preferably 0.5 to 10% by mass based on the total amount of the composition. Moreover, from the viewpoint of reducing or inhibiting turbidity or precipitation of cholesterol in an aqueous medium and stably dispersing cholesterol, the mass ratio of the surfactant having a sterol skeleton to cholesterol (surfactant having a sterol skeleton / cholesterol) is preferably 3 to 10, more preferably 4 to 10, further preferably 5 to 10, and particularly preferably 6 to 10. In some embodiments, the mass ratio (surfactant having a sterol skeleton / cholesterol) may be 4 to 8, or 5 to 9, or 6 to 8.

[0038] (c) water The water is not particularly limited, and for example, tap water, ion-exchanged water, distilled water, purified water, etc. can be used.

[0039] The content of water in the aqueous composition is not particularly limited, but from the viewpoint of transparency, it is preferably 60% by mass or more, more preferably 70% by mass or more, and even more preferably 80% by mass or more, based on the total amount of the composition. The upper limit is also not particularly limited, but is, for example, 99.5% by mass or less, or 97% by mass or less, or 95% by mass or less. In some embodiments, the content of water in the aqueous composition is preferably 60 to 99.5% by mass, more preferably 70 to 97% by mass, and even more preferably 80 to 95% by mass, based on the total amount of the composition. Furthermore, the mass ratio of cholesterol to water (cholesterol:water) is preferably from 0.1:99.9 to 3:97, more preferably from 0.1:99.9 to 2:98, and even more preferably from 0.1:99.9 to 1.5:98.5, from the viewpoints of transparency and stability of the composition. The aqueous composition may contain other aqueous components in addition to water. In terms of transparency, the proportion of water in the aqueous component is preferably 60% by mass or more, more preferably 70% by mass or more, and even more preferably 80% by mass or more.

[0040] In some embodiments, the main component of an aqueous composition is water and is a liquid.

[0041] (d) Other ingredients (aqueous component) The aqueous composition may contain a polyol. The polyol is amphiphilic, and the incorporation of the polyol can improve the dispersibility of cholesterol in water, thereby improving the transparency of the composition. The polyol is not particularly limited as long as it is one that is generally used in cosmetics. For example, propylene glycol, dipropylene glycol, 1,3-propanediol, 1,3-butylene glycol, isopentyl diol, pentylene glycol, hexanediol, caprylyl glycol, glycerin, diglycerin, polyglycerin, polyethylene glycol, polyoxyethylene methyl glucoside, etc. are included. Among them, from the viewpoint of improving transparency, it is preferable that the aqueous composition contains a diol (dihydric alcohol), such as propylene glycol, 1,3-propanediol, 1,3-butylene glycol, pentylene glycol, hexanediol, etc. The polyols may be used alone or in combination of two or more kinds.

[0042] The content of the polyol is not particularly limited, but from the viewpoint of transparency and stability, it is preferably 0.1% by mass or more, more preferably 1% by mass or more, and even more preferably 3% by mass or more, based on the total amount of the composition. The upper limit is also not particularly limited, but for example, it is preferably 10% by mass or less, more preferably 9% by mass or less, and even more preferably 8% by mass or less. In some embodiments, the content of the polyol in the aqueous composition is preferably 0.1 to 10% by mass, more preferably 1 to 9% by mass, and even more preferably 3 to 8% by mass, based on the total amount of the composition. The polyol is an aqueous component and, together with water, forms the external phase of the composition.

[0043] The aqueous composition may contain other aqueous components, such as wetting agents, natural polymers, synthetic polymers, preservatives, lower monoalcohols, etc. These may be used alone or in combination of two or more. In this specification, the aqueous component does not include a surfactant.

[0044] The humectant is not particularly limited, but examples thereof include sugars such as sorbitol, mannitol, glucose, xylitol, maltitol, and trehalose; amino acids such as arginine, proline, serine, lysine HCl, threonine, valine, leucine, histidine, alanine, glycine, and carnosine; sodium pyrrolidone carboxylate, betaine, urea, sodium lactate, plant extracts, and seaweed extracts.

[0045] Other aqueous components that may be used include, but are not limited to, natural polymers such as guar gum, carrageenan, xanthan gum, sodium hyaluronate, pullulan, agar, and alginates; synthetic polymers such as hydroxyethyl cellulose, hydroxymethyl cellulose, carboxyvinyl polymer, polyacrylates, polyvinyl alcohol, and acrylic acid / alkyl methacrylate copolymers; and preservatives such as phenoxyethanol.

[0046] The aqueous component may contain a lower monoalcohol, for example, a primary alcohol having 1 to 5 carbon atoms, preferably ethanol. The content of the lower monoalcohol as the aqueous component is preferably small from the viewpoint of safety. The content of the lower monoalcohol is preferably 0 to 5% by mass, more preferably 0 to 3% by mass, and further preferably 0 to 1% by mass, based on the total amount of the composition. In one embodiment, the aqueous composition does not contain a lower monoalcohol. In addition, when a lower monoalcohol is contained, the mass ratio of the lower monoalcohol to (c) water (lower monoalcohol / water) is preferably 0.3 or less, more preferably 0.2 or less, and even more preferably 0.1 or less.

[0047] (Oily ingredients) The aqueous composition may contain an oily component other than cholesterol. In this specification, the oil component means a component generally used as an oil component in the field of cosmetics. For example, it means a component having a solubility in water of less than 0.1% by mass (less than 1 g / L) at 25°C. In this specification, the oil component does not include surfactants.

[0048] In the case of an aqueous composition, if the amount of oily components other than cholesterol is large, it may adversely affect the skin permeability, transparency, stability, etc. Therefore, cholesterol is preferably 95% by mass or more of the total mass of the oily components, more preferably 98% by mass or more, and even more preferably substantially 100% by mass. In some embodiments, the aqueous composition is substantially free of oily components other than cholesterol (such as non-water-soluble solvents). In some embodiments, the aqueous composition is free of oily components other than cholesterol. By configuring the composition to contain a small amount or no oily components other than cholesterol, transparency and stability can be improved.

[0049] The oily component other than cholesterol is not particularly limited, but examples thereof include higher monoalcohols, hydrocarbons, ester oils, fats and oils, fatty acids, waxes, silicone oils, etc. These may be used alone or in combination of two or more.

[0050] The higher monoalcohol is not particularly limited, but is preferably a straight-chained one, more preferably a straight-chained higher monoalcohol having 12 to 22 carbon atoms (more preferably 14 to 22 carbon atoms, and even more preferably 16 to 22 carbon atoms). Specific examples include cetyl alcohol, stearyl alcohol, cetostearyl alcohol, behenyl alcohol, and the like. The hydrocarbon is not particularly limited, and examples thereof include liquid paraffin, squalane, α-olefin oligomer, petrolatum, and microcrystalline wax. The ester oil is not particularly limited, and examples thereof include cetyl 2-ethylhexanoate, 2-ethylhexyl palmitate, 2-hexyldecyl 2-ethylhexanoate, 2-octyldodecyl myristate, 2-hexyldecyl myristate, isononyl isononanoate, 2-ethylhexyl isononanoate, isodecyl pivalate, isostearyl pivalate, 2-hexyldecyl stearate, isostearyl isostearate, isopropyl isostearate, pentaerythritol tetraisostearate, glyceryl tri-2-ethylhexanoate, caprylic / capric triglyceride, myristyl myristate, and cetyl palmitate. The oils and fats are not particularly limited, and examples thereof include olive oil, shea butter, grape seed oil, and the like. The wax is not particularly limited, and examples thereof include jojoba oil, beeswax, candelilla wax, and the like. The fatty acid is not particularly limited, and examples thereof include myristic acid, palmitic acid, and stearic acid.

[0051] In some embodiments, the aqueous composition is free of higher monoalcohols. In some embodiments, the aqueous composition is free of hydrocarbons. In some embodiments, the aqueous composition is free of ester oils. In some embodiments, the aqueous composition is free of fats and oils. In some embodiments, the aqueous composition is free of fatty acids. In some embodiments, the aqueous composition is free of waxes. In some embodiments, the aqueous composition is free of silicone oils.

[0052] (Other Surfactants) The aqueous composition may contain surfactants other than the surfactant having a sterol skeleton (other surfactants) as necessary. Other surfactants are not particularly limited, but may be, for example, alkyl ether surfactants such as polyoxyethylene alkyl ether and polyoxyethylene polyoxypropylene alkyl ether; polyoxyethylene fatty acid ester; sorbitan fatty acid esters such as polyoxyethylene sorbitan fatty acid ester and monopalmitic acid sorbitan; polyoxyethylene castor oil, polyoxyethylene hydrogenated castor oil, polyoxyethylene hydrogenated castor oil isostearate, polyoxyethylene sorbitol tetra fatty acid ester, polyoxyethylene phytosterol, glycerin fatty acid esters such as lipophilic monostearate glyceryl and monostearate tetraglyceryl, glycerin fatty acid esters such as polyglycerin fatty acid ester, sucrose fatty acid ester, polyethylene glycol distearate, etc., may be used alone or in combination of two or more. Among these, it is preferable to use at least one of monopalmitic acid sorbitan, lipophilic monostearate glyceryl, and monostearate tetraglyceryl.

[0053] The content of the other surfactant is not particularly limited, but from the viewpoints of transparency and stability, it is preferably 0 to 3 mass %, more preferably 0 to 1 mass %, based on the total amount of the composition. In one embodiment, the aqueous composition does not contain any other surfactant.

[0054] In addition, raw materials may be selected as necessary to prepare the aqueous composition. In addition to the above components, the aqueous composition may contain any components contained in ordinary cosmetics, quasi-drugs, or pharmaceuticals. For example, it may contain moisturizing agents, antioxidants, UV absorbers, thickeners, pH adjusters, powder components, metal sequestering agents, coloring materials, various skin nutrients, drugs, etc., as necessary.

[0055] (Production method of the composition) The aqueous composition can be prepared according to a conventional method. For example, (a) cholesterol, (b) a surfactant having a sterol skeleton, (c) water, and, if necessary, (d) other components (other aqueous components, other oily components, other surfactants) may be mixed and stirred in an appropriate order, heated as necessary, and homogenized.

[0056] (Dosage form / use) The aqueous composition can be in various forms, including, but not limited to, a mist, a spray, a lotion, and the like.

[0057] The aqueous composition can be used as a cosmetic. Examples of the aqueous composition include skin care cosmetics such as lotion, milky lotion, cream, beauty essence, massage agent, pack agent, hand cream, body lotion, body cream, and sunscreen, finishing cosmetics such as foundation, eye shadow, eye liner, mascara, eyebrow, concealer, lipstick, and lip balm, and hair cosmetics such as hair mist, treatment, hair cream, hair shampoo, and hair rinse. Methods of use include using the composition on hands, fingers, or cotton, and impregnating nonwoven fabrics and the like.

[0058] The aqueous composition is also suitable for use in quasi-drugs, pharmaceuticals, and the like. For example, the aqueous composition can be used as a skin external preparation. Examples of the aqueous composition include external liquid preparations, external gel preparations, cream preparations, ointments, liniments, lotions, poultices, plasters, sprays, aerosols, etc. The method of use can be the same as that of the above-mentioned cosmetics.

[0059] In some embodiments, the aqueous composition is solubilized. As used herein, "solubilized" refers to a microemulsion state. Aqueous compositions in the form of a microemulsion are preferred in terms of skin permeability.

[0060] Although the anti-inflammatory composition has been described above in the form of an aqueous composition, the anti-inflammatory composition can be used in various forms other than an aqueous composition.

[0061] The anti-inflammatory composition of the present invention can be used for all applications in which it is meaningful to exert an anti-inflammatory effect. EXAMPLES

[0062] The present invention will be described in more detail below with reference to examples, but the present invention is not limited to these examples. The blending amount and content of each component in the table are converted to the pure amount, and are expressed as "mass %" unless otherwise specified.

[0063] The abbreviations used in the examples are conventional abbreviations well known to those skilled in the art. Some abbreviations are listed below. FBS: Fetal bovine serum DMEM: Dulbecco's modified Eagle's medium CO2: Carbon dioxide HBSS: Hanks Balanced Salt Solution SDS: Sodium dodecyl sulfate UVB: Ultraviolet B rays

[0064] [Test Example 1] Using a human epidermal keratinocyte cell line, we investigated whether a cholesterol-containing formulation suppresses the inflammatory response induced by UVB irradiation.

[0065] (1) Preparation of cholesterol-containing formulations Preparation method: Cholesterol, CS-20, pentylene glycol, and 1,3-butylene glycol were dissolved by heating at 80°C, and then a small amount of water was added, and the cholesterol solution was subjected to high-pressure emulsification using a high-pressure emulsification machine. The mixture was then cooled to 35°C to obtain a concentrated liquid (concentrate) in which cholesterol was dispersed within the solution. A Nanomizer was used as the high-pressure emulsification machine, and the treatment was performed at a treatment temperature of 80°C and a pressure of 150 MPa. The concentrate was diluted with water to finally obtain an aqueous microemulsion (cholesterol aqueous emulsion 1) containing 1% cholesterol by mass. The composition of each component is shown in Table 1. [Table 1]

[0066] (2) Cell experiments Human epidermal keratinocyte cell line HaCaT (AddexBio #T0020001) cultured in DMEM (Gibco #11995-065) supplemented with 10% FBS (Gibco #10270-106) and penicillin-streptomycin solution (Nacalai Tesque #09367-34) was added to 2.5 × 10 5 The cells were cultured in an incubator at 37°C with 5% CO2. The cell medium was removed using an aspirator and replaced with HBSS(+) (Nacalai Tesque #09735-75), and then incubated at 20 mJ / cm 2 The UVB-irradiated group was wrapped in aluminum foil and placed in the irradiator at the same time. After irradiation, the HBSS(+) was removed and replaced with a medium prepared by adding cholesterol aqueous emulsion 1 so that the final cholesterol concentration was 0.05% to 0.001% by mass, and the medium was cultured for 15 minutes. The original medium was then returned and the medium was cultured for 6 hours. The group composition used in the study is shown below. [Table 2]

[0067] (Result: Cell safety confirmation (RNA concentration quantification)) RNA was extracted from cells using RNeasy Mini Kit (QIAGEN #74106) according to the standard protocol. Figure 1 shows the RNA concentration in each group. The RNA concentration was calculated from the absorbance at 260 nm using NanoDrop2000 (Thermo Fisher Scientific #ND-2000). Figure 2 shows micrographs of cells before sample collection for the non-UVB non-irradiated non-treatment group (left, (UVB(-), Non-treatment), UVB non-irradiated non-treatment group (center, (UVB(+), Non-treatment), and UVB irradiated plus treated with cholesterol aqueous emulsion at a final concentration of 0.05% by mass (right, (UVB(+)+0.05%Chaq). As shown in Figure 1, the amount of recovered RNA was significantly reduced in the group treated with 0.05% cholesterol by mass, and as shown in Figure 2, the group treated with 0.05% cholesterol by mass showed a change in which the cells appeared to shrink (become rounded). Therefore, it is believed that a concentration of less than 0.05% by mass is preferable. The following gene expression quantification was performed for the groups with final concentrations of 0.01% by mass to 0.001% by mass, excluding 0.05% by mass.

[0068] (3) Quantitative gene expression cDNA synthesis from RNA solution was performed using the Verso cDNA Synthesis Kit (Thermo Fisher Scientific #AB1453A) according to the standard protocol. Quantitative PCR was performed using TaqMan Fast Universal PCR Master Mix (Thermo Fisher Scientific #4352042) and TaqMan Gene Expression Assay (Thermo Fisher Scientific #4331182) as primers, using Thermo Fisher Scientific's Step One Plus PCR kit. TM The expression of various genes was quantified using the primers shown in Table 3. The expression levels were evaluated as relative gene expression levels using the ΔΔCt method and normalized to the actin gene as a housekeeping gene. [Table 3]

[0069] (Statistical analysis) The non-treated group without UVB irradiation (Non-treatment) and the non-treated group with UVB irradiation (UVB(+)) were analyzed by Student's t-test, and the non-treated group with UVB irradiation (UVB(+)) and the group treated with cholesterol aqueous emulsion in addition to UVB irradiation (UVB(+)+Ch) were analyzed by Dunnett's test using the non-treated group with UVB irradiation (UVB(+)) as the control group. The significance level was 5%, *P<0.05, **P<0.01.

[0070] (Results: Confirmation of the inhibitory effect on UVB irradiation-induced inflammatory response (quantification of inflammatory cytokine gene expression)) Figure 3 shows the expression levels of inflammatory cytokine (IL-1α, IL-6, and TNFα) genes in each group. As shown in Figure 3, cholesterol aqueous emulsion significantly suppressed the inflammatory response induced by UVB irradiation in HaCaT. As shown in Figure 3, UVB irradiation significantly increased the expression levels of inflammatory cytokine (IL-1α, IL-6, and TNFα) genes, but the expression levels of these inflammatory cytokine (IL-1α, IL-6, and TNFα) genes were significantly decreased in the group treated with cholesterol aqueous emulsion in addition to UVB irradiation (UVB(+)+Ch) compared to the group not treated with UVB irradiation (UVB(+)). From this, it was confirmed that the addition of cholesterol aqueous emulsion suppressed the expression of inflammatory cytokine (IL-1α, IL-6, and TNFα) genes induced by UVB irradiation in HaCaT. In particular, it is suggested that the aqueous cholesterol emulsion has a large inhibitory effect on the expression of inflammatory cytokine (IL-6 and TNAα) genes (Figures 3(b)(c)).

[0071] These results confirmed that the cholesterol-containing preparation can suppress the production of inflammatory cytokines induced by exposure of the skin to ultraviolet rays, and can suppress inflammation (particularly inflammation in the epidermis). It has been shown that cholesterol can function as an inhibitor of the production of inflammatory cytokines (IL-1α, IL-6, and TNFα) induced by skin irritation due to exposure to ultraviolet light.

[0072] [Test Example 2] Using a three-dimensional epidermal culture model, we investigated whether a cholesterol-containing formulation could suppress the inflammatory response to a stimulant (sodium dodecyl sulfate (SDS)).

[0073] (1) Preparation of cholesterol-containing formulations Preparation method: Cholesterol, CS-20, and phenoxyethanol were dissolved by heating at 80°C to obtain a concentrate. The concentrate was diluted with water to finally obtain a microemulsion of 1% by mass of cholesterol in water (cholesterol aqueous emulsion 2). The composition of each component is shown in Table 4. [Table 4]

[0074] (2) Cell experiments (Acclimatization of the skin model) 2 mL of assay medium (SkinEthic EPISKIN TM A 3D epidermal culture model (SkinEthic EPISKIN TM A large one was then placed in the cavity and pre-cultured for 15 to 30 hours to allow the skin model to acclimate. (Inflammation induction) 200μL of 0.1% SDS aqueous solution (SDS: Nacalai Tesque #31606-75) was added to the cultured epidermal surface of the skin model as a stimulant to induce inflammation, and the skin model was cultured for 1 hour in a 5% CO2 incubator at 37℃. The cultured epidermal surface was washed twice with PBS. (Test substance exposure) Before or after inflammation induction, 200 μL of cholesterol aqueous emulsion 2 was added as a test substance to the cultured epidermal surface of the skin model, and the skin model was cultured in a 5% CO2 incubator at 37°C for 3 hours. After removing the test substance, the cultured epidermal surface was washed twice with PBS. The group to which cholesterol was added before inflammation induction was designated "pre-treatment (Pre)", and the group to which cholesterol was added after inflammation induction was designated "post-treatment (Post)". (Sample collection) After the inflammation induction and the test substance exposure procedures were completed, the culture cups were transferred to a 12-well plate containing fresh assay medium, and then cultured for 72 hours, after which 1 mL of the assay medium from under the culture cups was collected.

[0075] (3) Quantification of inflammatory cytokine protein levels For IL-1α / β, the amount of each protein was quantified using an ELISA kit (proteintech #KE00123, KE00021) according to standard protocols.

[0076] (Statistical analysis) A Student's t-test was performed for the untreated group and the group exposed to 0.1% SDS and not given cholesterol aqueous emulsion 2, and a Dunnett's test was performed for the group exposed to 0.1% SDS and given cholesterol aqueous emulsion 2 (pre-treatment and post-treatment), using the group exposed to 0.1% SDS and not given cholesterol aqueous emulsion 2 as the control group. *P<0.05, **P<0.01, with a significance level of 5%.

[0077] (result) The quantitative results of inflammatory cytokines (IL-1α / β) are shown in Figure 4. Figure 4 shows the amount of cytokine released per skin model. The number of cases was N=3. As can be seen from FIG. 4, exposure of the skin model to 0.1% SDS significantly increased the production of inflammatory cytokines IL-1α / β, but the production of these cytokines was significantly suppressed by post-SDS exposure treatment of cholesterol aqueous emulsion 2.

[0078] These results confirmed that the cholesterol-containing preparation can suppress the production of inflammatory cytokines induced by skin irritation with an irritant (sodium dodecyl sulfate (SDS)), and can suppress inflammation (especially inflammation in the epidermis). It has been shown that cholesterol can function as an inhibitor of the production of inflammatory cytokines (IL-1α, IL-1β) induced by skin stimulation with an irritant (sodium dodecyl sulfate (SDS)).

[0079] [Test Example 3] The anti-inflammatory effects of a group of ingredients having a sterol skeleton (cholesterol, phytosterol, and dipotassium glycyrrhizinate) were compared.

[0080] (1) Preparation of the formulation Cholesterol-containing preparations Preparation method: Cholesterol and CS-20 were heated and dissolved at 80°C to obtain a concentrate. The concentrate was diluted with water to finally obtain a microemulsion of an aqueous solution of 1% cholesterol by mass (cholesterol aqueous emulsion 3). The composition of each component is shown in Table 5. [Table 5]

[0081] Phytosterol-containing preparations Preparation method: A mixture of sitosterol, stigmasterol and campesterol was used as phytosterol (Phy). Phytosterol and CS-20 were heated and dissolved at 80°C to obtain a concentrate. The concentrate was diluted with water to finally obtain a microemulsion of an aqueous solution of 1% by mass of phytosterol (Phy) (aqueous phytosterol emulsion). The composition of each component is shown in Table 6. [Table 6]

[0082] Preparations containing dipotassium glycyrrhizinate Preparation method: An aqueous solution of dipotassium glycyrrhizinate (GK2) was prepared using Otsuka distilled water (Otsuka Pharmaceutical Co., Ltd.) so that the concentration was 1% by mass.

[0083] (2) Cell experiments Human epidermal keratinocyte cell line HaCaT (AddexBio #T0020001) cultured in DMEM (Gibco #11995-065) supplemented with 10% FBS (Gibco #10270-106) and penicillin-streptomycin solution (Nacalai Tesque #09367-34) was added to 2.5 × 10 5 The cells were cultured in an incubator at 37°C with 5% CO2. The cell medium was removed using an aspirator and replaced with HBSS(+) (Nacalai Tesque #09735-75), and then incubated at 20 mJ / cm 2 The non-UVB-irradiated group was wrapped in aluminum foil and placed in the UVB irradiator at the same time. After UVB irradiation, the HBSS(+) was removed and replaced with a medium prepared by adding aqueous emulsions (cholesterol aqueous emulsion 3, phytosterol aqueous emulsion, or dipotassium glycyrrhizinate aqueous emulsion) so that the final concentration of cholesterol (Ch), phytosterol (Phy), or dipotassium glycyrrhizinate (GK2) was 0.0025% by mass to 0.001% by mass, and the medium was cultured for 15 minutes.The original medium was then returned to and the medium was cultured for 6 hours. The group composition used in the study is shown below. [Table 7]

[0084] (3) Quantitative gene expression cDNA synthesis from RNA solution was performed using the Verso cDNA Synthesis Kit (Thermo Fisher Scientific #AB1453A) according to the standard protocol. Quantitative PCR was performed using TaqMan Fast Universal PCR Master Mix (Thermo Fisher Scientific #4352042) and TaqMan Gene Expression Assay (Thermo Fisher Scientific #4331182) as primers, using Thermo Fisher Scientific's Step One Plus PCR kit. TM The expression of various genes was quantified using the primers shown in Table 8. The expression levels were evaluated as relative gene expression levels using the ΔΔCt method and normalized to the actin gene as a housekeeping gene. [Table 8]

[0085] (Statistical analysis) A Dunnett's test was performed on the groups treated with UVB irradiation and aqueous phytosterol emulsion (UVB(+)+Phy) and UVB irradiation and aqueous dipotassium glycyrrhizinate emulsion (UVB(+)+GK2), with a control group treated with aqueous cholesterol emulsion (UVB(+)+Chaq). The significance level was 5%, *P<0.05, **P<0.01.

[0086] (Results: Confirmation of the inhibitory effect on UVB irradiation-induced inflammatory response (quantification of inflammatory cytokine gene expression)) 5(a) to (c) show the expression levels of inflammatory cytokine (IL-1α, IL-6, and TNFα) genes in each group. As shown in Figure 5, the gene expression levels of inflammatory cytokines under UVB irradiation were significantly lower in the cholesterol aqueous emulsion addition group at all concentrations compared to the dipotassium glycyrrhizinate addition group, and IL-6 was significantly lower and IL-1α and TNFα tended to be lower compared to the phytosterol aqueous emulsion addition group. Therefore, compared to dipotassium glycyrrhizinate, which is known as a conventional active ingredient for preventing rough skin, these results indicate that cholesterol may be effective as an anti-inflammatory ingredient at lower concentrations. [Industrial Applicability]

[0087] The anti-inflammatory composition of the present invention can suppress the production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα, and can suppress inflammation in the epidermis. In some forms, the anti-inflammatory composition can suppress inflammation caused by exposure of the skin to ultraviolet rays or irritation to the skin. The anti-inflammatory composition of the present invention can be suitably used, for example, as a cosmetic or topical skin preparation.

Claims

1. An anti-inflammatory composition comprising cholesterol as an active ingredient.

2. The composition according to claim 1, which suppresses inflammation in the epidermis.

3. The composition according to claim 2 , which suppresses inflammation caused by light exposure to the skin or irritation to the skin.

4. The composition according to claim 3, which suppresses inflammation caused by exposure of the skin to ultraviolet light.

5. The composition according to claim 1, which suppresses production of at least one inflammatory cytokine selected from interleukin-1α (IL-1α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and tumor necrosis factor (TNFα).

6. The composition according to claim 5, which suppresses production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα in the epidermis.

7. The composition according to any one of claims 1 to 6, wherein the cholesterol content is 0.1 to 3% by mass based on the total amount of the composition.

8. The composition according to any one of claims 1 to 6, which is in the form of an aqueous composition.

9. The composition according to claim 8 , further comprising a surfactant having a sterol skeleton.

10. The composition according to claim 9, wherein the surfactant having a sterol skeleton comprises at least one of polyoxyethylene cholesteryl ether and polyoxyethylene phytosteryl ether.

11. The composition of claim 8 further comprising a polyol.

12. The composition of claim 11 , wherein the polyol comprises a diol.

13. The composition according to any one of claims 1 to 6, which is a cosmetic preparation or an external skin preparation.

14. The composition according to any one of claims 1 to 6, which is in the form of a mist, spray or lotion.

15. The composition according to any one of claims 1 to 6, which is administered transdermally at a concentration of 0.001 to 0.01% by mass as cholesterol.

16. A cholesterol-containing inhibitor of the production of at least one inflammatory cytokine selected from IL-1α, IL-1β, IL-6, and TNFα.

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