Il-36γ / il-37 expression ratio increase suppressor

Abstract

To provide an IL-36γ / IL-37 expression ratio increase suppressor that is effective in prevention or improvement of skin symptoms resulting from fatty acid-induced increase of IL-36γ and decrease of IL-37.SOLUTION: An IL-36γ / IL-37 expression ratio increase suppressor comprising Anthemis Nobilis or an extract thereof as an active ingredient.SELECTED DRAWING: None

Description

[Technical Field]

[0001] The present invention relates to an agent for suppressing an increase in the IL-36γ / IL-37 expression ratio induced by fatty acids. [Background technology]

[0002] Recently, proteome analysis of tape-stripped stratum corneum (SC) revealed that the SC contains high levels of the inflammatory cytokine interleukin-36γ (IL-36γ) and the anti-inflammatory cytokine interleukin-37 (IL-37). Furthermore, the IL-36γ / IL-37 ratio was shown to be positively correlated with the facial erythema index (Non-Patent Document 1).

[0003] Furthermore, in vitro studies have shown that the IL-36γ / IL-37 ratio is increased by fatty acids, particularly oleic acid, and that the facial IL-36γ / IL-37 ratio is positively correlated with the amount of sebum, particularly the content of unsaturated fatty acids (C18:1) such as oleic acid in sebum (Non-Patent Document 2).

[0004] This strongly suggests a relationship between the increased IL-36γ / IL-37 expression ratio due to fatty acid induction and skin redness, and controlling the increase in the IL-36γ / IL-37 expression ratio may be useful in improving skin redness.

[0005] Sebum has various beneficial functions, including regulating epidermal moisture, inhibiting the growth of gram-positive bacteria, and preventing invasion from the outside world. However, qualitative and quantitative changes in sebum can also contribute to the development of inflammatory diseases such as seborrheic dermatitis. Seborrheic dermatitis, also known as seborrheic eczema, is a type of dermatitis characterized by redness and dandruff in areas of active sebum secretion, such as the scalp and face. Seborrheic dermatitis is thought to be caused by a combination of multiple factors, one of which is Malassezia. Malassezia breaks down sebum, and the breakdown products, free fatty acids, particularly oleic acid, induce inflammation (Non-Patent Document 3). Therefore, controlling oleic acid-induced inflammation is also important in seborrheic dermatitis. Malassezia is also thought to be a cause or exacerbating factor for Malassezia folliculitis and atopic dermatitis.

[0006] On the other hand, Roman chamomile (also known as Roman chamomile) flower extract has an apple-like fragrance, which is known to have a calming and relaxing effect, and has long been incorporated into cosmetics and foods. Roman chamomile flower extract has also been recognized to have an anti-glycation effect by inhibiting the Maillard reaction, and it has been reported that it can be used to inhibit skin aging and pigmentation (Patent Document 1).

[0007] However, nothing is known about the effect of Roman chamomile extract on the expression of IL-36γ and IL-37, or its effectiveness in improving skin redness and seborrheic eczema. [Prior art documents] [Patent documents]

[0008] [Patent Document 1] Japanese Patent Application Laid-Open No. 2003-212770 [Non-patent literature]

[0009] [Non-Patent Document 1] J Dermatol Sci. 2023 Dec 8:S0923-1811(23)00251-7. [Non-patent document 2] J Cosmet Dermatol. 2023 Aug;22(8):2308-2317. [Non-patent document 3] Exp Dermatol. 2019 Sep;28(9):991-1001. Summary of the Invention [Problem to be solved by the invention]

[0010] The present invention relates to providing an agent for suppressing an increase in the IL-36γ / IL-37 expression ratio, which is useful for preventing or ameliorating skin symptoms caused by fatty acid-induced increase in IL-36γ or decrease in IL-37. [Means for solving the problem]

[0011] The present inventors investigated natural substances that affect the expression of IL-36γ and IL-37 and found that an extract of Roman chamomile is effective in suppressing an increase in the IL-36γ / IL-37 expression ratio and can improve skin redness.

[0012] That is, the present invention relates to the following 1) and 2). 1) An agent for suppressing an increase in the IL-36γ / IL-37 expression ratio, containing Roman chamomile or its extract as an active ingredient. 2) An agent for preventing or improving sebum-induced skin redness, containing Roman chamomile or its extract as an active ingredient. [Effects of the Invention]

[0013] The present invention provides a medicine, quasi-drug, or cosmetic that is useful for preventing or ameliorating skin symptoms caused by an increase in IL-36γ or a decrease in IL-37, such as sebum-induced skin redness and seborrheic eczema, and that can be used safely. [Brief explanation of the drawings]

[0014] [Figure 1] Inhibitory effect of Roman chamomile extract on the increase in the IL-36γ / IL-37 expression ratio. DETAILED DESCRIPTION OF THE INVENTION

[0015] The term "Roman chamomile" used in the present invention refers to Anthemis Nobilis of the genus Chamaemelum in the family Asteraceae, and is also known as Roman chamomile (Chamaemelum nobile).

[0016] In the present invention, Roman chamomile can be used in the form of the whole plant, leaves, stems, buds, flowers, buds, roots, rhizomes, or seeds, or a combination of these, but it is preferable to use the flowers as they are or after crushing. Roman chamomile can be used as it is, or as juice obtained by squeezing it, or as a dried or pulverized product of the plant itself, or as an extract extracted from these, but it is preferably used as an extract, and more preferably as a Roman chamomile flower extract.

[0017] Examples of extracts include various solvent extracts obtained by known extraction methods, such as extracting Roman chamomile plant matter at room temperature or under heating, or extracting using an extraction device such as a Soxhlet extractor, as well as diluted solutions, concentrated solutions, and dried powders thereof. Known extraction methods include, for example, steeping, decoction, percolation, reflux extraction, supercritical extraction, ultrasonic extraction, and microwave extraction.

[0018] The extraction solvent used to obtain the extract may be either a polar solvent or a non-polar solvent, such as water; alcohols such as methanol, ethanol, propanol, and butanol; polyhydric alcohols such as propylene glycol and butylene glycol; ketones such as acetone and methyl ethyl ketone; esters such as methyl acetate and ethyl acetate; linear and cyclic ethers such as tetrahydrofuran and diethyl ether; polyethers such as polyethylene glycol; haloethers such as dichloromethane, chloroform, and carbon tetrachloride. Examples of suitable solvents include fluorinated hydrocarbons; hydrocarbons such as hexane, cyclohexane, and petroleum ether; aromatic hydrocarbons such as benzene and toluene; and pyridines, which can be used alone or in combination. Among these, water, alcoholic (alcohols and / or polyhydric alcohols) solvents, and water-alcohol mixed solvents are preferred, with water-alcohol mixed solvents being more preferred. Furthermore, preferred alcohols include methanol, ethanol, propanol, and butanol, with ethanol being more preferred. Furthermore, preferred polyhydric alcohols include butylene glycol and propylene glycol, with 1,3-butylene glycol being more preferred. A more suitable water-alcohol mixed solvent is an aqueous solution of 1,3-butylene glycol at a concentration of 20% (v / v) or more, preferably 50% (v / v) or more, and 95% (v / v) or less, preferably 90% (v / v) or less, such as a 20 to 95% (v / v) aqueous solution of 1,3-butylene glycol, preferably a 50 to 90% (v / v) aqueous solution of 1,3-butylene glycol.

[0019] Extraction using the above-mentioned extraction solvent can be carried out, for example, by using 5 to 60 parts by mass of solvent, preferably 5 to 30 parts by mass of solvent per 1 part by mass of plant body, and extracting at 4°C to 100°C for 1 hour to 30 days, preferably 10°C to 30°C for 10 days to 25 days.

[0020] The extracts described above can be used as they are, or they can be diluted, concentrated, or freeze-dried to prepare powders or pastes. They can also be freeze-dried and diluted with solvents commonly used in extractions, such as water, ethanol, propylene glycol, butylene glycol, water-ethanol mixtures, water-propylene glycol mixtures, and water-butylene glycol mixtures, before use. They can also be encapsulated in vesicles such as liposomes or microcapsules.

[0021] The Roman chamomile extract of the present invention may be a crude product as long as it meets the standards acceptable for use as a cosmetic, quasi-drug, or pharmaceutical product and exhibits the effects of the present invention, and the purity of the crude product obtained may be further increased by appropriately combining known separation and purification methods, such as organic solvent precipitation, centrifugation, ultrafiltration membrane, high performance liquid chromatography, and column chromatography.

[0022] As shown in the Examples below, Roman chamomile extract suppresses the increase in the IL-36γ / IL-37 expression ratio induced by oleic acid, and therefore, by administering an effective amount of the extract to humans, it is possible to suppress the increase in the IL-36γ / IL-37 expression ratio in vivo. Therefore, Roman chamomile or an extract thereof can be an agent for suppressing an increase in the IL-36γ / IL-37 expression ratio and can be used to suppress an increase in the IL-36γ / IL-37 expression ratio. This use may be in humans or non-human animals, or in specimens derived therefrom, and may be therapeutic or non-therapeutic. Here, "non-therapeutic" does not include medical procedures, i.e., does not include methods of surgery, therapy, or diagnosis performed on humans; more specifically, it does not include methods of surgery, therapy, or diagnosis performed on humans by physicians or those acting under the direction of a physician. Furthermore, Roman chamomile or an extract thereof can be used to produce an agent for suppressing an increase in the IL-36γ / IL-37 expression ratio.

[0023] In the present invention, IL-36γ belongs to the IL-1 family. Upon binding to the IL-36 receptor, it activates the NF-kB pathway and MAPK pathway via MyD88, increasing the expression of proinflammatory cytokines (PLoS One. 2015 Nov 12;10(11):e0138423). It is known that expression is elevated in the skin of patients with inflammatory skin diseases such as psoriasis and atopic dermatitis (JAMA Dermatol. 2019 Dec 1;155(12):1358-1370, Clin Exp Immunol. 2016 May;184(2):159-173). IL-37 is thought to exhibit broad anti-inflammatory effects and is considered a negative regulator of the IL-1 family (Front Immunol. 2021 Mar 16;12:652846.). In healthy individuals, IL-37 mRNA is primarily expressed in the skin, particularly in keratinocytes in the granular layer (Sci Rep. 2017 Dec 12;7(1):17446.).

[0024] In the present invention, the IL-36γ / IL-37 expression ratio refers to the ratio of the expression level of IL-36γ to the expression level of IL-37. Here, the expression level may be at either the gene level or the protein level. That is, the expression levels of IL-36γ and IL-37 may be, for example, the amounts of mRNA encoding IL-36γ and IL-37, or the amounts of IL-36γ protein and IL-37 protein, extracted from a cell line such as normal human epidermal keratinocytes. The amount of mRNA can be measured using, for example, PCR or Northern hybridization, and the amount of protein can be measured using, for example, ELISA or Western blotting.

[0025] In the present invention, "inhibiting an increase in the IL-36γ / IL-37 expression ratio" specifically means inhibiting the increase in the IL-36γ / IL-37 expression ratio induced by fatty acids, particularly oleic acid (C18:1). Here, "inhibiting" encompasses both suppression and reduction of the increase.

[0026] As mentioned above, the IL-36γ / IL-37 expression ratio is increased by fatty acids, and the facial IL-36γ / IL-37 expression ratio is known to be positively correlated with the amount of sebum, particularly the amount of oleic acid (C18:1) (Non-Patent Document 2). Furthermore, it is known that an increase in the IL-36γ / IL-37 expression ratio is accompanied by an increase in the facial erythema index (Non-Patent Document 1). Furthermore, it is known that free fatty acids, which are sebum degradation products, can cause seborrheic dermatitis (Patent Document 3). Therefore, an agent that inhibits an increase in the IL-36γ / IL-37 expression ratio can be useful for preventing or ameliorating sebum-induced skin redness and seborrheic dermatitis. In fact, when Roman chamomile extract was applied to the skin, it was found to have an effect of reducing redness (Example 2 described below).

[0027] Facial erythema is a common dermatological complaint resulting from dilation of cutaneous blood vessels and increased blood flow to the skin. While some facial erythema is a transient, normal neurological response to strong emotion, exercise, or heat exposure, others are clinical manifestations of inflammatory skin diseases such as rosacea, acne, and seborrheic dermatitis (Clin Cosmet Investig Dermatol. 2021 Jun 8;14:601-614, Actas Dermosifiliogr. 2015 Jun;106(5):427-9), and non-inflammatory erythema, such as post-acne erythema, manifests as dilated changes in the microvasculature associated with wound healing (J Cosmet Dermatol. 2022 Apr;21(4):1379-1392). Here, sebum-induced skin redness refers to the reddish skin tone that occurs when inflammation of the skin is caused by sebum components such as oleic acid, which leads to dilation of skin blood vessels and increased blood flow. Seborrheic dermatitis is a skin inflammation that occurs in areas where sebum is secreted, mainly on the scalp and face, and causes redness and dandruff.Seborrheic dermatitis that occurs within the first few months of life due to excessive sebum secretion is specifically called infantile seborrheic dermatitis.

[0028] The above-mentioned agent for suppressing an increase in the IL-36γ / IL-37 expression ratio suppresses an increase in the IL-36γ / IL-37 expression ratio, and can therefore be used as a cosmetic, drug, or quasi-drug for preventing or ameliorating symptoms associated with an increase in the IL-36γ / IL-37 expression ratio, such as an increase in IL-36γ or a decrease in IL-37, such as sebum-induced skin redness and seborrheic dermatitis, or as a material or preparation to be incorporated into these.

[0029] The above-mentioned pharmaceuticals or quasi-drugs can be administered in any dosage form. Administration may be oral or parenteral. Examples of dosage forms for oral administration include solid preparations such as tablets, coated tablets, granules, powders, and capsules, as well as liquid preparations such as elixirs, syrups, and suspensions. Examples of dosage forms for parenteral administration include injections, infusions, topical preparations, external preparations, transdermal preparations, transmucosal preparations, nasal preparations, enteral preparations, inhalants, suppositories, boluses, and patches. Preferably, the pharmaceuticals or quasi-drugs are in a dosage form for parenteral administration, and more preferably in the form of external preparations for skin.

[0030] The above-mentioned preparations can be formulated with a pharmaceutically acceptable carrier. Examples of such carriers include excipients, coating agents, binders, bulking agents, disintegrants, lubricants, diluents, osmotic pressure adjusters, pH adjusters, dispersants, emulsifiers, preservatives, stabilizers, antioxidants, colorants, UV absorbers, moisturizers, thickeners, activity enhancers, anti-inflammatory agents, disinfectants, fragrances, flavorings, and odor enhancers. Furthermore, the pharmaceuticals and quasi-drugs may contain other active ingredients or pharmacological ingredients, as long as the inhibitory effect on the increase in the IL-36γ / IL-37 expression ratio is not lost.

[0031] The above-mentioned cosmetics can be administered in any parenteral form. Examples of formulations for parenteral administration include topical preparations, external preparations, transdermal preparations, transmucosal preparations, and patches. Preferably, the cosmetics are in the form of external preparations for skin. They can be produced by conventional methods by combining Roman chamomile or an extract thereof, and, if necessary, cosmetically acceptable carriers and / or other active ingredients or cosmetic ingredients. Examples of such carriers include excipients, coating agents, binders, bulking agents, disintegrants, lubricants, diluents, osmotic pressure adjusters, pH adjusters, dispersants, emulsifiers, preservatives, stabilizers, antioxidants, colorants, UV absorbers, moisturizers, thickeners, activity enhancers, anti-inflammatory agents, disinfectants, fragrances, flavorings, and odor enhancers. Furthermore, the cosmetic product may contain other active ingredients or cosmetic ingredients, such as moisturizers, whitening agents, UV protection agents, cell activators, cleansers, keratolytic agents, and makeup ingredients (e.g., makeup base, foundation, face powder, powder, blush, lipstick, eye makeup, eyebrow makeup, mascara, and the like), as long as the effect of promoting elastic fiber formation of the present invention is not lost. When used as a cosmetic product, the cosmetic product may take any form that can be used in cosmetics, such as creams, emulsions, lotions, suspensions, gels, powders, packs, sheets, patches, sticks, and cakes.

[0032] The content of Roman chamomile or its extract in each of the above preparations (drugs, quasi-drugs, cosmetics) is, for example, 0.00001% by mass or more, preferably 0.0001% by mass or more, and 5% by mass or less, preferably 1% by mass or less, calculated as dry solid content, of the total mass of the preparation. Also, the content may be 0.00001 to 5% by mass, preferably 0.0001 to 1% by mass.

[0033] When the Roman chamomile or extract thereof of the present invention is used as a pharmaceutical or incorporated into a pharmaceutical, the dosage may vary depending on the patient's condition, weight, sex, age, and other factors. However, the daily oral dosage for an adult is typically 0.0001 mg / kg or more, preferably 0.001 mg / kg or more, and 100 mg / kg or less, preferably 10 mg / kg or less, and is 0.0001 to 100 mg / kg, preferably 0.001 to 10 mg / kg, in terms of Roman chamomile (dry matter).

[0034] Furthermore, the subjects for administration or ingestion are not particularly limited as long as they are animals that need or desire it, and examples include humans that need or desire prevention or improvement of symptoms associated with an increase in the IL-36γ / IL-37 expression ratio, such as an increase in IL-36γ or a decrease in IL-37, such as sebum-induced skin redness and seborrheic dermatitis.

[0035] In relation to the above-described embodiment, the present invention further discloses the following aspects. <1> An agent for suppressing an increase in the IL-36γ / IL-37 expression ratio, which contains Roman chamomile or its extract as an active ingredient. <2> An agent for preventing or improving sebum-induced skin redness, which contains Roman chamomile or its extract as an active ingredient. <3> Sebum-induced skin redness is redness induced by oleic acid <2> An agent for preventing or improving skin redness as described above. <4> A preventive or ameliorating agent for seborrheic dermatitis, containing Roman chamomile or its extract as an active ingredient. <5> The Roman chamomile extract is a solvent extract of Roman chamomile, <1> ~ <4> The preventive or ameliorative agent according to any one of the preceding claims. <6> The solvent is a water-alcohol mixed solvent. <5> The preventive or improving agent described above. <7> The water-alcohol mixed solvent is a 1,3-butylene glycol aqueous solution, preferably a 50 to 90% (v / v) 1,3-butylene glycol aqueous solution. <6> The preventive or improving agent described above. <8> The Roman chamomile or its extract is a Roman chamomile flower or its extract. <1> ~ <7> The preventive or ameliorative agent according to any one of the preceding claims.

[0036] <9> Use of Roman chamomile or an extract thereof for producing an agent for suppressing an increase in the IL-36γ / IL-37 expression ratio. <10> Use of Roman chamomile or an extract thereof for producing an agent for preventing or improving sebum-induced skin redness. <11> Sebum-induced skin redness is redness induced by oleic acid <10> Use as described. <12> Use of Roman chamomile or an extract thereof for producing an agent for preventing or improving seborrheic dermatitis. <13> The Roman chamomile extract is a solvent extract of Roman chamomile, <9> ~ <12> Use as described in any of the above. <14> The solvent is a water-alcohol mixed solvent. <13> Use as described. <15> The water-alcohol mixed solvent is a 1,3-butylene glycol aqueous solution, preferably a 50 to 90% (v / v) 1,3-butylene glycol aqueous solution. <14> Use as described. <16> The Roman chamomile or its extract is a Roman chamomile flower or its extract. <9> ~ <15> Use as described in any of the above.

[0037] <17> Roman chamomile or an extract thereof for use in suppressing an increase in the IL-36γ / IL-37 expression ratio. <18> Roman chamomile or an extract thereof for use in preventing or improving sebum-induced skin redness. <19> Sebum-induced skin redness is redness induced by oleic acid <10> The Roman chamomile or its extract described above. <20> 1. Roman chamomile or an extract thereof for use in preventing or improving seborrheic dermatitis.

[0038] <21> A method for suppressing an increase in the IL-36γ / IL-37 expression ratio, comprising administering Roman chamomile or an extract thereof to a subject in need thereof. <22> A method for preventing or improving sebum-induced skin redness, comprising applying Roman chamomile or an extract thereof to a subject in need thereof. <23> Sebum-induced skin redness is redness induced by oleic acid <22> The prevention or improvement method described above. <24> A method for preventing or improving seborrheic dermatitis, comprising applying Roman chamomile or an extract thereof to a subject in need thereof. <25> The Roman chamomile extract is a solvent extract of Roman chamomile, <21> ~ <24> The method for prevention or improvement described in any one of the above. <26> The solvent is a water-alcohol mixed solvent. <25> The prevention or improvement method described above. <27> The water-alcohol mixed solvent is a 1,3-butylene glycol aqueous solution, preferably a 50 to 90 (v / v)% 1,3-butylene glycol aqueous solution. <26> The prevention or improvement method described above. <28> The Roman chamomile or its extract is a Roman chamomile flower or its extract. <21> ~ <27> The method for prevention or improvement described in any one of the above.

[0039] <29> The aforementioned <17> ~ <19> In either case, the use is a non-therapeutic use. <30> The aforementioned <21> ~ <23> In either case, the method is a non-therapeutic method. <31> The aforementioned <22> ~ <28> In either case, the subject in need thereof is a human who needs or desires prevention or amelioration of sebum-induced skin redness or seborrheic dermatitis that occurs in association with an increased IL-36γ / IL-37 expression ratio. [Example]

[0040] Production Example 1 Preparation of Roman chamomile extract 2000g of 60wt% butanediol was added to 100g of crushed flowers of Roman chamomile (Anthemis Nobilis) (from Poland) of the Asteraceae family, and the mixture was extracted at 40℃ for 20 hours and filtered. 126g of 60wt% butanediol was added to this extract to prepare an evaluation sample (solid content 1.1w / w%).

[0041] Example 1: Inhibitory effect of Roman chamomile extract on oleic acid-induced increase in IL-36γ / IL-37 expression ratio (1) Method Normal human epidermal keratinocytes (NHEKs) were purchased from Kurabo and cultured in a culture medium containing 0.06 mM Ca2+. 2+The cells were cultured in HuMedia-KG2 (Kurabo) containing insulin (10 μg / ml), human recombinant epidermal growth factor (hEGF) (0.1 ng / ml), hydrocortisone (0.67 μg / ml), bovine pituitary extract (BPE) (0.4%), gentamicin (50 μg / ml), and amphotericin B (50 ng / ml) at 37°C in a 5% CO environment. Keratinocytes (NHEKs) were seeded (1 x 10) onto 12-well plates. 5 When the cells reached subconfluence (cells / well), the medium was replaced with the above-mentioned medium without growth additives (hereinafter referred to as (-) medium) and pre-cultured for 16 hours. The medium was then replaced with (-) medium containing 50 μM oleic acid and 0, 0.03, 0.1, or 0.3 w / w% Roman chamomile extract (Production Example 1) at final concentrations of 0, 0.03, 0.1, or 0.3 w / w%, respectively. After 8 hours of incubation, the cells were harvested. RNA was extracted from the harvested cells using an RNeasy Mini Kit (QIAGEN). cDNA was synthesized from the extracted RNA using a High Capacity RNA-to-cDNA Kit (Applied Biosystems). An amount equivalent to 20 ng of total RNA was subjected to real-time PCR to measure the gene expression levels of IL-36γ and IL-37, and the IL-36γ / IL-37 ratio was calculated. The results are shown in Figure 2.

[0042] For real-time PCR, the TaqMan Gene Expression Assay (Applied Biosystems) probes used to specifically detect the target genes were IL36G (Hs00219742_m1) and IL37 (Hs00367201_m1). Expression levels of the target genes were normalized using the expression level of the internal control gene RPLP0 (Hs99999902_m1). Analysis was performed using a 7500 Real Time PCR System (Applied Biosystems).

[0043] (2) Results As shown in Figure 1, the Roman chamomile extract exhibited a concentration-dependent inhibitory effect on the increase in the IL-36γ / IL-37 expression ratio induced by oleic acid.

[0044] Example 2: Skin-improving effect of Roman chamomile extract (1) Method A parallel-group comparative study was conducted on 15 healthy men (ages: 20s-50s) and 15 healthy women (ages: 20s-50s) in which a lotion containing Roman chamomile extract or a placebo lotion, as shown in Table 1, was used twice a day, morning and night, for four weeks. After continuous use, participants were asked to complete a questionnaire regarding the degree of improvement in the skin concern of "uneven redness of the skin."

[0045] [Table 1]

[0046] (2) Results The survey results are shown in Table 2 below.

[0047] [Table 2]

[0048] A large percentage of people found that the lotion containing Roman chamomile extract had an effect on improving uneven redness.

Claims

1. An agent for suppressing an increase in the IL-36γ / IL-37 expression ratio, comprising Roman chamomile or an extract thereof as an active ingredient.

2. An agent for preventing or improving sebum-induced skin redness, which contains Roman chamomile or its extract as an active ingredient.

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