Protein expression enhancer

Abstract

To provide a novel protein expression enhancer having an action of increasing the expression level of protein which is a component of the skin ligament.SOLUTION: A protein expression enhancer is used to increase the expression level of at least one protein selected from Mimecan, Biglycan, Vinculin and Microfibrillar-associated protein 4 (MFAP4) in a tendon cell, the protein expression enhancer containing plant extract as an active ingredient.SELECTED DRAWING: None

Description

Technical Field

[0001] The present invention relates to a protein expression enhancer that increases the expression level of skin retinaculum components.

Background Art

[0002] Skin sagging and wrinkles that become prominent with aging are considered to be caused in part by a decrease in skin elasticity. Conventionally, as methods for improving skin elasticity, for example, techniques for promoting the production or suppressing the degradation of extracellular matrix such as collagen and hyaluronic acid, which play important roles in maintaining skin elasticity (Patent Document 1), and techniques for promoting the proliferation of dermal fibroblasts that produce extracellular matrix (Patent Document 2) are known. However, there has been no report on a method for improving sagging and wrinkles by improving the structure of the subcutaneous tissue existing directly under the skin.

[0003] The present inventors focused on a reticular fiber structure called the retinacula cutis existing in the subcutaneous tissue and examined its relationship with facial sagging. As a result, they found that the loosening of the retinacula cutis causes a decrease in the elasticity of the deep skin and may cause sagging (Non-Patent Document 1). In addition, it is known that the density of the retinacula cutis decreases in the subcutaneous tissue directly under the deep wrinkle sites on the forehead and the outer corners of the eyes (Non-Patent Document 2). From the descriptions of Non-Patent Documents 1 and 2, it is presumed that increasing the density of the retinacula cutis will lead to the improvement of sagging and wrinkles. However, until now, there has been no knowledge about the constituent components of the retinacula cutis, its producing cells, and the mechanism of changes in the density of the retinacula cutis, and no component that increases the cause of changes in the density of the retinacula cutis has been known. In addition, there has been no report on a retinacula cutis improving agent that increases the expression level of the constituent components of the retinacula cutis.

Prior Art Documents

Patent Documents

[0004]

Patent Document 1

Patent Document 2

Non-Patent Literature

[0005]

Non-Patent Literature 1

Non-Patent Literature 2

Summary of the Invention

Problems to be Solved by the Invention

[0006] An object of the present invention is to provide a novel skin retinaculum improving agent having an action of increasing the expression level of a protein which is a skin retinaculum component in cells.

Means for Solving the Problems

[0007] As a result of intensive research, the inventors of the present invention have identified the constituent components of the cutaneous ligament and the cell mass that produces the constituent components. Furthermore, it has been found that mimecan, biglycan, vinculin, microfibrillar-associated protein 4 (MFAP4), and annexin A2, which are proteins constituting the cutaneous ligament, decrease with aging. Furthermore, it has been found that the cutaneous ligament can be improved by a plant extract having an action of increasing the expression levels of mimecan, biglycan, vinculin, and microfibrillar-associated protein 4 (MFAP4). Based on the above findings, the inventors of the present invention have completed the present invention.

[0008] The present invention for solving the above problems is a cutaneous ligament improver comprising a plant extract having an action of increasing the expression level of at least one protein selected from mimecan, biglycan, vinculin, and microfibrillar-associated protein 4 (MFAP4) in cells. According to the present invention, the cutaneous ligament can be improved by increasing the expression level of the protein.

[0009] In a preferred embodiment of the present invention, it comprises two or more plant extracts selected from plant extracts having an action of increasing the expression levels of different proteins among the above proteins. By adopting a form of increasing the expression levels of different proteins, the improvement effect on the cutaneous ligament can be enhanced.

[0010] The inventors of the present invention have found that the above proteins constituting the cutaneous ligament are mainly produced in tendon cells and fibroblasts. Therefore, the present invention is preferably used to increase the expression level of the protein in tendon cells and / or fibroblasts.

[0011] Moreover, the present invention particularly preferably takes the form of a plant extract having an effect of increasing the expression level of the total protein in tendon cells.

[0012] The present invention preferably takes the form of a plant extract selected from extracts of plants belonging to the following. Geraniaceae, Geranium; Rosaceae, Rosa; Theaceae, Camellia; Fabaceae, Aspalathus; Rubiaceae, Uncaria.

[0013] Furthermore, the present invention preferably takes the form of an extract of a plant selected from plants belonging to the following. Geraniaceae, Geranium, Geranium thunbergii; Rosaceae, Rosa, Rosa multiflora; Theaceae, Camellia, Camellia sinensis; Fabaceae, Aspalathus, Aspalathus linearis; Rubiaceae, Uncaria, Uncaria gambir. By using extracts of these plants as the plant extract, the expression level of the total protein can be increased, and a higher skin support improvement effect can be obtained.

[0014] The present invention preferably takes the form of an external preparation or an oral preparation.

Effects of the Invention

[0015] The skin retinaculum improver of the present invention is excellent in the action of increasing the expression level of proteins, which are skin retinaculum components, in cells.

Mode for Carrying Out the Invention

[0016] <1> Skin retinaculum components In the subcutaneous tissue directly under the skin, there is a reticular fiber structure called the retinacula cutis, and as its constituent components, mimecan, annexin A2, biglycan, vinculin, microfibrillar-associated protein 4 (MFAP4), collagen type 6, collagen type 1, annexin A5, decorin, lumican, prolargin, etc. are known.

[0017] <2> Plant extract (active ingredient) The skin retinaculum improver of the present invention is composed of a plant extract that increases the expression level of at least one protein selected from mimecan, biglycan, vinculin, and microfibrillar-associated protein 4 (MFAP4) among these proteins. The plant extract in the present invention is not particularly limited as long as it has the effect of increasing the expression level of the protein. Examples of the genus to which the plant used in the present invention belongs include Geranium of the Geraniaceae family, Rosa of the Rosaceae family, Crataegus of the Rosaceae family, Sophora of the Fabaceae family, Humulus of the Cannabaceae family, Vaccinium of the Ericaceae family, Saxifraga of the Saxifragaceae family, Oenothera of the Onagraceae family, Averrhoa of the Oxalidaceae family, Arctiumu of the Asteraceae family, Camellia of the Theaceae family, Phyllostachys of the Poaceae family, Hypericum of the Guttiferae family, Citrus of the Rutaceae family, Aspalathus of the Fabaceae family, Uncaria of the Rubiaceae family, Scutellaria of the Lamiaceae family, Rubus of the Rosaceae family, and Cinchona of the Rubiaceae family. Preferably, it is Geranium of the Geraniaceae family , Rosa of the Rosaceae family, Camellia of the Theaceae family, Aspalathus of the Fabaceae family, Uncaria of the Rubiaceae family, Coptis of the Ranunculaceae family, Pterocarpus of the Faboideae subfamily, and Vitaceae of the Vitaceae family can be exemplified.

[0018] In addition, specific plant species include Geranium thunbergii of the Geraniaceae family, Rosa multiflora of the Rosaceae family, Crataegus cuneata of the Crataegus genus in the Rosaceae family, Sophora flavesens of the Sophora genus in the Fabaceae family, Humulus lupulus of the Humulus genus in the Cannabaceae family, Vaccinium myrtillus of the Vaccinium genus in the Ericaceae family, Saxifraga stolonifera of the Saxifraga genus in the Saxifragaceae family, Oenothera biennis of the Oenothera genus in the Onagraceae family, Averrhoa carambola of the Averrhoa genus in the Oxalidaceae family, Arctium lappa of the Arctium genus in the Asteraceae family, Camellia sinensis of the Camellia genus in the Theaceae family, Phyllostachys edulis of the Phyllostachys genus in the Poaceae family, Hypericum erectum of the Hypericum genus in the Guttiferae family, Citrus reticulata of the Citrus genus in the Rutaceae family, Aspalathus linearis of the Aspalathus genus in the Fabaceae family, Uncaria gambir of the Uncaria genus in the Rubiaceae family, ScutellariaScutellaria baicalensis, Rubus idaeus of the Rosaceae family, Cinchona pubescens of the Rubiaceae family, Coptis japonica of the Ranunculaceae family, Pterocarpus marsupium of the Faboideae subfamily, and Vitis spp of the Vitaceae family can be exemplified.

[0019] The extracts of the above plants have the effect of improving the expression of at least one of Mimecan, Biglycan, Vinculin, and Microfibrillar-associated protein 4 (MFAP4). Tables 1 and 2 show which protein expression each plant extract improves in fibroblasts or tendon cells.

[0020]

Table 1

[0021]

Table 2

[0022] As shown in Tables 1 and 2, there are some that show an expression-improving effect only on some of the four types of proteins. In the present invention, it is preferable to combine two or more plant extracts that show an expression-improving effect only on some of the four types of proteins. That is, the present invention preferably takes an embodiment comprising a combination of two or more plant extracts having the effect of increasing the expression levels of different proteins among the above-mentioned proteins. More specifically, among the plant extracts shown in Tables 1 and 2, it is preferable to adopt an embodiment comprising a combination of two or more plant extracts having an effect of increasing the expression levels of different proteins. By adopting such an embodiment, a higher skin support improvement effect can be realized.

[0023] In order to realize a higher skin support improvement effect, it is preferable to adopt a form using a plant extract having an effect of increasing the expression level of the total protein. As shown in Table 2, plant extracts of Geranium thunbergii of the family Geraniaceae, Rosa multiflora of the genus Rosa of the family Rosaceae, Camellia sinensis of the genus Camellia of the family Theaceae, Aspalathus linearis of the genus Aspalathus of the family Fabaceae, and Uncaria gambir of the genus Uncaria of the family Rubiaceae have the effect of improving the expression of all of mimecan, biglycan, vinculin, and microfibril-associated protein 4. Therefore, it is particularly preferable to adopt an embodiment using at least one of these plant extracts.

[0024] The present inventors have found that fibroblasts or tendon cells are cells constituting the skin support, and these cells produce the above-mentioned proteins which are components of the skin support. Therefore, the present invention is preferably used in the form of increasing the expression level of the protein in tendon cells and / or fibroblasts, particularly preferably in tendon cells.

[0025] The extract of the plant in the present invention can be prepared by using plants that grow wild or are cultivated in Japan, or those produced in Japan and sold as raw materials for Kampo medicine. It is also possible to purchase and use commercially available extracts sold by companies dealing with plant extracts such as Maruzen Co., Ltd.

[0026] The plant extract means not only the plant extract itself, but also fractions of the extract, purified fractions, and the total name of the solvent-removed products of the extract or fractions and purified products. Plant-derived extracts include extracts using plants that grow wild or are cultivated, those sold as raw materials for Kampo medicine, and commercially available extracts. For the extraction operation, in addition to using the whole herb of the plant part, parts such as the whole plant, aerial part, rhizome part, trunk part, leaf part, stem part, spike, and flower bud can be used. However, it is preferable to pulverize or finely cut them in advance to improve the extraction efficiency. As the extraction solvent, one or more selected from polar solvents such as water, alcohols such as ethanol, isopropyl alcohol, and butanol, polyhydric alcohols such as 1,3-butanediol and polypropylene glycol, ketones such as acetone and methyl ethyl ketone, and ethers such as diethyl ether and tetrahydrofuran can be preferably exemplified. As a specific extraction method, for example, 1 to 30 parts by mass of the solvent is added to 1 part by mass of the part used for the extraction of the plant body or its dried product, immersed at room temperature for several days or at a temperature near the boiling point for several hours, cooled to room temperature, and then, if desired, insoluble substances and / or the solvent are removed, and fractionation and purification are performed by column chromatography or the like.

[0027] <3>Skin girdle improver The skin girdle improver of the present invention is preferably made into the form of an external preparation or an oral preparation by appropriately combining it with any component used in the formulation. Examples of external preparations include, for example, forms such as cosmetics, quasi-drugs, and topical skin medications. Also, their dosage forms are not particularly limited. Among them, from the perspective of the use of promoting the expression of skin supporting components, a form of cosmetics that can be continuously used is preferred, and among them, forms such as lotions, essences, milks, creams, gels, and sun care products are preferred.

[0028] When used as an oral preparation, it is preferably in the form of a food composition containing the skin supporting improvement agent of the present invention as an active ingredient. More specifically, it is preferably in the form of a supplement having a dosage form such as general food, tablets, granules, and drink preparations.

[0029] The content of the plant extract in the external preparation (in the case of the extract, the dry mass) is usually 0.00001% by mass or more, preferably 0.0001% by mass or more, more preferably 0.001% by mass or more, and usually 80% by mass or less, preferably 30% by mass or less, more preferably 10% % by mass or less. By setting it within the above range, it can preferably exhibit effects of improving wrinkles, improving dullness, preventing loss of firmness, and preventing reduction of skin elasticity. Also, the types of the expression promoter of skin supporting components may be not only one type but also two or more types.

[0030] In the case of an oral preparation, depending on the dosage form, the intake amount per time is usually 0.1 mg or more, preferably 1 mg or more, more preferably 10 mg or more, and usually 2000 mg or less, preferably 1000 mg or less, more preferably 500 mg or less, as the dry mass of the extract.

[0031] Skin support plays an important role in maintaining the shape of the skin. Therefore, the skin support improvement agent of the present invention can be used as an agent for improving the subcutaneous tissue structure.

[0032] In addition, the density of dermal ligaments has a correlation with age and the degree of skin sagging. Furthermore, among the proteins that make up dermal ligaments, Mimecan, Biglycan, Vinculin, and Microfibrillar-associated protein 4 (MFAP4) are components that decrease with aging, and the inventors have found that the decrease in these proteins is the cause of wrinkles and sagging. Therefore, the dermal ligament improving agent of the present invention is useful as an anti-aging agent, and is particularly preferably used as an agent for improving wrinkles, improving sagging, preventing loss of firmness, and preventing loss of skin elasticity.

[0033] When the dermal ligament improving agent is in the form of a cosmetic, it is possible to widely incorporate components usually used in cosmetics, and its dosage form and use are not limited in any way. Hereinafter, when applied to cosmetics, other components that can be contained in the cosmetics will be described.

[0034] In cosmetics, in addition to the above-described plant extracts, whitening components, other wrinkle-improving components, anti-inflammatory components, etc. can be incorporated. As the whitening ingredient, there is no particular limitation as long as it is generally used in cosmetics. For example, 4-n-butylresorcinol, ascorbyl glucoside, 3-O-ethylascorbic acid, tranexamic acid, arbutin, 1-triphenylmethylpiperidine, 1-triphenylmethylpyrrolidine, 2-(triphenylmethyloxy)ethanol, 2-(triphenylmethylamino)ethanol, 2-(triphenylmethyloxy)ethylamine, triphenylmethylamine, triphenylmethanol, triphenylmethane, and aminodiphenylmethane, N-(o-toluoyl)cysteic acid, N-(m-toluoyl)cysteic acid, N-(p-toluoyl)cysteic acid, N-(p-methoxybenzoyl)cysteic acid, etc. can be mentioned. Further, as other whitening ingredients, N-benzoyl-serine, N-(p-methylbenzoyl)serine, N-(p-ethylbenzoyl)serine, N-(p-methoxybenzoyl)serine, N-(p-fluorobenzoyl)serine, N-(p-trifluoromethylbenzoyl)serine, N-(2-naphthoyl)serine, N-(4-phenylbenzoyl)serine, N-(p-methylbenzoyl)serine methyl ester, N-(p-methylbenzoyl)serine ethyl ester, N-(2-naphthoyl)serine methyl ester, N-benzoyl-O-methylserine, N-(p-methylbenzoyl)-O-methylserine, N-(p-methylbenzoyl)-O-acetylserine, N-(2-naphthoyl)-O-methylserine, etc. can be mentioned.

[0035] Some of these whitening ingredients are already commercially available, and some can also be obtained by synthesis. For example, 3-O-ethylascorbic acid can be synthesized by the known method described in JP-A-8-134055. Since there are also commercially available products (manufactured by Nippon Fine Chemical Co., Ltd., "VC Ethyl"), it is possible to obtain and use these. 1-triphenylmethylpiperidine, 1 - Triphenylmethylpyrrolidine, 2-(triphenylmethyloxy)ethanol, 2-(triphenylmethylamino)ethanol, 2-(triphenylmethyloxy)ethylamine, triphenylmethylamine, triphenylmethanol, triphenylmethane, aminodiphenylmethane are disclosed in Patent Document WO2010-074052 Pamphlet. N-(o-Toluoyl)cysteic acid, N-(m-toluoyl)cysteic acid, N-(p-toluoyl)cysteic acid, N-(p-methoxybenzoyl)cysteic acid, N-(4-phenylbenzoyl)cysteic acid, N-(p-toluoyl)homocysteic acid are disclosed in WO2011-058730 Pamphlet. N-Benzoyl-serine, N-(p-methylbenzoyl)serine, N-(p-ethylbenzoyl)serine, N-(p-methoxybenzoyl)serine, N-(p-fluorobenzoyl)serine, N-(p-trifluoromethylbenzoyl)serine, N-(2-naphthoyl)serine, N-(4-phenylbenzoyl)serine, N-(p-methylbenzoyl)serine methyl ester, N-(p-methylbenzoyl)serine ethyl ester, N-(2-naphthoyl)serine methyl ester, N-benzoyl-O-methylserine, N-(p-methylbenzoyl)-O-methylserine, N-(p-methylbenzoyl)-O-acetylserine, N-(2-naphthoyl)-O-methylserine, etc. are disclosed in WO2011 / 074643 Pamphlet. Since the synthesis methods thereof are each disclosed, they can be synthesized according to the disclosure. The content of the whitening component in the cosmetic is usually 0.0001 to 30% by mass, preferably 0.001 to 10% by mass, and more preferably 0.01 to 5% by mass (in the case of an extract, it is the dry mass).

[0036] There are no particular limitations on the wrinkle-improving ingredients as long as they are generally used in cosmetics. However, it is preferable to use ingredients that have a wrinkle-improving effect through a mechanism different from the promotion of the expression of dermal ligaments from the perspective of fully utilizing the effects of each ingredient. For example, retinol, retinal, retinoic acid, tretinoin, isotretinoin, retinyl tocopherol, retinol palmitate, and retinol acetate as vitamin A or its derivatives can be mentioned. In addition, benzyl ursonate, phosphate ester of ursolic acid, benzyl betulinate, and phosphate ester of benzoic acid can be mentioned. The content of other wrinkle-improving ingredients in addition to the promoter of the expression of dermal ligaments in cosmetics is usually 0.0001 to 30% by mass, preferably 0.001 to 10% by mass, and more preferably 0.01 to 5% by mass (in the case of extracts, it is the dry mass).

[0037] Examples of the anti-inflammatory ingredients include clarinol, glabridin, glycyrrhizic acid, glycyrrhetinic acid, pantothenyl alcohol, etc., and preferably, glycyrrhizic acid and its salts, alkyl glycyrrhetinates and their salts, and glycyrrhetinic acid and its salts are preferably mentioned. The content of the anti-inflammatory ingredient in the cosmetic is usually 0.01 to 30% by mass, preferably 0.1 to 10% by mass, and more preferably 1 to 5% by mass (in the case of extracts, it is the dry mass).

[0038] Furthermore, it is preferable to use extracts derived from animals and plants that are generally used in pharmaceuticals, cosmetics, foods, and the like.For example, extracts such as Akebia extract, Asnaro extract, Asparagus extract, Avocado extract, Amacha extract, Almond extract, Arnica extract, Aronia extract, Apricot extract, Ginkgo extract, Wikyō extract, Udo extract, Ezo-kogigi extract, Enmei-sō extract, Ōbaku extract, Otane carrot extract, Odoriko-sō extract, Kakyo extract, Kakkōn extract, Chamomile extract, Carrot extract, Kawara-yomogi extract, Kanzō extract, Kiwifruit extract, Cucumber extract, Guava extract, Gardenia extract, Kumazasa extract, Walnut extract, Black rice extract, Chlorella extract, Kwa extract, Keikettō extract, Getto-yō extract, Gentiana extract, Rice extract, Fermented rice extract, Rice bran fermented extract, Rice germ oil, Sage extract, Soapwort extract, Sasa extract, Sansha extract, Sanshō extract, Shiitake extract, Dioscorea extract, Shikon extract, Perilla extract, Shinano-oki extract, Shimotsuke-sō extract, Shōkyō extract, Shōbu root extract, Sugina extract, Stevia extract, Fermented stevia, English ivy extract, English plantain extract, English mint extract, Sage extract, Zeniaoiy extract, Senkyū extract, Senburi extract, Sōhakuhi extract, Daiō extract, Soybean extract, Taisō extract, Thyme extract, Dandelion extract, Clove extract, Chili pepper extract, Tōki extract, Tōkin-senka extract, Tōnin extract, Tomato extract, Nattō extract, Carrot extract, Garlic extract, Hibiscus extract, Bakumondō extract, Lotus extract, Parsley extract, Birch extract, Hamamelis extract, Hikioji extract, Hinoki extract, Loquat extract, Fuki-tanpopo extract, Fuki-no-tō extract, Bukuryō extract, Bottle gourd extract, Peppermint extract, Bodaiju extract, Pine extract, Mizubashō extract, Melissa extract, Mozuku extract, Peach extract, Yaguruma-giku extract, Eucalyptus extract, Lily extract, Yokuin-in extract, Mugwort extract, Lavender extract, Apple extract, Rooibos tea extract, Jujube extract, Lettuce extract, Kelp extract, Renge-sō extract, Rosemary extract, Roman chamomile extract, Royal jelly extract, Waremokō extract, etc. are preferably listed.

[0039] The content (dry mass) of any of the above-mentioned extracts derived from animals and plants in the cosmetic is usually 0.01 to 30% by mass, preferably 0.1 to 10% by mass, and more preferably 0.3 to 3% by mass. The content (dry mass) of any of the above-mentioned extracts derived from animals and plants in the food is usually 0.01 to 80% by mass, preferably 0.1 to 50% by mass, and more preferably 1 to 30% by mass.

[0040] In addition to the aforementioned active ingredients, optional ingredients commonly used in cosmetics include polyols such as polyethylene glycol, glycerin, 1,3-butylene glycol, erythritol, sorbitol, xylitol, maltitol, propylene glycol, dipropylene glycol, diglycerin, isoprene glycol, 1,2-pentanediol, 2,4-hexylene glycol, 1,2-hexanediol, 1,2-octanediol, etc.; fatty acid soaps (such as sodium laurate, sodium palmitate, etc.); anionic surfactants such as potassium lauryl sulfate, triethanolamine alkyl sulfate ether, etc.; cationic surfactants such as stearyl trimethyl ammonium chloride, benzalkonium chloride, lauryl amine oxide, etc.; imidazoline-based amphoteric surfactants (such as 2-cocoyl-2-imidazolinium hydroxide-1-carboxyethyloxy 2 sodium salt, etc.); betaine-based surfactants (alkyl betaine, amide betaine, sulfobetaine, etc.); amphoteric surfactants such as acylmethyl taurine; sorbitan fatty acid esters (such as sorbitan monostearate, sorbitan sesquioleate, etc.); glycerin fatty acids (such as glycerin monostearate, etc.); propylene glycol fatty acid esters (such as propylene glycol monostearate, etc.); hydrogenated castor oil derivatives; glycerin alkyl ethers; POE sorbitan fatty acid esters (such as POE sorbitan monooleate, polyoxyethylene sorbitan monostearate, etc.); POE sorbit fatty acid esters (such as POE-sorbitan monolaurate, etc.); POE glycerin fatty acid esters (such as POE-glycerin monoisostearate, etc.); POE fatty acid esters (such as polyethylene glycol monooleate, POE distearate, etc.); POE alkyl ethers (such as POE 2-octyldodecyl ether, etc.); POE alkyl phenyl ethers (such as POE nonyl phenyl ether, etc.); pluronic (registered trademark) types; POE·POP alkyl ethers (such as POE·POP 2-decyltetradecyl ether, etc.); tetronic types; POE castor oil·hydrogenated castor oil derivatives (such as POE castor oil, POE hydrogenated castor oil, etc.); sucrose fatty acid esters; nonionic surfactants such as alkyl glucoside; sodium pyrrolidone carboxylate, lactic acid,Moisturizing ingredients such as sodium lactate, mica, talc, kaolin, synthetic mica, calcium carbonate, magnesium carbonate, anhydrous silicic acid (silica), aluminum oxide, barium sulfate and other powders which may be surface-treated, inorganic pigments such as cobalt oxide, ultramarine, dark blue, zinc oxide which may be surface-treated, composite pigments such as iron oxide titanium dioxide sintered body which may be surface-treated, pearl agents such as mica titanium, fish scale foil, bismuth oxychloride which may be surface-treated, organic pigments such as Red No. 202, Red No. 228, Red No. 226, Yellow No. 4, Blue No. 404, Yellow No. 5, Red No. 505, Red No. 230, Red No. 223, Orange No. 201, Red No. 213, Yellow No. 204, Yellow No. 203, Blue No. 1, Green No. 201, Purple No. 201, Red No. 204 which may be lake-colored, organic powders such as polyethylene powder, polymethyl methacrylate, nylon powder, organopolysiloxane elastomer, lower alcohols such as ethanol, isopropanol, vitamin A or its derivatives, vitamin B6 hydrochloride, vitamin B6 tripalmitate, vitamin B6 dioctanoate, vitamin B2 or its derivatives, vitamin B12, vitamin B15 or its derivatives and other vitamin Bs, vitamin Es such as α-tocopherol, β-tocopherol, γ-tocopherol, vitamin E acetate, vitamin Ds, vitamin H, pantothenic acid, pantethine, pyrroloquinoline quinone and other vitamins are included.

Example

[0041] <Test Example 1> Influence of plant extracts on gene expression levels in dermal-derived fibroblasts Human normal dermal fibroblasts were seeded in a 24-well plate at 2.0×10 4 cells / well in 10% FBS / DEME medium and cultured at 37°C in a 5% CO2 environment for 24 hours. Twenty-four hours after seeding, plant extracts prepared using the extraction solvents shown in Tables 3 to 5 were added (final concentration 0.5% by mass).

[0042] After 24 hours, the medium was removed and the cells were washed with PBS. mRNA was recovered using the FastLane Cell RT-PCR Kit (QIAGEN), and cDNA was synthesized using the QuantiTect Reverse Transcription Kit (QIAGEN). Quantitative RT-PCR was performed using the QuantiTect Primer Assay (QIAGEN) to measure the mRNA levels of genes encoding the skin extracellular matrix proteins biglycan, vinculin, and microfibrillar-associated protein 4 (MFAP4), which decrease with aging. The above-mentioned mRNA expression levels were calculated by the comparative CT method using β-actin as an endogenous control, and the results are shown in Tables 3 - 5 as relative values when the mRNA expression level of the target gene in normal human dermal fibroblasts without the test substance was set to "1".

[0043]

Table 3

Table 4

Table 5

[0044] As shown in Tables 3 - 5, the addition of Ganoderma lucidum extract increased the expression ratio of the Biglycan gene to 2.34 and the expression ratio of the MFAP4 gene to 2.20. The addition of Coptis japonica extract increased the expression ratio of the Biglycan gene to 1.28. Furthermore, the addition of Asparagus racemosus extract increased the expression ratio of the MFAP4 gene to 1.41, the addition of Gambierdiscus toxicus extract increased the expression ratio of the Vinculin gene to 1.93 and the expression ratio of the MFAP4 gene to 2.75, and the addition of pterostilbene increased the expression ratio of the Biglycan gene to 2.25 and the expression ratio of the MFAP4 gene to 2.28. From these results, it was found that the above plant extracts can improve the expression level of at least one of the genes of Biglycan, Vinculin, and MFAP4 in dermal fibroblasts.

[0045] <Test Example 2> Influence of Plant Extracts on Gene Expression Levels in Tendon Cells Human tendon-derived tendon cells were seeded on a collagen I-coated 24-well plate at 2.0×10 4 cells / well using a dedicated medium and cultured for 24 hours in an environment of 37°C and 5% CO2. Twenty-four hours after seeding, various plant extracts extracted using the extraction solvents shown in Tables 6 to 9 were added (final concentration 0.5% by mass). After 24 hours, the medium was removed, and the cells were washed with PBS. mRNA was recovered using the FastLane Cell RT-PCR Kit (manufactured by QIAGEN), and cDNA was synthesized using the QuantiTect Reverse Transcription Kit (manufactured by QIAGEN). Quantitative RT-PCR was performed using the QuantiTect Primer Assay (manufactured by QIAGEN) to measure the mRNA amounts of the genes encoding mimecan, biglycan, vinculin, and microfibrillar-associated protein 4 (MFAP4), which are skin ligament constituent proteins that decrease with aging. The above mRNA expression levels were calculated by the comparative CT method using β-actin as an endogenous control, and the results are shown in Tables 6 to 9 as relative values when the mRNA expression level of the target in human tendon-derived cells without the test substance was set to "1".

[0046] [Table 6] [Table 7] [Table 8]

Table 9

[0047] From Tables 6 to 9, it was found that when ganoderma lucidum extract, rhubarb extract, tea leaf extract, asparagus racemosus extract, and gambeer no ki extract were added, the expression ratio of all of Mimecan, Biglycan, Vinculin, and MFAP4 genes could be increased. It was also found that when other plant extracts were added, the expression ratio of at least one of the genes of Mimecan, Biglycan, Vinculin, and MFAP4 could be increased. It became clear that the above extracts increase the expression ratio of at least one of the genes of Mimecan, Biglycan, Vinculin, and MFAP4 in tendon cells and were found to be suitable as an active ingredient of a skin ligament improving agent.

Industrial Applicability

[0048] The present invention can be applied to external preparations and oral preparations.

Claims

1. For use in increasing the expression levels of all proteins of mimecan, biglycan, vinculin, and microfibrillar-associated protein 4 (MFAP4) in tendon cells, A protein expression enhancer, characterized by comprising all of (A)' to (D)' as active ingredients: (A)' An extract obtained by using a polar solvent as an extraction solvent from one or more plants selected from the following group, which is used for increasing the expression level of mimecan in tendon cells, and the polar solvent is one or two or more selected from water, ethanol, and 1,3-butanediol. Protein expression enhancer. Sophhora flavescens of the genus Sophora in the family Fabaceae, Humulus lupulus of the genus Humulus in the family Cannabaceae, Vaccinium myrtillus of the genus Vaccinium in the family Ericaceae, Arctium lappa of the genus Arctium in the family Asteraceae, Hypericum erectum of the genus Hypericum in the family Hypericaceae, Citrus reticulata of the genus Citrus in the family Rutaceae, Rubus idaeus of the genus Rubus in the family Rosaceae (B)' An extract obtained by using a polar solvent as an extraction solvent from one or more plants selected from the following group, which is used for increasing the expression level of biglycan in tendon cells, and the polar solvent is one or two or more selected from water, ethanol, and 1,3-butanediol. Protein expression enhancer. Crataegus cuneata of the genus Crataegus in the family Rosaceae, Saxifraga stolonifera of the genus Saxifraga in the family Saxifragaceae, Citrus reticulata, which belongs to the genus Citrus of the Rutaceae family, and Scutellaria baicalensis, which belongs to the genus Scutellaria of the Lamiaceae family A protein expression enhancer, which is composed of an extract obtained by using a polar solvent as an extraction solvent from one or more plants selected from the following group and is used to increase the expression level of vinculin in tendon cells, and the polar solvent is one or two or more selected from water, ethanol, and 1,3-butanediol. Crataegus cuneata, which belongs to the genus Crataegus of the Rosaceae family, Humulus lupulus, which belongs to the genus Humulus of the Cannabaceae family, Saxifraga stolonifera, which belongs to the genus Saxifraga of the Saxifragaceae family, Oenothera biennis, which belongs to the genus Oenothera of the Onagraceae family, Averrhoa carambola, which belongs to the genus Averrhoa of the Oxalidaceae family, Arctium lappa, which belongs to the genus Arctium of the Asteraceae family, and Rubus idaeus, which belongs to the genus Rubus of the Rosaceae family A protein expression enhancer, which is composed of an extract obtained by using a polar solvent as an extraction solvent from one or more plants selected from the following group and is used to increase the expression level of MFAP4 in tendon cells, and the polar solvent is one or two or more selected from water, ethanol, and 1,3-butanediol. Crataegus cuneata, which belongs to the genus Crataegus of the Rosaceae family, Humulus lupulus, which belongs to the genus Humulus of the Cannabaceae family, Saxifraga stolonifera, which belongs to the genus Saxifraga of the Saxifragaceae family, Evening Primrose (Oenothera biennis) of the genus Oenothera in the family Onagraceae, Star Fruit (Averrhoa carambola) of the genus Averrhoa in the family Oxalidaceae, Great Burdock (Arctium lappa) of the genus Arctium in the family Asteraceae, Moso Bamboo (Phyllostachys edulis) of the genus Phyllostachys in the family Poaceae, Hypericum erectum of the genus Hypericum in the family Hypericaceae, Mandarin Orange (Citrus reticulata) of the genus Citrus in the family Rutaceae, European Raspberry (Rubus idaeus) of the genus Rubus in the family Rosaceae, and Cinchona pubescens of the genus Cinchona in the family Rubiaceae [

2. ] The protein expression enhancer according to claim 1, which satisfies any one of the following conditions (1) to (14). (1) An extract of Sophora flavescens of the genus Sophora in the family Fabaceae obtained using a polar solvent as an extraction solvent, and the above (B)' to (D)', are included as active ingredients, and the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. (2) An extract of Humulus lupulus of the genus Humulus in the family Cannabaceae obtained using a polar solvent as an extraction solvent, and the above (B)', are included as active ingredients, and the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. (3) An extract of Vaccinium myrtillus of the genus Vaccinium in the family Ericaceae obtained using a polar solvent as an extraction solvent, and the above (B)' to (D)', are included as active ingredients, and the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. (4) An extract of Arctium lappa of the genus Arctium (Asteraceae) obtained using a polar solvent as the extraction solvent, the above (B)', and containing as an active ingredient, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. (5) An extract of Hypericum erectum of the genus Hypericum (Guttiferae) obtained using a polar solvent as the extraction solvent, the above (B)' and (C)', and containing as an active ingredient, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. (6) An extract of Citrus reticulata of the genus Citrus (Rutaceae) obtained using a polar solvent as the extraction solvent, the above (C)', and containing as an active ingredient, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. (7) An extract of Rubus idaeus of the genus Rubus (Rosaceae) obtained using a polar solvent as the extraction solvent, the above (B)', and containing as an active ingredient, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. (8) An extract of Crataegus cuneata of the genus Crataegus (Rosaceae) obtained using a polar solvent as the extraction solvent, the above (A)', and containing as an active ingredient, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. (9) An extract of Saxifraga stolonifera of the genus Saxifraga (Saxifragaceae) obtained using a polar solvent as the extraction solvent, the above (A)', and containing as an active ingredient, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. An extract of Scutellaria baicalensis of the Lamiaceae family, Scutellaria genus, obtained using a polar solvent as the extraction solvent, the above (A)', (C)' and (D)', and containing them as active ingredients, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. An extract of Oenothera biennis of the Onagraceae family, Oenothera genus, obtained using a polar solvent as the extraction solvent, the above (A)' and (B)', and containing them as active ingredients, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. An extract of Averrhoa carambola of the Oxalidaceae family, Averrhoa genus, obtained using a polar solvent as the extraction solvent, the above (A)' and (B)', and containing them as active ingredients, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. An extract of Phyllostachys edulis of the Poaceae family, Phyllostachys genus, obtained using a polar solvent as the extraction solvent, the above (A)' to (C)', and containing them as active ingredients, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol. An extract of Cinchona pubescens of the Rubiaceae family, Cinchona genus, obtained using a polar solvent as the extraction solvent, the above (A)' to (C)', and containing them as active ingredients, wherein the polar solvent is one or more selected from water, ethanol, and 1,3-butanediol.

3. The extract of Sophora flavescens, the extract of Humulus lupulus, the extract of Hypericum erectum, the extract of Citrus reticulata, the extract of Crataegus cuneata, and the extract of Scutellaria baicalensis are extracts obtained by using one or more selected from water and ethanol as an extraction solvent, The extract of Vaccinium myrtillus, the extract of Arctium lappa, the extract of Rubus idaeus, the extract of Saxifraga stolonifera, the extract of Oenothera biennis, the extract of Averrhoa carambola, the extract of Phyllostachys edulis, and the extract of Cinchona pubescens are extracts obtained by using one or more selected from water and 1,3-butanediol as an extraction solvent. The protein expression enhancer according to claim 1 or 2.

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