HK1178571B - Platelet-derived growth factor-bb production promoter, and mesenchymal stem cell production promoter, stem cell stabilizer and dermis regenerator each comprising same - Google Patents

Description

Platelet-derived growth factor-BB production enhancer, and mesenchymal stem cell production promoter, stem cell stabilizer and dermal regeneration agent each comprising same

Technical Field

The present invention relates to a platelet-derived growth factor-BB (PDGF-BB) production enhancer, and a mesenchymal stem cell production promoter, a stem cell stabilizer and a dermal regeneration agent each comprising the PDGF-BB production enhancer.

Background

Stem cells are cells having 2 properties of having both the ability to differentiate into multiple cells and the ability to self-replicate by cell division to produce cells similar to the cells. Stem cells derived from embryos at an early developmental stage of fertilized eggs are called embryonic stem cells (ES cells). Although human ES cells are expected to be used in regenerative medicine, the use of fertilized eggs is an ethical problem, and therefore, the production of new human ES cells is not allowed.

In recent years, as cells having properties similar to ES cells, artificial pluripotent stem cells (iPS cells) have also received attention. However, production of iPS cells has many problems from the viewpoints of canceration of cells, production efficiency, and the like. On the other hand, since somatic stem cells having the ability to differentiate into specific tissues can be obtained from the patient's own body tissue such as bone marrow, there is no ethical problem like embryonic stem cells.

It is known that epidermal stem cells are present in the epidermal basal layer of the skin (non-patent document 1), and hair follicle epithelial stem cells and skin pigment stem cells are reported to be present in a region called a hair follicle bulge region (non-patent document 2) and skin pigment stem cells (non-patent document 3). On the other hand, although fibroblasts in the form of a slender spindle exist in a fibrous component mainly composed of collagen in the dermis, it is not clear whether stem cells exist in the fibroblasts in the dermis. Further, although skin-derived precursor cells (SKP) that differentiate into a plurality of cell lines such as fat, glial, cartilage, muscle and the like are known to exist in dermis (non-patent document 4), the relationship between dermal fibroblasts and SKP is not clear.

Since mesenchymal stem cells (non-patent document 5) isolated from bone marrow as fibroblast precursor cells are differentiated into various cells belonging to the mesenchymal system (bone cells, muscle cells, chondrocytes, tendon cells, adipocytes, etc.), they are expected to be applied to regenerative medicine such as reconstruction of bone, blood vessels, muscle, and the like. Recently, it has become clear that there is a possibility that a large amount of tissue having mesenchymal tissue exists, and mesenchymal stem cells have been isolated from fat, umbilical cord blood, placenta, and the like (non-patent documents 6 to 8).

It has been known from recent findings that mesenchymal stem cells are present in systemic blood vessels as perivascular cells (percytes) and play a role in stabilizing blood vessels and maintaining tissue homeostasis (non-patent documents 9 and 10).

In addition, if blood vessels are destroyed at or near a tissue injury site, mesenchymal stem cells, which are perivascular cells (percytes), detach from the blood vessels and proliferate, supply the lost cells (non-patent documents 11 to 14), and release bioactive factors to protect tissues (non-patent documents 15 to 19), thereby playing a role in repair and regeneration of the damaged tissues. These secreted factors have been reported to have potent immune-suppressive effects (non-patent documents 21 and 22), and T-cell and B-cell mediated destruction of damaged tissues (non-patent documents 9 and 22), in addition to angiogenesis and anti-apoptosis effects.

It is also known that mesenchymal stem cells exhibit an anti-fibrotic effect (non-patent documents 23 and 24) and/or an effect on multiple sclerosis and diabetes (non-patent document 9).

On the other hand, it has been clarified that chronic inflammation is a basic disease state common to various diseases (for example, metabolic syndrome, arteriosclerotic disease, cancer, neurodegenerative disease, autoimmune disease, and the like) (non-patent document 25). For example, it has been reported that endothelial cell dysfunction and insulin resistance are induced by chronic inflammation, and the induced inflammation causes various diseases such as diabetes and arteriosclerotic diseases (non-patent document 26). Furthermore, it has been revealed that adipose tissue itself of obesity is involved in inflammatory changes (non-patent documents 27 to 29). Since chronic inflammation occurs around blood vessels, it is considered that interaction between mesenchymal stem cells, which are perivascular cells (pericytes), and blood vessels is also hindered in chronic inflammation.

Based on the above findings, it is considered that if the promotion and/or stabilization of the production of mesenchymal stem cells can be achieved, the promotion and/or stabilization of mesenchymal stem cells is extremely effective in various applications such as the stabilization of blood vessels, the maintenance of tissue homeostasis, the repair and/or regeneration of damaged tissues, the prevention and/or treatment of various diseases such as fibrosis, multiple sclerosis, and diabetes, and the prevention and/or improvement of various states of chronic inflammation such as metabolic syndrome.

The present inventors reported that mesenchymal stem cells are present in the dermis, and established a method for efficiently isolating mesenchymal stem cells from the dermis (Japanese patent application No. 2009-213291). Considering the above-described effects of mesenchymal stem cells, it is considered that the stabilization and/or enhancement of the production of mesenchymal stem cells in the dermis is effective also in the improvement and/or regeneration of the state of the dermis.

Further, the present inventors have clarified in more detail the existence site of mesenchymal stem cells in dermis and subcutaneous fat, and have found that platelet-derived growth factor-BB (PDGF-BB) is involved in the localization of mesenchymal stem cells, and that the enhancement of PDGF-BB production in vascular endothelial cells contributes to the enhancement and stabilization of mesenchymal stem cell production (japanese patent application 2010-209705).

Platelet-derived growth factor (PDGF) is a growth factor involved in regulation of migration, proliferation, and the like of mesenchymal stem cells such as fibroblasts, smooth muscle cells, glial cells, and the like, and is produced by various cells such as epithelial cells, endothelial cells, and the like. PDGF-A, PDGF-B, PDGF-C and PDGF-D are present in at least 4 of PDGFs, but 3 isomers (PDGF-AA, PDGF-AB, PDGF-BB) are present because a homodimer or heterodimer structure is obtained by disulfide bonding of the A chain and the B chain. PDGF is known to exhibit its physiological action via PDGF receptors (PDGFR) which are tyrosine kinase-coupled types. The PDGF-B gene is known and has been cloned (non-patent document 30).

If a component effective for enhancing PDGF-BB production can be found, it is considered that this component can be used to promote the production of mesenchymal stem cells and/or stabilize stem cells, and can be effectively used for various applications as described above.

Non-patent document 1: WattFM, JDermatolSci, 28: 173-180, 2002

Non-patent document 2: cotsarelis, ge, Cell, 57: 201-209, 1989

Non-patent document 3: nishimuraeket, Nature, 416: 854-860, 2002

Non-patent document 4: wongceal, JCellBiol, 175: 1005-1015, 2006

Non-patent document 5: pittenger mfet, Science, 284: 143-147, 1999

Non-patent document 6: parkkweital, CellMetab, 8: 454-457, 2008

Non-patent document 7: flynenetal, Cytotherapy, 9: 717-726, 2007

Non-patent document 8: igurakey, Cytotherapy, 6: 543-553, 2004

Non-patent document 9: dasilva meirellesleal, StemCells, 2008 Sep; 26(9): 2287-2299

Non-patent document 10: dasilva meirelles, leital, JCellSci, 2006; 119: 2204-2213

Non-patent document 11: daiwdeal, Circulation, 2005; 112: 214-223

Non-patent document 12: fazelSetal, JThoracCardiovascSurg, 2005; 130: 1310-1318

Non-patent document 13: noiseuxnet al, MolTher, 2006; 14: 840-850

Non-patent document 14: zhaolretal, ExpNeurol, 2002; 174: 11-20

Non-patent document 15: gneckimetal, NatMed, 2005; 11: 367-368

Non-patent document 16: kinnaird tetal, CircRes, 2004; 94: 678-685

Non-patent document 17: kinnaird tea, Circulation, 2004; 109: 1543-1549

Non-patent document 18: tangyletal, ann thoracsurg, 2005; 80: 229-237

Non-patent document 19: zhangmetal, FASEBJ, 2007; 21: 3197-3207

Non-patent document 20: lebancketal, jintermed, 2007; 262: 509-525

Non-patent document 21: uccelliaetal, TrendsImmunol, 2007; 28: 219-226

Non-patent document 22: caplanaietal, JCellBiochem, 2006; 98: 1076-1084

Non-patent document 23: fangbjetal, Transplantation, 2004; 78: 83-88

Non-patent document 24: orizzlaetal, procnatl acadsiiusa, 2003; 100: 8407-841

Non-patent document 25: xiaochuan jiahong, experimental medicine, 28: 1680-1687, 2010

Non-patent document 26: MedzhitovR, Nature, 454: 428-35, 2008

Non-patent document 27: hitamisligilgs, Nature, 444 (7121): 860-7, 2006

Non-patent document 28: wellenkeetal, JClinInvest, 115 (5): 1111-9, 2005

Non-patent document 29: xiapaxianxiata, experimental medicine, 28: 1717-1723, 2010

Non-patent document 30: dalla-fronterameter, Nature, 292: 31-35, 1981

Disclosure of Invention

Problems to be solved by the invention

The present invention has been made in view of the above-mentioned circumstances, and an object thereof is to provide a preparation effective for enhancing PDGF-BB production and a preparation effective for promoting mesenchymal stem cell production and/or stabilizing the same using the preparation.

Means for solving the problems

The present inventors have conducted extensive studies on various materials and screened agents for enhancing the production of PDGF-BB, and as a result, have found that plant sources of Vaccinium myrtillus, Shikimia japonica and Scutellaria baicalensis, and inositol phosphate each exhibit a significant PDGF-BB production enhancing effect, leading to completion of the present invention.

Accordingly, the present application includes the following inventions:

[1] a platelet-derived growth factor-BB (PDGF-BB) production enhancer comprising 1 or 2 or more members selected from the group consisting of a blueberry source, a mangosteen source, a scutellaria baicalensis source, inositol and inositol phosphate as active ingredients.

[2] The PDGF-BB production enhancer of [1], which comprises a yeast extract together with inositol.

[3] The PDGF-BB production enhancer of [1] or [2], wherein the inositol is an inositol phosphate derived from rice bran.

[4] A mesenchymal stem cell production promoter comprising the PDGF-BB production enhancer of any one of [1] to [3].

[5] A stem cell stabilizer comprising the PDGF-BB production-enhancing agent according to any one of [1] to [3].

[6] A dermal regeneration agent comprising the PDGF-BB production-enhancing agent according to any one of [1] to [3].

ADVANTAGEOUS EFFECTS OF INVENTION

The present invention provides an agent effective for enhancing PDGF-BB production and an agent effective for promoting and/or stabilizing the production of mesenchymal stem cells by using the agent.

Detailed Description

[ cowberry fruit source ]

Vaccinium myrtillus (scientific name: Vaccinium vitis-idaeaaL.) is a plant of the genus Vaccinium of the family Ericaceae. In the present invention, as the source of blueberry, there can be used a material obtained by crushing and squeezing a flower, a flower ear, a fruit peel, a fruit, a stem, a leaf, a branch and leaf, a trunk, a bark, a rhizome, a root bark, a root, a seed or a whole plant of blueberry, a material obtained by extracting with a solvent or the like after crushing, or a material obtained by decomposing by a treatment such as an enzyme treatment or a mechanical treatment after crushing.

In the case of extracting cranberries, the extraction solvent may be any solvent that can be used for extraction, and may be used alone or in combination with, for example, water, alcohols such as methanol, ethanol, propylene glycol, 1, 3-butylene glycol, and glycerol, and organic solvents such as chloroform, dichloroethane, carbon tetrachloride, acetone, and ethyl acetate.

The extraction may be performed at normal temperature or under heating (for example, using a heated solvent such as warm water or hot water).

The extract obtained by the above-mentioned solvent extraction or the concentrated extract may be used as it is, or the extract may be subjected to impurity removal by an adsorption method such as ion exchange resin, or may be adsorbed by a column of a porous polymer (e.g., アンバ - ライト XAD-2) and then eluted with methanol or ethanol, followed by concentration. Further, an extract obtained by extraction by a known partition method (for example, a partition method using water/ethyl acetate or the like) may be used.

In the case of decomposing vaccinium vitis-idaea, a "unicellular treatment" capable of selectively decomposing the intercellular substance and maintaining the cell morphology may be performed. For example, the method described in japanese patent laid-open No. 2002-. Specifically, the following treatment is performed: in plants, the intercellular substance is selectively decomposed or destroyed by enzymolysis, mechanical decomposition or the like without substantially decomposing the cell wall, and the object is decomposed into cell units in a state of maintaining the cell morphology, and made into porridge, liquid composition or freeze-dried product. For the above selective decomposition, enzyme treatment may be used. For example, an enzyme isolated or purified by a generally known method from a zymogen material of Rhizopus (Rizopus), Aspergillus (Aspergillus), or the like may be used, or a commercially available product may be used. When these single-cell enzymes are allowed to act on cranberries, it is preferable to allow these enzymes to act under optimum conditions such as the optimum temperature and optimum pH of the enzymes used, and the minimum amount is necessary. The optimum temperature of the unicellular enzyme is 30-45 ℃, and the optimum pH value is 4-6. The residue after the enzymatic reaction can be removed by using a sieve of about 20 meshes, for example, to prepare a cranberry source that can be contained in the external composition.

In addition to the suspension of the unicellular plant, a unicellular paste obtained by dehydrating the suspension by a method such as centrifugation or the like, or a dried product thereof may be used in the present invention.

Regarding the blueberry, there have been hitherto used examples as an external preparation containing the blueberry after the unicellular treatment (Japanese patent laid-open publication No. 2002-193734), a cosmetic for sterilization, antibacterial and dandruff prevention (Japanese patent laid-open publication No. 61-238719), an elastase activity inhibitor (Japanese patent laid-open publication No. 2002-363088), an active enzyme scavenger (Japanese patent laid-open publication No. 2002-363027), a melanogenesis inhibitor (Japanese patent laid-open publication No. 2002-305367), a lipid peroxidation inhibitor (Japanese patent laid-open publication No. 2002-363089), a collagenase activity inhibitor (Japanese patent laid-open publication No. 2003-12531), a hyaluronidase activity inhibitor (Japanese patent laid-open publication No. 2003-73287), a metal protease expression inhibitor (Japanese patent laid-open publication No. 2002-193738), an anti-glycation agent (Japanese patent laid-open publication No. 2004-505007), and an anti-skin injury agent against skin injury such as, however, it has not been known that a bilberry source has PDGF-BB production promoting activity, mesenchymal stem cell production promoting activity, stem cell stabilizing activity and dermal stabilizing activity, and these activities have been found by the present inventors for the first time.

[ Shikimchis source ]

Mangosteen (scientific name: Garciniamangostana) is a plant of the genus Garcinia of the family Guttiferae. In the present invention, as the mangosteen source, there can be used a material obtained by crushing and squeezing the fruit ear, fruit peel, fruit, stem, leaf, branch and leaf, trunk, bark, rhizome, root bark, root, seed or whole plant of mangosteen, or a material obtained by crushing and extracting the crushed material with a solvent or the like.

When mangosteen is extracted, any extraction solvent may be used as long as it can be used for extraction, and for example, the extraction solvent can be obtained by extracting with various organic solvents such as a lower alcohol such as methanol, ethanol, propanol, isopropanol, butanol, and isobutanol, a polyhydric alcohol such as aqueous lower alcohol, propylene glycol, and 1, 3-butanediol, or aqueous polyhydric alcohol, acetone, and ethyl acetate, and distilling off the solvent.

The extraction may be performed at normal temperature or under heating (for example, using a heated solvent such as warm water or hot water).

In addition, an enzyme may be added to the solvent to perform the extraction treatment. By adding the enzyme, the cell tissue of the fruit can be destroyed, and thus the extraction efficiency can be further improved. As the enzyme, a cell tissue macerating enzyme is preferably used. Examples of such enzymes include pectinase, cellulase, hemicellulase, α -amylase, and phytase. These enzymes may be used alone in 1 kind, or in a mixture of 2 or more kinds.

The extract obtained by using the above-mentioned extraction solvent may be used as it is, or the extraction solvent may be distilled off and dried as necessary before use.

Regarding shikimia, there have been examples of use as ultraviolet absorbers (Japanese patent laid-open No. 9-87155), MMPs inhibitors (Japanese patent laid-open No. 2003-252745), and fibroblast activators (Japanese patent laid-open No. 2006-249051), and it is known that it has effects of xanthone derivative production and collagen production (Japanese patent laid-open No. 2009-84169), but shikimia sources have PDGF-BB production promoting effect, mesenchymal stem cell production promoting effect, stem cell stabilizing effect, and dermal stabilizing effect, which have been found by the present inventors for the first time.

[ Baikal skullcap root source ]

Scutellaria baicalensis Georgi (Labiatae) is a plant of the genus Scutellaria of the family Labiatae. In the present invention, as the source of scutellaria, a substance obtained by crushing and squeezing the cluster, pericarp, fruit, stem, leaf, branch and leaf, trunk, bark, rhizome, root bark, root, seed or whole plant of scutellaria, or a substance obtained by crushing and extracting with a solvent or the like can be used.

In the case of extracting scutellaria baicalensis, any extraction solvent may be used as long as it is a solvent that can be generally used in plant extraction and the like, and examples thereof include water, lower alcohols such as methanol, ethanol, isopropanol, and n-butanol, polyhydric alcohols such as propylene glycol and 1, 3-butanediol, aqueous solutions of these alcohols, and hydrocarbon solvents such as n-hexane and toluene. These solvents may be used alone or in combination of any two or more thereof. Among them, lower alcohols such as methanol and/or ethanol are preferably used.

The extraction may be performed at normal temperature or under heating (for example, using a heated solvent such as warm water or hot water).

In addition, an enzyme may be added to the solvent to perform the extraction treatment. By adding the enzyme, the cell tissue of the fruit can be destroyed, and thus the extraction efficiency can be further improved. As the enzyme, a cell tissue macerating enzyme is preferably used. Examples of such enzymes include pectinase, cellulase, hemicellulase, α -amylase, and phytase. These enzymes may be used alone in 1 kind, or in a mixture of 2 or more kinds.

The extract obtained by using the above-mentioned extraction solvent may be used as it is, or the extract may be used after the extraction solvent is distilled off and dried as necessary.

As for scutellaria baicalensis, there have been hitherto used examples as a papilla activator (japanese patent application laid-open No. 11-240823), a hyaluronic acid production ability enhancer (japanese patent application laid-open No. 10-95735), an immunoactivator for preventing ultraviolet skin immune function reduction (japanese patent application laid-open No. 11-71295), an anti-inflammatory agent (japanese patent application laid-open No. 2006-8536), and an antioxidant (japanese patent application laid-open No. 5-238925), but scutellaria baicalensis-derived substances have been hitherto unknown as having PDGF-BB production promoting action, mesenchymal stem cell production promoting action, stem cell stabilizing action, and dermal stabilizing action, and these actions have been found by the present inventors for the first time.

[ inositol, inositol-phosphate ]

Inositol is considered to be one of vitamin B, which is also known to be biosynthesized from glucose in the organism. Inositol is also contained in plants such as grains, bran, beans, and fruits. Inositol often exists in a form in which its hydroxyl group is phosphorylated (inositol phosphate). In the present invention, the above-mentioned inositol phosphates may also be used. Inositol for use in the present invention includes various stereoisomers of inositol. As stereoisomers of inositol, cis-inositol (1, 2, 3, 4, 5, 6/0-inositol), epi-inositol (1, 2, 3, 4, 5/6-inositol), iso-inositol (1, 2, 3, 4/5, 6-inositol), myo-inositol (1, 2, 3, 5/4, 6-inositol, muco-inositol (1, 2, 4, 5/3, 6-inositol), neo-inositol (1, 2, 3/4, 5, 6-inositol), chiro-inositol (1, 2, 4/3, 5, 6-inositol) and scyllo-inositol (1, 3, 5/2, 4, 6-inositol) are mentioned, and as inositol phosphate, a compound in which any 1 or 2 or more of 6 hydroxyl groups of inositol are phosphorylated is mentioned, phytic acid is preferably phytic acid which is a compound in which all of the 6 hydroxyl groups of inositol are phosphorylated. The inositol or inositol phosphate used in the present invention is preferably derived from rice bran. In addition, any salt of inositol phosphates may also be used in the present invention.

According to a certain embodiment of the present invention, inositol and inositol phosphate can be used by dissolving the respective powders in a medium to be appropriately diluted.

Inositol and inositol phosphates may also enhance PDGF-BB production by combination with other substances. Examples of the other substances include yeast extracts, and バイオダイン (registered trademark) EMPP (manufactured by ArchPersonal careproducts L.P.) can be used. When the extract is used, about 0.1% of the extract can be added to the medium for use by converting the concentration of the dry residue of the extract.

As regards inositol and inositol phosphate, examples have hitherto been used as cosmetic compositions for aged skin and/or stressed skin containing inositol (Japanese patent publication No. 204-501069), cosmetic compositions for reducing or preventing symptoms of fatty edema containing inositol phosphate (in particular, inositol hexaphosphate and salts thereof) (Japanese unexamined patent publication No. Hei 8-253406), and compositions for skin moisturizing and/or skin protection and/or skin anti-aging (Japanese unexamined patent publication No. 2010-150258). Further, as the vitamin B group, there are used an external preparation for skin for activating cells (Japanese patent application laid-open No. 9-241146) containing vitamin B2 and B6, and an antioxidant containing vitamin B (Japanese patent application laid-open No. 7-277939, Japanese patent application laid-open No. 205306831, etc.). However, the vitamin B group or inositol phosphate having PDGF-BB production promoting action, mesenchymal stem cell production promoting action, stem cell stabilizing action and dermal stabilizing action, which have been found by the present inventors for the first time, have not been known so far.

[ PDGF-BB production enhancer, mesenchymal stem cell production promoter, stem cell stabilizer and dermal regeneration agent ]

The PDGF-BB production enhancer of the present invention comprises 1 or 2 or more members selected from the group consisting of a blueberry source, a mangosteen source, a scutellaria baicalensis source, inositol and inositol phosphate as active ingredients. The mesenchymal stem cell production promoter, stem cell stabilizer and dermal regeneration agent of the present invention contain the PDGF-BB production enhancer of the present invention containing the above-described active ingredient. The PDGF-BB production enhancer, the mesenchymal stem cell production promoter, the stem cell stabilizer, and the dermal regeneration agent of the present invention (hereinafter, these agents are collectively referred to as "the preparation of the present invention") may contain any 1 of the above active ingredients alone, or may contain any 2 or more of the above active ingredients in any combination and ratio.

As described above, according to the findings of the present inventors, PDGF-BB is involved in localization of mesenchymal stem cells and enhances PDGF-BB production in vascular endothelial cells, and thus it is possible to enhance and stabilize the production of mesenchymal stem cells (Japanese patent application No. 2010-209705). That is, the PDGF-BB production enhancer of the present invention can be used very effectively for the purpose of enhancing and stabilizing the production of mesenchymal stem cells.

As described above, it is found that enhancement and stabilization of mesenchymal stem cell production are extremely effective for applications such as stabilization of blood vessels, maintenance of tissue homeostasis, repair and/or regeneration of damaged tissues (particularly, regeneration of dermis), prevention and/or treatment of various diseases such as fibrosis, multiple sclerosis and/or diabetes, and prevention and/or improvement of various states of chronic inflammation such as metabolic syndrome. Therefore, the mesenchymal stem cell production promoter and stem cell stabilizer of the present invention using the PDGF-BB production enhancer of the present invention can be very effectively used for these applications.

The preparation of the present invention may be a composition comprising the above-mentioned active ingredient in combination with 1 or 2 or more other ingredients, for example, excipients, carriers, diluents and the like. The composition and form of the composition are arbitrary, and may be appropriately selected depending on the conditions such as the active ingredient and the use. The composition can be produced by a conventional method according to a formulation in which the composition is appropriately combined with an excipient, a carrier and/or a diluent, and other components, depending on the dosage form.

The preparation of the present invention can be incorporated into various foods, drinks, feeds (pet foods, etc.) and ingested by humans and animals. Furthermore, the compound can be used in cosmetics and the like for humans and animals, or can be administered as a pharmaceutical preparation to humans and animals.

Specifically, when the preparation of the present invention is incorporated into foods, drinks, feeds, and the like, the amount of the plant or the extract thereof to be incorporated (dry mass) can be determined as appropriate depending on the type, purpose, form, method of use, and the like of the plant or the extract thereof. For example, the plant or extract thereof may be formulated so that the daily intake of an adult is about 0.5mg to 1g (dry residue) in the case of an extract of emblic leafflower fruit (amla) and about 0.5mg to 3g in the case of an extract of cranberry. Particularly, when the extract is used as a health food or beverage, it is preferable to contain the extract in an amount of 10mg to 500mg (dry residue) in the case of an emblic leafflower fruit extract and 10mg to 1.5g (dry residue) in the case of a blueberry extract, so that the predetermined effect of the active ingredient of the present invention can be sufficiently exhibited by an adult daily intake.

The form of the food, drink or feed may be any form, and may be, for example, a pellet, a granule, a paste, a gel, a solid or a liquid. In these forms, known various substances that are allowed to be contained in foods, drinks, and the like, for example, excipients such as binders, disintegrants, thickeners, dispersants, resorption accelerators, taste correctives, buffers, surfactants, solubilizing agents, preservatives, emulsifiers, isotonic agents, stabilizers, pH adjusters, and the like can be appropriately contained.

When the preparation of the present invention is applied to cosmetics, the amount of plant or extract thereof to be blended (dry mass) can be determined appropriately according to the type, purpose, form, method of use, and the like of the plant or extract thereof. For example, the emblic extract and the cranberry extract may be added to the total amount of the cosmetic composition in an amount of 0.00001% to 50% (in terms of dry mass), respectively, and preferably 0.0001% to 5% (in terms of dry mass), respectively.

In addition to the above components, components generally used in skin external preparations such as cosmetics and medicines, for example, antioxidants, oil components, ultraviolet ray protection agents, surfactants, thickeners, alcohols, powder components, coloring materials, aqueous components, water, various skin nutrients, and the like may be appropriately blended as necessary within a range not to impair the effects of the present invention.

Further, disodium ethylenediaminetetraacetate, trisodium ethylenediaminetetraacetate, sodium citrate, sodium polyphosphate, sodium metaphosphate, a metal ion chelating agent such as gluconic acid, a preservative such as methyl paraben, ethyl paraben, and butyl paraben, a medicament such as caffeine, tannin, verapamil, tranexamic acid and its derivatives, a licorice extract, glabridin, a hot water extract of papaya fruits, various crude drugs, tocopherol acetate, glycyrrhizic acid and its derivatives or its salts, a whitening agent such as vitamin C, magnesium ascorbyl phosphate, ascorbyl glucoside, arbutin, and kojic acid, and a saccharide such as glucose, fructose, mannose, sucrose, and trehalose may be appropriately blended.

The skin external preparation can be widely used in cosmetics, quasi drugs, etc. applied to the outer skin, and particularly preferably in cosmetics. The formulation may be any formulation such as a solution system, a solubilization system, an emulsification system, a powder dispersion system, a water-oil two-layer system, a water-oil-powder three-layer system, an ointment, a cosmetic water, a gel, and an aerosol, as long as the formulation can be applied to the skin.

When the preparation of the present invention is used as a cosmetic, it is preferably used in the form of a lotion, an emulsion, a foundation, a lipstick, a lip balm, a cleansing cream, a massage cream, a pack, a hand cream, a hand powder, a body shampoo, a body lotion, a body cream, a bath cosmetic, etc., and in this case, components usually added and blended when these forms are prepared, for example, components such as a moisturizer, a perfume, a solubilizer, a stabilizer, an ultraviolet absorber, an ultraviolet scattering agent, etc., may be appropriately blended.

When the preparation of the present invention is applied as an external preparation, the amount of plant or extract thereof to be added (dry mass) may be determined appropriately depending on the type, purpose, form, method of use, and the like of the plant or extract. For example, the content is preferably 0.00001% to 50% (in terms of dry mass) in the total amount of the cosmetic, and is preferably 0.0001% to 5% (in terms of dry mass) in other foods and beverages.

When the preparation of the present invention is used as a quasi-drug, the preparation can be suitably used orally or parenterally (intravenous administration, intraperitoneal administration, etc.). The dosage form is also arbitrary, and for example, it may be a solid preparation for oral administration such as a tablet, granule, powder, capsule, etc., a liquid preparation for oral administration such as an oral liquid, syrup, etc., or a liquid preparation for non-oral administration such as an injection, etc., and these forms can be appropriately prepared by a known method. In these pharmaceutical preparations, excipients such as a binder, a disintegrant, a thickener, a dispersant, a resorption accelerator, a corrigent, a buffer, a surfactant, a solubilizer, a preservative, an emulsifier, an isotonic agent, a stabilizer, a pH adjuster, and the like, which are generally used, can be appropriately used.

In the case of an external preparation, the preparation can be widely used in the form of various ointments and the like, but is preferably used in the form of a lotion, a suspension, and/or an emulsion, a liquid, an ointment, a patch and the like. In addition, the form that the formulation of the present invention can take is not limited to these dosage forms and forms.

The expression of PDGF-BB gene in mesenchymal stem cells when the preparation of the present invention is administered can be evaluated by, for example, determining the expression level by measuring the PDGF-BB level. Preferably, the measurement can be carried out by a method known in the art, for example, an immunostaining method using a fluorescent substance, a dye, an enzyme, or the like, a western blotting (western blotting) method, an immunoassay method such as an ELISA method, a RIA method, or the like, using an antibody specific to PDGF-BB. In addition, for example, total RNA in mesenchymal stem cells may be extracted, and the expression level may be determined by measuring the amount of mRNA encoding PDGF-B, thereby evaluating the expression level. Extraction of mRNA and measurement of the amount thereof are also well known in the art, and for example, quantification of RNA can be performed by a quantitative Polymerase Chain Reaction (PCR) method, such as real-time polymerase chain reaction (RT-PCR). Primers suitable for RT-PCR can be selected by methods well known to those skilled in the art.

Examples

The present invention will be described in more detail by way of examples. In addition, the present invention is not limited thereto.

[ sample to be evaluated ]

The following samples were used as the evaluation target samples for the PDGF-BB production enhancing effect.

Inositol:

commercially available inositol powder (manufactured by Wako pure chemical industries, Ltd.: myo-inositol) was dissolved in PBS and used in an amount of 10ppm based on the medium described later.

Phytic acid:

commercially available phytic acid (manufactured by ナカライテスク, 50% aqueous solution) was used in an amount of 10ppm based on the medium described later.

Scutellaria baicalensis extract:

an extract obtained by extracting roots of scutellaria baicalensis after the removal of pericarp with 70 vol% ethanol (a mixed solution of water and ethanol at a volume ratio of 3: 7) was used. The extract was stored in a dry state, dissolved by adding 70 vol% ethanol immediately before use, and used in an amount of 15ppm relative to a medium (in terms of the dry weight of the extract) described later.

Cranberry CRS (cell release system):

a suspension obtained by adding 1, 3-BG40 vol% to a cell suspension obtained by treating leaves of Vaccinium myrtillus with a cell lytic enzyme (マセロチ - ム A) was used. The suspension was stored as a liquid and used in an amount of 30ppm relative to a culture medium (in terms of the dry weight of the extract) described later.

Mangosteen bark extract:

an extract obtained by extracting bark of mangosteen with 70 vol% 1, 3-BG (mixed solution of water and 1, 3-BG at a volume ratio of 3: 7) is used. The extract was stored in a dry state, and immediately before use, 1, 3-BG was dissolved in an amount of 70 vol% and then used in an amount of 10ppm based on a culture medium (in terms of the dry weight of the extract) described later.

[ evaluation of PDGF-BB production-enhancing action in vascular endothelial cells ]

HUVEC of human vascular endothelial cells were subcultured with EGM-2 medium (Sanguang pure drug), cells at passage 4 were suspended in Humedia-EG2 medium (クラボウ) without VEGF-A, seeded at a rate of 20,000 in collagen-coated 24-well multi-well plates (Asahi glass) and seeded at 5% CO₂In the presence of the culture medium, the cells were cultured at 37 ℃ for 3 to 5 days until the cells became dense. The above samples were added at the above concentrations or replaced by the addition of the evaluation pairs for dissolutionHumedia-EG2 medium (クラボウ) like the solvent of the sample, and then cultured for another 2 days. mRNA was extracted and purified from the cultured cells according to the protocol provided using an RNA extraction reagent MagNAPurceLCmRNAHSKit (Roche) and an automatic nucleic acid extraction apparatus MagNAPurceLC1.0 Instrument (Roche). For each sample, quantitative real-time (RT) -PCR was carried out by one-step method using the primer set of SEQ ID Nos. 1 and 2, the reaction reagent QuantiFastSYBRGreenRT-PCRKirt (Qiagen), and the reaction apparatus LightCycler (Roche) described later, using mRNA of the same volume as a template. The composition conditions were according to the protocol of Qiagen. In addition, RT-PCR conditions were RT reaction at 50 ℃ for 20 minutes, initial denaturation at 95 ℃ for 15 minutes, denaturation at 94 ℃ for 15 seconds, annealing at 60 ℃ for 20 seconds, and extension at 72 ℃ for 30 seconds. G3PDH was used as an internal standard (primer set of SEQ ID NOS: 3 and 4), and the amount of mRNA in the control group was corrected using the internal standard.

PDGF-B：

A forward primer: 5'-CCTGGCATGCAAGTGTGA-3' (Serial number 1)

Reverse primer: 5'-CCAATGGTCACCCGATTT-3' (Serial number 2)

G3PDH：

A forward primer: 5'-GCACCGTCAAGGCTGAGAAC-3' (SEQ ID NO. 3)

Reverse primer: 5'-ATGGTGGTGAAGACGCCAGT-3' (Serial number 4)

[ evaluation results ]

The ratio of the expression amount of PDGF-BB mRNA obtained from each of the above samples to the expression amount obtained from a control (solvent for dissolving each sample to be evaluated) according to the above evaluation procedure is shown in the following table. As is clear from the results below, these components have an activity of enhancing PDGF-BB expression.

[ Table 1]

Claims (9)

1. Use of 1 or more than 2 ingredients selected from cowberry fruit source, radix Scutellariae source, inositol and phytic acid in preparation of PDGF-BB production enhancer,

the cowberry fruit source is cowberry fruitVacciniumvitis-idaeaL.The cell-decomposing enzyme-decomposed solution of leaf (1),

the mangosteen source is mangosteenGarciniamangostanaThe 1, 3-butylene glycol or aqueous 1, 3-butylene glycol extract of bark of the tree,

the source of Scutellariae radix is yellowRoot of large-flowered skullcapScutellariabaicalensisGeorgiThe root of (a) or an aqueous ethanol extract.

2. Use of 1 or more than 2 components selected from cowberry fruit source, shikimia source, Scutellariae radix source, inositol and phytic acid in preparing mesenchymal stem cell localizer,

the cowberry fruit source is cowberry fruitVacciniumvitis-idaeaL.The cell-decomposing enzyme-decomposed solution of leaf (1),

the mangosteen source is mangosteenGarciniamangostanaThe 1, 3-butylene glycol or aqueous 1, 3-butylene glycol extract of bark of the tree,

the source of radix Scutellariae is radix ScutellariaeScutellariabaicalensisGeorgiThe root of (a) or an aqueous ethanol extract.

3. Use of 1 or more than 2 components selected from cowberry fruit source, radix Scutellariae source, inositol and phytic acid in preparing stem cell stabilizer,

the cowberry fruit source is cowberry fruitVacciniumvitis-idaeaL.The cell-decomposing enzyme-decomposed solution of leaf (1),

the mangosteen source is mangosteenGarciniamangostanaThe 1, 3-butylene glycol or aqueous 1, 3-butylene glycol extract of bark of the tree,

the source of radix Scutellariae is radix ScutellariaeScutellariabaicalensisGeorgiThe root of (a) or an aqueous ethanol extract.

4. Use of 1 or more than 2 components selected from cowberry fruit source, mangosteen source and inositol in preparing dermis regeneration agent,

the cowberry fruit source is cowberry fruitVacciniumvitis-idaeaL.The cell-decomposing enzyme-decomposed solution of leaf (1),

the mangosteen source is mangosteenGarciniamangostanaThe bark of (A) is 1, 3-butylene glycol or an aqueous 1, 3-butylene glycol extract.

5. Use of PDGF-BB production enhancer, which is platelet-derived growth factor-BB, comprising 1 or more than 2 members selected from the group consisting of a bilberry source, a mangosteen source, a Scutellaria baicalensis source, inositol and phytic acid, for producing a mesenchymal stem cell-localizing agent,

the cowberry fruit source is cowberry fruitVacciniumvitis-idaeaL.The cell-decomposing enzyme-decomposed solution of leaf (1),

the mangosteen source is mangosteenGarciniamangostanaThe 1, 3-butylene glycol or aqueous 1, 3-butylene glycol extract of bark of the tree,

the source of radix Scutellariae is radix ScutellariaeScutellariabaicalensisGeorgiThe root of (a) or an aqueous ethanol extract.

6. Use of PDGF-BB production enhancer comprising 1 or more than 2 members selected from cowberry fruit source, shikimia source, Scutellaria baicalensis source, inositol and phytic acid for preparing stem cell stabilizer,

the cowberry fruit source is cowberry fruitVacciniumvitis-idaeaL.The cell-decomposing enzyme-decomposed solution of leaf (1),

the mangosteen source is mangosteenGarciniamangostanaThe 1, 3-butylene glycol or aqueous 1, 3-butylene glycol extract of bark of the tree,

the source of radix Scutellariae is radix ScutellariaeScutellariabaicalensisGeorgiThe root of (a) or an aqueous ethanol extract.

7. Use of PDGF-BB production enhancer comprising 1 or more than 2 members selected from cowberry fruit source, mangosteen fruit source and inositol for preparing dermal regeneration agent,

the cowberry fruit source is cowberry fruitVacciniumvitis-idaeaL.The cell-decomposing enzyme-decomposed solution of leaf (1),

the mangosteen source is mangosteenGarciniamangostanaThe bark of (A) is 1, 3-butylene glycol or an aqueous 1, 3-butylene glycol extract.

8. Use according to any one of claims 1 to 7, which comprises a yeast extract in addition to inositol or phytic acid.

9. Use according to any one of claims 1 to 7, wherein the inositol or phytic acid is derived from rice bran.