JP2009149547A - Agent for promoting production of integrin and vinculin - Google Patents

Abstract

<P>PROBLEM TO BE SOLVED: To provide an agent for improving sagging skin and elasticity, which improves the sagging skin due to aging and imparts an elastic touch to the skin, and to provide a composition which has an effect for inhibiting the proliferation and metastasis of cancer cells. <P>SOLUTION: Glucose-1-phosphate having an effect for promoting the production of integrin and vinculin is contained as an active ingredient, and thereby the interaction of skin fibroblasts with an extracellular matrix is activated so that the elastic touch is imparted to the skin. Thus, there are provided the composition which improves the sagging and elasticity of the skin, and the composition having the effect for inhibiting the proliferation and metastasis of cancer cells. <P>COPYRIGHT: (C)2009,JPO&INPIT

Description

The present invention relates to an integrin and vinculin production promoter containing glucose-1-phosphate as an active ingredient, and as a field of use thereof, for example, by using it for cosmetics, foods and drinks, quasi drugs, pharmaceuticals, and the like. The present invention relates to a composition that improves skin sagging and imparts a feeling of elasticity, and a composition that has an effect of inhibiting cancer cell growth and metastasis.

The skin is always exposed to external stimuli such as dryness and ultraviolet rays, and as it ages, it loses its elasticity and elasticity, resulting in sagging. Conventionally, as a composition that improves the sagging of the skin and gives elasticity to the skin, the adhesion between the epidermis and the basement membrane is improved and strengthened, or the production of integrins in the epidermis cells is promoted to promote skin wrinkles, sagging, Proposals have been made to improve the texture structure. Moreover, what expects the effect which tightens skin with an astringent is known. Patent Document 1 exemplifies a composition for promoting the production of integrin α6β4 for keratinocytes containing plants of kizota, echinacea, pumpkin, dandelion, and angelica or extracts thereof.
Patent Document 2 also includes a laminin 5 production promoter that is a skin basement membrane activation component for keratinocytes and an integrin α6β4 production promoter that is a skin basal cell activation component. Examples are compositions that prevent, prevent and improve.
Furthermore, Patent Document 3 also discloses an integrin production promoting substance for keratinocytes, which is a hyssop belonging to the genus Willow genus, stevia belonging to the genus Stevia, broccoli belonging to the genus Brassica, mallows belonging to the genus Mallow, and hyacinths. Agents are illustrated.
JP2003-171225A JP 2003-226655 A Japanese Patent Application Laid-Open No. 2004-262861

However, improving and strengthening the adhesion between the epidermis and the basement membrane, and promoting the production of integrins by epidermal cells are not sufficient to improve skin wrinkles and sagging, and tighten the skin with astringents. The effect was only a temporary action and could not continuously improve the sagging / elasticity of the skin.

Therefore, the present inventors paid attention to the interaction between dermal fibroblasts and an extracellular matrix such as collagen. In other words, it acts on skin fibroblasts to promote the production of fibroblast integrins and vinculin, thereby strengthening adhesion to the dermal extracellular matrix and improving skin sagging and elasticity. Is to provide.

The skin is mainly divided into three layers: epidermis, dermis, and subcutaneous tissue. Among them, the dermis is extremely important for maintaining the structure of the skin, and it is made firmly and flexibly by fibers such as collagen, fibronectin, and elastin. Forming an organization. Human skin cells, particularly human skin fibroblasts, control the state of connective tissue by interacting with fibers such as collagen, fibronectin, and elastin. Therefore, the interaction between dermal fibroblasts and extracellular matrix such as collagen is deeply related to the maintenance of skin sagging and elasticity.

Therefore, the present inventors have focused on integrin and vinculin, which are adhesion proteins that have not attracted much attention in skin cells, and by increasing the amount of integrin and vinculin protein, skin cells, in particular skin fibroblasts, are obtained. It was found that sagging and elasticity of skin can be improved by activating the interaction between cells and extracellular matrix.

On the other hand, it is known that integrin, particularly integrin β-1 subunit is strongly expressed in many cancer cells. Integrins are biomolecules that form a family of molecules that form α and β heterodimers, and are known to bind to cell surface ligands and extracellular matrix. Integrins are further classified into β-1, β-2, and β-3 subgroups by their β subunits. It has also been reported that the expression level of integrin is closely related to the malignancy such as growth rate and metastasis of cancer cells in vivo. For example, it has been reported that α ₅ β ₁ belonging to the β-1 subgroup no longer shows tumor formation when its expression level increases (Cell. 60, 849-859 (1990)). Thus, the importance of integrins has increased in recent years, and application to the treatment and prevention of leukocyte adhesion inhibition, platelet aggregation inhibition, cancer metastasis inhibition, myocardial infarction, arteriosclerosis, osteolytic disease and the like can be expected. In addition, vinculin is best known as one of the integrin-lined proteins, and it is known to enhance adhesion by enhancing expression and suppress cell movement.

That is, the present invention provides a human skin cell integrin and vinculin production promoter containing glucose-1-phosphate as an active ingredient.

In the present invention, the integrin and vinculin are not particularly limited as long as they are expressed in human skin cells, but those expressed in human skin fibroblasts are particularly preferable. If it is expressed in human skin fibroblasts, a particularly excellent skin tightening effect can be exhibited.

Integrins are composed of α subunits and β subunits, α subunits further include α1 to α5, αL, etc., and β subunits include β1, β2, β3, etc., but collagens present in various connective tissues , Vitronectin, fibronectin, laminin, etc., and the interaction between the cells present in the connective tissue such as fibroblasts, it is preferable that the expression of α1 subunit, α2 subunit, and β1 subunit is promoted. Furthermore, it is more desirable that expression of α2 subunit is promoted and expression of β1 subunit is promoted at the same time. In particular, regarding dermal fibroblasts, expression of α2β1 integrin is preferably promoted from the viewpoint of interaction with collagen.

The integrin and vinculin production promoter of the present invention contains glucose-1-phosphate as an active ingredient. Glucose-1-phosphate is a glucose metabolite in vivo, and UDP-D-glucuronic acid important for detoxification and excretion of organic substances is produced. UDP-D-glucuronic acid is used for detoxification by enhancing renal excretion of foreign substances such as phenol and drugs. It is also an important substance as a precursor of D-glucuronic acid, an acidic polysaccharide such as hyaluronic acid and heparin.

Glucose-1-phosphate used in the present invention may be a commercially available reagent, or may be obtained by extraction from a microorganism such as yeast using a solvent.

The extraction solvent for obtaining the extract used in the present invention may be selected in consideration of the intended purpose and type of the product to be provided, or the processing performed later. For example, water; methyl alcohol, ethyl alcohol Lower monohydric alcohols such as: liquid polyhydric alcohols such as glycerin, propylene glycol, and 1,3-butylene glycol; ketones such as acetone and methyl ethyl ketone; alkyl esters such as ethyl acetate; hydrocarbons such as benzene and hexane; These can be extracted and purified using one or more of halogenated alkanes such as dichloromethane and chloroform.

In the case of extracting glucose-1-phosphate from a microorganism or the like, after drying the microorganism, 1 to 1000 times by weight, particularly 10 to 100 times the amount of solvent is used. In particular, it is preferably performed at 20 ° C. to 40 ° C. for 1 hour or longer, particularly 3 to 7 days. Moreover, you may heat-extract at 60-100 degreeC for 1 hour.

The above extract obtained under the above conditions may be used as an extracted solution, but if necessary, use a product that has been refined, filtered, concentrated, and powdered as appropriate. I can do it.

The form of glucose-1-phosphate used in the present invention may be in any form such as liquid, solid, powder, paste, gel, etc., and the optimum shape for configuring the final product is arbitrary. Can be selected.

The dosage form of the composition of the present invention is arbitrary, and examples of the external preparation for skin include pharmaceuticals or pharmaceuticals such as liquids, suspensions, emulsions, creams, ointments, gels, liniments, lotions, and poultices. An quasi-drug is preferred. Furthermore, it can mix | blend and use for pharmaceuticals, quasi-drugs, cosmetics, or food-drinks. In particular, it is applied to external preparation compositions such as pharmaceuticals, quasi drugs, and cosmetic compositions applied to the outer skin.

Specific use forms of the present invention include aqueous components, oily components, plant extracts, animal extracts, powders, excipients, surfactants, oils, alcohols, pH adjusters, preservatives, antioxidants, A lotion, emulsion, cream, pack, powder, spray, ointment, dispersion, cleaning agent for external preparation composition by mixing thickeners, sweeteners, pigments, fragrances and the like as necessary. In addition, various dosage forms such as liquid, paste, capsule, powder, and tablet can be used.

The amount of glucose-1-phosphate added to the skin composition according to the present invention is slightly different depending on the expected degree of action and is not particularly limited, but is usually 0.0001% by mass or more in terms of solid content in the total amount of the preparation, It is effective to make the concentration range preferably 0.01 to 20.0% by mass.

Hereinafter, test examples relating to the effects of the integrin and vinculin production promoter according to the present invention will be shown and application prescription examples to external preparations using the materials will be described, but the examples are not limited thereto. Needless to say.

Example 1
Cell culture cells: HFSKF-II (RIKEN)
Medium: D-MEM + FBS 15%
Normal human fibroblast HFSKF-II cells were cultured in D-MEM medium supplemented with 15% FBS. The cells were seeded and cultured on a 50 cm ² plate at about 50% confluence. On the next day, glucose-1-phosphate (reagent obtained from Wako Pure Chemical Industries) was added at each concentration. After the addition, the cells were collected 48 hours later and used as Western blotting samples.

Western blotting 7.6% acrylamide gel electrophoresis, transfer to PVDF membrane, Integrinβ-1 (M-106) sc-8978 (SANTA CRUZ BIOTECHNOLOGY, INC) Human (Rabbit polyclonal), vinculin antibody as integrin antibody Vinculin (H-10) sc-25336 (SANTA CRUZ BIOTECHNOLOGY, INC) Human (mouse monoclonal), ECL ^TM Anti-rabbit IgG as secondary antibody, Horseradish Peroxidase linked whole
antibody (from donkey) NA934V (Amersham
Bioscience UK Limited), ECL ^TM Anti-mouse IgG, Horseradish Peroxidase linked whole antibody (from sheep) NA931V (Amersham Bioscience UK Limited), Western blotting confirmed 130KDa integrin protein and 117KDa vinculin protein did.

Measurement of integrin and vinculin protein amounts Band intensities obtained by Western blotting were analyzed by Dolphin-DOC (WEALTEC). The area of the integrin protein band intensity in the non-sample-added group was defined as 100, and the integrin band intensity area in each extract-added group was proportionally calculated to obtain the integrin protein amount. Vinculin protein was also treated in the same manner as integrin.

The experimental results are shown in Table-1. As a result of analysis of the band intensity area, when the control group is 100, the integrin β1 production amount is 122 in the glucose-1-phosphate 2 mM addition group and 130 in the 4 mM addition group, and the integrin β1 by glucose-1-phosphate is shown. Production promotion effect was confirmed. Moreover, about the vinculin protein, the production amount of 118 was shown in the glucose-1-phosphate 2mM addition group, and 124 was produced in the 4mM addition group, and the vinculin production promotion effect by glucose-1-phosphate was confirmed.

(Example 2) Production of various compositions Various compositions according to the present invention were produced. Although the formulation example is shown below, this invention is not necessarily limited to these.
Examples of cosmetic formulations

(1) Cream composition
(weight%)
a) Beeswax
2.0
b) Stearyl alcohol
5.0
c) Stearic acid
8.0
d) Squalane
10.0
e) Self-emulsifying glyceryl monostearate
3.0
f) Polyoxyethylene cetyl ether (20E.O.)
1.0
g) Glucose-1-phosphate
2.0
h) 1,3-butylene glycol
5.0
i) Preservatives and antioxidants
Appropriate amount
j) Purified water
The rest
k) Potassium hydroxide
0.3
The process up to production methods a) to f) are dissolved by heating and kept at 80 ° C. G) to j) are heated and dissolved, kept at 80 ° C., and added to a) to f) to emulsify. Then, k), which has been dissolved in part of j) in advance, is added and cooled to 40 ° C. with stirring.

(2) Emulsion composition
(weight%)
a) Beeswax
0.5
b) Petrolatum
2.0
c) Squalane
8.0
d) Sorbitan sesquioleate
0.8
e) Polyoxyethylene oleyl ether (20E.O.) 1.2
f) Glucose-1-phosphate
5.0
g) 1,3-butylene glycol
7.0
h) Carboxyvinyl polymer
0.2
i) Potassium hydroxide
0.1
j) Purified water
The rest
k) Preservatives and antioxidants
Appropriate amount
l) Ethanol
7.0
The process up to production methods a) to e) are dissolved by heating and kept at 80 ° C. f) to k) are dissolved by heating,
Maintain at 80 ° C., emulsify in addition to a) to e), and cool to 50 ° C. with stirring. At 50 ℃
Add l) and cool to 40 ° C.

(3) Lotion-like composition
(weight%)
a) Glucose-1-phosphate

1.0
b) α-Linolenic acid 0.1
c) Glycerin
5.0
d) Polyoxyethylene sorbitan monolaurate (20E.O.) 1.0
e) ethanol
6.0
f) Fragrance
Appropriate amount
g) Preservatives and antioxidants
Appropriate amount
h) Purified water
Mix the remainder of the production methods a) to h) and dissolve uniformly.

(4) Packing agent
(weight%)
a) Glucose-1-phosphate
3.0
b) Vinyl acetate resin emulsion
15.0
c) Polyvinyl alcohol
10.0
d) Cuui seed oil
3.0
e) Glycerin
5.0
f) Titanium oxide
8.0
g) Kaolin
7.0
h) ethanol
8.0
i) Fragrance
Appropriate amount
j) Preservatives and antioxidants
Appropriate amount
k) Purified water
Mix the rest of the production methods a) to k), stir and disperse well, and make uniform.

(5) Ointment (wt%)
a) Liquid paraffin 20.0
b) White petrolatum 50.0
c) Cetanol 10.0
d) Polyoxyethylene (60) hydrogenated castor oil 18.0
e) Glucose-1-phosphate 2.0
Production method: a) to e) are added and mixed while heating to prepare an ointment.

(6) Gel agent (wt%)
a) Polyoxyethylene (60) hydrogenated castor oil 9.0
b) Carbopol 941 0.5
c) Ethanol 22.0
d) Triethanolamine 1.5
e) Glucose-1-phosphate 4.0
f) Purified water 63.0
Production method: While heating, add a), b) and e) to f) and dissolve, then add d) and c) and cool with stirring to prepare a gel.

(7) Solution (wt%)
a) Polyoxyethylene (60) hydrogenated castor oil 10.0
b) Ethanol 20.0
c) Glucose-1-phosphate 5.0
d) Purified water 65.0
Production method: a) to d) are added and dissolved to prepare a solution.

(8) Tablet (wt%)
a) Starch 72.0
b) Sucrose fatty acid ester 8.0
c) Glucose-1-phosphate 20.0
Production method: Raw materials of a) to c) were mixed and molded with a tableting machine to produce an 8 mm diameter uncoated tablet. This uncoated tablet was film-coated by a conventional method to complete a tablet.

(9) Capsule a) Lactose 190mg
b) Magnesium stearate 10mg
c) Glucose-1-phosphate 10 mg
210mg total
Production method: a) to c) were mixed uniformly, passed through a 60 mesh sieve, and the whole powder was filled into a No. 2 gelatin capsule.

(10) Drink a) Lemon juice 20mL
b) Propolis 0.3g
c) Vitamin C 0.2g
d) Honey 13.0g
e) Glucose-1-phosphate 1.0 g
f) Water proper amount total 100mL
Production method: After sufficiently stirring a) to e), f) was added to make the total amount 100 mL.

As described above, the integrin / vinculin production promoter according to the present invention enhances the adhesion between dermal fibroblasts and dermal extracellular matrix by being used in cosmetics, foods and drinks, quasi drugs, pharmaceuticals, etc. The present invention provides a composition having an effect of improving skin sagging and elasticity, and can be used for a composition having an effect of inhibiting the growth of cancer cells and metastasis.

Claims (4)

Integrin production promoter containing glucose-1-phosphate as an active ingredient The integrin production promoter according to claim 1, wherein the integrin of claim 1 is integrin β1. The integrin production promoter according to claim 1, wherein the integrin of claim 1 is integrin α2β1. A vinculin production promoter containing glucose-1-phosphate as an active ingredient.