KR20120002965A - Composition for promoting the differentiation of human mesenchymal stem cell to adipocytic cells - Google Patents

Abstract

PURPOSE: A composition which promotes differentiation of human mesenchymal stem cells into adipocytes is provided to enhance skin volume and elasticity and to prevent wrinkles. CONSTITUTION: An external use skin composition for anti-aging contains extract of Spatholobus suberectus Dunn., Alpinia officinarum Hance, or Angelica pubescens for. biserrata Shan Etyuan as an active ingredient. The composition is used for anti-wrinkling or elasticity improvement. The mesenchymal stem cells are derived from human. A pharmaceutical composition for promoting differentiation of adipocytes contains extract of Mucunae Caulis, Alpiniae officinari Rhizoma or Araliae cordatae Radix as an active ingredient.

Description

Composition for promoting the differentiation of human mesenchymal stem cell to adipocytic cells}

The present invention relates to a composition for promoting the differentiation of human mesenchymal stem cells into adipocytes, and more specifically, human mesenchymal cells containing at least one or more of the extracts such as cinnamon, high-volume rivers, venom and cheonnyeonja as an active ingredient By promoting differentiation of stem cells into adipocytes, the present invention relates to a composition for producing lipid droplets to promote skin volume and elasticity and improve wrinkles.

Skin is the largest organ in the human body, occupying about 16% of our body volume, 1.8 m ² , 2 to 4 mm thick, and weighing about 3 kg. Epidermis, dermis and subcutaneous fat It is composed of layers. The skin is in direct contact with the external environment and serves as an important protective film to protect the human body from various harmful factors such as human harmful microorganisms and external stimuli such as ultraviolet rays, while maintaining the biochemical functions necessary for metabolism of the whole body. It is an indispensable organ. In particular, the subcutaneous fat layer under the dermis is composed of the lobules of adipocytes, which act as a cushion to absorb and protect the organs inside the skin against external heat. It is stored in organelles called lipid droplets and used as an energy source if necessary. However, with age, the skin is damaged by natural factors and various external stimuli. In particular, the activity and expression of collagen and elastic fiber (elastin), which are responsible for elasticity in the dermis, are reduced and elasticity is reduced. At the same time, in the subcutaneous fat, the production of fat decreases, and eventually, the constituent cells of the entire skin do not function properly, resulting in skin aging such as wrinkles caused by loss of elasticity.

In the modern world, especially for women, skin is a major indicator of their health, so skin that loses its elasticity as it ages becomes a sensitive issue for women. In particular, the skin on the face is thinner than other skin areas, and it is easy to sag and lose elasticity due to factors such as weight and age. In addition, the decrease in facial elasticity and fat caused by metabolic abnormalities such as hormones, blood, fat, and bone, which occur mainly in postmenopausal women, make women feel a great loss of femininity. Increasing the volume of adipose tissue is a solution to the problems that are very sensitive to women in modern society, and many women try to maintain or increase the firmness of the skin through plastic surgery with side effects and economic burden. do.

Currently, a widely used method for improving the decrease in elasticity of the skin is liposuction. Lipograft surgery is a procedure in which a tulip needle-shaped needle is taken using a syringe having a needle that is not sharp and transplanted to each part of the skin where elasticity is poor. These fat grafts can be applied to fill and improve the wrinkles of aging skin as it ages, to correct the lip, jaw, and forehead lines, skin impregnated by nose shaping and burns or wounds, and areas lost due to cancer resection. The results are showing. However, it has been reported that 40-60% of the graft volume is absorbed back into the body after liposuction (S Eremia et al., Dermatol . Sur . 2000, 26: 1150-1158; Fulton JE et al., Dermatol . Clin 2001, 19 (3): 523-530), fat transplantation of skin is thin subcutaneous fat area is less raised a problem of which the cell mass touched, actually the transplanted fat survival and survival expectancy There is a case in which replanting is not necessary. Therefore, there is an increasing demand for a composition that can sufficiently increase the subcutaneous fat layer while replacing lipostomy with these problems.

Most of the cosmetics that have a skin elasticity promoting effect currently on the market using ingredients having the general efficacy, such as skin wrinkle removal or strengthening the dermis. Decreased skin elasticity is caused by decreased production or destruction of collagen and elastin caused by aging or ultraviolet rays. Various substances have been developed and used for the purpose of suppressing the reduction of collagen and elastin, which causes the decrease in elasticity. Among them, retinoids such as retinol and retinoic acid are typically used. Retinoids have an excellent elasticity-improving effect (Dermatology therapy 1998, 16, 357-364), but they have the disadvantage that irritation occurs even when only a small amount is applied to the skin. Therefore, in recent years, in order to solve the skin irritation problem of these raw materials, there is a high demand for the development of cosmetics using natural vegetable raw materials that are safer and have no skin irritation.

Meanwhile, since the basic structure of the skin is maintained by the subcutaneous fat tissue, and the subcutaneous fat tissue plays a role in determining the volume and strength of the skin, elasticity is applied to the dermis or epidermis of the outer skin layer that has been used as described above. increasing the volume of adipose tissue than how to give it can be a good solution to maintain voluminous and wrinkles of the skin, was recently published related (Kim WS et al ., J. Dermatol . Sci . 2009, 53 (2): 96-102; Trottier V et al ., Stem Cells . 2008 26 (10): 2713-23; Park BS et al ., Dermatol. Surg . 2008, 34 (10): 1323-1326).

Research using fat cells has been actively conducted in various fields. Preadipocytes generally used are widely used cells because they have the property of differentiating into adipocytes by differentiation inducing substances such as insulin. Recently, attention has been drawn in the study using the adipocytes, such as mesenchymal stem cells (adipocyte derived stem cells). These stem cells can generally be differentiated using methods similar to those used to differentiate adipocytes into adipocytes (Xu Y et. al ., Exp. Cell Res. 2006, 312, 1856-1864), widely used in the study of adipocyte differentiation (Pittenger MF et al ., Science 1999, 284, 143-146).

Therefore, the present inventors have tried to find a herbal extract that promotes the differentiation of fat cells forming adipose tissue, when the mesenchymal stem cells are treated with mesenchymal stem cells such as cinnamon, high-volume river, venom and cheonnyeonja extract, respectively The present invention has been found to effectively promote the process of differentiation into a.

Accordingly, it is an object of the present invention to provide a composition that contains herbal extracts that promote differentiation of human mesenchymal stem cells into adipocytes, thereby increasing skin volume and elasticity and improving wrinkles.

In order to achieve the above object, the present invention provides a skin external preparation composition and pharmaceutical composition for promoting the differentiation of fat cells containing one or more extracts selected from the group consisting of cinnamon, high amounts of steel, venom and cheonnyeon as an active ingredient.

The composition according to the present invention is useful for increasing the adipose tissue for cosmetic or cosmetic purposes by promoting the differentiation of mesenchymal stem cells into adipocytes by containing at least one of cinnamon, high-volume, poisonous, and physician extract as an active ingredient. Therefore, it helps to increase skin volume and elasticity and improve skin wrinkles.

Fig. 1 shows that the microscopic observation was performed after treatment of cinnamon, high doses, venom or Stellar extract under IDXT conditions, and after inducing adipocyte differentiation of mesenchymal stem cells under IDXT conditions, respectively. The picture shown.
Figures 2a to 2c is a graph showing the relative fat differentiation value compared to IDXT of mesenchymal stem cells after treatment, such as cinnamon, high doses, venom or cheonnyeonja extract.

The present invention relates to a composition for promoting adipocyte differentiation, which contains at least one extract selected from the group consisting of cinnamon, high dose steel, venom and cheonnyeon as an active ingredient.

As used herein, the term “fat cell differentiation” refers to the differentiation of adipocytes or mesenchymal stem cells from animals, preferably mammals into adipocytes, and more preferably human mesenchymal stem cells. Preferably, mesenchymal stem cells derived from human bone marrow differentiate into adipocytes.

The herbal extract used in the present invention is as follows.

Spatholobus suberectus Dunn.) is used as a medicine that acts on the heart and spleen to help blood circulation, replenish blood, and eliminate fish blood. In addition, it has the effect of leveling menstruation, and it is also used for menstrual irregularities, dysmenorrhea and amenorrhea. Other uses include degenerative diseases of elderly and women, atrophy of hands and feet, lethargy and dizziness, leukopenia of tumor patients due to radiation therapy.

Alpinia officinarum Hance has an unusual aroma, tastes great, and is warm. It is effective for abdominal pain, vomiting and diarrhea caused by cold, and it is effective for weakness in kidney function, diarrhea due to cold symptoms, diarrhea and dysentery. Chronic gastritis, loss of appetite, and drink when the stool is not cool. Pharmacological action has been reported to inhibit the development of hemolytic streptococci, diphtheria bacteria and pneumococci.

Melia azedarach var. japonica Makino has a peculiar smell, tastes bitter and is cold and slightly poisonous. It is used for narrowing, abdominal pain, testicular pain, etc. It is used for removing parasites and psoriasis dermatitis. Circulates the liver, relieves pain and has a pesticidal effect. Pharmacological action has been reported to have skin fungal and staphylococcus aureus inhibitory action, insecticidal action such as roundworm, earthworm, leech.

Angelica pubescens for. biserrata Shan Etyuan) has a peculiar smell and tastes a bit bitter and spicy. The effect is used for muscle pain, arthritis, back pain, knee and lower back pain caused by wind and humidity, and it is effective for skin itching, external fever, headache, and so on. Pharmacological effects have been reported for sedation, hypnosis, analgesic, anti-inflammatory, hypotensive, anticonvulsion, anti-ulcer, sun allergy.

Cinnamon lamp, high-volume river, poison and cheonnja extract used in the present invention can be extracted by conventional extraction methods well known in the art using an organic solvent such as water or ethanol.

The composition of the present invention promotes the differentiation of Mesenchymal Stem Cells (MSCs), thereby constituting the subcutaneous fat layer or providing fat cells to provide volume and elasticity to the skin and improve the wrinkles, It may be provided as a topical skin composition or a pharmaceutical composition.

When the composition according to the present invention is used as a skin external preparation composition, it is not particularly limited in the formulation according to the body part. Specifically, for example, it may be a cosmetic composition having a formulation of a softening lotion, nourishing lotion, massage cream, nutrition cream, pack, gel or skin adhesive type cosmetics, and may also be a lotion, ointment, gel, cream, patch or spray Transdermal dosage forms.

The external preparation composition for skin according to the present invention contains one or more extracts selected from the group consisting of cinnamon, high dose steel, venom, and cheonnyang as an active ingredient in an amount of 0.0001 to 30% by weight based on the total weight of the external preparation composition for skin.

In addition, in the external preparation composition for skin according to each formulation, other components other than cinnamon, high dose steel, venom or puncture extract of the present invention as described above are appropriately selected by those skilled in the art without difficulty according to the formulation or use purpose of other external preparations for skin. In this case, synergistic effect may occur when applied simultaneously with other raw materials.

In addition, when the composition according to the present invention is used as a pharmaceutical composition, it further contains a pharmaceutical adjuvant such as preservatives, stabilizers, hydrating or emulsifying accelerators, salts and / or buffers for controlling osmotic pressure and other therapeutically useful substances. And may be formulated in various oral or parenteral dosage forms according to conventional methods.

Formulations for oral administration include, for example, tablets, pills, hard and soft capsules, solutions, suspensions, emulsifiers, syrups, granules, etc. These formulations may contain diluents (e.g., lactose, dextrose, sucrose, Mannitol, sorbitol, cellulose and glycine) and glidants such as silica, talc, stearic acid and its magnesium or calcium salts and polyethylene glycols. Tablets may also contain binders such as magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose and polyvinylpyrrolidine, optionally starch, agar, alginic acid or its sodium salt Pharmaceutical additives such as disintegrants, absorbents, colorants, flavors, and sweeteners. Tablets may be prepared by conventional mixing, granulating or coating methods. Also representative of parenteral formulations are injectable formulations, preferably aqueous isotonic solutions or suspensions. However, it should be understood that the actual dosage of the active ingredient should be determined in light of several relevant factors such as the severity of the symptom, the route of administration chosen, the subject's age, sex, weight and health condition, and therefore the dosage of the herbal extracts. Does not limit the scope of the invention in any way. Generally, the dosage of the herbal extracts ranges from 0.001 mg / kg / day to approximately 2000 mg / kg / day. More preferred dosages are from 0.5 mg / kg / day to 2.5 mg / kg / day.

The site of application of the cosmetic or cosmetic composition for promoting the differentiation of mesenchymal stem cells according to the present invention includes all the adipose tissue of the body, but is not limited to a specific site, for example, breasts, hips of women with a lot of fat layers Site may be included. In addition, the increasing effect of the adipose tissue, but the effect of increasing the adipose tissue for cosmetic or cosmetic purposes, but is not limited thereto.

Hereinafter, the present invention will be described in more detail with reference to the following examples. However, the following examples are only for illustrating the present invention, and the scope of the present invention is not limited thereto.

Reference Example 1 Preparation of Herbal Medicine Extract

30g each of blood, light, high-yield, poisonous, bark, jujube, jujube or deodeok in 200ml of ethanol (95.0 ~ 95.5v / v%), and then 1500psi at 50 ℃ using ASE300 ACCELERATED SOLVENT EXTRACOR (DIONEX Corporation) Pulverized for 20 minutes under pressure. In addition, using an ultrasonic cleaner (BRANSON Ultrasonics corporation) sonicator device was further ground for 3 days at 50 ℃. Each of the pulverized extracts were dried at 40 ° C. for 24 hours using a module spin 40 (Modul spin 40; Biotron corporation) instrument to prepare each extract sample.

Experimental Example 1 Efficacy and Quantitative Measurement of Human Bone Marrow-derived Mesenchymal Stem Cells into Adipocytes

Human bone marrow mesenchymal stem cells (hBM-MSCs) were purchased from Lonza, Inc. Walkersville, MD, USA and cultured according to Lonza's instructions. Adipocyte differentiation was achieved using the method recommended by Lonza, except that Troglitazone was used instead of indomethacin to induce adipocyte differentiation of mesenchymal stem cells. Specifically, 1 μg / ml of insulin, 1 μM of dexamethasone (DEXA) and 0.5 mM of isobutyl methyl xanthine (IBMX) were cultured in the culture of mesenchymal stem cells to differentiate stem cells into adipocytes. At the time of induction of adipocyte differentiation, 10 ppm of cinnamon, high dose, venom or Stellar extract were added to the medium under IDXT conditions. In addition, jujube, saengpung or deodeok extract was added to the medium by 10 ppm each for comparison. As a positive control, Troglitazone (TRO) 2 μM was used and treated with IDX medium. After 14 days of adipocyte differentiation of mesenchymal stem cells, the cell culture was removed, the cells were washed with 10 ml of Phosphate Buffered Saline (PBS), and then 0.2 ml / cm of 10% formalin / PBS solution. Cells were fixed for 10 minutes using ² . The fixed cells were washed once with 0.5 ml / cm ² of 60% isopropanol solution, and stained with lipids in adipocytes with 0.2 ml / cm ² stained with Oil Red O in 60% isopropanol for 10 minutes. It was. Subsequently, re-staining with hematoxylin (hematoxylin) to make the cell nuclei easily distinguished and observed under a microscope, the results of the treatment of cinnamon lamp, high dose steel, venom or cheonnja extract is shown in FIG.

Figure 1 shows that the differentiation of adipocyte differentiation of mesenchymal stem cells under IDX or IDXT conditions, and then the addition of extracts such as cinnamon, high doses, venom or Stellar under IDXT conditions, followed by adipocyte differentiation of mesenchymal stem cells. Induction of differentiation of fat cells can be confirmed to occur more effectively.

After dyeing the oil redo with 100% isopropanol solution, the absorbance was measured at a wavelength of 500 nm, the results are shown in Figures 2a to 2c. At this time, each data value was corrected with an insulin control. This increase was statistically significant, and the statistics were verified by the two-tailed Student's t test, indicating that there was a significant difference when the p value was less than 0.05. In addition, the increase in fat differentiation-promoting rate compared to IDXT of mesenchymal stem cells when treated with cinnamon, high-volume river, venom, cheonnyeonja extract, jujube, jujube and Deodeok extract, respectively, is shown in Table 1 below.

Herbal medicine Scientific name IDXT vs. Growth Rate Blood clots Mucunae Caulis 44% High volume steel Alpiniae Officinari Rhizoma 31% Curious Meliae Fructus 37% Poisonous Araliae Cordatae Radix 41% Jujube Zizyphi Fructus 3% reduction Satire Hydnocarpi Semen 15% Deodeok Codonopsitis Lanceolatae Radix 0.09%

As can be seen from Table 1 and the results of Figs. 2a to 2c, the extracts of cinnamon, high-volume, venomous or cheonngi according to the present invention, respectively, induce differentiation of adipocytes without inducing cell death of mesenchymal stem cells. It can be seen that it can promote the differentiation of adipocytes much more effectively than when treated with jujube, saengpung or deodeok extract.

Test Example 2 Measurement of Skin Volume Enhancement Effect

In order to find out the effect of improving the volume and wrinkles of human skin when using the blood of the present invention, such as cinnamon, high doses, venom or cheonngija extract, a survey was conducted on 20 clinical participants.

Specifically, each of the extracts as shown in Table 2 was applied to the skin of the neck area once a day for 8 weeks, each time containing a nourishing cream containing 2.0% by weight based on the total weight of the composition, afterwards to increase the volume and wrinkles A survey was conducted on the degree of improvement. When the effect is good, ○, the normal case is △, the case where there is no effect is represented by ×, the results are shown in Table 3 below.

Compounding ingredient Content (% by weight) Purified water To 100 Cinnamon, high dose, poison or extract 2.0 Vegetable hydrogenated oil 1.5 Stearic acid 0.6 Glycerol stearate 1.0 Cetearyl alcohol 2.0 Polyglyceryl-10 Pentastearate & Behenyl Alcohol & Sodium Stearoyl Lactylate 1.0 Arachidyl behenyl alcohol 1.0 Cetylaryl alcohol & cetearyl glucoside 2.0 PEG-100 Stearate & Glycerololate & Propylene Glycol 1.5 Caprylic / Carlic Triglycerides 11.0 Cyclomedicon 6.0 Preservative, incense Quantity Triethanolamine 0.1

Herbal medicine Increase volume and
Wrinkle improvement effect (person) ○ △ X Blood clots 16 2 2 High volume steel 12 One 7 Curious 13 One 6 Poisonous 15 2 3 Jujube One 5 14

As can be seen in Table 3, when the jujube extract is used, but the skin volume improvement and wrinkle improvement effect is not good, while most of the test when using the cinnamon, such as the present invention, high-volume steel, venom or cheonnja extract Participants were found to have good skin volume and wrinkle improvement.

From the results of the experiment, the composition containing the cinnamon lamp, the high-strength, poisonous or natural extract, respectively, is applied to the skin of the face, invading or taking the skin, and the active ingredient of the extract increases the volume of adipose tissue. It can be seen that the effect of increasing the volume and elasticity of the wrinkles and reducing wrinkles can be obtained.

Examples of preparations for the compositions of the present invention are illustrated below.

Preparation Example 1 Tablet

Tablets were prepared by mixing 100 mg of extracts of one type of cinnamon lamp, high-strength, poisonous, and medicinal herbs according to the present invention, 400 mg of lactose, 400 mg of corn starch, and 2 mg of magnesium stearate, and then tableting according to a conventional method for preparing tablets. .

Preparation Example 2 Capsule

100 mg extract, one 400 mg lactose, 400 mg corn starch, and 2 mg magnesium stearate were mixed, and then filled into gelatin capsules according to a conventional method for preparing a capsule. Capsules were prepared.

[Example 3] Softening Cosmetic (Skin Lotion)

Compounding ingredient Content (% by weight) Purified water To 100 Extract of one of cinnamon, high amount river, poisonous leaf 0.1-0.2 Butylene glycol 2.0 Propylene glycol 2.0 Carboxy Vinyl Polymer 0.1 Fiji-12 nonylphenyl ether 0.2 Polysorbate 80 0.4 ethanol 10.0 Triethanolamine 0.1 Preservative, coloring, fragrance Quantity

[Example 4] Nutritional Longevity (Milk Lotion)

Compounding ingredient Content (% by weight) Purified water To 100 Extract of one of cinnamon, high amount river, poisonous leaf 0.1-0.2 Beeswax 4.0 Polysorbate 60 1.5 Sorbitan sesquioleate 1.5 Liquid paraffin 0.5 Caprylic / capric triglyceride 5.0 glycerin 3.0 Butylene glycol 3.0 Propylene glycol 3.0 Carboxy Vinyl Polymer 0.1 Triethanolamine 0.2 Preservative, coloring, fragrance Quantity

[Example 5] Nutritional Cream

Compounding ingredient Content (% by weight) Purified water To 100 Extract of one of cinnamon, high amount river, poisonous leaf 0.1-0.2 Beeswax 10.0 Polysorbate 60 1.5 Sebum 60 Cured Castor Oil 2.0 Sorbitan sesquioleate 0.5 Liquid paraffin 10.0 Squalane 5.0 Caprylic / capric triglyceride 5.0 glycerin 5.0 Putylene glycol 3.0 Propylene glycol 3.0 Triethanolamine 0.2 Preservative, coloring, fragrance Quantity

[Example 6] Massage Cream

Compounding ingredient Content (% by weight) Purified water To 100 Extract of one of cinnamon, high amount river, poisonous leaf 0.1-0.2 Beeswax 10.0 Polysorbate 60 1.5 Sebum 60 Cured Castor Oil 2.0 Sorbitan sesquioleate 0.8 Liquid paraffin 40.0 Squalane 5.0 Caprylic / capric triglyceride 4.0 glycerin 5.0 Butylene glycol 3.0 Propylene glycol 3.0 Triethanolamine 0.2 Preservative, coloring, fragrance Quantity

[Example 7] Pack

Compounding ingredient Content (% by weight) Purified water To 100 Extract of one of cinnamon, high amount river, poisonous leaf 0.1-0.2 Polyvinyl alcohol 13.0 Sodium Carboxymethyl Cellulose 0.2 glycerin 5.0 Allantoin 0.1 ethanol 6.0 Fiji-12 nonylphenyl ether 0.3 Polysorbate 60 0.3 Preservative, coloring, fragrance Quantity

Claims (11)

Anti-aging skin external composition comprising one or more extracts selected from the group consisting of cinnamon, high amounts of steel and venom as an active ingredient. According to claim 1, wherein the composition is a topical skin composition for improving wrinkles. According to claim 1, wherein the composition is an external skin composition for improving elasticity. The composition for external application for skin according to claim 1, wherein the composition promotes differentiation of mesenchymal stem cells into adipocytes. The composition for external application of skin according to claim 4, wherein the mesenchymal stem cells are human mesenchymal stem cells. The composition for applying the external of the skin according to claim 4, wherein the composition increases fat tissue. The composition for applying the external of the skin according to claim 1, wherein the composition is contained in an amount of 0.0001 to 30% by weight based on the total weight of one or more extract compositions selected from the group consisting of cinnamon, high amounts of steel and venom. The composition for applying the external of the skin according to claim 1, wherein the composition increases skin volume. A pharmaceutical composition for promoting adipocyte differentiation, which comprises at least one extract selected from the group consisting of cinnamon, high doses and venom as an active ingredient. The pharmaceutical composition of claim 9, wherein the composition promotes differentiation of mesenchymal stem cells into adipocytes. The pharmaceutical composition according to claim 9, wherein the composition has a dosage of at least one extract selected from the group consisting of cinnamon, high doses and venom.

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