KR20150068865A - Composition for rejuvenation of senescent cell comprising of Ginsenoside Rg3 as an active ingredient - Google Patents

Abstract

The present invention relates to a composition for aging cell rejuvenation comprising ginsenoside Rg3 as an active ingredient, and more particularly, to a composition for aging cell rejuvenation comprising ginsenoside Rg3 as an active ingredient, Compositions, external skin preparations, food compositions and pharmaceutical compositions.
The ginsenoside Rg3 of the present invention has the effect of restoring the aged cells with cell cycle arrest to the cell cycle of the young cell level and rejuvenating the aging cells.

Description

TECHNICAL FIELD The present invention relates to a composition for aging cell rejuvenation comprising ginsenoside Rg3 as an active ingredient,

The present invention relates to a composition for aging cell rejuvenation comprising ginsenoside Rg3 as an active ingredient, and more particularly, to a composition for aging cell rejuvenation comprising ginsenoside Rg3 as an active ingredient, Compositions, external skin preparations, food compositions and pharmaceutical compositions.

Cell senescence refers to the process of cell degeneration and functional degeneration followed by cell death or proliferation arrest. When normal proliferating cells approach the cell's lifespan by approaching the cell's longevity, it becomes impossible to divide due to the loss of its ability to divide. There are two kinds of metabolism: aging and regeneration of cells by non-dividing cells In general, it points to the former.

Cells enter an irreversible growth arrest or terminally non-diving state called senescence after a defined number of cell divisions. The aging of these cells ultimately affects the aging process of life. Aging cells actively express proliferative inhibitors that inhibit cell proliferation and cell senescence is thought to be closely related to cell cycle regulation.

To date, the cause of aging has not been clear yet, but more than a hundred hypotheses have been published and will increase in the future. However, many of these hypotheses can be largely classified into three groups. The first is the genetic program theories of aging, in which the aging and lifespan of the organism is predicted by the internal gene, and the second is the accumulation of damage to the biomaterial by various harmful factors, Theories of aging related to primary damage. Finally, it is the unifying model of the programmed and the stochastic theories of aging that combines the two hypotheses and develops into a hypothesis.

Aging scholars who advocate aging suggest that aging, the next step in growth, will be controlled by genetic information as well, as embryos are regulated by genes all the way to development and growth. This hypothesis sees senescence as an extension of normal genetic signal flow from embryos to the growth of life. As the ages grow, the biochemical metabolic pathway slows down or stops, And so-called senescence-associated genes, which indicate a decrease in white hair, menopause, and athletic performance. Reproduction of normal cells Since the aging phenomenon is known and the fact that factors regulating it are present on the chromosome has been known, many attempts have been made to find aging genes in cultured cells. It has been found that hybrid cells formed by fusing normal cells with a finite lifespan and immortalized cells have a finite life span. This means that the phenotype indicating the finite life span in the in vitro culture is dominant and the phenotype indicating the immortality in which the cells can divide forever is fervent. Pereira-Smith et al. Have divided the deficiency of aging genes, which represent immortality of these cells, into four complementary groups (complementation groups A, B, C, and D) through a fusion experiment between immortal cells. In other words, if the hybrid cells formed by the fusion of two immortal cells die and die, the two immortal cells are due to the deficiency of different aging genes. If the hybrid cells remain immortal cells, the two immortal cells are due to the same genetic deficiency And were classified as the same complementary group. To date, approximately 40 immortalized cells have been divided into four complementary groups, which means that immortal cells are not aged by a limited number of aging genes or lack of genetic pathways. Recently, it has been found that p53 (chromosome 17) and pRb (chromosome 13) play an important role as regulators of the cell cycle. Shay et al. Claim that p53 is an aging-related gene when cells transfected with SV40 large T antigen or HPV-E6 / E7 into normal cells prolong their life without prolonged life. Both of these proteins are known to bind to p53 and lose its function.

The harmful factor damage theory is a hypothesis that the damage accumulation of biomaterials caused by internal or external damage factors that cause irreversible changes in cells or tissues is considered as a main cause of aging, and there are many claims according to scholars, And the free radical theory of aging proposed by Harman in 1956. It is the theory that biomass is oxidatively damaged by various active oxygen which is generated incidentally during normal metabolism and accumulation of these damages leads to aging.

Since these two types of hypothesis alone can not account for aging phenomena enough, Papacons- tantinou proposed an aging hypothesis that incorporated two hypotheses in 1994. According to this hypothesis, the biological characteristics of aging are due to the age-related changes in the intrinsic life processes in the body, which are promoted by external environmental factors. Such intrinsic factors include cytokines and oxidative metabolites, and external factors include active oxygen, ultraviolet light, heavy metals, and air pollutants. When they activate the transcription factors such as NF-kB, AP-1, bZIP, C / EBP and HSF (heat shock factors) through the receptor of the cell membrane or directly stimulate intracellular signal transduction systems, response genes to protect cells. However, as age increases, changes in the homeostasis of intrinsic factors and exacerbation of these changes by exogenous factors lead to the inability to regulate the production of stress-responsive genes, resulting in the inability to respond to external stresses or to exacerbate aging characteristics .

Panax ginseng CA Meyer) is a perennial plant of the dicotyledonous Araliacaeae and is known to be the most important medicinal medicine used mainly in medicine. Ginseng contains major efficacy components such as saponin, essential oil, plant sterol, polyacetylene, phenolic substance, polysaccharide component and so on, and it is antidiabetic, anti-cardiovascular, antiarteriosclerosis, central nervous system, Have been reported to have efficacy such as brain activity promotion and nerve cell protection, anti-cancer and antioxidant activity, immune function regulation, sexual dysfunction improvement, diet efficacy and the like.

Ginsenoside refers to saponin in ginseng. Saponin is a kind of chemical compound called glycoside. Ginseng saponin has a unique chemical structure that is different from saponin found in other plants. Its pharmacological effect is also unique and is called ginsenoside (ginsenoside). At present, about 30 chemical structures of ginseng saponin are known. Depending on the chemical structure, protopanaxadiol (PPD) (19 kinds), protopanaxatriol (PPT) And Oleanane (1 kind) saponin. The ginseng content of Korean ginseng varies depending on the variety of ginseng. Our Korean ginseng contains many kinds of ginsenosides which have pharmacological activity, such as Rhodophyta, Rhg2, Rg3, Rf, etc. are present independently and are recognized as the best quality among various species.

Many studies have been conducted on the effect of ginsenoside on the aging phenomenon for a long time. As a known technique related to ginsenoside and aging, Korean Patent Laid-Open No. 10-2005-0038463; Cosmetic composition containing ginsenoside Rg3 ', Korean Patent Registration No. 10-1146104, Skin wrinkle improving agents containing ginsenoside Rg3, Rg6 and Rg2 as main components, and skin wrinkle improving agent containing them ', Korean Patent Laid-Open Publication No. 10-2009-0015127; Ginsenoside, and the concentration of the extract containing them ", and many other documents are available.

These techniques are mainly technologies to prevent aging of cells and promote cell proliferation. In other words, it is important to reduce the concentration of substances that affect cell damage or aging, or to help cells actively divide.

Recently, however, much research has been conducted on the rejuvenation of aging cells in order to prove the reversibility of aging cells in which cell cycle arrest is not limited to simple aging prevention techniques. In one study, cells of 100-year-olds were degenerated in vitro using 6 gene cocktails to produce iPSCs (inducible pluripotent stem cells), which were then cleared of the signs of aging to form human embryonic stem cells Laure Lapasset et al 11, 2011, Rejuvenating senescent and centenarian human cells by reprogramming through the pluripotent state, GENES & DEVELOPMENT, vol 25, 2248-2253.). The resulting iPSCs are capable of differentiating into all human cells and have been shown to have physiological characteristics of young cells in terms of proliferative capacity and cellular metabolism. Because senescent cells are a stumbling block to de-differentiation, it has been known that iPSC technology is difficult to apply to the oldest elderly people, demonstrating the reversibility of the cell senescence process by eliminating these obstacles.

However, since such techniques are performed in vitro using molecular genetic techniques, there is a problem that their use is limited in practice. On the other hand, there is a continuing research to find a substance having cell rejuvenation effect from a natural product which has few side effects and is widely applicable in vivo.

Thus, the present inventors confirmed the aging cell rejuvenation activity of ginsenoside Rg3 and completed the present invention.

Accordingly, an object of the present invention is to provide a composition for rejuvenation of aging cells containing ginsenoside Rg3 represented by the following formula (1) as an active ingredient.

≪ Formula 1 >

In order to achieve the above object, the present invention provides a composition for rejuvenating aged cells comprising ginsenoside Rg3 represented by the following general formula (1) as an active ingredient.

≪ Formula 1 >

Hereinafter, the present invention will be described in detail.

The ginsenoside Rg3 of the present invention has a structure represented by the following general formula (1), and can be purified by separation from nature, commercially available, or by chemical synthesis methods known in the art.

≪ Formula 1 >

For example, the ginsenoside Rg3 of the present invention can be prepared by a compound extraction method known from the natural art. For example, the ginsenoside Rg3 of the present invention can be obtained from processed ginseng obtained by heat treatment of ginseng or ginseng leaves at high temperature, or acid hydrolyzate of ginseng extract. Ginsenoside Rg3 is a plant of the genus Panax, which contains ginseng saponin, for example Panax ginseng), Panax Noto jinseng (Panax notoginseng), Panax kwinkwe poly Titanium (Panax quinquefolium , Panax japonicus , etc., or leaves of these plants, tissue cultures of these plants, or water or lower alcohols thereof, are heated at a temperature of 110 to 180 ° C for 0.5 to 20 hours And extracting the obtained processed ginseng with water or an appropriate organic solvent such as methanol, ethanol or a mixed solvent thereof, then concentrating the extract under reduced pressure, suspending it in water, adding a non-polar organic solvent such as hexane, ether , Extracted with dichloromethane, chloroform, ethyl acetate or a mixed solvent thereof, and the remaining water layer is extracted with a polar organic solvent such as butanol, and this extract is subjected to chromatography to obtain a fraction containing Rg3. At this time, if the chromatograph is repeatedly performed, the content of ginsenoside Rg3 can be further increased, and the fraction having a higher content can be fractionated in a suitable solvent system such as a solvent system such as water, lower alcohol, lower ketone, chloroform, Crystallization yields pure ginsenoside Rg3. In this method, the ginsenosides Ra, Rb1, Rb2, Rc, and Rd, which are pyrazine diol saponins present in ginseng in the process of heat treatment of ginseng, . In some cases, the step of heating treatment and the step of extracting organic solvent may be changed in the above-mentioned process, or the step of heating at a temperature of, for example, 30 to 100 ° C, preferably 70 ° C under mild conditions, , The same result can be obtained when the acid is treated with a weak acid such as hydrochloric acid, nitric acid, perchloric acid, or the like, or a lower organic acid such as acetic acid, tartaric acid, oxalic acid and the like. Also, in the above process, components such as ginsenosides Ra, Rb1, Rb2, Rc, and Rd, or fractions thereof, which are known compounds instead of ginseng, are directly heated or acid hydrolyzed as described above, Can be obtained.

The ginsenosides Rg3 of the present invention include all stereoisomers, preferably two forms of S or R forms, more preferably 20 (S) -ginnenosides Rg3.

The term 'aging' includes both normal division of the proliferating cells and proliferation of the proliferative cells, which are approaching the cell's longevity, and metabolic pathways in which the cells regress as the intracellular metabolism of the non-dividing cells continues. Preferably, the cell refers to an irreversible proliferation arresting cell, that is, a cell cycle arrest, wherein the cell stops the proliferation after a certain number of cell division.

The term 'aging cell' means a state in which all kinds of cells known to a person skilled in the art (for example, muscle cells, connective cells, skin cells, etc.) are aged, preferably skin aging cells.

The term 'rejuvenation' refers to the process of aging, or any other process that can damage macromolecules, cells, tissues and organs, including skin, or cause damage accumulation (eg, abrasions caused by falls, burns, etc.) Although reversal or mitigation means, the rejuvenation in the present invention preferably means that the cell cycle arrested cells are regenerated again to the cell cycle level of the young cell level.

The 'young cell' refers to a cell in which the cell continues to undergo a cell cycle without loss of normal proliferative capacity, and preferably a cell cycle is not stopped.

The ginsenoside Rg3 of the present invention has the effect of rejuvenating aging cells.

Such an effect is well illustrated in an embodiment of the present invention.

In one embodiment of the present invention, the rejuvenation phenomenon of senescent cells when Rg3 was treated was compared with the expression level of senescence-associated β-galactosidase (senescence-associated β-galactosidase). As a result, when Rg3 was treated with aging cells, β-galactosidase expression was reduced by about 30-40% compared with the cells not treated with Rg3. In addition, aging cells were treated with Rg3 to measure changes in cell cycle. As a result, it was confirmed that most of the senescent cells were stopped in S period before Rg3 treatment. When Rg3 was treated with aging cells, G1 group was increased to 72% level, and the cell cycle was regained again to the young cell level.

The ginsenoside Rg3 of the present invention has an aging cell rejuvenation effect superior to other saponins. Accordingly, the present invention provides a composition for aging cell rejuvenation comprising ginsenoside Rg3 as an active ingredient.

The composition may be prepared into any formulation according to a method known in the art by additionally containing a pharmaceutically acceptable carrier, but it may preferably be a skin external preparation, a cosmetic composition, a food composition and a pharmaceutical composition.

Examples of the form of the external preparation for skin include, but are not limited to, liquid coating agents, spray agents, lotions, gels, pastes, ointments, aerosols, powders and transdermal absorbers.

Pharmaceutically acceptable carriers in the external preparation of the present invention include, but are not limited to, hydrocarbons such as vaseline, liquid paraffin, gelling hydrocarbons (plastid base); Vegetable oils such as medium-chain fatty acid triglycerides, lard, hard fat, cacao butter and the like; Higher fatty alcohols such as cetanol, stearyl alcohol, stearic acid and isopropyl palmitate; fatty acids and esters thereof; Water-soluble bases such as polyethylene glycol, 1,3-butylene glycol, glycerol, gelatin, white sugar, sugar alcohol and the like; Emulsifiers such as glycerin fatty acid esters, polyoxyl stearate, and polyoxyethylene hardened castor oil; Acrylic acid ester, and sodium alginate; Jet agents such as liquefied petroleum gas and carbon dioxide; And preservatives such as paraoxybenzoic acid esters. In addition to these, a stabilizer, a flavoring agent, a coloring agent, a pH adjuster, a diluent, a surfactant, a preservative, an antioxidant, and the like can be optionally incorporated. The use of the external preparation of the present invention is preferably applied to cells aged by a conventional method.

In addition, the external preparation according to the present invention can be used by being adhered to a solid support such as a wound dressing cover of a conventional bandage. Adhesion can be achieved by saturating the ginsenoside Rg3 of the present invention with a solid support followed by dehydration Preferably the solid support is first coated with a tackifier to improve adhesion of the ginsenoside Rg3 of the present invention to the solid support have. Examples of the pressure-sensitive adhesive include polyacrylate and cyanoacrylate. This type of formulation is commercially available, for example, a bandage (Smith & Nephew Ltd) having a non-adhesive wound peel-off cover in the form of a perforated plastic film; Johnson &Johnson's thin strips, patches, spots, band-aids in the form of plastic strips (BAND-AID); Curity CURAD Ouchless from Colgate-Palmolive Co. (Kendall) Bandage; And STIK-TITE elastic strips from American White Cross Laboratories Inc. The ginsenoside Rg3 of the present invention can be applied as an active ingredient to this type of formulation.

The cosmetic composition can be easily prepared according to known methods in the art, including one or more excipients and additives commonly used in the field of cosmetic composition production together with ginsenoside Rg3 or salts thereof.

The cosmetic composition may be prepared in liquid or solid form using bases, adjuvants and additives conventionally used in the field of cosmetics. Liquid or solid form of cosmetics may include, for example, but not limited to, lotions, creams, lotions and bath salts. At this time, the bases, adjuvants and additives commonly used in the cosmetics field are not particularly limited and may include, for example, water, alcohol, propylene glycol, stearic acid, glycerol, cetyl alcohol and liquid paraffin.

More specifically, the cosmetic composition of the present invention contains the ginsenoside Rg3 of the present invention or a salt thereof as an active ingredient, together with a dermatologically acceptable excipient, together with a basic cosmetic composition (lotion, cream, essence, cleansing foam and cleansing water (Shampoos, rinse, hair conditioner, hair gel), functional cosmetic compositions (anti-wrinkle agents, anti-wrinkle agents), and hair cosmetic compositions Soap and the like. Such excipients include, but are not limited to, emollients, skin penetration enhancers, colorants, perfumes, emulsifiers, thickeners and solvents. In addition, it may further contain flavors, pigments, bactericides, antioxidants, preservatives, moisturizers and the like, and may include thickeners, inorganic salts and synthetic polymeric substances for the purpose of improving physical properties.

For example, in the case of producing a cleanser and a soap containing ginsenoside Rg3 of the present invention, ginsenoside Rg3 can be easily added to a common cleanser and soap base. Cream can be prepared by adding ginsenoside Rg3 to a cream base of a typical water-in-oil type (O / W). A synthetic or natural material such as a flavor, a chelating agent, a coloring matter, an antioxidant, an antiseptic, and a protein, a mineral, and a vitamin for the purpose of improving a physical property may be further added.

The content of ginsenoside Rg3 contained in the cosmetic composition of the present invention may be 0.0001 to 50% by weight, preferably 0.01 to 10% by weight based on the total weight of the cosmetic composition.

The food composition includes all forms such as functional food, nutritional supplement, health food and food additives. Food compositions of this type may be prepared in a variety of forms according to conventional methods known in the art.

For example, as a health food, the ginsenoside Rg3 of the present invention can be prepared in the form of tea, juice, and drink and liquefied, granulated, encapsulated, and powdered to be ingested. In addition, it can be prepared in the form of a composition by mixing together the ginsenoside Rg3 of the present invention and a known substance or active ingredient known to be effective for aging cell rejuvenation.

Functional foods also include but are not limited to beverages (including alcoholic beverages), fruits and processed foods (e.g., canned fruits, jams, maamarade, etc.), fish, meat and processed foods (Eg butter, chewing), edible vegetable oil, margarine (for example, sausage, noodles, etc.), breads and noodles (eg udon, buckwheat noodles, ramen noodles, spaghetti, macaroni, , Vegetable protein, retort food, frozen food, various kinds of seasonings (for example, soybean paste, soy sauce, sauce, etc.) by adding ginsenoside Rg3.

In addition, in order to use the ginsenoside Rg3 of the present invention in the form of a food additive, it may be prepared in the form of powder or concentrate.

The preferable content of the ginsenoside Rg3 in the food composition of the present invention is not particularly limited, but is preferably 0.01 to 50% by weight in the finally prepared food.

The pharmaceutical composition may be a composition comprising ginsenoside Rg3 or a pharmaceutically acceptable salt thereof as an active ingredient.

The ginsenoside Rg3 according to the present invention may be used as such or in the form of a pharmaceutically acceptable salt. As used herein, the term " pharmaceutically acceptable " means physiologically acceptable and does not normally cause an allergic reaction or similar reaction when administered to humans, and the salt includes a pharmaceutically acceptable free acid ) Are preferred. The free acid may be an organic acid or an inorganic acid. The organic acids include, but are not limited to, citric, acetic, lactic, tartaric, maleic, fumaric, formic, propionic, oxalic, trifluroacetic, benzoic, gluconic, methosulfonic, glycolic, succinic, Glutamic acid and aspartic acid. In addition, the inorganic acid includes, but is not limited to, hydrochloric acid, bromic acid, sulfuric acid, and phosphoric acid.

The pharmaceutical composition according to the present invention may contain, alone or in combination with one or more pharmaceutically acceptable carriers, excipients or diluents, ginsenoside Rg3 or a pharmaceutically acceptable salt thereof.

The pharmaceutically acceptable carrier may further include, for example, a carrier for oral administration or a carrier for parenteral administration. Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate, stearic acid, and the like. In addition, the carrier for parenteral administration may contain water, a suitable oil, a saline solution, an aqueous glucose and a glycol, and may further contain a stabilizer and a preservative. Suitable stabilizers include antioxidants such as sodium hydrogen sulfite, sodium sulfite or ascorbic acid. Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben and chlorobutanol. Other pharmaceutically acceptable carriers can be found in Remington's Pharmaceutical Sciences, 19th ed., Mack Publishing Company, Easton, Pa., 1995).

The pharmaceutical composition for aging cell rejuvenation of the present invention can be administered to mammals including humans by any method. For example, it can be administered orally or parenterally. Parenteral administration methods include, but are not limited to, intravenous, intramuscular, intraarterial, intramedullary, intrathecal, intracardiac, transdermal, subcutaneous, intraperitoneal, intranasal, enteral, topical, sublingual or rectal administration Lt; / RTI > Preferably, the pharmaceutical composition of the present invention can be transdermally administered. The term 'transdermal administration' as used herein refers to the administration of the pharmaceutical composition of the present invention to cells or skin to allow the active ingredient contained in the pharmaceutical composition for aging cell reuptake to be delivered into the skin. For example, the pharmaceutical composition of the present invention can be administered in a scanning type formulation, pricking the skin lightly with a 30 gauge needle, or directly applying it to the skin.

In addition, the pharmaceutical composition may be formulated into oral or parenteral formulations according to the route of administration as described above.

In the case of a preparation for oral administration, the composition of the present invention may be formulated into a powder, a granule, a tablet, a pill, a sugar, a tablet, a liquid, a gel, a syrup, a slurry, . For example, an oral preparation can be obtained by combining the active ingredient with a solid excipient, then milling it, adding suitable auxiliaries, and then processing the mixture into a granular mixture. Examples of suitable excipients include, but are not limited to, sugars including lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol and maltitol, and starches including corn starch, wheat starch, rice starch and potato starch, Cellulose such as methylcellulose, sodium carboxymethylcellulose and hydroxypropylmethyl-cellulose and the like, fillers such as gelatin, polyvinylpyrrolidone and the like. In addition, crosslinked polyvinylpyrrolidone, agar, alginic acid, or sodium alginate may optionally be added as a disintegrant. Further, the pharmaceutical composition of the present invention may further comprise an anti-coagulant, a lubricant, a wetting agent, a flavoring agent, an emulsifying agent and an antiseptic agent.

In the case of a preparation for parenteral administration, it can be formulated by a method known in the art in the form of injection, cream, lotion, external ointment, oil, moisturizer, gel, aerosol and nasal aspirate. These formulations are described in Remington's Pharmaceutical Science, 15th edition, 1975. Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Blaug, Seymour, a commonly known formulary for all pharmaceutical chemistries.

The total effective dose of ginsenoside Rg3 of the present invention may be administered to a patient in a single dose and administered by a fractionated treatment protocol administered over a prolonged period of time in multiple doses have. In the pharmaceutical composition of the present invention, the content of the active ingredient may be varied depending on the degree of the disease. Preferably, the preferred total dose of ginsenoside Rg3 of the present invention may be from about 0.01 to 1,000 mg, and most preferably from 0.1 to 100 mg, per kg of patient body weight per day. However, the dosage of ginsenoside Rg3 is not limited to the effective route of administration to the patient in consideration of various factors such as age, weight, health, sex, severity of disease, diet and excretion rate, Given this fact, one of ordinary skill in the art will be able to determine the appropriate effective dose of ginsenoside Rg3 according to the particular use of aging cell rejuvenation. The pharmaceutical composition according to the present invention is not particularly limited to its formulation, administration route and administration method as long as the effect of the present invention is exhibited.

The ginsenoside Rg3 of the present invention has the effect of restoring the aged cells with cell cycle arrest to the cell cycle of the young cell level and rejuvenating the aging cells.

FIG. 1 shows the expression level of β-galactosidase as an aging marker when treating ginsenoside Rg3 with aging cells (old HDF) (Y HDF: young human diploid fibroblast, O HDF: old human diploid fibroblast , And the numbers in parentheses indicate concentration of ginsenoside Rg3 treatment (unit μM)).
FIG. 2 shows the ratio of β-galactosidase-stained HDF cells treated with ginsenoside Rg3 (Y HDF: young human diploid fibroblast, O HDF: old human diploid fibroblast, the numbers in parentheses indicate ginsenoside Rg3 Means the treatment concentration (unit: μM)).
FIG. 3 shows the cell cycle measured by DAPI-staining of aging cells treated with ginsenoside Rg3 (Y HDF: young human diploid fibroblast, O HDF: old human diploid fibroblast, numbers in parentheses indicate ginsenoside Rg3 treatment Concentration (unit: μM)).

Hereinafter, the present invention will be described in detail.

However, the following examples are illustrative of the present invention, and the present invention is not limited to the following examples.

≪ Example 1 >

Preparation of sample

<Example 1-1> Preparation of ginsenoside Rg3

20 (S) -ginsenoside Rg3 (purity> 98%) was purchased from Ambo Laboratories (Daejeon, Republic of Korea). 20 (S) -ginsenoside Rg3 of this experiment was dissolved in DMSO (dimethyl sulfoxide) and treated by concentration.

&Lt; Example 1-2 > Cell culture

Human diploid fibroblast (HDF) was purchased from the Department of Biochemistry, Seoul National University College of Medicine. (Dulbecco's Modified Eagles Medium, Sigma Chemical Co) containing 10% fetal bovine serum (GIBCO), Penicillin (100 units / ml), Streptomycin (100 μg / ml) and Amphotericin B (0.25 μg / ml) CO ₂ at 37 ° C. At this time, the culture medium was changed every two days and the cells were cultured in Petri dish (100 mm in diameter). The cells were treated with 0.05% trypsin / 0.481 mM EDTA and subcultured at a ratio of 1: 4 to pass passage 8 (young HDF) and passage 34 (old HDF).

&Lt; Example 2 >

Measurement of aging marker β-galactosidase (SA-β-gal) expression

When aging cells were treated with Rg3 using a β-galactosidase staining kit (Sigma, St. Louis, Mo.), the aging marker β-galactosidase senescence-associated β-galactosidase, SA-β-gal). Young HDF and old HDF were dispensed into a 12-well plate at a density of 2 x 10 ⁴ / well, respectively, for overnight incubation. Then, Rg3 was treated with 10, 30 μM for 48 hours, washed with Phosphate Buffer Solution, and fixed. The chromogenic substrate at pH 6.0 was incubated at 37 ° C Lt; / RTI &gt; Three independent experiments were repeated and color changes were observed with a magnification of 100X using an optical microscope. Total number of cells in five fields and number of cells stained in blue were measured for one well, and the number of cells stained with blue color was calculated as a percentage of the total number of cells, Diego, CA) program.

As shown in FIG. 1 and FIG. 2, when aged cells were compared with young cells, it was confirmed that the number of cells stained with β-galactosidase was about 70-80% higher in aged cells. In addition, the expression of β-galactosidase was reduced by about 30-40% in Rg3-treated cells compared with cells not treated with Rg3.

&Lt; Example 3 >

Cell cycle measurement

Cell cycle measurements were performed using the NucleoCounter NC-3000 ^™ kit (Chemometec, France) according to the manufacturer's protocol. The cells were treated with 0.05% Trypsin / 0.481 mM EDTA for 48 hours and the cells were fixed in 70% ethanol for 2 hours. The immobilized cells were incubated in DAPI solution for 5 min at 37 ° C, absorbance was measured at 365 nm using a NucleoCounter NC-3000 ^™ instrument, and data were analyzed using Nucleoview NC-3000 software.

As shown in FIG. 3, most of the senescent cells were observed to have cell cycle arrest during S phase. However, when Rg3 was treated with aging cells, the G1 phase increased to 72%, indicating that the cell cycle was regressed to the level of the young cell again, and that the aging was reversed.

As described above, the present invention relates to a composition for aging cell rejuvenation comprising ginsenoside Rg3 as an active ingredient, and more particularly, to a composition for aging cells containing ginsenoside Rg3 as an active ingredient, A skin cosmetic composition, a skin external preparation, a food composition, and a pharmaceutical composition. The ginsenoside Rg3 of the present invention recovers aged cells with a cell cycle arrest again to a cell cycle level of a young cell level, and has the effect of rejuvenating aging cells, thus being highly industrially applicable.

Claims (6)

A composition for rejuvenation of aging cells comprising ginsenoside Rg3 represented by the general formula (1) as an active ingredient.
&Lt; Formula 1 >

The composition of claim 1, wherein the composition is a cosmetic composition.
The composition according to claim 1, wherein the composition is an external preparation for skin.
The composition of claim 1, wherein the composition is a food composition.
2. The composition of claim 1, wherein the composition is a pharmaceutical composition.
The composition according to claim 1, wherein the aging of the aging cells regenerates aged cells having cell cycle arrest back to the cell cycle of a young cell level.

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