Hair growth composition

Abstract

To provide a novel hair growth composition. [Solution] A hair growth composition containing CHI3L-1 protein and / or CXCL5 protein is provided, and by applying ingredients that promote the secretion of CHI3L-1 protein and / or CXCL5 protein, such as soapberry extract, to the scalp, the secretion of CHI3L-1 protein and / or CXCL5 protein is promoted, thereby promoting the transition from the resting phase to the growth phase in the hair cycle and promoting hair growth.

Description

Technical Field

[0001] The present invention relates to a hair growth composition.

Background Art

[0002] The hair tissue repeats growth and regression by a hair cycle (hair cycle) having a growth phase of 3 to 6 years, a regression phase of about 2 weeks, and a resting phase of about 3 months. It is the dermal papilla cells that control the hair cycle, and in the hair tissue, there are hair follicles surrounding it. In the growth phase, by activating the dermal papilla cells and hair follicles, the hair matrix cells proliferate and the hair grows. Thereafter, apoptosis is induced by the transition to the regression phase, and the hair falls out during the resting phase. The hair grows thick and long in proportion to the number of hair follicle cells and dermal papilla cells growing in the growth phase and the length of the growth phase.

[0003] In people with alopecia regardless of male or female pattern, due to abnormalities in the hair cycle caused by sex hormones etc., the "growth phase" becomes short, so the hair cannot grow sufficiently continuously and becomes thin and short hair. In addition, since the "resting phase" becomes long, the hair becomes easy to fall out, and its appearance is thin hair.

[0004] This hair growth and hair cycle regulation is controlled by signals secreted from dermal papilla cells which are the command tower of the hair tissue, and these signals are targeted for alopecia treatment. For example, (1) activation of dermal papilla cells and hair follicle cells via various nutrient provision and VEGF which is an angiogenesis promoting factor, IGF, HGF, FGF-2 etc. which are growth factors, (2) promotion of conversion to the growth phase via FGF-7 which is a conversion signal and Wnt which is a differentiation inducing factor, (3) normalization of the hair cycle by suppressing the conversion to the regression phase via conversion signals such as FGF-5 fibroblast growth factor 5 and suppression of male hormone secretion etc., aiming to improve alopecia and maintain hair.

[0005] Specifically, Patent Document 1 describes how horse chestnut extract contributes to hair nourishment and growth by promoting peripheral blood flow in the scalp and activating hair follicle cells. Patent Document 2 describes how inhibiting fibroblast growth factor 5 (FGF-5) contributes to the prevention of stress-induced alopecia and thinning hair. It is also a well-known fact that minoxidil promotes hair growth by influencing the growth and resting phases. Patent Document 3 describes how administering VEGF, which promotes regenerative blood flow around hair follicles, promotes the transition from the resting to the growth phase of hair follicles and extends the growth phase of hair follicles, thereby promoting hair growth. Furthermore, Patent Document 4 describes a hair composition containing anti-inflammatory substances such as safflower extract and soapberry extract, and antioxidants such as scutellaria baicalensis extract and horse chestnut extract. This hair composition aims to prevent hair loss by inhibiting hair follicle apoptosis, which is the cause of the transition to the regression phase. Thus, various hair growth ingredients have been proposed for thinning hair from the perspective of the above-mentioned mechanisms of action.

[0006] On the other hand, while it is clear that CHI3L-1 protein (Chitinase-3-like protein 1) and CXCL5 protein (CXC motif chemokine ligand 5) can be used as cancer markers, their effects on the hair cycle and hair tissue have not been known until now. [Prior art documents] [Patent Documents]

[0007] [Patent Document 1] Japanese Patent Application Publication No. 4-308523 [Patent Document 2] Japanese Patent Publication No. 2004-91411 [Patent Document 3] Japanese Patent Publication No. 2004-35443 [Patent Document 4] Japanese Patent Publication No. 2007-22923 [Overview of the Initiative] [Problems that the invention aims to solve]

[0008] Incidentally, the inventors of the present invention, seeking an effective means of combating hair loss from a perspective different from the aforementioned mechanism of action, investigated the effects of external stimuli (damage from the outside) on hair tissue. They discovered that a certain factor activates hair follicle cells and promotes the transition from the resting phase to the growth phase in the hair cycle, thus leading to the present invention. In other words, the present invention provides a hair growth composition and its main component based on a novel mechanism of action. It also provides a method for discovering novel hair growth promoting components. [Means for solving the problem]

[0009] The hair growth stimulant according to the present invention contains CHI3L-1 protein and / or CXCL5 protein as active ingredients. [Effects of the Invention]

[0010] The present invention provides a composition containing a hair growth ingredient based on a previously unknown mechanism of action. [Brief explanation of the drawing]

[0011] [Figure 1] Figure 1 shows the effect of damaged dermal papilla cells on the proliferation of hair follicle cells in the presence of other hair follicle cells. ** indicates a statistically significant difference compared to the control at a 1% significance level, and * indicates a statistically significant difference compared to the control at a 5% significance level. [Figure 2] Figure 2 shows the expression rates of specific genes expressed in cultured damaged dermal papilla cells. (A) shows the expression rate of the gene encoding the CHI3L-1 protein, and (B) shows the expression rate of the gene encoding the CXCL5 protein. ** indicates a statistically significant difference compared to the control at a 5% significance level. [Figure 3]Figure 3 shows the effects of CHI3L-1 protein and CXCL5 protein on hair follicle cell proliferation. ** indicates a statistically significant difference compared to the control at a 1% significance level. [Figure 4] Figure 4 shows the fluctuations in the secretion levels of CHI3L-1 protein and CXCL5 protein during the hair cycle. (A) shows the secretion level of CHI3L-1 protein, and (B) shows the secretion level of CXCL5 protein. [Figure 5] Figure 5 shows the changes in the secretion levels of CHI3L-1 and CXCL5 proteins in mouse skin samples in which the hair cycle transition was suppressed by hormones during the hair cycle. (A) shows the secretion level of CHI3L-1 protein, and (B) shows the secretion level of CXCL5 protein. ** indicates a statistically significant difference between the controlled group and the untreated group at a 1% significance level, and * indicates a statistically significant difference at a 5% significance level. [Figure 6] Figure 6 shows the effects of various plant extracts on the secretion levels of CHI3L-1 and CXCL5 proteins in dermal papilla cells. (A) shows the secretion level of CHI3L-1 protein, and (B) shows the secretion level of CXCL5 protein. ** indicates a statistically significant difference compared to the control at a 1% significance level, and * indicates a statistically significant difference compared to the control at a 5% significance level. [Figure 7] Figure 7 shows the effect of various plant extracts on the number of days required for the transition from the resting phase to the growth phase. [Figure 8] Figures 8(A) and 8(B) show the effects of simultaneous administration of various plant extracts on the number of days required to transition from the resting phase to the growth phase, respectively. [Modes for carrying out the invention]

[0012] The present invention relates to a method for utilizing CHI3L-1 protein and / or CXCL5 protein for hair growth promotion and hair restoration. CHI3L-1 protein is also known as YKL-40 or CGP-39 and is known as a various cancer marker protein. On the other hand, CXCL5 protein is also known as LIX, CGP-2, or ENA-78 and is known as an inflammation marker protein.

[0013] The hair growth stimulant according to the present invention contains CHI3L-1 protein and / or CXCL5 protein as active ingredients. This hair growth stimulant may not consist solely of CHI3L-1 protein and / or CXCL5 protein, but may also contain components, whether liquid or solid, such as water or lower alcohols, to facilitate the preparation of the composition. This hair growth stimulant promotes hair growth by contact with the skin, preferably the scalp, through application or other means. The skin can be that of a mammal, but human skin is particularly preferred. Furthermore, it may be administered to animals, including humans, by oral administration or injection in some cases.

[0014] The hair growth promoter can be applied directly to humans or other mammals, but is usually used for the production of hair growth compositions. The hair growth promoter and hair growth composition according to the present invention are used for the purpose of promoting hair growth, and by promoting hair growth, not only recovery from thinning hair but also prevention of thinning hair can be achieved. Therefore, the subjects to which they are applied can include not only those in a state of thinning hair for promoting hair growth but also those in a healthy hair growth state such as prevention of thinning hair for hair growth. Whether the hair growth composition is a composition as a cosmetic, a composition as a quasi-drug, or a composition as a pharmaceutical is not a concern. In addition to the CHI3L-1 protein and / or CXCL5 protein, the hair growth composition may contain bases for use in preparing the composition, such as water, alcohols, starches, oils and fats, solubilizing aids such as surfactants, stabilizers, pH adjusters, and other components necessary for formulation, such as preservatives, fragrances, and coloring agents. The dosage form of the hair growth composition is not particularly limited, and forms as external preparations such as liquids, creams, aerosols, ointments, tapes, patches, shampoos, and rinses, as well as oral preparations such as tablets, capsules, syrups, and oral liquids, and injections are also possible. Further, the production method is not particularly limited and can be produced by a so-called conventional method appropriate for each dosage form.

[0015] The hair growth composition may preferably contain an amount of the CHI3L-1 protein and / or CXCL5 protein that exhibits a hair growth promoting effect by itself, but an amount that exhibits an additive or synergistic effect due to coexistence with other components is also acceptable and can be appropriately determined by those skilled in the art according to the dosage form, administration method, usage method, etc.

[0016] The hair growth promoter according to the present invention promotes the activation and proliferation of hair follicle cells, particularly the activation from the telogen phase to the anagen phase, and promotes hair growth by promoting the transition of the hair cycle from the telogen phase to the anagen phase. In addition to promoting hair growth by imparting CHI3L-1 protein or / and CXCL5 protein, hair growth can also be promoted by contacting a component that promotes the secretion of CHI3L-1 protein or / and CXCL5 protein (secretion-promoting component) with the skin of mammals, particularly the human scalp. Examples of such secretion-promoting components include extracts of various animals and plants. For example, plant extracts that promote the secretion of CHI3L-1 protein include extracts of plants such as iris, elm, and zibopsow. In addition, plant extracts that promote the secretion of CXCL5 protein include extracts of plants such as mugwort and amatachazuru. The parts used for the plant extracts are not limited, and can be the leaves, flowers, fruits, roots, etc. of each plant, or even the whole plant. The extraction solvent used for the extract is also not particularly limited, and hydrophilic solvents such as water, lower alcohols with about 1 to 5 carbon atoms such as ethanol and isopropanol (regardless of whether they are straight-chain alcohols or branched alcohols), polyhydric alcohols such as ethylene glycol and glycerin, and various solvents used for the production of animal and plant extracts regardless of other lipophilic solvents can be used. The extraction method is also not particularly limited, and methods using supercritical fluids or other general extraction methods can be used. In addition, the secretion-promoting component can also be the animal or plant itself (plant bodies or animal bodies, and these can also be parts thereof) or a chemical substance.

[0017] In this application, the aforementioned secretion-promoting components, including these plant extracts, can also be used as hair growth stimulants according to the present invention. One or more of the secretion-promoting components may be used as hair growth stimulants according to the present invention and may be applied directly to the skin of a mammal, preferably the scalp, or a hair growth stimulant composition containing the secretion-promoting components as active ingredients may be applied to the skin. As illustrated above, the hair growth stimulant composition can be used in various forms. In some cases, it may also be administered orally or by injection. As a hair growth stimulant, it may be used by combining only components that promote the secretion of CHI3L-1 protein, or by combining only components that promote the secretion of CXCL5 protein, or by combining both components that promote the secretion of CHI3L-1 protein and components that promote the secretion of CXCL5 protein. For example, a combination of soapberry (plant) and / or its extract and horse chestnut (plant) and / or its extract. This combination is expected to synergistically promote the hair cycle, especially the transition from the resting phase to the growth phase, resulting in a more effective hair growth effect. Furthermore, in addition to the secretion-promoting components mentioned above, CHI3L-1 protein and / or CXCL5 protein may be combined and used as a hair growth stimulant or hair growth stimulant composition. The lower limit of the hair growth stimulant content in the composition may be, for example, 0.00001 w / v%, 0.0001 w / v%, and 0.001 w / v%, as the total amount of CHI3L-1 protein and / or CXCL5 protein. In the case of plant extracts, the lower limit may be 0.001 w / v%, 0.01 w / v%, 0.1 w / v%, and preferably 1.0 w / v%, as the total amount of plant extracts. When combining plant extracts in a quantity ratio that yields a synergistic effect, for example, the mass ratio of soapberry extract to horse chestnut extract is 0.1 or more, preferably 0.5 or more, and preferably soapberry extract:horse chestnut extract = 2:8 to 8:2.

[0018] The screening method according to the present invention includes the steps of culturing dermal papilla cells in a culture medium containing the test substance, and measuring the secretion amount of CHI3L-1 protein and / or CXCL5 protein before and after culturing. If the secretion amount increases compared to when the cells are cultured in a culture medium without the test substance or in a culture medium containing a known secretion-promoting component (a reference secretion-promoting component), it can be determined that the substance is a secretion-promoting substance that promotes the secretion of CHI3L-1 protein and / or CXCL5 protein. The secretion amount of each protein can be measured, for example, by an ELISA method using antibodies against these proteins. Furthermore, if the secretion amount decreases compared to when the cells are cultured in a culture medium without the test substance or in a culture medium containing a known secretion-promoting component (a reference secretion-promoting component), it can be inferred that the substance is a hair growth-inhibiting component. Thus, the screening method according to the present invention is a method that can screen not only hair growth-promoting components but also hair growth-inhibiting components, and is a method that can detect a wide range of hair growth-related components (components that have some effect on hair growth).

[0019] The conditions for culturing dermal papilla cells can be any general conditions used for culturing dermal papilla cells. For example, culture conditions of 37°C and 5% CO2 concentration in D-MEM medium (containing 10% FBS) are shown. The dermal papilla cells used for screening are preferably cells that have been pre-damaged. This is because damaged dermal papilla cells have enhanced proliferative capacity in the hair matrix cells co-cultured with them, and it is expected that hair growth-related components can be identified more effectively. Damage to dermal papilla cells can be inflicted as a pretreatment before contact with the test substance. Examples include culturing under nutrient-deficient conditions, i.e., culturing in a state where some nutrients (components involved in proliferation) in the culture medium for dermal papilla cells are completely absent, or where some nutrients are present at low concentrations. Alternatively, culturing in the presence of various sex hormones (both male and female hormones) or components that suppress cell proliferation, such as anticancer drugs (cell proliferation inhibitors), can also be used. Damage to dermal papilla cells can be inflicted by these methods. Damage can also be inflicted by culturing in the presence of various chemical components or plant and animal extracts. Of course, screening can also be performed using hair papilla cells cultured under normal conditions without inflicting damage. Culture conditions for inflicting damage can be determined as appropriate by those skilled in the art, but prolonged culture may cause hair papilla cells to die. For this reason, although it varies depending on the type and amount (concentration) of damage, it is preferable to limit the duration of damage to approximately 2 to 24 hours, and no longer than 72 hours. For example, in the case of inflicting damage with sex hormones, cell proliferation inhibitors, or animal and plant extracts, a culture medium is used in which the concentration in the medium is at least 0.0001 w / v%, preferably 0.001 to 0.01 w / v%.

[0020] Contact with the test substance is achieved by culturing dermal papilla cells and the test substance together in a culture medium. The concentration of the test substance in the culture medium and the contact time can be appropriately determined by those skilled in the art. Culture conditions and culture time can also be appropriately determined by those skilled in the art, but for example, 24 to 72 hours at 37°C. The test substance is not particularly limited and can be, for example, a plant extract or various chemical substances. In addition, hair follicle cells may be present during contact with the test substance.

[0021] Hair growth-related components can also be screened using the expression levels of the gene encoding the CHI3L-1 protein and / or the gene encoding the CXCL5 protein as indicators. Specifically, hair papilla cells are cultured in contact with the test substance, and the expression levels of these genes expressed in the hair papilla cells are measured to screen for hair growth-related components. There are no particular restrictions on the gene expression levels, and various known methods can be used.

[0022] The dermal papilla cells according to the present invention are obtained by a process of damaging dermal papilla cells and culturing the dermal papilla cells in a state where the damage has been removed. In other words, the damaged dermal papilla cells are brought closer to a so-called recovered state, or a normal state, by being damaged once and then cultured in a normal culture medium. After inflicting damage, it is preferable to immediately transfer the cells to a normal culture medium and culture them, but in order to completely remove the effects of the damage, the damaged cells may be washed with culture medium, water, or a buffer such as PBS.

[0023] Damage can be inflicted, as mentioned above, by culturing dermal papilla cells in the presence of one or more cell proliferation inhibitors such as anticancer drugs or sex hormones, or by culturing dermal papilla cells in a less nutrient-rich medium than that normally used for culture, that is, in a state where some of the nutrients in the dermal papilla cell culture medium are completely absent or at a low concentration. Of course, damage can also be inflicted by culturing in the presence of various other chemical components or plant and animal extracts.

[0024] These recovered dermal papilla cells have the property of increasing the proliferation of epithelial cells such as hair follicle cells compared to undamaged dermal papilla cells. Therefore, by using recovered dermal papilla cells, it is possible to investigate the effect of these cells on hair growth. For example, by co-culturing dermal papilla cells damaged by the above method with dermal papilla cells damaged by other methods or undamaged dermal papilla cells with hair follicle cells and comparing the proliferative capacity of the hair follicle cells after culturing, it is possible to search for factors that affect hair growth. If the proliferative capacity is equivalent or increased as a result of co-culturing, the factors damaged by other methods may be usable as factors that contribute to promoting hair growth, and if the proliferative capacity is inferior, these factors are presumed to be factors that negatively affect healthy hair growth. In addition, by comparing gene expression in undamaged dermal papilla cells with gene expression in recovered dermal papilla cells, it is possible to find out which gene expressions are activated or suppressed. The effects on hair growth include not only those that promote hair growth, but also those that inhibit it. This provides clues to how cells that have recovered from damage are related to hair growth, and how damage affects hair papilla cells. Ultimately, it can lead to the discovery of components that contribute to promoting hair growth, hair development, and overall hair health.

[0025] Thus, the present invention is based on the finding that CHI3L-1 protein and / or CXCL5 protein are factors involved in promoting hair growth, and hair growth can be promoted by stimulating the secretion of either or both of these proteins.

[0026] The present invention will be described in more detail below, but it goes without saying that the present invention is not limited to the following embodiments. [Examples]

[0027] [The effects of stimulation on hair papilla cells] First, we investigated the effects of stimulation during the growth phase. We observed how the proliferation of hair follicle cells changed when hair follicle cells and dermal papilla cells that had been temporarily damaged were co-cultured.

[0028] (1) External damage was inflicted on dermal papilla cells by culturing them at 37°C for 2 hours in a medium containing mitomycin C as a cell proliferation inhibitor, (2) for 24 hours in a medium containing the male hormone (testosterone), and (3) for 24 hours in a medium containing the female hormone (17β-estradiol). D-MEM medium (containing 10% FBS) was used as the culture medium. Alternatively, (4) damage was inflicted by culturing under nutrient-deficient conditions by culturing at 37°C for 24 hours in D-MEM medium without FBS. After inflicting damage, the dermal papilla cells were washed with PBS and cultured in D-MEM medium (containing 10% FBS) for 24 hours to obtain dermal papilla cells that had recovered from the damage.

[0029] Subsequently, the hair papilla cells (5 × 10 3 (cell / well) and hair follicle cells, specifically outer root sheath cells (8×10 4 Cells (manufactured by Cosmo Bio Co., Ltd.) were co-cultured in MCDB153 medium at approximately 37°C for 5 days, and the proliferation rate was determined from the number of viable hair follicle cells. As shown in Figure 1, it was found that temporarily damaged dermal papilla cells have an effect that promotes the proliferation of hair follicle cells.

[0030] Next, the dermal papilla cells restored under the above conditions were cultured in a standard culture medium (containing nutrients in the correct amounts and free from mitomycin C, male hormones, and female hormones) at approximately 37°C for 4 days, and the changes in gene expression during this period were examined using microarray analysis. Total RNA was extracted using Trizol reagent (Thermo Fisher Science) and the RNeasy Mini Kit (QIAGEN), and gene expression levels were evaluated using the 3D-Gene Human Oligo chip 25k (Toray) according to the protocol for the chip. As a result, as shown in Figure 2, an increase in the expression rate of genes encoding several proteins, including CHI3L-1 protein and CXCL5 protein, was observed. In addition, increased expression of several inflammation and bone formation-related factors was confirmed, but no significant changes were observed in the expression levels of the genes encoding VEGF protein, IGF protein, HGF protein, FGF protein, and Wnt protein, which have been considered hair growth factors.

[0031] [Hair follicle cell proliferation effect by CHI3L-1 protein and CXCL5 protein] Hair follicle cell proliferation was attempted using MCDB153 medium containing CHI3L-1 protein and CXCL5 protein. Hair follicle cells (8 × 10⁴) were placed in medium containing 5 ng / ml or 20 ng / ml of each protein. 4 Cells were added (cell / well) and cultured at approximately 37°C for 5 days. The cell proliferation rate was determined from the number of cells after culture, and the results are shown in Figure 3. As a result, it was found that CHI3L-1 protein and CXCL5 protein promote the proliferation of hair follicle cells, respectively.

[0032] [Fluctuations in the secretion levels of CHI3L-1 protein and CXCL5 protein during the hair cycle] The two experiments described above suggested that the secretion of CHI3L-1 and CXCL5 proteins has some influence on the hair cycle and is related to hair growth. Therefore, the secretion levels of CHI3L-1 and CXCL5 proteins in mouse skin samples were measured during the early, mid, and late stages of the growth phase, as well as three weeks before and after the growth phase. Observation of hair cycle changes in 11-week-old C3H / He female mice (Japanese SLC) revealed that the mice were in a resting phase from 11 to 14 weeks of age, transitioning from the resting phase to the growth phase around 14-15 weeks of age (determined by pigmentation, where the skin changes color), with the growth phase continuing for approximately 20 days before transitioning back to the resting phase. Therefore, the backs of 11, 12, and 13-week-old C3H / He female mice in the resting phase were shaved, and six skin samples of the same area were collected using a biopsy trephine (φ6mm). These samples were stored at -80°C as samples representing the transition from the resting phase to the growth phase of the hair cycle, and 1, 2, and 3 weeks prior, respectively. Furthermore, six skin samples were similarly collected from mice on days 3, 9, and 15 after the transition to the hair cycle and stored at -80°C. Similarly, six skin samples were also collected from mice 1, 2, and 3 weeks after the transition from the growth phase to the resting phase and stored at -80°C as samples. Total protein was extracted from the skin samples using T-PER reagent (Thermo Fisher Science) and bead homogenization, and the protein levels of CHI3L-1 protein and CXCL5 protein were measured using an ELISA kit (Funakoshi). The changes in the secretion levels of each protein are shown in Figure 4. The secretion levels of both CHI3L-1 protein and CXCL5 protein increased from the resting phase to the growth phase, peaked in the early growth phase, and then decreased towards the resting phase.

[0033] [Changes in the secretion levels of CHI3L-1 protein and CXCL5 protein in damaged mice] Eight-week-old C3H / He female mice (Japanese SLC) had their backs shaved (approximately 5cm x 2cm), were treated with a thioglycolic acid-containing depilatory cream, and were induced to enter the growth phase. From the following day, a male hormone solution (1mg / ml) or a female hormone solution (0.3mg / ml) was applied to only half of the depilated area for nine consecutive days (100μL / mouse). After euthanasia, three skin samples each from the treated and untreated areas were collected from each individual using a biopsy trephine (φ6mm), and the secretion levels of CHI3L-1 protein and CXCL5 protein in the mouse skin samples were measured in the same manner as described above. The results are shown in Figure 5. As a result, it was observed that the secretion levels of CHI3L-1 protein and CXCL5 protein were lower in the resting phase compared to the growth phase due to the damage.

[0034] These results suggest that increased secretion of CHI3L-1 and CXCL5 proteins promotes the proliferation of hair follicle cells and facilitates the transition from the resting phase to the growth phase. [Examples]

[0035] [Screening of plant extracts] Since the secretion of CHI3L-1 and CXCL5 proteins is predicted to promote the transition of hair tissue to the growth phase and promote the proliferation of hair follicle matrix cells, hair growth stimulants were screened using the amount of CHI3L-1 and CXCL5 protein secretion promotion as an indicator. For the screening, concentrated extracts obtained using aqueous ethanol or aqueous 1,3-butylene glycol were used from approximately 100 types of plants, including iris (root), horse chestnut (seed), soapwort (leaf), aloe (leaf), thyme (above ground), Gynostemma pentaphyllum (leaf), soapberry (pericarp), Echinacea (leaf), and rosehip (pericarp). A predetermined amount of each plant extract was added to D-MEM medium, and dermal papilla cells were cultured for 48 hours. Subsequently, the amounts of CHI3L-1 protein and CXCL5 protein in the medium were measured in the same manner as the ELISA method described above. The results are shown in Figure 6. As a result, it was confirmed that the extracts of iris, horse chestnut, and soapwort (extracted with aqueous ethanol: raw material manufacturer's product) promoted the secretion of CHI3L-1 protein, while the extracts of Gynostemma pentaphyllum and Sapindus mukorossi (extracted with aqueous ethanol: raw material manufacturer's product) promoted the secretion of CXCL5 protein.

[0036] [The effects of plant extracts on promoting hair cycle regeneration] The effects of plant extracts (iris, horse chestnut, soapwort, Gynostemma pentaphyllum, and soapberry) that showed secretion-promoting effects of CHI3L-1 protein or CXCL5 protein in the aforementioned screening study on hair cycle regeneration.

[0037] The backs of 8-week-old C3H / He female mice were shaved (approximately 5 cm x 2 cm), and starting the following day, test solutions of each extract (each extract diluted in 50% ethanol, containing 1.5 w / v%) were continuously applied to the test site. The number of days until 80% of the test site transitioned to the growth phase was measured. The transition from the resting phase to the growth phase was determined by visual observation of pigmentation, which is a change in skin color. The results are shown in Figure 7. As a result, application of these plant extracts was observed to accelerate the transition from the resting phase to the growth phase.

[0038] Next, the effects of two types of extracts, soapberry extract and horse chestnut extract, on the hair cycle were investigated in the same manner as described above. The test solution used contained 1.5 w / v% of the total amount of plant extracts, and 50% ethanol and a solution containing 1.5 w / v% of each extract were used as controls. The results are shown in Figure 8. As shown in the figure, when both soapberry extract and horse chestnut extract were used, the transition to the growth phase was synergistically promoted (the number of days until the transition was reduced) compared to when both soapberry extract and iris extract were used. [Examples]

[0039] [Hair growth effect from plant extracts] Horse chestnut, iris, and soapwort are plant extracts that promote the secretion of CHI3L-1. Soapberry and Gynostemma pentaphyllum are plant extracts that promote the secretion of CXCL5. We investigated whether a synergistic hair growth effect could be observed by combining plant extracts with different mechanisms of action. Hair follicles were isolated from 4-week-old C57 / BL female mice using a scalpel and forceps under a stereomicroscope. They were then cultured in William's E medium (containing 1% penicillin-streptomycin-amphotericin B, 10 ng / mL hydrocortisone (SIGMA), 2 mM L-glutamine, and 10 μg / mL insulin) at 37°C under 5% CO2 in a gelatin sponge (24-well plate). Test substances and controls were added on the day of hair follicle isolation (Day 0), and the culture medium was changed every two days. Hair follicles were photographed under a stereomicroscope on Day 0 and Day 3, and hair length was measured from the photographs using image analysis software. The difference between the Day 3 measurement and the Day 0 measurement was calculated as the amount of hair shaft elongation. Compared to the control group of 50% ethanol, the length increased by the addition of the extract was calculated as the amount of hair shaft elongation.

[0040] Each plant was heat-extracted with aqueous ethanol or aqueous 1,3-butylene glycol, and the concentrated, dried plant extracts were used as test substances. When each plant extract was used individually, it was added to the culture medium so that the concentration of each plant extract in the culture medium was 20 μg / mL. When the plant extracts were used in combination, each plant extract was added to the culture medium so that the concentration of each plant extract in the culture medium was 10 μg / mL (the total concentration of plant extracts in the culture medium was 20 μg / mL). Furthermore, the same experiment was conducted by changing the mixing ratio when horse chestnut and soapberry extracts, which were found to have a synergistic effect, were used together. The results are shown in Tables 1 and 2. As a result, it was found that each plant extract has a hair shaft elongation effect, and a synergistic hair shaft elongation effect was obtained by using horse chestnut extract and soapberry extract together. [Table 1] Table 2

Claims

1. A hair growth promotion method that stimulates hair growth by promoting the secretion of CHI3L-1 protein and / or CXCL5 protein.

2. A method for promoting hair growth according to claim 1, comprising bringing a substance that promotes the secretion of CHI3L-1 protein and / or CXCL5 protein into contact with the skin, preferably the scalp.

3. The hair growth promoting method according to claim 1, wherein the secretion-promoting substance is one or more plants from horse chestnut, soapwort, and mukorossi, and / or extracts of these plants.

4. A hair growth stimulant containing CHI3L-1 protein and / or CXCL5 protein as active ingredients.

5. A hair growth composition comprising CHI3L-1 protein and / or CXCL5 protein.

6. A method for using CHI3L-1 protein and / or CXCL5 protein in the production of a hair growth composition.

7. The process involves culturing hair papilla cells in a culture medium containing the test substance, A method for screening hair growth-related components, comprising the step of measuring the amount of CHI3L-1 protein secreted and / or the amount of CXCL5 protein secreted before and after culture, or the gene expression level of CHI3L-1 protein and / or the gene expression level of CXCL5 protein.

8. The screening method according to claim 7, wherein the hair papilla cells cultured in a culture medium containing the test substance are either damaged cells or undamaged cells.

9. The screening method according to claim 8, wherein the hair papilla cells in which the aforementioned damage has been inflicted are obtained by culturing in a nutrient-poor composition, or by culturing in the presence of at least one of sex hormones or cell proliferation inhibitors.

10. The process of damaging the hair papilla cells, Hair papilla cells obtained by a process of culturing the hair papilla cells in a state where damage has been removed.

11. The step of inflicting the aforementioned damage is: A method for culturing hair papilla cells in the presence of one or more cell proliferation inhibitors or sex hormones, or The hair papilla cells according to claim 10, which is a step of damaging the hair papilla cells by culturing them in a nutrient-lower environment than that used in normal culture media.

12. A method for creating hair papilla cells that have recovered from damage, The process of damaging the hair papilla cells, A method comprising the step of culturing the hair papilla cells in a state where damage has been removed.

13. The step of inflicting the aforementioned damage is: A method for culturing hair papilla cells in the presence of one or more cell proliferation inhibitors or sex hormones, or The hair papilla cells according to claim 12, which is a step of damaging the hair papilla cells by culturing them in a nutrient-lower environment than that used in normal culture media.

14. This is a method for investigating the effect of damaged hair papilla cells on hair growth. The process of damaging the hair papilla cells, The process involves culturing the hair papilla cells in a state where damage has been removed, A method comprising the step of comparing the state of hair papilla cells cultured after the aforementioned damage has been removed with the state of hair papilla cells that have not been damaged.

15. The step of inflicting the aforementioned damage is: A method for culturing hair papilla cells in the presence of one or more cell proliferation inhibitors or sex hormones, or The method according to claim 14, which involves a step of damaging hair papilla cells by culturing them in a nutrient-lower environment than that used in normal culture media.

16. The method according to claim 14 or 15, which evaluates the effect from gene expression in dermal papilla cells.

17. A hair growth promoting composition containing a substance that promotes the secretion of CHI3L-1 protein and / or CXCL5 protein.

18. The hair growth promoting composition according to claim 17, wherein the secretion-promoting substance is one or more plants selected from horse chestnut, soapwort, and mukorossi, and / or extracts of these plants.

19. The hair growth promoting composition according to claim 17 or 18, wherein the secretion-promoting substance is horse chestnut or / and horse chestnut extract and soapberry or / and soapberry extract.

20. A hair growth composition containing horse chestnut or / and horse chestnut extract, and soapberry or / and soapberry extract as active ingredients.

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