Application of L-carnosine in resisting solar elastic tissue degeneration

Abstract

The present invention relates to a non-therapeutic use of L-carnosine for increasing the amount of fibril-1 in the dermis after irradiation of the skin with infrared radiation. Furthermore, the present invention relates to a non-therapeutic use of an L-carnosine for increasing the ratio of fibril-1 and troelastin in the skin of a subject. Furthermore, the present invention relates to a non-therapeutic use of L-carnosine for increasing the size and / or amount of elastic fibers in the skin of a subject. Furthermore, the present invention relates to a non-therapeutic use of an L-carnosine for strengthening an elastic fibrous network in the skin of a subject. Furthermore, the present invention relates to a non-therapeutic use of an L-carnosine for preventing or reducing the symptoms of solar elastic tissue degeneration. Furthermore, the present invention relates to a cosmetic composition, said cosmetic composition comprising L-carnosine.

Description

Technical Field

[0001] The present invention relates to a non-therapeutic use of L-carnosine for increasing the amount of fibrillin-1 in the dermis after irradiating the skin with infrared radiation. In addition, the present invention relates to a non-therapeutic use of L-carnosine for increasing the ratio of fibrillin-1 and procollagen in the skin of a subject. In addition, the present invention relates to a non-therapeutic use of L-carnosine for increasing the size and / or amount of elastic fibers in the skin of a subject. In addition, the present invention relates to a non-therapeutic use of L-carnosine for strengthening the elastic fiber network in the skin of a subject. In addition, the present invention relates to a non-therapeutic use of L-carnosine for preventing or alleviating the symptoms of solar elastosis. In addition, the present invention relates to a cosmetic composition comprising L-carnosine.

[0002] Elastic fibers are an essential component of the extracellular matrix in the skin. Elastic fibers consist of bundles of elastin, which are produced by many different cell types including fibroblasts.

[0003] Elastin is the main component (90%) of the elastic fibers in the skin (Ross et al., 1973. Journal of Histochemistry and Cytochemistry 21:199-208; Ross and Bornstein, 1969. Journal of Biological Chemistry 2:366-381). It is a protein of the connective tissue responsible for the elastic properties of the skin.

[0004] Proelastin is the soluble monomer precursor of elastic fibers. After being excreted from cells, proelastin self-associates by aggregation into ~200 nm particles. These particles continue to grow and deposit onto the fibrillin microfibril scaffold.

[0005] As described above, fibrillin is a glycoprotein necessary for the formation of elastic fibers. Fibrillin is secreted by fibroblasts into the extracellular matrix and forms microfibrils, which then provide a scaffold for the deposition of (pro)elastin.

[0006] Fibrillin-1 is a 340 kDa calcium-binding glycoprotein and a key component of the dermal elastic network that exhibits a highly complex molecular organization. In the skin, fibrillin-1 is produced by keratinocytes and fibroblasts (Haynes et al., 1997; British Journal of Dermatology 137:17–23; Kielty and Shuttleworth, 1993. Journal of Cell Science 106:167-173). BACKGROUND OF THE INVENTION

[0007] Solar elastosis is a cosmetic condition that describes skin wrinkling resulting from prolonged and excessive exposure of a subject to sunlight. It has been found that in solar elastosis, abnormal material accumulations of elastic tissue occur in the skin of the subject. In particular, abnormal elastic fibers have been found in these accumulations, which are mainly located in the dermis of the skin of the subject.

[0008] It is hypothesized that this accumulation of abnormal elastic fibers may be related to increased elastin production in photo-damaged skin. In addition, it has been found that prolonged and excessive exposure to sunlight leads to increased tropoelastin gene expression, which may contribute to increased elastin production and abnormal elastic fiber accumulation in photo-damaged skin. These abnormal fibers are dysfunctional compared to elastic fibers with properly deposited elastin and fibrillin-1. In addition, increased fibrillin expression and deposition in photo-damaged skin have been reported.

[0009] However, a detailed study, namely Chen et al., "Heat Modulation of Tropoelastin, Fibrillin-1, and Matrix Metalloproteinase-12 in Human Skin In Vivo", THE JOURNAL OF INVESTIGATIVE DERMATOLOGY, 124: January 1, 2005 revealed increased tropoelastin mRNA and protein expression in the epidermis and dermis due to heat. It was also found that fibrillin-1 mRNA and protein expression in the epidermis increased due to heat, but decreased in the dermis. This indicates that this abnormal production of tropoelastin and fibrillin-1 and in particular the abnormal ratio of tropoelastin and fibrillin-1 contribute to abnormal elastin accumulation in the dermis and thus to the formation of solar elastosis.

[0010] As described above, it has been reported by Chen et al. that heat induces an increase in tropoelastin synthesis, and this increase in tropoelastin production results in a higher amount of tropoelastin being deposited on the fibrillin microfibril scaffold. However, at the same time, heat reduces the total amount of fibrillin-1, especially in the dermis, which leads to a reduction in the microfibrils on which tropoelastin needs to be deposited. This process results in dysfunctional elastic fibers.

[0011] Therefore, a reduced ratio of fibrillin-1 to tropoelastin is observed in the skin of subjects with solar elastosis.

[0012] Several treatment options for solar elastosis have been provided, such as dermabrasion, which is the surgical controlled deeper abrasion of the upper to middle layers of the skin with any kind of strong abrasive device, including wire brushes, diamond grinding wheels or reamers, sterile sandpaper, salt crystals or other mechanical components. However, this surgical procedure should only be the last treatment option.

[0013] In addition, active substances such as hyaluronic acid can be injected into the dermis. However, this option requires injection, which is often refused by the subjects, especially since the symptoms of solar elastosis are only related to aesthetics.

[0014] In addition, other active substances can also be applied topically. However, these treatments and the above treatments have variable efficacy.

[0015] Therefore, there is a need to restore an appropriate elastic fiber network to prevent or alleviate the symptoms of solar elastosis, while reducing or avoiding the above disadvantages of the current options.

[0016] Therefore, the main object of the present invention is to provide an improved possibility for restoring an appropriate elastic fiber network, thereby preventing or alleviating the symptoms of solar elastosis. Summary of the Invention

[0017] The main object of the present invention is achieved by the non-therapeutic use of L-carnosine for increasing the amount of fibrillin-1 in the dermis after irradiating the skin with infrared radiation.

[0018] Unexpectedly, it has been found that L-carnosine is able to strongly increase the amount of fibrillin-1 in the dermis after infrared irradiation. At the same time, unexpectedly, when L-carnosine is applied, the amount of elastin after infrared irradiation is not significantly different from the case without infrared irradiation.

[0019] Accordingly, the present invention also relates to a non-therapeutic use of L-carnosine, said L-carnosine being used to increase the ratio of fibrillin-1 and procollagen in the skin of a subject, preferably in the dermis of the subject, after irradiating the skin of the subject with infrared radiation.

[0020] The amount of fibrillin-1 and / or procollagen can be determined by standard methods for determining the amount of nucleic acid or protein in a sample. Such methods are well known to those skilled in the art. Preferably, the term "amount" in this context refers to the amount of protein.

[0021] Preferably, the term "ratio" refers to a weight ratio. Preferably, the term "ratio" refers to a ratio with respect to the amount of substance.

[0022] As used herein, the term "irradiation" includes exposing, for example, the skin to natural sunlight or to artificial light such as an IR lamp. Preferably, the artificial light provides radiation having an intensity comparable to that of natural sunlight. Thus, preferably, artificial light such as an IR lamp provides infrared irradiation having an intensity in the range of 10 mW / cm 2 to 200 mW / cm 2 and preferably in the range of 15 mW / cm 2 to 150 mW / cm 2 of intensity.

[0023] As described above, infrared irradiation (heat) causes the formation of abnormal elastic fibers. Thus, after infrared irradiation, the size and amount of proper elastic fibers are reduced.

[0024] Preferably, as used herein, the term "skin cell" refers to skin cells in vivo, i.e., in the skin of a subject, or in vitro, i.e., in a culture as a cell culture or a biopsy, including fibroblasts and keratinocytes, each as described herein.

[0025] Preferably, the term "elastic fiber" refers to a fiber comprising an amorphous elastin core, preferably an elastin core surrounded by one or more glycosaminoglycans, preferably heparan sulfate, and one or more proteins selected from microfibril-associated glycoproteins, fibrillin, fibulin, and elastin receptors.

[0026] Unexpectedly, it has been found that L-carnosine can reverse this reduction in the size and amount of elastic fibers. Unexpectedly, an increase in the amount of fibrillin-1 in the dermis causes an increase in the amount of proper elastic fibers because there are more fibrillin-1 microfibrils on which procollagen can be deposited. This effect causes an increase in the size and amount of elastic fibers.

[0027] Accordingly, the present invention also relates to a non-therapeutic use of L-carnosine for increasing the size and / or amount of elastic fibers in the skin after irradiating the skin of a subject with infrared radiation.

[0028] The size and amount of elastic fibers can be determined by well-known methods, such as visual or software-based image analysis of microscopic images of tissue staining, preferably where the images are taken by confocal laser scanning microscopy.

[0029] Likewise, an increase in the amount of fibrillin-1 in the dermis and an increase in the amount of appropriate elastic fibers cause an increase in the connectivity of the elastic fibers and an increase in the organization of the elastic fibers in the connective tissue in the skin, following a reduction in these parameters by infrared irradiation.

[0030] Accordingly, the present invention also relates to a non-therapeutic use for strengthening the elastic fiber network in the skin after irradiating the skin of a subject with infrared radiation.

[0031] The strengthened elastic fiber network in the skin provides increased resistance to tensile forces in the tissue, which can be determined by atomic force microscopy of skin biopsies or by measurements using a cutometer.

[0032] Furthermore, the strengthened elastic fiber network is represented by an increased organization of the elastic fibers in the connective tissue, which can be determined, for example, by visual or software-based image analysis of microscopic images of tissue staining, preferably where the images are taken by confocal laser scanning microscopy.

[0033] The term "increase" as used in the context of the non-therapeutic use according to the present invention preferably describes a comparison of a tissue or tissue sample to which L-carnosine has been administered with an untreated control. Preferably, the term describes a comparison of a subject suffering from solar elastosis before and after administration of L-carnosine.

[0034] As described above, abnormal elastic fibers are found in the dermal deposits of subjects suffering from solar elastosis. Typically, the symptoms of solar elastosis are or include the formation of wrinkles. As described above, it has been found that L-carnosine helps to restore proper elastic fiber production and thus also reduces wrinkles.

[0035] Accordingly, the present invention also relates to a non-therapeutic use of L-carnosine for preventing or alleviating the symptoms of solar elastosis, wherein the symptoms of solar elastosis to be prevented or alleviated are or include skin wrinkling.

[0036] The term "preventing or reducing" skin wrinkling preferably refers to the number, length, and / or depth of skin wrinkles. The number and length of wrinkles can be determined by counting and measuring in images taken of or directly from the subject. The depth of skin wrinkles can be determined by software-based high-resolution image analysis, such as by using VAM software from Canfield Scientific Inc., such as version 5.9.7, or by high-definition fast optical in vivo topometry of human skin (FOITS HD ), or by using the PRIMOS from Canfield Scientific Inc CR system.

[0037] L-carnosine can be administered to the skin cells or skin of a subject in any possible way. However, L-carnosine is typically administered topically to the skin of the subject. The skin of the subject consists of different layers, and the outer layer of the skin provides very weak permeability of L-carnosine. Therefore, substances for enhancing the penetration of L-carnosine into the skin are often administered. Such substances are well known to those skilled in the art, and the enhanced penetration can be determined by measuring the penetration depth of L-carnosine, such as by visual microscopy of tissue staining obtained especially from skin biopsies as described herein.

[0038] Therefore, preferably, in the non-therapeutic use according to the present invention, L-carnosine is administered topically to the skin of the subject or to a skin biopsy,

[0039] wherein L-carnosine is administered together with a substance that enhances the penetration of L-carnosine into the skin or skin biopsy, preferably into the dermis.

[0040] It is also preferred that the use is an in vitro use, and wherein the skin of the subject refers to a skin biopsy of the subject,

[0041] preferably wherein L-carnosine is administered topically to the skin biopsy, and wherein L-carnosine is administered together with a substance that enhances the penetration of L-carnosine into the skin biopsy, preferably into the dermis.

[0042] As used herein, the term "skin biopsy" preferably refers to removed cells or portions of a subject's skin. This skin biopsy can be obtained by shaving and collecting the top layer of the skin (epidermis and a portion of the dermis). Additionally, this skin biopsy can be obtained by using a punch tool to remove a small core of the skin (including deeper layers (epidermis, dermis, and superficial fat)). Additionally, this skin biopsy can be obtained by cutting with a knife or similar tool (e.g., a scalpel) to remove an entire area of the skin (including a portion of the skin down through or across the skin fat layer).

[0043] Generally, any substance can be used that enhances the penetration of L-carnosine into the skin or a skin biopsy. However, it has been found that 1,2 - hexanediol, 1,2 - heptanediol, 1,3 - propanediol, pentylene glycol, butylene glycol, propylene glycol, ethoxydiglycol, dipropylene glycol, isopropyl myristate, dimethyl isosorbide, ethanol, octylene glycol, and phenoxyethanol provide particularly favorable penetration of L-carnosine.

[0044] Thus, preferably, in the non - therapeutic use according to the present invention, the one, two, three, or more or all substances that enhance the penetration of L-carnosine into the skin or a skin biopsy, preferably into the dermis, are selected from the group consisting of: 1,2 - hexanediol, 1,2 - heptanediol, 1,3 - propanediol, pentylene glycol, butylene glycol, propylene glycol, ethoxydiglycol, dipropylene glycol, isopropyl myristate, dimethyl isosorbide, ethanol, octylene glycol, phenoxyethanol, and combinations thereof.

[0045] Preferably, L-carnosine is applied to cosmetic formulations.

[0046] Different concentrations of L-carnosine have been tested, and higher concentrations provide a stronger effect.

[0047] Thus, preferably, L-carnosine is applied to cosmetic formulations, wherein, based on the total weight of the cosmetic formulation, preferably based on the total weight of the dry matter of the cosmetic formulation, the amount of L-carnosine contained in the cosmetic formulation is in the range of 0.05 wt.-% to 10 wt.-%, preferably 0.075 wt.-% to 7.5 wt.-%, preferably 0.1 wt.-% to 5 wt.-%, preferably 0.125 wt.-% to 4 wt.-%, preferably 0.15 wt.-% to 3 wt.-%, preferably 0.175 wt.-% to 2.5 wt.-%.

[0048] Unexpectedly, it has been found that, based on the total weight of the cosmetic formulation, preferably based on the total weight of the dry matter of the cosmetic formulation, an amount of at least 0.2 wt.-% of L-carnosine provides particularly favorable results.

[0049] Therefore, it is preferred that L-carnosine is applied to a cosmetic preparation, wherein, based on the total weight of the cosmetic preparation, preferably based on the total weight of the dry matter of the cosmetic preparation, the amount of L-carnosine comprised in the cosmetic preparation is in the range of 0.15 wt.-% to 10 wt.-%, preferably 0.15 wt.-% to 7.5 wt.-%, preferably 0.15 wt.-% to 5 wt.-%, preferably 0.175 wt.-% to 4 wt.-%, preferably 0.175 wt.-% to 3 wt.-%, preferably 0.175 wt.-% to 2.5 wt.-%, preferably at least 0.15 wt.-%, preferably at least 0.175 wt.-%, preferably at least 0.2 wt.-%.

[0050] Furthermore, it is preferred that the cosmetic preparation is applied in such a way that the application amount of L-carnosine is in the range of 1 μg to 7.5 μg per square centimeter of the skin surface or the surface of a skin biopsy.

[0051] It has been found that L-carnosine can be applied to prevent the negative effects of infrared irradiation. Therefore, L-carnosine can be applied before irradiation.

[0052] Therefore, it is preferred that, before exposing the skin or a skin biopsy to infrared irradiation,

[0053] preferably within the range of 1 minute to 48 hours before said exposure, preferably within the range of 2 minutes to 24 hours, preferably within the range of 5 minutes to 12 hours, preferably within the range of 10 minutes to 6 hours, preferably within the range of 15 minutes to 4 hours, preferably within the range of 20 minutes to 2 hours, preferably within the range of 30 minutes to 1 hour, L-carnosine is applied to the skin or a skin biopsy.

[0054] Similarly, it has also been found that L-carnosine can be applied to reduce the negative effects of infrared irradiation. Therefore, L-carnosine can be applied after irradiation.

[0055] Therefore, it is preferred that, after exposing the skin or a skin biopsy to infrared irradiation,

[0056] preferably within the range of 1 minute to 48 hours after said exposure, preferably within the range of 2 minutes to 24 hours, preferably within the range of 5 minutes to 12 hours, preferably within the range of 10 minutes to 6 hours, preferably within the range of 15 minutes to 4 hours, preferably within the range of 20 minutes to 2 hours, preferably within the range of 30 minutes to 1 hour, L-carnosine is applied to the skin or a skin biopsy.

[0057] Furthermore, the present invention relates to a cosmetic composition, said cosmetic composition comprising

[0058] (i) L-carnosine, and

[0059] (ii) one, two, three or more substances that enhance the penetration of L-carnosine into the skin or skin biopsy

[0060] wherein the amount of L-carnosine in the composition is sufficient to

[0061] - increase the amount of fibrillin-1 in the dermis after irradiating the skin with infrared radiation, and / or

[0062] - increase the ratio of fibrillin-1 to procollagen in the skin of the subject, preferably in the dermis of the subject, after irradiating the skin of the subject with infrared radiation, and / or

[0063] - increase the size and / or amount of elastic fibers in the skin after irradiating the skin of the subject with infrared radiation, and / or

[0064] - strengthen the elastic fiber network in the skin after irradiating the skin of the subject with infrared radiation, and / or

[0065] - prevent or alleviate the symptoms of solar elastosis, wherein the symptoms of solar elastosis prevented or alleviated are or include skin wrinkling.

[0066] Advantageously, this composition can be used for non-therapeutic uses according to the present invention.

[0067] To determine whether the corresponding increase, enhancement, prevention or alleviation is achieved by the amount of L-carnosine in the composition, the composition can be administered so as to determine whether the corresponding effect of the non-therapeutic use as described herein is achieved.

[0068] In addition, to determine whether the corresponding increase, enhancement, prevention or alleviation is achieved by the amount of L-carnosine in the composition, an in vitro method can be performed, wherein skin cells or skin biopsies are provided and cultured and wherein the corresponding parameters are measured. Subsequently, a corresponding amount of L-carnosine (or a composition containing the corresponding amount of L-carnosine) is administered, and the skin cells or skin biopsies are irradiated with infrared radiation. Thereafter, the above parameters are measured again and compared with the first measurement results.

[0069] Thus, preferably, a sufficient amount of L-carnosine in the cosmetic composition describes the amount that causes the corresponding increase, enhancement, prevention or alleviation when performing the following in vitro analysis:

[0070] i) Provide and culture a cell culture containing skin cells or skin biopsies,

[0071] ii) Measure the amount of fibrillin-1 and / or procollagen in the cell culture or skin biopsy, and / or

[0072] Measuring the size and / or amount of elastic fibers in a skin biopsy, and / or

[0073] Measuring the connectivity of elastic fibers in a skin biopsy, and / or

[0074] Measuring the amount of fibrillin-1 and / or procollagen in a cell culture or a skin biopsy and determining the ratio of fibrillin-1 to procollagen, and / or

[0075] Measuring the number, length, and / or depth of skin wrinkles in a skin biopsy,

[0076] iii) Administering a predetermined amount of L-carnosine to the cell culture or the skin biopsy,

[0077] iv) Irradiating the skin cells or the skin biopsy,

[0078] Preferably with infrared radiation having an irradiance in the range of 10 mW / cm 2 to 200 mW / cm 2 and preferably in the range of 15 mW / cm 2 to 150 mW / cm 2 and preferably with infrared radiation having an irradiance in the range of 500 J / cm

[0079] Preferably with infrared radiation having an irradiance in the range of 500 J / cm 2 to 800 J / cm 2 and preferably in the range of 600 J / cm 2 to 750 J / cm 2 and preferably with infrared radiation having an irradiance in the range of 600 J / cm

[0080] v) Repeating step ii) and comparing the measurement results obtained with the measurement results obtained in step ii).

[0081] Preferably, step iv) of the in vitro method is performed after step iii) of the in vitro method. Preferably, step iv) of the in vitro method is performed before step iii) of the in vitro method.

[0082] Preferably, as used herein, the term "cell culture" refers to a two-dimensional cell culture containing the corresponding cells, and the term "skin biopsy" preferably refers to a three-dimensional culture obtained and cultured from a subject.

[0083] Furthermore, preferably, in the cosmetic composition according to the present invention, the one, two, three or more or all substances that enhance the penetration of L-carnosine into the skin or skin biopsy are selected from the group consisting of: 1,2 - hexanediol, 1,2 - heptanediol, 1,3 - propanediol, pentylene glycol, butylene glycol, propylene glycol, ethoxydiglycol, dipropylene glycol, isopropyl myristate, dimethyl isosorbide, ethanol, octylene glycol, phenoxyethanol, and combinations thereof.

[0084] Preferably, the terms "increase", "enhance", and "reduce" describe a difference in the corresponding parameter of at least 5%, preferably at least 7.5%, preferably at least 10%, preferably at least 12.5%, preferably at least 15%, preferably at least 20% between the untreated sample and the treated sample, or between before and after treatment, based on the untreated sample or based on the measurement results before treatment.

[0085] Furthermore, the present invention relates to a method for increasing the amount of fibrillin - 1 in the dermis after irradiating skin cells with infrared radiation, the method comprising the steps of:

[0086] i) identifying a patient in need of increasing the amount of fibrillin - 1 in the dermis, and

[0087] ii) administering L - carnosine to the skin cells.

[0088] Furthermore, the present invention relates to a method for increasing the ratio of fibrillin - 1 and procollagen in the skin, preferably in the dermis, after irradiating the skin of a subject with infrared radiation, the method comprising the steps of:

[0089] i) identifying a patient in need of increasing the ratio of fibrillin - 1 and procollagen in the skin, preferably in the dermis, and

[0090] ii) administering L - carnosine to the skin cells.

[0091] Furthermore, the present invention relates to a method for increasing the size and / or amount of elastic fibers in the skin after irradiating the skin of a subject with infrared radiation, the method comprising the steps of:

[0092] i) identifying a patient in need of increasing the size and / or amount of elastic fibers in the skin, and

[0093] ii) administering L - carnosine to the skin cells.

[0094] Furthermore, the present invention relates to a method for strengthening the elastic fiber network in the skin after irradiating the skin of a subject with infrared radiation, the method comprising the steps of:

[0095] i) identifying a patient in need of strengthening the elastic fiber network in the skin, and

[0096] ii) Administer L-carnosine to skin cells.

[0097] Furthermore, the present invention relates to a method for preventing or alleviating the symptoms of solar elastosis, the method comprising the steps of:

[0098] i) Identifying a patient in need of preventing or alleviating the symptoms of solar elastosis, and

[0099] ii) Administering L-carnosine to skin cells,

[0100] wherein the symptoms of solar elastosis to be prevented or alleviated are or include skin wrinkling. Description of the Drawings

[0101] Figure 1 Shows the percentage of surface positive for fibrillin-1 according to Example 2.2, where untreated control (1; batch C1), treated with Composition A (2; batch C2), treated with Composition B (3; batch T), untreated control after IR irradiation (4; batch IR), treated with Composition A after IR irradiation (5; batch IR-C2), treated with Composition B after IR irradiation (6; batch IR-T), error bars represent standard deviation, where **: p < 0.01.

[0102] Figure 2 Shows the percentage of surface positive for tropoelastin according to Example 2.3, where untreated control (1; batch C1), treated with Composition B (2; batch T) and treated with Composition B after IR irradiation (3; batch IR-T), error bars represent standard deviation.

[0103] Other aspects and advantages of the present invention result from the following description of the preferred embodiments. Detailed Description

[0104] Example

[0105] Example 1: Research Conditions

[0106] Example 1.1: Test Composition

[0107] In the following studies, the following compositions were provided and tested.

[0108] Composition A (comparative)

[0109]

[0110] Composition B (according to the present invention)

[0111]

[0112]

[0113] The composition is provided as follows:

[0114] The polymer is sprayed onto Phase A and allowed to swell, then heated to 80 °C. Phase C is heated to 80 °C. Phase B is dispersed into Phase A and Phase C is added to the mixture of A and B under high shear. Neutralize with Phase D and cool. Where applicable: at a temperature of <40 °C, Phase E is added. Set the pH to 6.

[0115] Before, during, and after the study period, the composition has been stored at room temperature.

[0116] Example 1.2: Skin Sample

[0117] Human skin explants with an average diameter of 12 mm (±1 mm) are prepared during abdominoplasty of a 54-year-old Caucasian female with Fitzpatrick skin type II. The explants are kept viable in BEM medium (explants medium of BIO-EC) at 37 °C in a humidified, 5%-CO2 atmosphere.

[0118] The skin samples are divided into batches as follows

[0119] Batch Name Number of Samples Sampling Time C0 Tissue Control 3 Day 0 C1 Untreated Control 3 Day 7 C2 Composition A 3 Day 7 T Composition B 3 Day 7 IR IR-Irradiation 3 Day 7 IR-C2 IR-Irradiation, Composition A 3 Day 7 IR-T IR-Irradiation, Composition B 3 Day 7

[0120] On days 0, 1, 2, 3, and 6 (30 minutes before exposure to infrared irradiation), compositions A and B are applied at a rate of 2 μL / 1 cm 2 skin sample (≈2 mg / cm 2 ) and spread using a spatula. Control sample C0 and control sample C1 did not receive any treatment other than renewal of the medium. Half of the medium (1 ml / well) is renewed on days 2, 3, and 6.

[0121] Example 1.3: Irradiation

[0122] On day 6, the medium of all batches is replaced with HBSS (Hank's balanced salt solution; 1 ml per explant). Batches IR, IR-C2, and IR-T are irradiated using an infrared lamp (Dr FISCHER 1000W, 235V 2500K; 760 - 3000 nm) for 1:24 hours (720 J / cm 2 ; 140 mW / cm 2 ). The non-irradiated batches are kept in HBSS in the dark. At the end of irradiation, all explants are returned to 2 mL of fresh BEM medium.

[0123] On day 0, 3 samples of batch C0 were collected and cut into two parts. Half were fixed in buffered formalin while the other half was frozen at -80 °C. On day 7, 24 hours after exposure to infrared, 3 samples of each batch were collected and processed in the same manner as the samples of C0 on day 0.

[0124] Example 1.4: Histological Processing

[0125] After fixation in buffered formalin for 24 hours, the samples were dehydrated and impregnated in paraffin using a Leica PEARL automatic dehydrator. The samples were embedded using a Leica EG 1160 embedding station. 5-μm thick sections were made using a Leica RM 2125 Minot-type microtome and the sections were mounted on histological slides. Microscopic observations were achieved using a Leica DMLB, Olympus BX43 or BX63 microscope. The pictures were digitized using a digital DP72 or DP74 Olympus camera with cellSens storage software.

[0126] Cell viability analysis:

[0127] After Masson's trichrome staining (Goldner variant), cell viability of epidermal and dermal structures was observed on formaldehyde-fixed paraffin-embedded (FFPE) skin sections. Cell viability was evaluated by microscopic observation. Batches of interest: all.

[0128] Fibrillin-1 immunostaining:

[0129] Fibrillin-1 was stained on frozen skin sections with a polyclonal antifibrillin-1 antibody (Sigma Aldrich, ref. HPA021057), diluted 1:200 in PBS-BSA 0.3%, incubated overnight at room temperature, and visualized using AF488 (Life technologies, A11008). The nuclei were counterstained with propidium iodide. The staining was evaluated by microscopic observation and semi-quantified by image analysis. Batches of interest: all.

[0130] Proelastin immunostaining:

[0131] Proelastin was stained on frozen skin sections with monoclonal anti-elastin antibody (Chemicon, ref. MAB2503), diluted 1:50 in PBS-BSA 0.3%, incubated overnight at room temperature, and visualized using AF488 (Life technologies, A11001). Nuclei were counterstained with propidium iodide. Staining was evaluated by microscopic observation and semi-quantified by image analysis. Batches of interest: All.

[0132] Image analysis:

[0133] Fibrillin-1 immunostaining was evaluated by microscopic observation of skin sections in the context of semi-quantification by image analysis using the software cellSens (Olympus).

[0134] Batches of interest: All.

[0135] Analysis area: ROI, including papillary dermis.

[0136] Number of analyzed images per batch: 9.

[0137] Statistical test: Student T-test.

[0138] Proelastin immunostaining was evaluated by microscopic observation of skin sections in the context of semi-quantification by image analysis using the software cellSens (Olympus).

[0139] Batches of interest: All.

[0140] Analysis area: ROI, including papillary dermis.

[0141] Number of analyzed images per batch: 9.

[0142] Statistical test: Student T-test.

[0143] Example 2: Research Results

[0144] Example 2.1: Cell Viability

[0145] As described in Example 1.4, cell viability of different batches was determined. The following results were obtained:

[0146]

[0147] In the epidermis, without irradiation:

[0148] At day 0, on the blank batch (C0), cell viability was good. At day 7, on the blank batch (C1), cell viability was fairly good.

[0149] Comparison with C1:

[0150] Composition A (C2) induced slight epidermal alterations, and composition B (T) did not induce modifications.

[0151] Comparison with C2:

[0152] Composition B (T) induced slight improvements.

[0153] In the epidermis, there was irradiation:

[0154] Comparison of IR with C1:

[0155] Infrared irradiation did not induce modifications in cell viability.

[0156] Comparison with IR:

[0157] Composition A (C2) induced very slight epidermal alterations. Composition B (T) induced very slight epidermal alterations.

[0158] Comparison with C2:

[0159] Composition B (T) did not induce modifications.

[0160] In the dermis:

[0161] No alterations were detected in any of the samples.

[0162] Conclusion:

[0163] The test conditions and the treatments applied did not reduce cell viability in the dermis, and only slight modifications were observed in the epidermis.

[0164] Example 2.2: Fibrillin-1

[0165] As described in Example 1.4, the percentage of the surface positive for fibrillin-1 immunostaining in the papillary dermis of different batches was determined. The following results (including the standard deviation, SD) were obtained:

[0166]

[0167] The results are also shown in Figure 1 .

[0168] Comparison of IR with C1:

[0169] Infrared irradiation induced a significant 61% decrease in fibrillin-1 expression in the papillary dermis **(where**: p < 0.01).

[0170] Comparison with IR:

[0171] Composition A did not result in a significant decrease in the expression of profibrillin-1. However, Composition B induced a significant increase of 140% (where: p < 0.01).

[0172] Comparison with IR-C2:

[0173] Composition B induced a significant increase of 101% (where: p < 0.01).

[0174] Example 2.3: Proelastin

[0175] As described in Example 1.4, the surface percentage positive for immunoreactivity to tropoelastin in the nipple dermis of different batches was determined. The following results (including standard deviation, SD) were obtained:

[0176]

[0177] No significant differences were observed between the untreated control (C1), the samples treated with Composition B (T), and the samples irradiated and treated with Composition B (IR-T).

[0178] The results are also shown in Figure 2 .

Claims

1. A non-therapeutic use of L-carnosine, wherein the L-carnosine is used to increase the amount of fibrillin-1 in the dermis after irradiating the skin with infrared radiation.

2. A non-therapeutic use of L-carnosine, wherein the L-carnosine is used to increase the ratio of fibrillin-1 to procollagen in the skin after irradiating the skin of a subject with infrared radiation.

3. A non-therapeutic use of L-carnosine, wherein the L-carnosine is used to increase the size and / or amount of elastic fibers in the skin after irradiating the skin of a subject with infrared radiation.

4. A non-therapeutic use of L-carnosine, wherein the L-carnosine is used to strengthen the elastic fiber network in the skin after irradiating the skin of a subject with infrared radiation.

5. A non-therapeutic use of L-carnosine, wherein the L-carnosine is used to prevent or alleviate the symptoms of solar elastosis, wherein the symptoms of solar elastosis to be prevented or alleviated are or include skin wrinkling.

6. The non-therapeutic use according to any one of claims 1 to 5, wherein the L-carnosine is topically applied to the skin of a subject or to a skin biopsy, wherein the L-carnosine is applied together with a substance that enhances the penetration of L-carnosine into the skin or skin biopsy, preferably into the dermis.

7. The non-therapeutic use according to claims 1 to 4, wherein the use is an in vitro use, and wherein the skin of the subject refers to a skin biopsy of the subject, preferably wherein the L-carnosine is topically applied to the skin biopsy, and wherein the L-carnosine is applied together with a substance that enhances the penetration of L-carnosine into the skin biopsy, preferably into the dermis.

8. The non-therapeutic use according to claim 6 or 7, wherein the one, two, three or more or all substances that enhance the penetration of L-carnosine into the skin or skin biopsy, preferably into the dermis, are selected from the group consisting of: 1,2-hexanediol, 1,2-heptanediol, 1,3-propanediol, pentylene glycol, butylene glycol, propylene glycol, ethoxydiglycol, dipropylene glycol, isopropyl myristate, dimethyl isosorbide, ethanol, octylene glycol, phenoxyethanol and combinations thereof.

9. The non-therapeutic use according to claims 6 to 8, wherein the L-carnosine is applied to a cosmetic preparation, preferably wherein, based on the total weight of the cosmetic preparation, preferably based on the total weight of the dry matter of the cosmetic preparation, the amount of L-carnosine contained in the cosmetic preparation is in the range of 0.05 wt.-% to 10 wt.-%, preferably 0.075 wt.-% to 7.5 wt.-%, preferably 0.1 wt.-% to 5 wt.-%, preferably 0.125 wt.-% to 4 wt.-%, preferably 0.15 wt.-% to 3 wt.-%, preferably 0.175 wt.-% to 2.5 wt.-%.

10. The non-therapeutic use according to claim 9, wherein the cosmetic preparation is administered in such a way that the amount of L-carnosine administered is in the range of 1 μg to 7.5 μg per square centimeter of the skin surface or the surface of the skin biopsy.

11. The non-therapeutic use according to any one of the preceding claims, wherein before exposing the skin or skin biopsy to the infrared irradiation, preferably within the range of 1 minute to 48 hours before the exposure, preferably within the range of 2 minutes to 24 hours, preferably within the range of 5 minutes to 12 hours, preferably within the range of 10 minutes to 6 hours, preferably within the range of 15 minutes to 4 hours, preferably within the range of 20 minutes to 2 hours, preferably within the range of 30 minutes to 1 hour, L-carnosine is administered to the skin or skin biopsy.

12. The non-therapeutic use according to any one of claims 1 to 10, wherein after exposing the skin or skin biopsy to the infrared irradiation, preferably within the range of 1 minute to 48 hours after the exposure, preferably within the range of 2 minutes to 24 hours, preferably within the range of 5 minutes to 12 hours, preferably within the range of 10 minutes to 6 hours, preferably within the range of 15 minutes to 4 hours, preferably within the range of 20 minutes to 2 hours, preferably within the range of 30 minutes to 1 hour, L-carnosine is administered to the skin or skin biopsy.

13. A cosmetic composition comprising (i) L-carnosine; and (ii) one, two, three or more substances that enhance the penetration of L-carnosine into the skin or skin biopsy, wherein the amount of L-carnosine in the composition is sufficient to - increase the amount of fibrillin-1 in the dermis after irradiating the skin with infrared radiation, and / or - increase the ratio of fibrillin-1 and procollagen in the skin after irradiating the skin of a subject with infrared radiation, and / or - increase the size and / or amount of elastic fibers in the skin after irradiating the skin of a subject with infrared radiation, and / or - strengthen the elastic fiber network in the skin after irradiating the skin of a subject with infrared radiation, and / or - prevent or alleviate the symptoms of solar elastosis, wherein the symptoms of solar elastosis prevented or alleviated are or include skin wrinkling.

14. The cosmetic composition according to claim 13, wherein the sufficient amount of L-carnosine in the cosmetic composition is the amount that causes the corresponding increase, strengthening, prevention or alleviation when performing the following in vitro analysis: i) providing and culturing a cell culture comprising skin cells or a skin biopsy, ii) measuring the amount of fibrillin-1 and / or procollagen in the cell culture or the skin biopsy, and / or measuring the size and / or amount of elastic fibers in the skin biopsy, and / or measuring the connectivity of the elastic fibers in the skin biopsy, and / or Measure the amount of fibrillin-1 and / or profibrillin in the cell culture or the skin biopsy and determine the ratio of fibrillin-1 to profibrillin, and / or Measure the number, length, and / or depth of skin wrinkles in the skin biopsy, iii) Administer a predetermined amount of L-carnosine to the cell culture or the skin biopsy, iv) Irradiate the skin cells or the skin biopsy, Preferably used with an irradiance of infrared radiation in the range of 10 mW / cm 2 to 200 mW / cm 2 and preferably in the range of 15 mW / cm 2 to 150 mW / cm 2 is irradiated with infrared radiation in this range. Preferably used at an irradiance of infrared radiation in the range of 500 J / cm 2 to 800 J / cm 2 and preferably in the range of 600 J / cm 2 to 750 J / cm 2 ​ v) Repeat step ii) and compare the measurement results obtained with the measurement results obtained in step ii).

15. The cosmetic composition according to claim 13 or 14, wherein the one, two, three or more or all substances that enhance the penetration of L-carnosine into the skin or the skin biopsy are selected from the group consisting of: 1,2-hexanediol, 1,2-heptanediol, 1,3-propanediol, pentylene glycol, butylene glycol, propylene glycol, ethoxydiglycol, dipropylene glycol, isopropyl myristate, dimethyl isosorbide, ethanol, octylene glycol, phenoxyethanol, and combinations thereof.

Images