Barrier repair composition and application

Abstract

This application provides a barrier-repairing composition and its use. The composition comprises panthenol, niacinamide, folic acid, and cobalamin in a mass ratio of 3-7:1-5:0.0005-0.1:0.00005-0.01. This application also provides uses of the composition and products containing the same. The composition utilizes panthenol, niacinamide, folic acid, and cobalamin in combination, where the components complement and enhance each other, achieving a synergistic effect in upregulating BMAL2 gene expression. This enhances the DNA damage repair capacity of skin fibroblasts and potentially promotes barrier repair. Furthermore, the composition provided herein can significantly reduce the expression of capsaicin receptor 1 (TRPV1) and increase the expression of involucrin, PIEZO1, and TNXB, reducing skin discomfort and enhancing skin barrier function. Furthermore, the composition provided herein can reduce the expression of inflammatory factors, demonstrating its anti-inflammatory effect.

Description

Technical Field

[0001] The present invention relates to the technical field of daily chemical products, and in particular to a barrier repair composition and its application. Background Art

[0002] The core of skin repair lies in rebuilding and strengthening the skin's barrier function. The skin barrier, primarily composed of the stratum corneum, acts as a "brick wall" to resist external stimuli and prevent water loss. When this barrier is damaged, transepidermal water loss increases, leading to dry and sensitive skin and the entry of harmful substances such as bacteria and allergens. Effective skin repair products must replenish essential skin lipids, such as ceramides, fatty acids, and cholesterol, to repair the intercellular lipid structure. They also activate cell proliferation and differentiation, promote keratinocyte production, and accelerate skin metabolism. Furthermore, for inflammatory injuries, products must possess anti-inflammatory activity to inhibit the release of inflammatory factors and promote the repair and regeneration of damaged tissue.

[0003] Therefore, it is very necessary to develop a composition that has the functions of soothing, repairing, anti-inflammatory and stabilizing. Summary of the Invention

[0004] In view of this, the technical problem to be solved by the present invention is to provide a barrier repair composition, product and application thereof. The composition provided in this application achieves a synergistic effect in upregulating BMAL2 gene expression, has a barrier repair function, and can inhibit the expression of inflammatory factors and has an anti-inflammatory effect.

[0005] The present application provides a barrier repair composition comprising panthenol, niacinamide, folic acid and cobalamin in a mass ratio of 3-7:1-5:0.0005-0.1:0.00005-0.01.

[0006] Panthenol, also known as panthenol, is a provitamin B5 (pro-VB5). As a precursor to coenzyme A, it can be converted into pantothenic acid to participate in energy metabolism and fuel the repair of damaged skin. Penetrating deeply into the stratum corneum, panthenol stimulates the synthesis of hyaluronic acid, significantly improving skin hydration and alleviating dryness. In the early stages of inflammation, panthenol can rapidly inhibit inflammatory signaling pathways, reduce the release of inflammatory factors such as prostaglandin E2, alleviate redness, swelling, and stinging, and accelerate the synthesis of lipids such as ceramides and cholesterol in the skin barrier, strengthening the "brick wall" structure and enhancing the skin's ability to resist external irritants.

[0007] Niacinamide, also known as nicotinamide, is an amide compound of niacin (VB3). It inhibits protein-degrading enzymes in the stratum corneum, reducing the breakdown of lipids like ceramide at the source and directly strengthening the lipid structure of the skin barrier. Niacinamide has the ability to block the NF-κB signaling pathway, effectively inhibiting the release of inflammatory factors such as IL-6 and TNF-α, thereby alleviating the inflammatory response. Niacinamide also regulates melanosome transport, inhibits tyrosinase activity, reduces post-inflammatory hyperpigmentation, and improves uneven skin tone, achieving both whitening and barrier repair while combating inflammation.

[0008] Folic acid, a member of the vitamin B9 (VB9) family, is a key coenzyme in DNA synthesis. It accelerates keratinocyte mitosis, increases cell proliferation, and rapidly replaces damaged, aging cells after exposure to UV rays and other factors. Its metabolite, tetrahydrofolate, possesses strong antioxidant properties, neutralizing free radicals and reducing oxidative stress damage to skin cell DNA, proteins, and lipids. This prevents inflammatory chain reactions and helps maintain normal skin function and structural integrity during inflammation.

[0009] Cobalamin, also known as cyanocobalamin and vitamin B12 (VB12), participates in the methionine cycle to maintain methylation reactions, precisely regulates the expression of genes associated with cell differentiation, and guides the normal maturation of keratinocytes. It is particularly crucial for the repair of skin nerve endings. Damaged skin is often accompanied by dysfunctional nerve endings, leading to sensitivity and burning sensations. Cobalamin can improve nerve conduction and relieve discomfort. It works synergistically with other ingredients to alleviate paresthesia caused by inflammation at the neurological level, promoting overall skin repair.

[0010] This application uses a combination of panthenol, niacinamide, folic acid, and cobalamin. The ingredients work together and promote each other, achieving a synergistic effect in upregulating BMAL2 gene expression, thereby enhancing the DNA damage repair ability of skin fibroblasts and exerting a potential role in promoting barrier repair. At the same time, the composition provided in this application can significantly reduce the expression of capsaicin receptor 1 (TRPV1), increase the expression of involucrin, PIEZO1, and TNXB, reduce skin discomfort reactions, and make the skin barrier function more robust. In addition, the composition provided in this application can reduce the expression of inflammatory factors and has the effect of inhibiting inflammation.

[0011] In some specific implementations, the composition includes panthenol, niacinamide, folic acid, and cobalamin in a mass ratio of 4-6:2-4:0.001-0.05:0.0001-0.005.

[0012] In some specific implementations, the composition includes panthenol, niacinamide, folic acid, and cobalamin in a mass ratio of 4.5-5.5:2.5-3.5:0.005-0.015:0.0005-0.0015.

[0013] In some specific implementations, the composition includes panthenol, niacinamide, folic acid, and cobalamin in a mass ratio of 5:3:0.01:0.001.

[0014] The present application also provides the use of the above composition in the preparation of anti-inflammatory products.

[0015] The present application also provides the use of the above composition in preparing repair products.

[0016] The present application also provides the use of the above composition in preparing a product for upregulating the expression of the aryl hydrocarbon receptor nuclear translocation-like protein 2 (BMAL2) gene.

[0017] The present application also provides the use of the above composition in preparing a product for upregulating the expression of one or more of involucrin, PIEZO1 and TNXB.

[0018] The present application also provides the use of the above composition in preparing a product for inhibiting the expression of the capsaicin receptor 1 (TRPV1) gene.

[0019] The present application also provides the use of the above composition in preparing a product for inhibiting the expression of inflammatory factor genes. In some specific implementations, the inflammatory factors include IL-1α, IL-1β, IL-6, IL-8, etc.

[0020] The present invention also provides a product, comprising: the composition described in the above technical solution.

[0021] In some embodiments of the present invention, the above-mentioned products include but are not limited to personal care products, medicines, etc. Those skilled in the art will understand that, in addition to the composition described in the above technical solution, other excipients may also be included, such as pharmaceutical excipients or excipients for personal care products.

[0022] In some embodiments of the present invention, the personal care product includes: a basic care product and / or a makeup product.

[0023] In some embodiments of the present invention, the cosmetic products include but are not limited to:

[0024] (1) Foundation makeup:

[0025] Liquid foundation / cream: used to even out skin tone and cover blemishes;

[0026] BB Cream / CC Cream: A lightweight makeup base that combines skincare and finishing touches.

[0027] Concealer / pen: for covering local areas such as acne, dark circles, etc.

[0028] Loose powder / face powder: sets makeup and reduces facial shine.

[0029] (2) Eye makeup:

[0030] Eyeshadow: adds color and depth to the eyes;

[0031] Eyeliner pencil / liquid / gel: outline the eyeliner to make the eyes look more vivid;

[0032] Mascara: lengthens and thickens eyelashes, adds depth to eyes;

[0033] Eyebrow pencil / eyebrow powder / eyebrow gel: fill in the gaps in the eyebrows and shape the ideal eyebrows.

[0034] (3) Cheek makeup:

[0035] Blush: Add natural redness to cheeks and enhance complexion;

[0036] Contouring cake / stick: Use shading techniques to make the facial contours more three-dimensional;

[0037] (4) Lip makeup:

[0038] Lipstick / Lip Glaze / Lip Gloss: Change or emphasize the color of the lips;

[0039] Lip liner: Draw a clear lip boundary to prevent lipstick from overflowing.

[0040] (5) Multifunctional cosmetics:

[0041] Highlighter stick / liquid / powder: Brighten the high points of the face (such as the bridge of the nose and cheekbones) to create a radiant effect.

[0042] In addition, there are products designed specifically for special occasions, such as waterproof and sweat-proof eyeliner and long-lasting lipstick.

[0043] In some embodiments of the present invention, the basic care products include but are not limited to:

[0044] Facial Cleanser / Face Wash: Gently removes dirt, oil and makeup residue from the face;

[0045] Makeup remover oil / makeup remover water / makeup remover cream: specially designed to completely remove makeup, especially waterproof makeup;

[0046] Toner / Lotion: Use after cleansing to further cleanse residue on the skin surface, replenish moisture to the skin, restore the skin's pH balance, and lay a good foundation for the absorption of subsequent skin care products;

[0047] Essence: Contains high concentrations of active ingredients, providing deep nourishment and repair for specific skin problems (such as anti-aging, moisturizing, whitening, etc.);

[0048] Eye cream: specially designed for the sensitive area around the eyes, it can reduce fine lines, dark circles and tighten the skin around the eyes. The texture is usually light and easy to absorb.

[0049] Day / night lotion or cream: has the functions of protecting, repairing and nourishing, promoting cell regeneration, etc., which can provide the skin with necessary moisture and lock in the replenished moisture;

[0050] Sunscreen: used for UV protection, prevention of photoaging, etc.;

[0051] Facial masks: Provide extra nourishment and care to the skin, such as hydration, pore cleansing or brightening.

[0052] In some specific implementations, the personal care product includes 0.1wt%~10wt% of the composition described in the above technical solution, preferably includes 0.5wt%~9.5wt% of the composition described in the above technical solution, and more preferably includes 1wt%~9wt% of the composition described in the above technical solution.

[0053] This application uses a combination of panthenol, niacinamide, folic acid, and cobalamin. The ingredients work together and promote each other, achieving a synergistic effect in upregulating BMAL2 gene expression, thereby enhancing the DNA damage repair ability of skin fibroblasts and exerting a potential role in promoting barrier repair. At the same time, the composition provided in this application can significantly reduce the expression of capsaicin receptor 1 (TRPV1), increase the expression of involucrin, PIEZO1, and TNXB, reduce skin discomfort reactions, and make the skin barrier function more robust. In addition, the composition provided in this application can reduce the expression of inflammatory factors and has the effect of inhibiting inflammation. BRIEF DESCRIPTION OF THE DRAWINGS

[0054] Figure 1 The relevant expression level bar graph of the targeted genes provided in Experimental Example 1;

[0055] Figure 2 This is a fluorescence photograph of the capsaicin receptor 1 (TRPV1) protein in Experimental Example 2 of this application;

[0056] Figure 3 This is a bar graph of the capsaicin receptor 1 (TRPV1) protein test in Experimental Example 2 of this application. DETAILED DESCRIPTION

[0057] The present invention provides a barrier repair composition, product, and application thereof. Those skilled in the art can, drawing upon the disclosure herein, appropriately modify process parameters to achieve the desired results. The methods and applications of the present invention have been described using preferred embodiments. It is apparent that those skilled in the art can modify, alter, and combine the methods and applications herein to implement and apply the technology of the present invention without departing from the disclosure, spirit, and scope of the present invention.

[0058] The terms "comprising", "having" or "containing", including their grammatical synonyms, should generally be understood as open and non-restrictive, e.g., not excluding other unrecited elements or steps, unless otherwise specifically stated or understood from the context.

[0059] It should be understood that the order of steps or the order in which certain actions are performed are not important as long as the present invention remains operable. Additionally, two or more steps or actions may be performed simultaneously.

[0060] The use of any and all examples or exemplary language, such as "such as" or "including," herein is intended merely to better illustrate the invention and does not limit the scope of the invention unless otherwise claimed. No language in this specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0061] In addition, the numerical ranges and parameters used to define the present invention are approximate values. The relevant numerical values ​​in the specific examples have been presented as accurately as possible. However, any numerical value inherently inevitably contains standard deviations due to individual testing methods. Therefore, unless otherwise expressly stated, it should be understood that all ranges, amounts, values, and percentages used in this disclosure are modified by the word "about." As used herein, "about" generally means that the actual value is within plus or minus 10%, 5%, 1%, or 0.5% of a particular value or range.

[0062] The present application provides a barrier repair composition comprising panthenol, niacinamide, folic acid and cobalamin in a mass ratio of 3-7:1-5:0.0005-0.1:0.00005-0.01.

[0063] This application uses a combination of panthenol, niacinamide, folic acid, and cobalamin. The ingredients work together and promote each other, achieving a synergistic effect in upregulating BMAL2 gene expression, thereby enhancing the DNA damage repair ability of skin fibroblasts and exerting a potential role in promoting barrier repair. At the same time, the composition provided in this application can significantly reduce the expression of capsaicin receptor 1 (TRPV1), increase the expression of involucrin, PIEZO1, and TNXB, reduce skin discomfort reactions, and make the skin barrier function more robust. In addition, the composition provided in this application can reduce the expression of inflammatory factors and has the effect of inhibiting inflammation.

[0064] The present application is further described below with reference to the following embodiments.

[0065] In the following examples, the raw materials were purchased from the market.

[0066] Example 1, Comparative Examples 1-2

[0067] The composition was prepared according to the formula shown in Table 1:

[0068] Table 1 Formulas provided in the examples and comparative examples of this application

[0069]

[0070] Experimental Example 1 Effect of aryl hydrocarbon receptor nuclear translocation-like protein 2 (BMAL2) expression

[0071] Test method:

[0072] 1. Cell seeding

[0073] Fibroblasts were selected and after cell recovery, when the plating rate reached about 60%, the cells were seeded into 12-well plates and incubated overnight in a CO2 incubator (37°C, 5% CO2).

[0074] 2. Test Grouping

[0075] The experimental test groups were grouped according to Table 2.

[0076] Table 2 Test grouping for Test Example 1

[0077]

[0078] 3. Administer medication

[0079] The culture medium was replaced with the culture medium containing the sample to be tested and incubated for 24 hours as shown in Table 2. At the end of the incubation period, the cells were harvested and immediately dry-frozen at -80°C.

[0080] 4. RNA-seq analysis:

[0081] Total RNA was extracted from cells collected under each experimental condition using TriPure Isolation Reagent. Total RNA was then extracted using the MJzol Animal RNA Isolation Kit (Majorivd) according to the manufacturer's standard protocol. The purified total RNA was subjected to mRNA isolation, fragmentation, first-strand cDNA synthesis, second-strand cDNA synthesis, end-repair, 3' end A addition, adapter ligation, and enrichment to complete mRNA sequencing library construction. Sequencing was performed based on the effective library concentration and data output requirements. The sequencing platform used was the Illumina NovaSeq 6000, using the PE150 (pair-end 150 bp) sequencing mode, which involves sequencing 150 bp on each end. cDNA was sequenced using NGS (Next Generation Sequencing) technology on a high-throughput sequencer, such as the Illumina NovaSeq 6000.

[0082] 5. Data processing

[0083] First, Seqtk was applied to filter the initial data, and the splicedmapping algorithm of Hisat2 (version: 2.0.4) was used to perform genome mapping on the preprocessed reads. Stringtie (version: 1.3.0) was applied to count the number of fragments of each gene after Hisat2 alignment, and then the TMM (trimmed mean of M values) method was used for normalization. Finally, the FPKM value of each gene was calculated using a Perl script.

[0084] The calculation formula is as follows:

[0085]

[0086] in:

[0087] Total exon fragments: the number of fragments mapped to the gene exon (fragment: a pair of reads);

[0088] Exon length: the total length of gene exon;

[0089] Mapped reads: the total number of reads mapped to the reference genome;

[0090] We used edgeR to analyze differentially expressed genes between samples. After obtaining the P-value, we performed multiple hypothesis testing correction. The P-value threshold was determined by controlling the FDR (False Discovery Rate). The corrected P-value is the q-value. Furthermore, the FPKM value was used to calculate the fold-change of differential expression.

[0091] The differentially expressed genes were screened for the following conditions:

[0092] 1) Q-value ≤ 0.05 (when the total number of differentially expressed genes in all comparisons is less than 50, the P-value will be automatically adjusted to <= 0.05);

[0093] 2)|log2FC|≥1.

[0094] The results are shown in Table 3 and Figure 1 Table 3 shows the relative expression levels and enhancement rates of the targeted genes provided in Experimental Example 1. Figure 1 This is a bar graph of the relative expression levels of the targeted genes provided in Experimental Example 1.

[0095] Table 3 Relative expression levels and enhancement rates of targeted genes provided in Experimental Example 1

[0096]

[0097] In Table 3, when the t-test method was used for statistical analysis, compared with Example 1, significance was indicated by *, P-value < 0.05 was indicated by *, P-value < 0.01 was indicated by **, P-value < 0.001 was indicated by ***, and P-value < 0.0001 was indicated by ****.

[0098] Aryl hydrocarbon receptor nuclear transporter-like 2 (BMAL2) belongs to the core circadian clock gene family and is mainly expressed in basal stem cells in the skin. BMAL2 overexpression can enhance the DNA damage repair ability of skin fibroblasts. Once BMAL2 is lost, it will lead to keratinocyte cell cycle disorder and delayed wound healing. Figure 1 It can be seen that the composition provided by the present application has a significant upregulation effect on the gene expression of keratinocytes at the BMAL2 gene expression level. Moreover, compared with Comparative Examples 1 and 2, according to the King's method for calculating synergistic synergy, Example 1:

[0099] q=E A+B / (E A +E B -E A ×E B )

[0100] The q value is 4.21. Therefore, the composition provided in Example 1 achieves a synergistic effect in promoting the expression level of the BMAL2 gene compared to a single agent, that is, it can play a potential role in promoting barrier repair function by significantly upregulating BMAL2, an important gene regulated by the skin circadian clock, and is superior to panthenol and the combination of niacinamide, folic acid and cobalamin in this gene-level repair mechanism.

[0101] Experimental Example 2: Effect of Capsaicin Receptor 1 (TRPV1) Protein

[0102] Test method:

[0103] 1. Test system

[0104] The model used in this test is a 3D epidermal skin model (EpiKutis®), batch number: ES250502, provided by Guangdong Boxi Shaanxi Branch.

[0105] 2. Test Grouping

[0106] The groups were divided according to the conditions shown in Table 4.

[0107] Table 4 Test conditions for Test Example 2

[0108]

[0109] 3. Model drug administration

[0110] According to the test group, the models were transferred to a 6-well plate (pre-filled with 0.9 mL of EpiGrowth medium) and the test group number was labeled on the plate. 25 μL of 0.1% SLS solution was added to the surface of the NC, PC, and sample groups and incubated for 30 minutes. After the incubation period, the corresponding concentration of working solution was added to the PC group submerged in the solution. For the sample group, 12.5 μL of the working solution of the test sample was applied to the surface of the model, spread evenly, and incubated in a CO2 incubator (37°C, 5% CO2) for 24 hours. After the incubation period, any residual test substance on the surface of the model was washed with sterile PBS solution, and any residual liquid inside and outside the model was wiped clean with a sterile cotton swab.

[0111] 4. Immunofluorescence test

[0112] The model used for testing was removed by ring cutting and fixed with 4% paraformaldehyde for 24 hours before immunofluorescence detection. The model was photographed and observed under a microscope, and the images were collected and analyzed.

[0113] 5. Calculation of inhibition rate

[0114] Inhibition rates were calculated as follows:

[0115]

[0116] 6. Statistical analysis of results

[0117] Graphs were generated using GraphPad Prism, and the results are expressed as mean ± SD. Comparisons between groups were analyzed using the t-test. All statistical analyses were two-tailed. P < 0.05 was considered a significant difference, and P < 0.01 was considered a highly significant difference.

[0118] The results are shown in Table 5. Figure 2 and Figure 3 Table 5 shows the test results of capsaicin receptor 1 (TRPV1) protein in Experimental Example 2 of this application. Figure 2 This is a fluorescence photo of the capsaicin receptor 1 (TRPV1) protein in Experiment 2 of this application. Figure 3 This is a bar graph of the capsaicin receptor 1 (TRPV1) protein test in Experimental Example 2 of this application.

[0119] Table 5 Test results of capsaicin receptor 1 (TRPV1) protein in Experimental Example 2 of this application

[0120]

[0121] Note: When the t-test method was used for statistical analysis, significance was indicated by *, P-value < 0.05 was indicated by *, and P-value < 0.01 was indicated by **.

[0122] Result analysis:

[0123] Compared with the BC group, the TRPV1 content in the NC group increased significantly, indicating that the stimulation conditions of this test were effective.

[0124] Compared with the NC group, the TRPV1 content in the PC group decreased significantly, indicating that the positive control in this test was effective.

[0125] Compared with the NC group, the TRPV1 contents of Comparative Example 1 and Example 1 were significantly decreased, with inhibition rates of 22.22% and 54.25%, respectively.

[0126] In addition, compared with Comparative Example 1, Example 1 significantly inhibited the expression of capsaicin receptor TRPV1, reduced skin discomfort, and made the skin barrier function more robust.

[0127] Experimental Example 3: Promoting the expression of barrier repair-related proteins

[0128] Test method:

[0129] 1. Test system

[0130] The model used in this test is a 3D epidermal skin model (EpiKutis®), batch number: ES250502, provided by Guangdong Boxi Shaanxi Branch.

[0131] 2. Test Grouping

[0132] Perform group testing according to the conditions shown in Table 6:

[0133] Table 6 Experimental test groups for Test Example 3

[0134]

[0135] 3. Model drug administration

[0136] According to the test group, the models were transferred to a 6-well plate (pre-filled with 0.9 mL of EpiGrowth medium) and the test group number was marked on the 6-well plate. 25 μL of 0.1% SLS solution was added to the surface of the negative control (NC), positive control (PC), and sample groups and incubated for 30 minutes. After the incubation period, the corresponding concentration of working solution was added to the PC group solution. For the sample group, 12.5 μL of the working solution of the test sample was applied to the surface of the model, spread evenly, and incubated in a CO2 incubator (37°C, 5% CO2) for 24 hours. After the incubation period, any residual test substance on the surface of the model was washed with sterile PBS solution, and any residual liquid inside and outside the model was wiped clean with a sterile cotton swab.

[0137] 4. Immunofluorescence test

[0138] The model used for testing was removed by ring cutting and fixed with 4% paraformaldehyde for 24 hours before immunofluorescence detection. The model was photographed under a microscope and the images were collected and analyzed.

[0139] 5. Calculation of promotion rate

[0140] The improvement rate is calculated as follows:

[0141]

[0142] 6. Statistical analysis of results

[0143] Graphs were generated using GraphPad Prism, and the results are expressed as mean ± SD. Comparisons between groups were analyzed using the t-test. All statistical analyses were two-tailed. P < 0.05 was considered a significant difference, and P < 0.01 was considered a highly significant difference.

[0144] The results are shown in Table 7, which is the test results of Experiment 3 of this application.

[0145] Table 7 Test results of Experiment 3 of this application

[0146]

[0147] In Table 7: #Integrated Optical Density (IOD) test results. When statistical analysis was performed using the t-test method, significance is indicated by *, P-value < 0.05 is indicated by *, and P-value < 0.01 is indicated by **.

[0148] In the construction of the epidermal stratum corneum barrier, involucrin forms an insoluble stratum corneum mantle with loricrin and filaggrin through a transglutaminase-mediated cross-linking reaction, becoming the core functional unit of the physical barrier. This protein complex can effectively protect against mechanical stimulation, chemical corrosion and enzymatic damage, and together with the lipid bilayer, maintain the integrity of the stratum corneum barrier. In this experiment, compared with the BC group, the IVL content of the NC group decreased significantly, indicating that the stimulation conditions of this test were effective. Compared with the NC group, the IVL content of the PC group increased significantly, indicating that the positive control of this test was effective. Compared with the NC group, the IVL content of Example 1 increased significantly, with an increase rate of 116.67%. It can be seen that Example 1 can increase IVL expression and enhance the skin barrier function.

[0149] It is worth noting that in disease states such as atopic dermatitis and psoriasis, the expression level of epidermal involucrin shows a characteristic decrease, which in turn causes structural disorder of the keratin mantle and dysfunction of the barrier function, revealing its key role in the pathological mechanism of skin diseases.

[0150] Test Example 4: Verification of Anti-inflammatory Efficacy

[0151] Test Method

[0152] 1. Cell seeding

[0153] Immortalized keratinocytes HaCat were selected. After cell recovery, when the plating rate reached about 60%, the cells were seeded into 12-well plates and incubated overnight in a CO2 incubator (37°C, 5% CO2).

[0154] 2. Test Grouping

[0155] The group test was performed according to the conditions shown in Table 8, with 3 replicate wells set up in each group.

[0156] Table 8 Experimental test groups for Test Example 4

[0157]

[0158] 3. Administer medication

[0159] Replace the culture medium with the test sample according to the conditions shown in Table 8 and incubate for 24 hours. All experimental conditions were performed with an n = 3. At the end of the incubation period, harvest the cells and immediately freeze them at -80°C.

[0160] 4. q-PCR analysis:

[0161] Total RNA was extracted from cells collected under each experimental condition using TriPure Isolation Reagent, converted to cDNA, and the expression of target genes was detected by RT-qPCR. The primers used for the target genes are shown in Table 9. Each example and the untreated group had three replicate samples.

[0162] Table 9 Target gene primers

[0163]

[0164] 5. Statistical analysis of results

[0165] Graphs were generated using GraphPad Prism, and the results are expressed as mean ± SD. Comparisons between groups were analyzed using a two-tailed t-test. P < 0.05 was considered a significant difference, and P < 0.01 was considered a highly significant difference. See Table 10 for the results, which show the effects of the test samples on the expression of inflammatory factors in keratinocytes.

[0166] Table 10 Effects of test samples on the expression of inflammatory factors in keratinocytes

[0167]

[0168] IL-1α, IL-1β, IL-6, and IL-8, as key inflammatory mediators, play a central role in skin inflammation and barrier dysfunction. IL-1α and IL-1β activate the NF-κB signaling pathway in keratinocytes. This, on the one hand, inhibits the expression of barrier proteins such as filaggrin, involucrin, and loricrin, reducing stratum corneum lipid synthesis and cross-linking, leading to impaired skin barrier integrity. On the other hand, it induces the release of matrix metalloproteinases (MMPs), which degrade extracellular matrix components and weaken the skin's ability to resist external stimuli. IL-6, through the JAK-STAT3 pathway, interferes with the normal differentiation of keratinocytes, reduces lamellar body secretion, disrupts the intercellular lipid bilayer, and synergizes with other cytokines to exacerbate the inflammatory response. IL-8, a potent chemokine, recruits neutrophils to sites of inflammation, releasing reactive oxygen species (ROS) and proteases that directly damage the stratum corneum and amplify the inflammatory cascade, creating a vicious cycle of inflammation and barrier damage. These inflammatory factors work together to not only directly destroy the skin's physical barrier, but also interfere with lipid metabolism, cell differentiation and immune regulation, ultimately leading to skin barrier dysfunction and promoting the continued progression of inflammation.

[0169] As shown in Table 10, Example 1 can reduce the expression of inflammatory factors and has the effect of inhibiting inflammation.

[0170] Experimental Example 5: Promoting PIEZO1 and TNXB gene expression

[0171] Test method:

[0172] 1. Cell seeding

[0173] Human primary fibroblasts were selected. After cell recovery, when the plating rate reached about 60%, the cells were seeded into 12-well plates and incubated overnight in a CO2 incubator (37°C, 5% CO2).

[0174] 2. Test Grouping

[0175] The cells were grouped according to the conditions shown in Table 11, with one replicate well set up in each group.

[0176] Table 11 Experimental test groups for Test Example 5

[0177]

[0178] 3. Administer medication

[0179] The culture medium was replaced with the culture medium containing the sample to be tested and incubated for 24 hours as shown in Table 11. At the end of the incubation period, the cells were harvested and immediately frozen at -80°C.

[0180] 4. RNA-seq analysis:

[0181] Total RNA was extracted from cells collected under each experimental condition using TriPure Isolation Reagent. Total RNA was extracted using the MJzol Animal RNA Isolation Kit (Majorivd) according to the manufacturer's standard protocol. The purified total RNA was subjected to mRNA isolation, fragmentation, first-strand cDNA synthesis, second-strand cDNA synthesis, end-repair, 3' end A addition, adapter ligation, and enrichment to complete mRNA sequencing library construction. Sequencing was performed based on the effective library concentration and data output requirements. The sequencing platform used was the Illumina NovaSeq 6000, using the PE150 (pair-end 150 bp) sequencing mode, which involves sequencing 150 bp on each end. cDNA was sequenced using NGS (Next Generation Sequencing) technology on a high-throughput sequencer such as the Illumina NovaSeq 6000. (This testing was performed at Bohao Biotechnology Co., Ltd.)

[0182] 5. Data processing

[0183] First, Seqtk was applied to filter the initial data, and the splicedmapping algorithm of Hisat2 (version: 2.0.4) was used to perform genome mapping on the preprocessed reads. Stringtie (version: 1.3.0) was applied to count the number of fragments of each gene after Hisat2 alignment, and then the TMM (trimmed mean of M values) method was used for normalization. Finally, the FPKM value of each gene was calculated using a Perl script.

[0184] The calculation formula is as follows:

[0185]

[0186] in:

[0187] Total exon fragments: the number of fragments mapped to the gene exon (fragment: a pair of reads);

[0188] Exon length: the total length of gene exon;

[0189] Mapped reads: the total number of reads mapped to the reference genome;

[0190] We used edgeR to analyze differentially expressed genes between samples. After obtaining the P-value, we performed multiple hypothesis testing correction. The P-value threshold was determined by controlling the FDR (False Discovery Rate). The corrected P-value is the q-value. Furthermore, the FPKM value was used to calculate the fold-change of differential expression.

[0191] The differentially expressed genes were screened for the following conditions:

[0192] 1) Q-value <= 0.05 (when the total number of differentially expressed genes in all comparisons is less than 50, the P-value will be automatically adjusted to <= 0.05);

[0193] 2) |log2FC |>=1;

[0194] The results are shown in Tables 12 and 13. Table 12 shows the effect of the test sample in Experimental Example 5 on the gene expression profile of fibroblasts. Table 13 shows the relative improvement rate of the test sample in Experimental Example 5 on the gene expression of fibroblasts.

[0195] Table 12 Results of the effects of the test samples in Experimental Example 5 on the gene expression profile of fibroblasts

[0196]

[0197] Table 13 Relative enhancement rate of fibroblast gene expression by test samples

[0198]

[0199] BC is a blank control example, and Example 1 is a mixture obtained by dissolving panthenol, niacinamide, folic acid, and cobalamin in an aqueous solution at a ratio of 5:3:0.01:0.001. The results showed that the composition provided in Example 1 significantly upregulated the gene expression of fibroblasts in terms of the two indicators PEIZO1 and TNXB.

[0200] PIEZO1 is a mechanosensitive ion channel that is highly expressed by keratinocytes at the wound edge after skin injury. It senses tissue tension and cellular microenvironmental deformation, activating downstream signaling pathways that directly drive the directional migration and spreading of keratinocytes, promoting wound healing.

[0201] Tenascin-X (TNXB) is a key glycoprotein in the extracellular matrix (ECM), primarily expressed in dermal fibroblasts. It maintains the stability of the dermal-epidermal junction by anchoring collagen fibers (type I / III) and elastin, enhancing the tissue's resistance to mechanical stress. During the middle and late stages of repair, TNXB expression continues to increase, promoting collagen cross-linking and ECM remodeling, inhibiting excessive fibrosis, and reducing scar formation.

[0202] The above are only preferred embodiments of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A barrier repair composition comprising panthenol, niacinamide, folic acid, and cobalamin in a mass ratio of 3-7:1-5:0.0005-0.1:0.00005-0.

01.

2. The barrier repair composition according to claim 1, characterized in that It is composed of panthenol, niacinamide, folic acid and cobalamin in a mass ratio of 4~6:2~4:0.001~0.05:0.0001~0.

005.

3. The barrier repair composition according to claim 2, characterized in that It is composed of panthenol, niacinamide, folic acid and cobalamin in a mass ratio of 4.5~5.5:2.5~3.5:0.005~0.015:0.0005~0.0015.

4. The barrier repair composition according to claim 3, characterized in that It is composed of panthenol, niacinamide, folic acid and cobalamin in a mass ratio of 5:3:0.01:0.

001.

5. Use of the barrier repair composition according to any one of claims 1 to 4 in the preparation of anti-inflammatory products.

6. Use of the barrier repair composition according to any one of claims 1 to 4 in the preparation of repair products.

7. Barrier repair product, characterized in that, include: The barrier repair composition according to any one of claims 1 to 4.

8. The barrier repair product according to claim 7, characterized in that: Including personal care products.

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