Beauty and anti-aging mesenchymal stem cell exosome composition

Abstract

The invention relates to the technical field of exosomes, and particularly discloses a cosmetic anti-aging mesenchymal stem cell exosome composition, which comprises a therapeutically effective amount of mesenchymal stem cell-derived exosome, the functional carrier comprises a transdermal absorption enhancer and an antioxidant stabilizer; at least one synergistic active component selected from the group consisting of small molecule peptides, plant-derived antioxidants, hyaluronic acid or derivatives thereof; the small molecule peptide is one of oligopeptide-1 and palmitoyl tripeptide-5; the plant source antioxidant is one of resveratrol and ferulic acid; wherein the exosome and the synergistic active component are compounded through a phospholipid bilayer fusion technology; through a phospholipid bilayer fusion technology, the mesenchymal stem cell exosome and a synergistic active component are directionally compounded, an exosome membrane structure is used as a natural carrier, the active component is effectively prevented from being degraded, and meanwhile, the bioavailability is improved by utilizing the targeting property of the exosome membrane structure.

Description

Technical Field

[0001] This invention belongs to the field of exosome technology, specifically relating to a cosmetic and anti-aging mesenchymal stem cell exosome composition. Background Technology

[0002] Currently, mesenchymal stem cell exosomes are widely used as active ingredients in the field of beauty and anti-aging. With their low immunogenicity, high biocompatibility and ability to regulate cell regeneration, they have become a new alternative to traditional growth factors. Existing technologies mainly use single exosome preparations and physical mixing and compounding.

[0003] While such technologies can partially leverage the anti-aging effects of exosomes, the particle size and hydrophilicity of exosomes make it difficult for traditional transdermal agents to penetrate the stratum corneum barrier, resulting in many active ingredients remaining in the epidermis and failing to reach the target cells in the dermis. Summary of the Invention

[0004] The purpose of this invention is to provide a cosmetic and anti-aging mesenchymal stem cell exosome composition to solve the problems mentioned in the background art.

[0005] To achieve the above objectives, the present invention provides the following technical solution:

[0006] A cosmetic and anti-aging mesenchymal stem cell exosome composition comprising:

[0007] Therapeutic amounts of mesenchymal stem cell-derived exosomes;

[0008] Functional carriers, including transdermal absorption enhancers and antioxidant stabilizers;

[0009] At least one synergistic active component is selected from small molecule peptides, plant-derived antioxidants, hyaluronic acid or its derivatives;

[0010] The small molecule peptides are one of oligopeptide-1 and palmitoyl tripeptide-5;

[0011] The plant-derived antioxidant is one of resveratrol and ferulic acid;

[0012] Among them, exosomes and synergistic active components are compounded through phospholipid bilayer fusion technology.

[0013] Preferably, the exosomes derived from mesenchymal stem cells are umbilical cord mesenchymal stem cell exosomes with a particle size distribution of 30-150 nm and a positive expression level of surface markers CD9, CD63, and TSG101 ≥90%.

[0014] Preferably, the exosome concentration is 1×10⁻⁶. 8 ~1×10 11 The particles / mL ratio is 1:0.5 to 1:5 in mass ratio with the synergistic active component.

[0015] Preferably, the transdermal absorption enhancer is a phosphatidylcholine / glycolic acid copolymer micelle, accounting for 0.1% to 5% of the total mass of the composition.

[0016] Preferably, the composition is a staged release system, comprising:

[0017] Phase 1: Exosome-phospholipid complex enables rapid epidermal penetration;

[0018] Second phase: Synergistic active components are slowly released in the dermis through pH-responsive hydrogels.

[0019] Preferably, the antioxidant stabilizer comprises a complex system of reduced glutathione and trehalose in a molar ratio of 1:2 to 1:5, used to maintain the integrity and biological activity of exosome membranes.

[0020] Compared with the prior art, the beneficial effects of the present invention are:

[0021] (1) Mesenchymal stem cell exosomes are directionally compounded with synergistic active components through phospholipid bilayer fusion technology. The exosome membrane structure serves as a natural carrier, effectively protecting the active components from degradation, while its targeting enhances bioavailability. Combined with a staged release system, the exosomes and active components are precisely delivered in the epidermis and dermis of the skin, fully activating the exosome-mediated cell regeneration signal and the antioxidant pathway of plant components, resulting in significant synergistic effects in promoting collagen synthesis, inhibiting matrix degradation, and scavenging free radicals.

[0022] (2) The reduced glutathione-trehalose compound stabilization system is adopted. Through the dual action of intermolecular hydrogen bonds and thiol antioxidant mechanism, it effectively maintains the integrity of exosome membrane and the long-term stability of bioactive ingredients, so that the product can maintain its efficacy without harsh cold chain conditions, greatly reducing storage and transportation costs. In addition, the composition takes into account both high transdermal efficiency and sustained-release characteristics, avoiding the skin burden caused by frequent reapplication. Attached Figure Description

[0023] Figure 1 This is a process flow diagram of the present invention. Detailed Implementation

[0024] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0025] Example 1:

[0026] Exosome preparation includes the following steps:

[0027] Human umbilical cord mesenchymal stem cells were selected and cultured to passage P4. The cells were starved in serum-free medium for 48 hours, and the supernatant was collected. The cells were then centrifuged at differential speed (300g×10min→2000g×20min→10,000g×30min) and ultracentrifuged at 110,000g×70min, 4℃ to obtain crude exosomes. Impurities were removed by size exclusion chromatography column (Sepharose CL-4B), and the cells were eluted with PBS to obtain the exosomes.

[0028] Example 2:

[0029] A cosmetic and anti-aging mesenchymal stem cell exosome composition comprising:

[0030] Exosomes 1×10 10 Particles / mL, Palmitoyl Tripeptide-50.2 mg / mL, Ferulic Acid 0.05 mg / mL, Sodium Hyaluronate (10 kDa) 0.5 mg / mL, Phosphatidylcholine / glycolic acid copolymer 2% (w / v), GSH-Trehalose (1:3) 1.5 mM;

[0031] The preparation method includes the following steps:

[0032] Exosomes were mixed with palmitoyl tripeptide-5 and ferulic acid, and phosphatidylcholine ethanol solution (chloroform:methanol = 2:1) was added. The mixture was sonicated (40 kHz, 100 W, 10 min) to induce phospholipid bilayer fusion. The organic solvent was removed by dialysis (molecular weight cutoff 3.5 kDa).

[0033] The fusion compound was combined with sodium hyaluronate and GSH-trehalose, and then a pH-responsive chitosan / β-glycerophosphate hydrogel (gelation temperature 32℃) was added to obtain the composition.

[0034] Experimental Example 1:

[0035] The experimental materials are detailed in the table below:

[0036]

[0037]

[0038] The experimental steps are as follows:

[0039] Thaw the frozen skin at 4°C, rinse three times with PBS, remove subcutaneous fat tissue, cut into 3×3cm pieces, fix the skin between the supply chamber and the recipient chamber (with the stratum corneum facing the supply chamber), fill the recipient chamber with recipient solution preheated to 37°C, stir magnetically at 600 rpm to remove air bubbles, and check the seal.

[0040] 500 μL of sample (groups A / B / C, n=6) was added to the supply chamber at sampling times of 0.5, 1, 2, 4, 8, 12, 24, 36, and 48 h. 0.5 mL of receptor fluid was taken each time, and fresh receptor fluid of the same volume was added at the same time. The samples were immediately frozen at -80 °C for later testing.

[0041] Disassemble the diffusion cell, rinse the skin surface with PBS, separate the epidermis and dermis: freeze at -20℃ for 30 min and then peel off, homogenize the tissue (RIPA lysis buffer), centrifuge and take the supernatant to detect fluorescence intensity;

[0042] The cumulative transdermal permeability of exosomes (%, n=6) is shown in the table below:

[0043] Time (h) Group A Group B Group C 4 18.3±1.2 5.1±0.8 1.9±0.3 8 36.7±2.1 10.5±1.1 3.8±0.6 24 68.2±3.5 21.5±2.3 8.7±1.1 48 72.4±3.8 23.1±2.5 9.2±1.3 Time (h) Group A Group B Group C

[0044] The distribution of fluorescence intensity in different skin layers (24-hour fluorescence intensity percentage, n=3) is shown in the table below:

[0045] Group stratum corneum Epidermis dermis Group A 12.3％ 19.5％ 68.2％ Group B 47.6％ 31.9％ 20.5％ Group C 82.1％ 15.3％ 2.6％

[0046] As shown above, in Group A, the staged release system delivered >65% of exosomes to the dermis, and the concentration of exosomes in the dermis of Group A remained >60% for 24-48 hours.

[0047] Experimental Example 2:

[0048] Human dermal fibroblasts were cultured in DMEM medium containing 10% fetal bovine serum (37℃, 5% CO2) at an electrophoresis rate of 30 mJ / cm². 2 Cells were irradiated with a dose of sodium, and after 24 hours of recovery culture, they were divided into groups (6 replicates per group):

[0049] The blank control group was prepared with an equal volume of PBS buffer added.

[0050] The positive control group consisted of a 0.1% retinol solution;

[0051] Group A consists of the composition prepared in Example 2 (0.1 mg / mL);

[0052] Group B consists of unfused physical mixtures (equal components, 0.1 mg / mL);

[0053] After 72 hours of treatment, the collagen content in the cell supernatant was quantified using an ELISA kit (Human COL1A1 Kit, Abcam ab210966).

[0054] After 48 hours of treatment, total RNA was extracted from cells for qPCR analysis, and the inhibition rate was calculated based on the expression level of the UVB model group.

[0055] The DPPH radical scavenging assay was used to measure the absorbance at 515 nm and calculate the scavenging percentage.

[0056] The quantitative data on anti-aging efficacy (n=6, *p<0.01 vs. Group B) are shown in the table below:

[0057] detection indicators Blank control Positive control Group A Group B Type I collagen synthesis 100％ 130.5％ 148.3±5.2％* 118.2±4.1％ MMP-1 inhibition rate - 42.1％ 57.6±3.8％* 24.7±2.9％ DPPH removal rate 0％ 78.3％ 92.1±2.4％* 53.8±3.7％

[0058] Collagen synthesis kinetics (72h, ELISA pg / mL) are shown in the table below:

[0059] Time (h) Group A Group B Improvement rate 24 38.2±2.1 25.3±1.8 +51.0％ 48 112.5±6.3 74.6±4.2 +50.8％ 72 185.7±8.9 119.8±5.6 +55.0％

[0060] As shown above, group A significantly promoted type I collagen synthesis, reaching 148.3±5.2% of the blank control, which was 165% higher than group B and higher than the positive control group (130.5%). Kinetic data showed that at 72 hours, the collagen content in group A reached 185.7±8.9 pg / mL, which was 55% higher than group B.

[0061] The inhibition rate of matrix metalloproteinase MMP-1 gene expression in group A was 57.6±3.8%, which was significantly better than that in the positive control group and group B.

[0062] The DPPH free radical scavenging rate of group A reached 92.1±2.4%, which was 71% higher than that of group B.

[0063] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A cosmetic and anti-aging mesenchymal stem cell exosome composition, characterized in that, include: Therapeutic amounts of mesenchymal stem cell-derived exosomes; Functional carriers, including transdermal absorption enhancers and antioxidant stabilizers; At least one synergistic active component is selected from small molecule peptides, plant-derived antioxidants, hyaluronic acid or its derivatives; The small molecule peptides are one of oligopeptide-1 and palmitoyl tripeptide-5; The plant-derived antioxidant is one of resveratrol and ferulic acid; Among them, exosomes and synergistic active components are compounded through phospholipid bilayer fusion technology.

2. The cosmetic anti-aging mesenchymal stem cell exosome composition according to claim 1, characterized in that: The exosomes derived from mesenchymal stem cells are umbilical cord mesenchymal stem cell exosomes with a particle size distribution of 30-150 nm and positive expression levels of surface markers CD9, CD63, and TSG101 ≥90%.

3. The cosmetic anti-aging mesenchymal stem cell exosome composition according to claim 1, characterized in that: The exosome concentration was 1×10⁻⁶. 8 ~1×10 11 The particles / mL ratio is 1:0.5 to 1:5 in mass ratio with the synergistic active component.

4. The cosmetic anti-aging mesenchymal stem cell exosome composition according to claim 1, characterized in that: The transdermal absorption enhancer is a phosphatidylcholine / glycolic acid copolymer micelle, accounting for 0.1% to 5% of the total mass of the composition.

5. The cosmetic anti-aging mesenchymal stem cell exosome composition according to claim 1, characterized in that: The composition is a staged release system, comprising: Phase 1: Exosome-phospholipid complex enables rapid epidermal penetration; Second phase: Synergistic active components are slowly released in the dermis through pH-responsive hydrogels.

6. The cosmetic anti-aging mesenchymal stem cell exosome composition according to claim 1, characterized in that: The antioxidant stabilizer comprises a complex system of reduced glutathione and trehalose in a molar ratio of 1:2 to 1:5.

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