Collagen regeneration preparation

Abstract

The invention belongs to the technical field of pharmaceutical preparations, and discloses a collagen regeneration preparation. Comprising SRT1720 and auxiliary materials. The product can promote skin fibroblasts to synthesize more endogenous I-type and III-type collagen, increase the skin compactness and quickly improve the facial appearance.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical preparations, and in particular relates to a collagen regeneration preparation. Background Art

[0002] Currently, collagen injections are widely used in the medical beauty industry as an effective treatment for facial filling and rapid improvement of skin condition. [1] However, the problems faced by direct collagen injection are: the injected collagen is lost quickly in the body, regular injections are required, it can only provide temporary effects, and long-term use may face biocompatibility issues, thus causing allergic reactions and other safety issues. [2] Therefore, more and more research is beginning to focus on providing more lasting effects by activating endogenous collagen synthesis in the skin. [3,4] .

[0003] SRT1720 is a known SIRT1 activator that can regulate various physiological functions of cells by activating the SIRT1 pathway. SIRT1 is a NAD+ deacetylase that can regulate various physiological processes of cells, including DNA repair, energy metabolism, and the aging process. [5] . Summary of the invention

[0004] In order to overcome the deficiencies of the prior art, the present invention provides a collagen regeneration preparation, which can promote skin fibroblasts to synthesize more endogenous type I and type III collagen, increase skin firmness, and quickly improve facial appearance.

[0005] The above object of the present invention is achieved through the following technical solution: A collagen regeneration preparation, comprising SRT1720 and auxiliary materials, wherein the chemical structure of SRT1720 is:

[0006]

[0007] Furthermore, the carrier includes one or more of a filler, a diluent, a binder, a disintegrant, an emulsifier, and a drug-carrying carrier with no toxic or side effects.

[0008] Furthermore, the collagen regeneration preparation is selected from a microneedle preparation and a subcutaneous injection preparation.

[0009] Furthermore, the collagen regeneration preparation is prepared in the form of a single-dose drug.

[0010] Another object of the present invention is to protect the use of the above-mentioned collagen regeneration preparation, specifically to increase the expression of type I collagen and type III collagen through SRT1720, promote collagen synthesis, and inhibit collagen degradation.

[0011] The beneficial effects of the present invention compared with the prior art are as follows: the present invention aims to propose a novel collagen regeneration preparation based on endogenous mechanism by conducting an in-depth study on the role of SRT1720 in skin fibroblasts, aiming to provide a new method for skin care to enhance the natural collagen synthesis of the skin and improve the skin quality, and avoid the safety hazards and adverse reactions caused by exogenous collagen supplementation. The product can promote skin fibroblasts to synthesize more endogenous type I and type III collagen, increase the firmness of the skin, and quickly improve the facial appearance. SRT1720 increases the synthesis of collagen by regulating the TGF-β1 / Smad2 / 3 pathway, and regulates the synthesis and degradation of collagen by regulating the MMPs / TIMPs system. At the same time, SRT1720 can also promote the migration of skin fibroblasts and accelerate the reconstruction and repair of the extracellular matrix. The technical solution of the present invention is aimed at the needs of modern people to maintain a beautiful and youthful appearance, and provides an effective method for promoting collagen synthesis by endogenous means, avoiding the limitations of exogenous collagen injection, and can provide lasting firmness and elasticity improvement for the skin, and improve the skin's own collagen synthesis ability. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] The present invention will be further described below in conjunction with the accompanying drawings and specific embodiments.

[0013] Figure 1 Western blot was used to detect the protein expression levels of type I collagen (A) and type III collagen (B) in different experimental groups;

[0014] Figure 2 Western blot was used to detect the protein expression levels of TGF-β1 (A) and Smad2 / 3 (B) in different experimental groups;

[0015] Figure 3 Western blot was used to detect the protein expression levels of MMP-1 (A) and TIMP-1 (B) in different experimental groups;

[0016] Figure 4 The scratch test was used to detect the effect of SRT1720 on the migration ability of dermal fibroblasts. The scratch area of ​​the H2O2 treatment group and the H2O2+SRT1720 treatment group was photographed at 0, 6 and 12 hours (A), and the scratch area after cell migration was quantitatively analyzed (B). DETAILED DESCRIPTION

[0017] The present invention is described in detail below by specific examples, but the protection scope of the present invention is not limited. Unless otherwise specified, the experimental methods used in the present invention are all conventional methods, and the experimental equipment, materials, reagents, etc. used can be obtained from commercial channels.

[0018] Example 1

[0019] Human dermal fibroblasts were used and the following experiments were performed:

[0020] SRT1720 was dissolved in DMSO at the required concentration, filtered using a 0.22 μm sterile filter membrane, and the prepared stock solution was dispensed into small tubes and stored at -20°C (short-term storage).

[0021] 1. After plating, the cells were divided into two groups: H2O2 group and H2O2+SRT1720 group. H2O2 induced oxidative stress in the cells to simulate the oxidative damage caused by skin exposure to external environmental factors.

[0022] The H2O2 group was first treated with DMSO solvent for 2 hours, and then treated with 200μmol / L H2O2 for 2 hours; the H2O2+SRT1720 group was pretreated with 4μmol / L SRT1720 for 2 hours, and then treated with 200μmol / L H2O2 for 2 hours.

[0023] 2. Western blot detection of protein expression (1) Collect cells from the H2O2 group and the H2O2+SRT1720 group. (2) Add cell lysis buffer to extract cells, where RIPA:PMSF=10:1. (3) Protein quantification: Use the BCA protein quantification method to determine the concentration of the extracted protein. (4) SDS-PAGE electrophoresis: Use polyacrylamide gel electrophoresis (SDS-PAGE) to separate protein samples according to molecular weight. (5) Transfer: Assemble the gel, membrane and transfer buffer in a transfer device, and use a wet transfer device to transfer the separated proteins from the gel to the PVDF membrane. (6) Blocking: Place the membrane in a shaker containing 5% skimmed milk powder and incubate for 2 hours. (7) Primary antibody incubation: The primary antibody of the target protein is fully bound to the target protein in the membrane and incubated overnight. (8) Washing: Wash the membrane with TBST buffer 3 times, 5 minutes each time to remove unbound primary antibody. (9) Secondary antibody incubation: Incubate the secondary antibody corresponding to the source of the primary antibody (such as goat anti-mouse IgG) for 2 hours to allow it to bind to the primary antibody. (10) Washing: Wash the membrane 3 times with TBST buffer for 5 minutes each time to remove unbound secondary antibody. (11) Signal detection: Use the substrate chemiluminescence ECL method to detect the protein labeled with the secondary antibody, and use the imaging system to image and save the target protein. (12) Result analysis: Use ImageJ to quantitatively analyze the target protein, and use GraphPad Prism8.4 analysis software for statistical analysis.

[0024] In the above experimental groups, the protein expression levels of type I collagen and type III collagen in dermal fibroblasts were detected. Type I and type III collagen are the two most important types of collagen in the skin. Figure 1The results showed that compared with the H2O2 group, the expression levels of type I collagen and type III collagen in the H2O2+SRT1720 group were increased ( Figure 1 A, B), SRT1720 has collagen repair effect on H2O2-induced cell damage.

[0025] In the above experimental groups, the protein expression levels of TGF-β1 and Smad2 / 3 in dermal fibroblasts were detected. The TGF-β1 / Smad2 / 3 signaling pathway is a classic signaling pathway for collagen synthesis. Figure 2 The results showed that compared with the H2O2 group, the protein expression levels of TGF-β1 and Smad2 / 3 in the H2O2+SRT1720 group were significantly increased ( Figure 2 A, B), SRT1720 has a great promoting effect on collagen synthesis by dermal fibroblasts.

[0026] In the above experimental groups, the protein expression levels of MMP-1 and TIMP-1 in dermal fibroblasts were detected. MMP-1 is mainly responsible for the degradation of type I and type III collagen. TIMP-1 (tissue inhibitor of metalloproteinase 1) is a natural inhibitor of matrix metalloproteinases (MMPs). It binds to MMP-1, inhibits its activity, and thus reduces the degradation of collagen. Figure 3 The results showed that compared with the H2O2 group, the protein expression level of MMP-1 in the H2O2+SRT1720 group was significantly decreased ( Figure 3 A), while the protein expression level of TIMP-1 increased ( Figure 3 B). This suggests that SRT1720 can inhibit the degradation of collagen.

[0027] 3. Cell scratch test (1) Culture cells in a 6-well culture plate until the cells form a monolayer (usually reaching 70-90% confluence). (2) Use a sterile 10ul pipette tip to make uniform straight scratches on the cell monolayer. (3) Gently rinse with culture medium to remove cell debris and dead cells produced by the scratch. (4) Use a microscope to take pictures and record the initial state of the scratch (i.e., the width of the cell scratch). (5) Add drug-treated culture medium to each well. (6) Place the 6-well plate back into the CO2 incubator and continue to culture the cells. (7) After a predetermined time interval (6h, 12h), observe cell migration under a microscope and take pictures to record the scratch area observed each time. (8) Compare images at different time points to evaluate the speed and ability of cell migration.

[0028] The cell scratch test simulates the process of cell migration in vivo to a certain extent. Figure 4) to detect the effect of SRT1720 on the migration ability of dermal fibroblasts. After scratching the cell layer, the cells in the H2O2 group and the H2O2+SRT1720 group were cultured under the same conditions, and the scratch healing was recorded at 0, 6 and 12 hours. Compared with the H2O2 group, the migration ability of the cells in the H2O2+SRT172 group was significantly improved at 6 hours and 12 hours ( Figure 4 A, B). SRT1720 can partially restore the impaired migration ability caused by H2O2 and enhance the migration and repair ability of HSFs.

[0029] The above-described embodiments are only preferred embodiments of the present invention, but not all feasible embodiments of the present invention. For those skilled in the art, any obvious changes made thereto without departing from the principles and spirit of the present invention should be considered to be included in the scope of protection of the claims of the present invention.

[0030] References:

[0031] 1.He,N.,et al.,Preparation of injectable porcine skin-derivedcollagen and its application in delaying skin aging by promoting the adhesionand chemotaxis ofskin fibroblasts.Int J Biol Macromol,2023.253(0).

[0032] 2.Patricia,L.and G.David J,Complications of collagen fillers.FacialPlastSurg,2014.30(6).

[0033] 3.Ying,Z.,et al.,Lactate triggers KAT8-mediated LTBP1 lactylation atlysine752 to promote skin rejuvenation by inducing collagen synthesis infibroblasts.Int JBiol Macromol,2024.277(0).

[0034] 4.Yilei,C.,et al.,Rejuvenation of Aged Human Skin by InjectionofCross-linked Hyaluronic Acid.Plast Reconstr Surg,2020.147(0).

[0035] 5.Cui,C.,et al.,SIRT1 and aging related signaling pathways.MechAgeingDev,2020.187(0).

Claims

1. A collagen regeneration preparation, characterized in that: The ingredients include SRT1720 and auxiliary materials, wherein the chemical structure of SRT1720 is:

2. The collagen regeneration preparation according to claim 1, characterized in that The carrier includes one or more of a filler, a diluent, a binder, a disintegrant, an emulsifier, and a drug-carrying carrier without toxic or side effects.

3. The collagen regeneration preparation according to claim 1, characterized in that The collagen regeneration preparation is selected from a microneedle preparation and a subcutaneous injection preparation.

4. The collagen regeneration preparation according to claim 1, characterized in that The collagen regeneration preparation is prepared in the form of a single-dose medicine.

5. The use of the collagen regeneration preparation as claimed in claim 1, specifically increasing the expression of type I collagen and type III collagen through SRT1720, promoting collagen synthesis, and inhibiting collagen degradation.

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