Application of sialic acid and hyaluronic acid in preparation of medicine for promoting stem cell migration

Abstract

The invention relates to the technical field of medicine, in particular to application of sialic acid and hyaluronic acid in preparation of a medicine for promoting stem cell migration, the administration concentration of the sialic acid is 2.5 mM, and the administration concentration of the hyaluronic acid is 100-500 [mu] g / mL; the cell density of the SA group is increased, the migration ability is remarkably improved, compared with the SA group, the concentration of sialic acid in the SA + HA group is consistent, and along with the increase of the concentration of hyaluronic acid, the cell proliferation speed is increased, and the cell migration ability is remarkably improved. The sialic acid and the hyaluronic acid are mixed, so that the migration capability of the amniotic fluid mesenchymal stem cells can be improved, and the proliferation of the amniotic fluid mesenchymal stem cells can be better promoted.

Description

Technical Field

[0001] The present invention relates to the field of medical technology, and in particular to the use of bird's nest acid and hyaluronic acid in the preparation of drugs that promote stem cell migration. Background Art

[0002] Sialic acid (SA), also known as sialic acid, is a unique α-keto acid. Currently, more than 50 forms of sialic acid have been found in nature, the most important of which is N-acetylneuraminic acid. SA is naturally present in many animal foods, such as bird's nests, breast milk and eggs. Sialic acid has the effects of promoting brain development, anti-virus, anti-Alzheimer's disease and anti-hypertension. In addition, SA also has anti-aging and antioxidant effects.

[0003] Hyaluronic acid (HA), also known as hyaluronic acid, is a naturally occurring acidic mucopolysaccharide with excellent moisturizing properties. It is widely used in the pharmaceutical, cosmetic, and food industries. Studies have shown that HA, due to its biocompatibility, biodegradability, and ease of chemical modification, can promote wound healing by promoting hemostasis, regulating inflammation, and promoting epithelial regeneration. Studies have investigated the efficacy of HA combined with glutamic acid (Glu) in skincare formulations. Results show that the HA-Glu complex effectively alleviates inflammation-induced skin symptoms and skin aging when used in skincare formulations. Stem cells are a core tool in regenerative medicine. Amniotic fluid-derived mesenchymal stem cells (AFMSCs) have multipotential differentiation potential, capable of differentiating into fat, bone, and cartilage cells. They are also low immunogenic and have a low risk of allograft rejection. AFMMSCs are obtained from prenatal amniotic fluid and are ethically uncontroversial. Stem cell therapy has attracted significant attention in recent years, but stem cell treatments still face challenges such as low homing efficiency and poor microenvironmental adaptability. Improving the directional migration ability of stem cells, enhancing their adaptability to the environment, and prolonging their duration of action are key solutions. Summary of the Invention

[0004] In order to solve the above technical problems, an embodiment of the present invention provides the use of bird's nest acid and hyaluronic acid in the preparation of drugs that promote stem cell migration.

[0005] To achieve the above objectives, the embodiments of the present invention adopt the following technical solutions:

[0006] The present invention provides the use of bird's nest acid and hyaluronic acid in the preparation of a drug for promoting stem cell migration.

[0007] In some embodiments, the administration concentration of the sialic acid is 2.5 mM, and the administration concentration of the hyaluronic acid is 100-500 μg / mL.

[0008] In some embodiments, the stem cells are amniotic fluid mesenchymal stem cells.

[0009] In some embodiments, the above-mentioned drug includes a pharmaceutical preparation prepared with an effective amount of bird's nest acid and hyaluronic acid as active ingredients, and pharmaceutically acceptable carriers and / or excipients.

[0010] An effective amount refers to the amount of a drug, compound, or pharmaceutical composition necessary to obtain any one or more beneficial or desired therapeutic results. Beneficial or desired results include: clinical improvement (such as reducing morbidity, reducing mortality, improving one or more symptoms), reducing severity, delaying the onset of symptoms (including symptoms or their complications, intermediate pathological phenotypes presented during the development of the disease, biochemistry, histology, and / or behavioral symptoms). The preparation can be an injection, spray, aerosol, nasal drops, oral preparation, or a dosage form suitable for mucosal administration. In addition, the route of administration to the subject includes, but is not limited to, intramuscular, intravenous, subcutaneous, intradermal, oral, intranasal, respiratory, transmucosal, sublingual, and parenteral.

[0011] The present invention has the following beneficial effects compared to the prior art:

[0012] The combination of bird's nest acid and hyaluronic acid promotes the proliferation of amniotic fluid mesenchymal stem cells by enhancing their ability to migrate. When the concentration of bird's nest acid is consistent, as the concentration of hyaluronic acid increases, the cell proliferation rate becomes faster and the cell migration ability is significantly increased. BRIEF DESCRIPTION OF THE DRAWINGS

[0013] Figure 1 This is a bright field microscope image of amniotic fluid mesenchymal stem cells (AFMSCs);

[0014] Figure 2 This is a physical picture of Sialic acid powder (SA);

[0015] Figure 3 This is a physical picture of hyaluronic acid (HA);

[0016] Figure 4 The cell state diagram at 0h;

[0017] Figure 5 This is the cell status diagram at 24 hours;

[0018] Figure 6 This is the cell status diagram at 48h;

[0019] Figure 7 This is a statistical analysis chart of the cell scratch experiment. DETAILED DESCRIPTION

[0020] The following will be combined with the accompanying drawings to clearly and completely describe the technical solutions in some embodiments of the present disclosure. Obviously, the embodiments described are only some embodiments of the present disclosure, not all embodiments. Based on the embodiments provided by the present disclosure, all other embodiments obtained by ordinary technicians in this field are within the scope of protection of the present disclosure.

[0021] Unless otherwise specified, all raw materials, reagents, instruments and equipment used in this application can be purchased from the market or prepared by existing methods.

[0022] Experimental Materials:

[0023] (1) Cells

[0024] Amniotic fluid mesenchymal stem cells (AFMSCs) were derived from clinical amniotic fluid samples from the Third Affiliated Hospital of Guangzhou Medical University.

[0025] (2) Materials

[0026] Swallow acid (SA): purchased from MCE, CAS No.: 131-48-6.

[0027] Hyaluronic acid (HA): purchased from MCE, CAS No.: 9004-61-9.

[0028] DMEM culture medium: 1% fetal bovine serum + 1% P / S double antibody (final concentration of 50 U / mL) + 98% low-glucose DMEM basal culture medium.

[0029] Example

[0030] First, dissolve 100mg of bird's nest acid powder in 10mL of pure water to prepare a 32mM bird's nest acid solution. Filter through a 0.22μm disposable microporous membrane filter to remove impurities. Dilute the 100mM bird's nest acid solution to a 2.5mM solution using DMEM medium containing 1% FBS and 1% P / S / B. Dissolve 100mg of hyaluronic acid powder in 20mL of pure water until a transparent gel forms to prepare a 4.16μM hyaluronic acid solution. Dilute the 4.16μM hyaluronic acid solution using DMEM medium to obtain solutions with concentrations of 100μg / mL, 200μg / mL, 300μg / mL, 400μg / mL, and 500μg / mL, respectively.

[0031] Place 10 5Amniotic fluid-derived mesenchymal stem cells were evenly plated in a six-well plate. When the cell density reached 90%, a 200 μL pipette tip was used to scratch the wells of the six-well plate, maintaining a consistent width of 1 cm in each well. 2 mL of DMEM culture medium, 2 mL of 2.5 mM sialic acid solution, and 2 mL of hyaluronic acid solution at concentrations of 100 μg / mL, 200 μg / mL, 300 μg / mL, 400 μg / mL, and 500 μg / mL were prepared in advance. The solutions were added to the cells and incubated at 37°C in 5% CO2. The experiment was divided into seven groups: the control group did not add bird's nest acid and hyaluronic acid, that is, the concentrations of bird's nest acid and hyaluronic acid were 0mM and 0μg / mL; the SA group only added bird's nest acid, and its concentration was 2.5mM; the SA+HA group, that is, the concentration of bird's nest acid was 2.5mM, and the concentrations of hyaluronic acid were 100μg / mL, 200μg / mL, 300μg / mL, 400μg / mL, and 500μg / mL respectively.

[0032] like Figure 4 The cells were in the state of 0 h. At this time, a 1 cm wide scratch was made with a 200 μL pipette tip, and 2.5 mM bird's nest acid and different concentrations of hyaluronic acid were added. The cells were cultured at a temperature of 37 ° C and 5% CO2.

[0033] like Figure 5 The cells were cultured in a medium containing sialic acid and hyaluronic acid for 24 hours. The cell density changed, and the scratch width narrowed accordingly. Compared with the control group (sialic acid and hyaluronic acid concentrations of 0 mM and 0 μg / mL), the cell density and migration ability of the SA group increased. Compared with the SA group, the cell density and migration ability of the SA+HA group increased significantly with the increase of hyaluronic acid concentration.

[0034] like Figure 6 The state of the cells at 48 hours. At this time, the cells have been cultured in a medium containing sialic acid and hyaluronic acid for 48 hours. The cell density has changed more significantly, and the width of the scratch has become narrower. Compared with the control group (sialic acid and hyaluronic acid concentrations of 0mM and 0μg / mL), the cell density of the SA group increased, and the migrated cells almost filled the gap in the scratch. Compared with the SA group, the cell density of the SA+HA group almost grew to 100%, and the migrated cells filled the gap in the scratch.

[0035] like Figure 7 Statistical analysis was performed on the changes in the width of the scratches. The results showed that compared with the control group (bird's nest acid and hyaluronic acid concentrations of 0 mM and 0 μg / mL), the cell density of the SA group increased and the migration ability was significantly increased. Compared with the SA group, the SA+HA group had the same bird's nest acid concentration. With the increase of hyaluronic acid concentration, the cell proliferation rate became faster and the cell migration ability was significantly increased.

[0036] Through the cell scratch test, the effect of the mixture of bird's nest acid (SA) and hyaluronic acid (HA) on the proliferation and migration of amniotic fluid mesenchymal stem cells (AFMSCs) was explored. The experiment was conducted with bird's nest acid at a concentration of 2.5mM and hyaluronic acid at concentrations of 100μg / mL, 200μg / mL, 300μg / mL, 400μg / mL, and 500μg / mL. The results showed that the SA concentration was consistent. As the HA concentration increased, the migration of amniotic fluid mesenchymal stem cells was promoted. At 48h, the HA concentrations of 300μg / mL, 400μg / mL, and 500μg / mL made the migration rate of AFMSCs 100%. In summary, it can be concluded that the mixture of bird's nest acid and hyaluronic acid can enhance the migration ability of amniotic fluid mesenchymal stem cells and better promote the proliferation of amniotic fluid mesenchymal stem cells.

[0037] In the description of this specification, specific features, structures, materials or characteristics may be combined in an appropriate manner in any one or more embodiments or examples.

[0038] The above description is merely a specific embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any modifications or substitutions that can be easily conceived by a person skilled in the art within the technical scope disclosed in the present invention should be included in the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be based on the scope of protection of the claims.

Claims

1. Application of bird's nest acid and hyaluronic acid in the preparation of drugs that promote stem cell migration.

2. The use according to claim 1, characterized in that The administration concentration of the sialic acid is 2.5 mM, and the administration concentration of the hyaluronic acid is 100-500 μg / mL.

3. The use according to claim 1, characterized in that The stem cells are amniotic fluid mesenchymal stem cells.

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