Chitosan composite fiber material for tendon repair and preparation method and application thereof

Abstract

The present application relates to a kind of chitosan composite fiber material for tendon repair and its preparation method and application, belong to the technical field of biomedical materials.The chitosan composite fiber material is the acetylated chitosan / polyglyceryl sebacate composite fiber material with poly(2-methyl allyl oxyethyl phosphocholine) on the surface.The present application can effectively adjust the degradation rate of chitosan by the addition of polyglyceryl sebacate, obtain the elasticity matched with tendon activity, by the adjustment of the acetylation degree of chitosan composite fiber material, can promote the proliferation and regeneration of tendon stem cells, the surface coated with poly(2-methyl allyl oxyethyl phosphocholine) can prevent postoperative adhesion, has good effect in tendon repair.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biomedical materials, and particularly relates to a chitosan composite fiber material for tendon repair and a preparation method and application thereof. BACKGROUND

[0002] Tendon injury has become one of the common diseases in life, which can be caused by trauma, intense exercise or long-term labor, but the disease cannot be self-healed and can only be repaired by surgical or conservative treatment. Tendon healing can be divided into two types, one is endogenous healing, and the other is exogenous healing. However, the poor blood supply and limited regenerative capacity of the tendon lead to poor intrinsic healing ability. At present, the ligament / tendon injury usually needs surgical operation using autologous or allogeneic grafts.

[0003] Tendon adhesion has been considered as a component of tendon healing, but it is generally believed that the formation of adhesion is related to the superiority of extrinsic tendon healing. More specifically, the intrinsic healing through the proliferation of epitenon cells and endotenon cells contributes to better biomechanics and less dysfunction, while the external healing promotes the formation of adhesion through the invasion of surrounding sheath and synovial cells, thereby destroying the tendon sliding. In addition, in the early inflammatory response after tendon injury, the release of cytokines and the recruitment of fibroblasts greatly promote adhesion. At present, 7-15% of patients after tendon repair have complications such as scar formation, increased tendon adhesion, limited joint movement and the like in the early healing stage. Therefore, secondary surgery is needed for repair, resulting in dysfunction, ultimately leading to reduced work capacity and high cost.

[0004] Chitosan material is widely used in the field of tissue engineering, but the degradation speed of chitosan material is slow and does not match the repair speed of tendon, the elasticity is low and cannot match the elasticity required by tendon activity, and at the same time, the anti-adhesion effect of tendon material is poor, which limits the application of chitosan material in tendon repair. SUMMARY

[0005] (I) Technical problems to be solved

[0006] In view of the above-mentioned shortcomings and deficiencies of the prior art, the present application provides a chitosan composite fiber material for tendon repair and a preparation method and application thereof, which solves the technical problems of insufficient repair effect of tendon repair material on tendon and poor anti-adhesion effect.

[0007] (II) Technical scheme

[0008] In order to achieve the above-mentioned purposes, the main technical scheme adopted by the present application comprises:

[0009] In a first aspect, the present application provides a chitosan composite fiber material for tendon repair, wherein the chitosan composite fiber material is an acetylated chitosan / polyglycerol sebacate composite fiber material coated with poly(2-methyallyloxyethylphosphocholine) on the surface.

[0010] Optionally, the acetylation degree of the acetylated chitosan is 60-100%.

[0011] Preferably, the acetylation degree of the acetylated chitosan is 75-85%.

[0012] Optionally, the chitosan composite fiber material is in a sheet structure with micron-sized mesh.

[0013] Optionally, the coating thickness of the poly(2-methyallyloxyethylphosphocholine) is 0.1-0.2 mm.

[0014] In a second aspect, the present application provides a preparation method of a chitosan composite fiber material for tendon repair, comprising the following steps:

[0015] S1, dissolving polyglycerol sebacate and chitosan in a spinning solvent together, preparing a formed fiber material by electrospinning, and drying to remove the solvent;

[0016] S2, immersing the formed fiber material prepared in step S1 in a mixed solution of acetic anhydride and methanol with a volume ratio of 1-5:5-1, acetylating at 0-50℃ for 0.5-24h, then soaking in anhydrous ethanol for 6-48h, cleaning, drying, and obtaining an acetylated formed fiber material;

[0017] S3, spraying poly(2-methyallyloxyethylphosphocholine) on the surface of the acetylated formed fiber material, and obtaining the chitosan composite fiber material.

[0018] Optionally, in step S1, the spinning solvent is one or more of dichloromethane, trichloromethane, acetic acid, dimethyl sulfoxide, and acetone.

[0019] Preferably, in step S1, the spinning solvent is acetic acid with a mass concentration of 90%.

[0020] Optionally, in step S1, the weight ratio of polyglycerol sebacate to chitosan is 2-4:1.

[0021] Optionally, in step S1, the voltage of the electrospinning is 12.5-20kV.

[0022] In a third aspect, the present application provides the use of the chitosan composite fiber material or the chitosan composite fiber material prepared by the preparation method in tendon repair and anti-adhesion during the tendon repair process.

[0023] (Three) beneficial effects

[0024] The beneficial effects of the present application are: the present application is a kind of chitosan composite fiber material for tendon repair and its preparation method and application.The addition of polyglycerol sebacate can effectively regulate the degradation rate of chitosan, and the adjustment of acetylation degree can enhance the adsorption capacity of the material to tendon stem cells, and the surface coated with poly (2-methyl allyl oxyethyl phosphocholine) can prevent postoperative adhesion, and has good effect in tendon repair.

[0025] The present application combines PGS with CS, prepares a shaped fiber material with micron-sized mesh by electrospinning technology, and then acetylates the shaped fiber material.PGS can significantly improve the insufficient repair of CS as a tendon repair material, and the compounding of PGS can significantly improve the strength (UTS), stiffness and elasticity of the composite fiber material by slowing down the degradation rate of CS.And when the weight ratio of PGS to CS is 2-4:1, there is a significant effect.In the acetylation process, the acetylation degree is 60-100%, especially when the acetylation degree is 75%-85%, the ability of tendon stem cells to adhere to the fiber material is significantly improved.

[0026] The present application coats PMPC on the surface of the acetylated shaped fiber material, further improving the anti-adhesion performance of the composite fiber material.The spraying thickness of PMPC is 0.1-0.2mm, so that the composite fiber material retains part of the micron-sized mesh, has a large specific surface area, and further increases the adhesion capacity of tendon stem cells.

[0027] The PGS / CS / PMPC composite fiber material of the present application is used for tendon repair and prevention of adhesion during tendon repair process, and has significant effect and broad application prospect. BRIEF DESCRIPTION OF DRAWINGS

[0028] Figure 1 The mechanical test results of PGS / CS / PMPC composite fiber material; wherein, A is the linear region graph of the stress-strain curve of PGS / CS / PMPC composite fiber material in the first 15% of the strain region; B is the stiffness performance graph of PGS / CS / PMPC composite fiber material; C is the strength performance graph of PGS / CS / PMPC composite fiber material; D is the elasticity performance graph of PGS / CS / PMPC composite fiber material;

[0029] Figure 2 Cell adhesion PGS / CS / PMPC composite fiber material under different acetylation degrees;

[0030] Figure 3 PGS / CS / PMPC composite fiber material cell proliferation;

[0031] Figure 4 Results of fibroblast adhesion experiments. DETAILED DESCRIPTION

[0032] For a better understanding of the present application, and to enable an embodiment thereof to be described in detail, preferred embodiments will be described hereinbelow and explained with reference to the accompanying drawings, of which:

[0033] For a better understanding of the above technical solutions, the exemplary embodiments of the present application will be described in more detail with reference to the accompanying drawings. Although the exemplary embodiments of the present application are shown in the drawings, it should be understood that the present application can be implemented in various forms and should not be limited by the embodiments set forth herein. Rather, these embodiments are provided so that the present application can be more completely understood, and the scope of the present application can be accurately conveyed to those skilled in the art.

[0034] Example 1

[0035] A method for preparing a PGS / CS / PMPC composite fiber material with an anti-adhesion coating, comprising the following steps:

[0036] (1) Preparation of a polyglycerol sebacate (PGS) / chitosan (CS) formed fiber material:

[0037] PGS and CS were blended at a weight ratio of 4:1 and dissolved in a spinning solvent of 90 wt% concentrated acetic acid. A formed fiber was prepared by electrospinning technology. The voltage for electrospinning was 20 kV. One end of the electrospun thread was fixed on a rotating metal shaft, and after a period of time, a cylindrical fiber layer with a certain thickness (the fiber layer had micron-sized mesh holes) was obtained. The fiber layer was cut to obtain a sheet-shaped fiber film with bundle-shaped fibers. Other parameters of electrospinning, such as the distance between the needle and the collector (18 cm), the flow rate (2 mL / h), and the needle diameter (21G), were kept constant during the fiber production process. In each case, the scaffold was rotated for 30 minutes and dried overnight in a desiccator to remove any remaining solvent before further use.

[0038] (2) Acetylation fixation:

[0039] The formed fiber material prepared in step (1) was subjected to different degrees of acetylation. The degree of acetylation was controlled by adjusting the content of acetic anhydride and methanol, the acetylation time, and the acetylation temperature to obtain samples with different degrees of acetylation. Specifically, a solution of acetic anhydride and methanol was prepared at a volume ratio of 2:1. The prepared fiber scaffold was immersed in the mixed solution of acetic anhydride and methanol at 25°C for 3 hours, then soaked in anhydrous ethanol solution for 24 hours, and then washed, dried to obtain a formed fiber material with an acetylation degree of 80%.

[0040] (3) Preparation of PGS / CS / PMPC composite fiber with anti-adhesion coating:

[0041] Poly(2-methallyloxyethylphosphocholine) (PMPC) was nano-sprayed onto the surface of the acetylated fiber material obtained in step (2) to form a thin film with a thickness of 0.1 mm. The PGS / CS / PMPC composite fiber material with anti-adhesion coating was obtained.

[0042] Example 2

[0043] A method for preparing a PGS / CS / PMPC composite fiber material with anti-adhesion coating, comprising the following steps:

[0044] (1) Preparation of polyglycerol sebacate (PGS) / chitosan (CS) composite fiber material:

[0045] PGS and CS were blended at a weight ratio of 3:1 and dissolved in a spinning solvent of 90 wt% concentrated acetic acid. The formed fibers were prepared by electrospinning technology. The electrospinning voltage was 20 kV. The electrospun fiber line was fixed at one end of the rotating metal shaft. After a period of time, a cylindrical fiber layer with a certain thickness (the fiber layer has micrometer-sized mesh holes) was obtained. The fiber layer was cut to obtain a sheet-shaped fiber film with bundle-shaped fibers. Other parameters of electrospinning, such as the distance between the needle and the collector (18 cm), the flow rate (2 mL / h) and the needle diameter (21G) were kept constant during the fiber production process. The scaffold was rotated for 30 minutes in each case and dried overnight in a desiccator to remove any remaining solvent before further use.

[0046] Steps (2) and (3) are the same as in Example 1.

[0047] Example 3

[0048] A method for preparing a PGS / CS / PMPC composite fiber material with anti-adhesion coating, comprising the following steps:

[0049] (1) Preparation of polyglycerol sebacate (PGS) / chitosan (CS) composite fiber material:

[0050] PGS was blended with CS at a weight ratio of 2: 1 and dissolved in a spinning solvent of 90 wt% concentrated acetic acid to prepare the shaped fibers by electrospinning. The voltage for electrospinning was 20 kV. The electrospun thread was fixed at one end to a rotating metal shaft, and after a period of time, a cylindrical fiber layer with a certain thickness (the fiber layer has micron-sized mesh holes) was obtained. The fiber layer was cut to obtain a sheet-shaped fiber film with bundle-shaped fibers. Other parameters for electrospinning, such as the distance between the needle and the collector (18 cm), the flow rate (2 mL / h), and the needle diameter (21G), were kept constant during the fiber production process. The scaffold was rotated for 30 minutes in each case, and dried overnight in a desiccator to remove any remaining solvent before further use.

[0051] Steps (2) and (3) are the same as in Example 1.

[0052] Example 4

[0053] A method for preparing a PGS / CS / PMPC composite fiber material with an anti-adhesion coating, comprising the following steps:

[0054] (1) Preparation of a polyglycerol sebacate (PGS) / chitosan (CS) composite fiber material:

[0055] PGS was blended with CS at a weight ratio of 4: 1 and dissolved in a spinning solvent of 90 wt% concentrated acetic acid to prepare the shaped fibers by electrospinning. The voltage for electrospinning was 12.5 kV. The electrospun thread was fixed at one end to a rotating metal shaft, and after a period of time, a cylindrical fiber layer with a certain thickness (the fiber layer has micron-sized mesh holes) was obtained. The fiber layer was cut to obtain a sheet-shaped fiber film with bundle-shaped fibers. Other parameters for electrospinning, such as the distance between the needle and the collector (18 cm), the flow rate (2 mL / h), and the needle diameter (21G), were kept constant during the fiber production process. The scaffold was rotated for 30 minutes in each case, and dried overnight in a desiccator to remove any remaining solvent before further use.

[0056] Steps (2) and (3) are the same as in Example 1.

[0057] Example 5

[0058] A method for preparing a PGS / CS / PMPC composite fiber material with an anti-adhesion coating, comprising the following steps:

[0059] (1) Preparation of a polyglycerol sebacate (PGS) / chitosan (CS) composite fiber material:

[0060] PGS and CS were blended at a weight ratio of 4:1 and dissolved in a spinning solvent of 90 wt% concentrated acetic acid to prepare the shaped fibers by electrospinning. The voltage for electrospinning was 15 kV. The electrospun thread was fixed at one end to a rotating metal shaft, and after a period of time, a cylindrical fiber layer with a certain thickness (the fiber layer has micron-sized mesh holes) was obtained. The fiber layer was cut to obtain a sheet-shaped fiber film with bundle-shaped fibers. Other parameters for electrospinning, such as the distance between the needle 25 and the collector (18 cm), the flow rate (2 mL / h), and the needle diameter (21G), were kept constant during the fiber production process. The scaffold was rotated for 30 minutes in each case, and dried overnight in a desiccator to remove any remaining solvent before further use.

[0061] Steps (2) and (3) are the same as in Example 1.

[0062] Example 6

[0063] A method for preparing a PGS / CS / PMPC composite fiber material with an anti-adhesion coating, comprising the following steps:

[0064] (1) Preparation of a polyglycerol sebacate (PGS) / chitosan (CS) composite fiber material:

[0065] PGS and CS were blended at a weight ratio of 4:1 and dissolved in a spinning solvent of 90 wt% concentrated acetic acid to prepare the shaped fibers by electrospinning. The voltage for electrospinning was 17.5 kV. The electrospun thread was fixed at one end to a rotating metal shaft, and after a period of time, a cylindrical fiber layer with a certain thickness (the fiber layer has micron-sized mesh holes) was obtained. The fiber layer was cut to obtain a sheet-shaped fiber film with bundle-shaped fibers. Other parameters for electrospinning, such as the distance between the needle and the collector (18 cm), the flow rate (2 mL / h), and the needle diameter (21G), were kept constant during the fiber production process. The scaffold was rotated for 30 minutes in each case, and dried overnight in a desiccator to remove any remaining solvent before further use.

[0066] Steps (2) and (3) are the same as in Example 1.

[0067] Example 7

[0068] Steps (1) and (3) of this example are the same as in Example 1, and the difference lies in that step (2) is specifically:

[0069] The shaped fibrous material prepared in step (1) is subjected to acetylation treatment of different degrees, and the acetylation degree is controlled by adjusting the content of acetic anhydride and methanol, acetylation time and acetylation temperature to obtain samples with different acetylation degrees. Specifically, a solution of acetic anhydride and methanol with a volume ratio of 1:3 is configured, the prepared fibrous scaffold is immersed in the mixed solution of acetic anhydride and methanol at 40°C for 1 hour, then immersed in anhydrous ethanol solution for 6 hours, washed, dried to obtain a shaped fibrous material with an acetylation degree of 60%.

[0070] Example 8

[0071] Steps (1) and (3) of this example are the same as those of Example 1, and the difference lies in that step (2) is specifically:

[0072] The shaped fibrous material prepared in step (1) is subjected to acetylation treatment of different degrees, and the acetylation degree is controlled by adjusting the content of acetic anhydride and methanol, acetylation time and acetylation temperature to obtain samples with different acetylation degrees. Specifically, a solution of acetic anhydride and methanol with a volume ratio of 1:2 is configured, the prepared fibrous scaffold is immersed in the mixed solution of acetic anhydride and methanol at 30°C for 2 hours, then immersed in anhydrous ethanol solution for 24 hours, washed, dried to obtain a shaped fibrous material with an acetylation degree of 70%.

[0073] Example 9

[0074] Steps (1) and (3) of this example are the same as those of Example 1, and the difference lies in that step (2) is specifically:

[0075] The shaped fibrous material prepared in step (1) is subjected to acetylation treatment of different degrees, and the acetylation degree is controlled by adjusting the content of acetic anhydride and methanol, acetylation time and acetylation temperature to obtain samples with different acetylation degrees. Specifically, a solution of acetic anhydride and methanol with a volume ratio of 1:2 is configured, the prepared fibrous scaffold is immersed in the mixed solution of acetic anhydride and methanol at 30°C for 2 hours, then immersed in anhydrous ethanol solution for 24 hours, washed, dried to obtain a shaped fibrous material with an acetylation degree of 70%.

[0076] Example 10

[0077] Steps (1) and (3) of this example are the same as those of Example 1, and the difference lies in that step (2) is specifically:

[0078] The shaped fiber material prepared in step (1) is subjected to acetylation treatment of different degrees, and the acetylation degree is controlled by adjusting the content of acetic anhydride and methanol, acetylation time and acetylation temperature to obtain samples with different acetylation degrees. Specifically, a solution of acetic anhydride and methanol with a volume ratio of 2:1 is prepared, and the prepared fiber scaffold is immersed in the mixed solution of acetic anhydride and methanol at 25°C for 3.5 hours. Then, after being immersed in anhydrous ethanol solution for 36 hours, it is cleaned and dried to obtain a shaped fiber material with an acetylation degree of 85%.

[0079] Example 11

[0080] Steps (1) and (3) of this example are the same as those of Example 1, except that step (2) is specifically as follows:

[0081] The shaped fiber material prepared in step (1) is subjected to acetylation treatment of different degrees, and the acetylation degree is controlled by adjusting the content of acetic anhydride and methanol, acetylation time and acetylation temperature to obtain samples with different acetylation degrees. Specifically, a solution of acetic anhydride and methanol with a volume ratio of 2:1 is prepared, and the prepared fiber scaffold is immersed in the mixed solution of acetic anhydride and methanol at 25°C for 3.5 hours. Then, after being immersed in anhydrous ethanol solution for 36 hours, it is cleaned and dried to obtain a shaped fiber material with an acetylation degree of 85%.

[0082] Example 12

[0083] Steps (1) and (3) of this example are the same as those of Example 1, except that step (2) is specifically as follows:

[0084] The shaped fiber material prepared in step (1) is subjected to acetylation treatment of different degrees, and the acetylation degree is controlled by adjusting the content of acetic anhydride and methanol, acetylation time and acetylation temperature to obtain samples with different acetylation degrees. Specifically, a solution of acetic anhydride and methanol with a volume ratio of 2:1 is prepared, and the prepared fiber scaffold is immersed in the mixed solution of acetic anhydride and methanol at 25°C for 3.5 hours. Then, after being immersed in anhydrous ethanol solution for 36 hours, it is cleaned and dried to obtain a shaped fiber material with an acetylation degree of 85%.

[0085] Comparative Example 1

[0086] A shaped fiber material is prepared from chitosan by electrospinning technology, and the parameters of electrospinning are the same as those of Example 1. Then, acetylation treatment is performed to obtain chitosan with an acetylation degree of 80%. Then, PMPC is sprayed on the surface of the shaped fiber material by nano-spraying, and the thickness of the sprayed PMPC is 0.1 mm, to obtain a CS / PMPC composite fiber material with an anti-adhesion coating.

[0087] Comparative Example 2

[0088] The present comparative example provides a preparation method of a composite fiber material for tendon repair, comprising the following steps:

[0089] (1) PGS and CS were blended at a weight ratio of 4:1, dissolved in 90wt% concentrated acetic acid, and electrospun at 20V respectively to obtain shaped fibers. Other parameters such as the distance between the needle and the collector (18 cm), the flow rate (2 mL / h) and the needle diameter (21G) remained unchanged during the fiber production process. The scaffold was rotated for 30 minutes in each case and dried overnight in a desiccator to remove any remaining solvent before further use.

[0090] (2) The shaped fibers prepared in step (1) were subjected to acetylation treatment, and a sample with an acetylation degree of 50% was prepared by setting the volume ratio of acetic anhydride and methanol to 1:5, the acetylation time to 0.5 hours, the acetylation temperature to 50°C, and the soaking time in anhydrous ethanol solution to 6h.

[0091] (3) Poly(2-methylallyloxyethylphosphocholine) was sprayed on the surface of the material obtained in step (2) by nano-spraying to form a PMPC film with a thickness of 0.1 mm, thereby obtaining a composite fiber material for tendon repair.

[0092] Comparative Example 3

[0093] The method of the present comparative example is the same as that of Example 1, except that the electrospinning voltage is 25kV. The following performance tests were conducted on the composite fiber materials prepared in Examples 1-12 and Comparative Examples 1-3:

[0094] 1. Morphology

[0095] The shaped fibers prepared in Examples 1-12 and Comparative Examples 1-3 were observed by scanning electron microscopy. The arrangement degree, morphology and diameter of the shaped fibers of Examples 1-12 were influenced by the PGS:CS ratio, applied voltage and degree of chitosan acetylation. Under the same PGS:CS ratio, the applied voltage was increased from 12.5kV to 20kV, and the fiber surface was smoother at 20kV. However, when the applied voltage was increased to 25kV (Comparative Example 3), the fiber surface appeared to have a beaded structure. Therefore, 20kV was selected as the voltage value for subsequent experiments.

[0096] Under the same applied voltage, the fiber diameter increased significantly from 2.76±0.03μm for the material prepared from pure acetylated CS in Comparative Example 1 to 6.77±0.26μm for the material prepared from PGS:CS at a ratio of 4:1. This was due to the lower solution viscosity caused by the higher PGS ratio. In Comparative Example 2, the chitosan acetylation degree was insufficient, and the fiber material surface also had a beaded structure, which resulted in a non-smooth fiber surface and insufficient mechanical properties of the fiber.

[0097] Generally, electrospun fibers require a minimum amount of polymer chain entanglement and viscosity. At lower viscosities, more solvent molecules and less polymer chain entanglement result in a dominant surface tension effect, leading to beads or spindles.

[0098] 2. Test of mechanical properties

[0099] To characterize the mechanical properties of the composite fiber materials prepared in Examples 1-12 and Comparative Examples 1-3, uniaxial extension tests were performed using a mechanical tester. The composite fiber materials were dried under vacuum for at least 48 hours to ensure complete removal of solvent and cut into rectangles (5 x 15 mm). The specimen was inserted into the grips with a 10 mm gap as the initial size of the test region. During the uniaxial test, the initial strain rate was 25% of the original test region length, and the crosshead speed was constant at 2.5 mm / min. Five specimens of different batches were tested under each condition. The elastic modulus was calculated from the stress-strain curve in the 0-5% strain region. The ultimate strength (UTS) and the ultimate elongation (UE) were measured from the highest point of the stress-strain curve.

[0100] The results are shown in Figure 1 Fig. 1, which shows the mechanical test results of the composite fiber materials prepared with different PGS:CS ratios. CS represents the technical solution of Comparative Example 1, 2:1 represents the technical solution of Example 3, 3:1 represents the technical solution of Example 2, and 4:1 represents the technical solution of Example 1. Figure 1 A is the linear region of the stress-strain curve of the composite fiber materials in the first 15% of the strain region. The elastic modulus (EM) of the composite fiber materials of the CS, 2:1, and 3:1 groups was basically around 10 MPa, while the EM of the composite fiber materials of the 4:1 group was enhanced by about 3.8 times higher than the other three groups. It was calculated that the Young's modulus of the composite fiber materials of the 4:1 group could reach 38 MPa, which was better than the performance of the CS group ( Figure 1 B). Similar trends were observed in the ultimate tensile strength (UTS), which increased by about 1.5 times for the composite fiber materials of the 4:1 group ( Figure 1 C). At the same time, by testing the ultimate elongation (UE) of the composite fiber materials with different ratios, it was found that the UE of the composite fiber materials of the 4:1 group was about 480%, which was about 0.6 times higher than the UE of the CS group ( Figure 1D). By comparison, it was found that the stiffness (EM) and elasticity (elongation) of the composite fiber materials of the 2:1 and 3:1 groups were lower, while the strength (UTS), stiffness and elasticity of the composite fiber material of the 4:1 group were higher than those of the pure CS scaffold. EM, UTS and UE represent stiffness, strength and elasticity, respectively, thus it can be concluded that the composite fiber material prepared according to the PGS / CS weight ratio of 4:1 has good mechanical properties, and is significantly better than pure chitosan group.

[0101] 3. Cell attachment and proliferation

[0102] The Alamar Blue (AB) assay was used to quantitatively evaluate the attachment (after 8 hours) and proliferation of rat tendon stem cells. This test achieves a time course analysis of cell proliferation by measuring the ability of viable cells to reduce the resazurin dye (blue) to resorufin (pink); the decrease in AB is directly proportional to the number of viable cells. Briefly, cells were seeded into the composite fiber materials as described above. After 8 hours, the composite fiber materials with cells attached were transferred to new culture dishes containing fresh medium for rat tendon stem cells. Cells seeded on cell culture plates served as controls. AB dissolved in SD rat tendon liver cell culture (0.5 mL, 10% v / v) was added to each well, and incubated at 37°C for 3 h. Then the reduced AB (100 μL) was divided into three parts and transferred to a 96-well plate, and the absorbance was measured at 570 nm at 1 day, 3 days, 5 days and 7 days. The change in cell proliferation was calculated. The blank group was without any composite fiber material (TCP).

[0103] The attachment of tendon stem cells on different acetylation degree composite fiber materials (the weight ratio of PGS to CS of the composite fiber material was 4:1) was quantitatively analyzed by AB method after 8 h of inoculation.

[0104] The results are shown in Table 1. Figure 2As shown, at 7 days of culture, the cell adhesion was quantitatively analyzed by the AB method, which clearly showed that the composite fiber material of the CS group promoted the adhesion of cells. When the acetylation degree was 80%, the cell adhesion performance was the highest and was higher than that of the CS group, indicating that the adjustment of the acetylation degree can improve the adhesion of the material to the cells. With the further increase of the acetylation degree, the adhesion of the cells showed a decreasing trend. At the acetylation degree of 90%, the PGS / CS / PMPC composite fiber material had no significant difference compared with the cells grown on TCP, but at the acetylation degree of 80%, the cells grown on the PGS / CS / PMPC composite fiber material were much higher than those grown on TCP, which had obvious difference. With the decrease of the acetylation degree, when the acetylation degree was 50%, the cell adhesion was significantly reduced. It is shown that the acetylation degree of the application has a significant influence on the cell adhesion material. Therefore, the PGS / CS / PMPC composite fiber material with the acetylation degree of 80% has better cell growth conditions, and the composite fiber material with the acetylation degree of 80% is the best condition.

[0105] The AB experiment verified the performance of the composite fiber material in promoting the proliferation of tendon stem cells. From the figure, it can be seen that with the change of days, the number of cells grown on the PGS / CS / PMPC composite fiber material is more than that on the composite fiber material of the CS group. It is proved that the PGS of the application can promote the cell proliferation ability of CS, and the PGS / CS / PMPC composite fiber material of the application effectively improves the problem of insufficient repair ability of chitosan to tendon hepatocytes, and has the potential to promote tendon regeneration. Figure 3

[0106] 4、Promote tendon repair of rat tendon injury model

[0107] SD rats were used as animal models and were raised in a conventional feeding manner. The experiment was divided into 6 groups, each group had 3 parallel tests, which were: normal group (without implanting the scaffold), tendon injury group (control, the construction method was the general method in the art), CS scaffold group, PGS:CS ratio of 4:1 PGS / CS / PMPT scaffold group, PGS:CS ratio of 3:1 PGS / CS / PMPT scaffold group, PGS:CS ratio of 2:1 PGS / CS / PMPT scaffold group. The scaffold was implanted into the rat tendon injury reconstruction. The AB experiment and other methods were used for analysis.

[0108] ​The PGS / CS / PMPT composite fiber material can enhance the mechanical properties of the tendon repair scaffold. The implants harvested at 4 and 8 weeks after the operation show regional tendon-like luminous white new tissues at the injury site. At 8 weeks, the appearance of the extracted product is clearer, smooth and the tendon-like structure is more obvious, indicating that the new tissue is mature, while the control group shows less tendon-like structure, adhesion and swelling. Since the rats have better self-repairing ability, the control group can generate new tissues without the scaffold, but the recovery is not good compared with other groups with the scaffold. Compared with the PGS / CS / PMPT composite fiber material with different proportions of PGS and CS, the composite fiber material group of the CS group has poor repair ability and also has adhesion.

[0109] 5. Fibroblast adhesion experiment

[0110] The prepared composite fiber material is sterilized by ultraviolet, cut and placed in a 48-well cell culture plate, 1x10 4 fibroblasts are added to the well plate containing the composite fiber material, and placed in an incubator for different times (1, 3, 5, 7 days), and the unadhered cells in the experimental group are washed with PBS, and the number of cells in the well plate is detected by CCK8. The blank group is without any scaffold (TCP).

[0111] As Figure 4 , the results of the cell adhesion experiment show that the number of cells on the surface of the composite fiber material of the CS group is more than that of the blank group, and the number of cells on the surface of the PGS / CS / PMPC composite fiber material sprayed with the PMPC coating is less than that of the CS group, which indicates that the coating of PMPC can inhibit the growth of fibroblasts, and the growth of fibroblasts is a key factor for the adhesion of tendons and surrounding tissues. Therefore, the PGS / CS / PMPC composite fiber material can prevent adhesion.

[0112] In summary, the present application combines PGS and CS, prepares a shaped fiber material with micron-level mesh by electrospinning technology, and then acetylates the shaped fiber material. PGS can significantly improve the insufficient repair of CS as a tendon repair material. The compounding of PGS can significantly improve the mechanical properties of the composite fiber material by slowing down the degradation rate of CS. And when the weight ratio of PGS to CS is 4:1, it has a significant effect. In the acetylation process, when the acetylation degree is 80%, the ability of tendon stem cells to adhere to the fiber material is significantly improved.

[0113] The present application coats PMPC on the surface of the acetylated formed fiber material, further improves the anti-blocking performance of the composite fiber material. The spraying thickness of PMPC is about 0.1mm, so that the composite fiber material retains part of the micron-level mesh, has large specific surface area, and further increases the adhesion capacity of tendon stem cells.

[0114] The PGS / CS / PMPC composite fiber material of the present application is used for tendon repair and preventing adhesion in the tendon repair process, has significant effect and broad application prospect.

[0115] Finally, it should be noted that: the above examples are only used to illustrate the technical solutions of the present application, but not to limit them; although the present application has been described in detail with reference to the foregoing examples, those skilled in the art should understand that: it can still modify the technical solutions recorded in the foregoing examples, or make equivalent replacement for part or all of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present application.

Claims

1. A chitosan composite fiber material for tendon repair, characterized in that: The chitosan composite fiber material is an acetylated chitosan / polyglycerol sebacate composite fiber material coated with poly(2-methacryloyloxyethyl phosphorylcholine) on the surface; The acetylation degree of the acetylated chitosan is 75-85%.

2. The chitosan composite fiber material for tendon repair according to claim 1, characterized in that: The chitosan composite fiber material is a sheet structure with micron-level meshes.

3. The chitosan composite fiber material for tendon repair according to claim 1, characterized in that: The coating thickness of the poly (2-methacryloyloxyethyl phosphorylcholine) is 0.1-0.2 mm.

4. The method for preparing the chitosan composite fiber material for tendon repair according to any one of claims 1 to 3, characterized in that: The steps include: S1, dissolving polyglycerol sebacate and chitosan in a spinning solvent, preparing a formed fiber material by electrospinning, and drying to remove the solvent; S2. Immersing the shaped fiber material prepared in step S1 in a mixed solution of acetic anhydride and methanol in a volume ratio of 1-5:5-1, acetylizing at 0-50° C. for 0.5-24 h, then soaking in anhydrous ethanol for 6-48 h, washing, and drying to obtain an acetylated shaped fiber material; S3. Spraying poly (2-methacryloyloxyethyl phosphorylcholine) on the surface of the acetylated formed fiber material to obtain.

5. The preparation method according to claim 4, characterized in that In step S1, the spinning solvent is one or a combination of two or more of dichloromethane, chloroform, acetic acid, dimethyl sulfoxide, and acetone.

6. The preparation method according to claim 4, characterized in that In step S1, the weight ratio of polyglycerol sebacate to chitosan is 2-4:

1.

7. The preparation method according to claim 4, characterized in that In step S1, the voltage of the electrospinning is 12.5-20 kV.

8. Use of the chitosan composite fiber material according to any one of claims 1 to 3 or the chitosan composite fiber material prepared by the preparation method according to any one of claims 4 to 7 in tendon repair and as an anti-adhesion material during tendon repair.

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