A modified polyurethane, its method of preparation and use as a meniscal replacement material

Abstract

The application discloses a modified polyurethane preparation and application thereof in 3D printing as a meniscus replacement material. A prepolymer is synthesized through the reaction of sulfobetaine diol, polycarbonate diol and hexamethylene diisocyanate, and the prepolymer is chain-extended by adipic acid dihydrazide to build a multiple hydrogen bond cross-linked zwitterionic supramolecular polyurethane. The introduced zwitterionic side groups can significantly improve the toughness, printability and lubricity of the polyurethane elastomer by adjusting the hydrogen bond in the hard segment and the hydration in the water environment. The fused deposition modeling 3D printing technology is used to customize the zwitterionic polyurethane meniscus support with strong toughness and lubricity without any post-processing. The 3D printed zwitterionic polyurethane meniscus replacement is implanted into the knee joint of a rabbit, and the in-vivo result shows that the zwitterionic polyurethane meniscus replacement has a good application prospect as a meniscus replacement.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biomedical materials, more particularly to a modified polyurethane and its application in 3D printing as a meniscus replacement material. BACKGROUND

[0002] Due to the unique vascularization feature, the self-repairing ability of meniscus is limited. So far, meniscectomy is still the most commonly used method in clinical treatment of severe meniscal injury. However, the removal of meniscus can cause stress concentration and load transfer disorder of the knee joint, thereby accelerating the progression of osteoarthritis (OA), and allogeneic meniscus transplantation has problems such as insufficient graft source, immune rejection and risk of disease transmission. Therefore, it is urgent to develop an artificial meniscus substitute to reconstruct the function of meniscus, reduce cartilage wear and avoid the development of osteoarthritis after meniscectomy.

[0003] Polyurethane is a recognized biomedical material, which has been widely used as a substitute for internal tissues and organs of the human body due to its high degree of freedom in molecular design. Among various polyurethanes, polycarbonate polyurethane exhibits excellent oxidative degradation resistance and hydrolytic stability, and is expected to be used as a long-term implanted meniscus substitute. Generally, polyurethane has low hydrophilicity, which can cause cartilage wear problems when used as an implant. Some studies have confirmed that the introduction of zwitterions into the polyurethane system can significantly improve the hydrophilicity and lubricity of polyurethane.

[0004] The emerging 3D printing technology (e.g., fused deposition modeling (FDM)) is expected to achieve precise printing with high shape fidelity, which can well solve the design of meniscus microstructure and individual customization, thereby reducing the problems of shape mismatch and load transfer that may occur after transplantation. SUMMARY

[0005] The present application aims to overcome the shortcomings of the prior art, solve the problem that the current meniscus substitute is difficult to simulate the in vivo lubrication mechanism and faces the problem of cartilage wear in long-term implantation, and proposes a preparation method of modified polyurethane and 3D printed meniscus, which has a simple formula and easy preparation method.

[0006] The technical purpose of the present application is achieved by the following technical scheme.

[0007] A modified polyurethane and its preparation method are carried out according to the following steps:

[0008] Step 1, the zwitterionic diol, polyhexamethylene carbonate diol, and diisocyanate are uniformly dispersed in an organic solvent, a catalyst is added for prepolymerization, wherein the mass ratio of polyhexamethylene carbonate diol, zwitterionic diol, and diisocyanate is 1:(0.1-0.3):(0.5-0.8), the prepolymerization temperature is 70-90 degrees Celsius, and the time is 1-3 hours.

[0009] In step 1, the mass ratio of polyhexamethylene carbonate diol, zwitterionic diol, and diisocyanate is 1:(0.1-0.2):(0.5-0.6).

[0010] In step 1, the diisocyanate is hexamethylene diisocyanate hard segment.

[0011] In step 1, the number average molecular weight of polyhexamethylene carbonate diol is 1k-3k.

[0012] In step 1, the molecular formula of the zwitterionic diol is as follows

[0013] Prepared according to the following method: N-methyldiethanolamine (5.96 g, 50 mmol), 1,3-propane sultone (6.70 g, 55 mmol) and dichloromethane (200 mL) were added to a 500 mL round-bottom flask. The mixture was stirred at 400 rpm in an oil bath at 40°C under a nitrogen atmosphere for 24 hours. The reaction solution was rotated to about 25 mL using a rotary evaporator. The remaining solution was precipitated with acetone at a ratio of 1:20, then washed with acetone three times to obtain a white powder. Finally, the product was dried in a vacuum drying oven and weighed; Reference: Liu Q, Wang X, Chiu A, et al. Azwitterionic polyurethane nanoporous device with low foreign-body response for islet encapsulation [J]. Advanced Materials, 2021, 33(39): 2102852.

[0014] In step 1, the polyhexamethylene carbonate diol (solid state) and the zwitterionic diol (solid state) are selected to be added to a three-necked flask, a mechanical stirring device is assembled and simply sealed with plastic wrap at the interface, and then placed in a 120°C oil bath pot for vacuum drying for 1 hour to remove water from the raw materials and avoid violent polymerization during the prepolymerization stage.

[0015] In step 1, the catalyst is a commonly used catalyst in the polyurethane field, such as stannous 2-ethylhexanoate, which is added in an amount adjusted according to the amount of reactants and uniformly dropped.

[0016] In step 1, the organic solvent is a commonly used reaction solvent in the polyurethane field, and the type and / or amount are adjusted according to the amount of the reactants, such as dimethyl sulfoxide, dimethyl formamide or dimethyl acetamide.

[0017] In step 1, the prepolymerization temperature is 70-80 degrees Celsius, and the time is 1-2 hours.

[0018] In step 2, after prepolymerization, the temperature of the reaction system is reduced to 40-50 degrees Celsius, the chain extender adipic acid dihydrazide (ADH) is added dropwise into the reaction system, and the reaction is carried out under heat preservation. The entire reaction process from the start of the prepolymerization is carried out under an inert protective atmosphere, wherein the mass ratio of polyhexamethylene carbonate diol to adipic acid dihydrazide is 1:(0.3-0.5).

[0019] In step 2, the same solvent as in step 1 is selected to uniformly disperse the chain extender adipic acid dihydrazide, which is then added dropwise in batches into the reaction system to carry out the chain extension reaction (the amount of each batch is basically equal), during which the presence of the "climbing rod" phenomenon is observed.

[0020] In step 2, the inert protective atmosphere is nitrogen, helium or argon.

[0021] In step 2, the reaction temperature is 40-50 degrees Celsius, and the reaction time is 6-12 hours.

[0022] In step 2, the mass ratio of polyhexamethylene carbonate diol to adipic acid dihydrazide is 1:(0.3-0.4).

[0023] In step 2, after the reaction is completed, the reactants are added into diethyl ether in a volume ratio of 1:20 for sedimentation, the solid product obtained by sedimentation is suction filtered, and then washed with diethyl ether and pure water for 3 times in sequence before being placed into a vacuum drying box for drying. Finally, a dry white product is obtained, which is the modified polyurethane of the present application, and is named as PCU-MeSB.

[0024] The application of the modified polyurethane as a meniscus replacement material. The modified polyurethane is 3D printed by a biological 3D printer, the prepared modified polyurethane (PCU-MeSB) material is added to the high-temperature barrel of the printer, heated to 180-220℃, and heat preserved for a period of time, such as 10-20min, the printing parameters are set, and continuous printing is carried out. The extruded microfibers are deposited on a printing platform at 35-45℃; after the printing is completed, the printing product is soaked in a PBS buffer solution to swell to equilibrium, and a 3D printed PCU-MeSB scaffold with a specific shape is obtained.

[0025] In the technical scheme of the application, the printing mode is a linear filling mode, the printing speed is 6-10 mm / s, and the filament speed is 0.05-0.15 mm / s.

[0026] In the technical scheme of the application, the CAD / CAM software is used to design the 3D printing model, the meniscus model of the rabbit is obtained by reconstructing the Micro-CT data, and the design system architecture is constructed by using the layer-by-layer accumulation method.

[0027] The application starts from the design of hydrogen bond structure, adopts the combination of polyhexamethylene carbonate diol soft segment, hexamethylene diisocyanate hard segment and adipic acid dihydrazide chain extender to obtain a supramolecular polyurethane (PCU) with high symmetry and high rigidity, so as to ensure sufficient structural support in the subsequent printing process. On this basis, a zwitterionic diol (MeSB-diol) is synthesized and introduced into the polyurethane system to synthesize a zwitterionic polyurethane (PCU-MeSB). On the one hand, the introduced zwitterionic side group will help to adjust the intermolecular hydrogen bond interaction in the hard segment and the movement of the polymer chain, so as to make it have proper processing temperature, printability and toughness; on the other hand, the hydration of the zwitterionic group can improve the hydrophilicity of the PCU and provide a lubricating surface. Through the FDM 3D printing technology, the PCU-MeSB meniscus substitute with specific performance can be easily and directly prepared without any other post-processing (such as coating and filling of hydrogel). The application has simple formula, easy preparation method, and selects appropriate soft segment and hard segment and chain extender to construct a polyurethane framework with excellent mechanical properties, and then synthesizes and introduces a zwitterionic chain extender to improve the hydrophilicity and biocompatibility, so as to solve the problem of cartilage wear caused by high rigidity polyurethane. The modified polyurethane is added to the high-temperature barrel of the 3D printer, heated to 180-220 DEG C for melt deposition molding 3D printing, and the 3D printed meniscus scaffold is obtained; the 3D printed meniscus scaffold is soaked in a phosphate buffered saline solution to swell to equilibrium, and the 3D printed meniscus substitute for in vivo implantation is obtained. The method can realize excellent mechanical properties through hydrogen bond structure design, relieve the problem of cartilage wear by introducing a zwitterionic chain extender, and obtain a size-matched meniscus scaffold through 3D printing, which has great application prospect as a meniscus substitute. BRIEF DESCRIPTION OF DRAWINGS

[0028] Figure 1 is the synthesis route map in the application, wherein a is the synthesis route map of the zwitterionic diol MeSB-diol, and b is the synthesis route map of the PCU and PCU-MeSB polyurethane.

[0029] Figure 2are photographs of 3D printed PCU and PCU-MeSB in the present invention, wherein a is single filament photograph, b is the display of PCU-MeSB multi-layer square grid, meniscus, cylinder and snowflake shape.

[0030] Figure 3 are tensile stress-strain curves of 3D printed PCU and PCU-MeSB structures in the present invention.

[0031] Figure 4 are the corresponding test results bar charts of Young's modulus, tensile strength, elongation at break and toughness of 3D printed PCU-MeSB structures in the present invention.

[0032] Figure 5 are 1000-cycle tensile loading-unloading curves of 3D printed PCU-MeSB structures in the present invention.

[0033] Figure 6 are the corresponding final stress and dissipated energy curves of the 1st and every 100-cycle tensile loading-unloading of 3D printed PCU-MeSB structures in the present invention.

[0034] Figure 7 are 1000-cycle compression loading-unloading curves of 3D printed PCU-MeSB structures in the present invention.

[0035] Figure 8 are the corresponding final stress and dissipated energy curves of the 1st and every 100-cycle compression loading-unloading of 3D printed PCU-MeSB structures in the present invention.

[0036] Figure 9 are tear energy test results bar charts of 3D printed PCU and PCU-MeSB structures in the present invention.

[0037] Figure 10 are friction coefficient test curves of 3D printed PCU and PCU-MeSB structures in the present invention.

[0038] Figure 11 are implantation process and gross observation figures of 3D printed meniscus substitutes in rabbit models in the present invention, wherein a is the implantation process of 3D printed meniscus substitutes in rabbit models in animal experiments, (i) is the incision of skin tissue and fascial tissue, (ii) is the cutting of medial collateral ligament and opening of knee joint capsule, (iii) is meniscectomy, (iv) is the implantation of stent, (v) is the suture of medial collateral ligament and closing of knee joint capsule, (vi) is the suture of skin tissue and fascial tissue, b is the gross observation figures of femoral condyle (FC), tibial plateau (TP) and meniscus substitute at 4 and 12 weeks after surgery, scale: 5 millimeters.

[0039] Figure 12 are pictures of femoral condyle (FC) and tibial plateau (TP) staining at 4 and 12 weeks after implantation of 3D-printed meniscal substitutes in a rabbit model according to the present invention, wherein hematoxylin-eosin is H&E staining and safranin O-fast green is SOFG staining.

[0040] Figure 13 are bar graphs of the results of the International Cartilage Research Society (OARSI) score for assessing the degree of cartilage degeneration (1) according to the present invention.

[0041] Figure 14 are bar graphs of the results of the International Cartilage Research Society (OARSI) score for assessing the degree of cartilage degeneration (2) according to the present invention. DETAILED DESCRIPTION

[0042] The technical solutions of the present application are further described below through specific examples.

[0043] Example 1

[0044] Step 1, N-methyldiethanolamine (5.96 g, 50 mmol), 1,3-propane sultone (6.70 g, 55 mmol) and dichloromethane (200 mL) were added into a 500 mL round bottom flask. The mixture was stirred at 400 rpm in a 40 °C oil bath under nitrogen atmosphere for 24 hours. Then, the reaction solution was rotated to about 25 mL using a rotary evaporator. The remaining solution was precipitated with acetone at a ratio of 1:20, and then washed with acetone three times to obtain a white powder. Finally, the product was dried in a vacuum drying oven and weighed. 1 g of polyhexamethylene carbonate diol (Mn ~ 2000) and 0.121 g of MeSB-diol were weighed into a 100 mL three-necked flask, and a mechanical stirring device was assembled and simply sealed with plastic wrap at the interface before being placed in a 120 °C oil bath for vacuum drying for 1 hour to remove the moisture in the raw materials and avoid the occurrence of violent polymerization during the prepolymerization stage. After the temperature was reduced to 80 °C, 0.504 g of hexamethylene diisocyanate (HDI) was weighed into a 10 mL centrifuge tube, and 6 mL of dimethyl sulfoxide (DMSO) solvent and a few drops of 2-ethylhexanoic acid stannous catalyst were added in turn, and then added dropwise into the three-necked flask after mixing well, and the prepolymerization reaction was carried out for 1 hour. The temperature was reduced to 40 °C, and 0.369 g of adipic acid dihydrazide (ADH) chain extender was dissolved in 20 mL of DMSO solvent, and was added dropwise into the three-necked flask in two batches, and the “climbing rod” phenomenon was observed during the chain extension. After that, it was stirred at 40 °C overnight. The entire reaction process from the beginning of the prepolymerization was always kept under a nitrogen atmosphere. Then the reaction was precipitated in ether at a ratio of 1:20, and the solid product obtained by precipitation was suction filtered and washed with ether and pure water in turn for 3 times, and then placed in a vacuum drying oven for drying, and finally a dry white product was obtained, which was named PCU-MeSB. The same method was used to prepare a polyurethane control group with the same content of soft and hard segments, which was named PCU.

[0045] Example 2

[0046] Step 1, N-methyldiethanolamine (5.96 g, 50 mmol), 1,3-propane sultone (6.70 g, 55 mmol) and dichloromethane (200 mL) were added into a 500 mL round bottom flask. The mixture was stirred at 400 rpm in a 40 °C oil bath under nitrogen atmosphere for 24 hours. Then, the reaction solution was rotated to about 25 mL using a rotary evaporator. The remaining solution was settled with acetone at a ratio of 1:20, and then washed with acetone three times to obtain a white powder. Finally, the product was dried in a vacuum drying oven and weighed. 1 g of polyhexamethylene carbonate diol (Mn ~ 2000) and 0.121 g of MeSB-diol were weighed into a 100 mL three-necked flask, and a mechanical stirring device was assembled and simply sealed with plastic wrap at the interface before being placed in a 120 °C oil bath for vacuum drying for 1 hour to remove the moisture in the raw materials and avoid the occurrence of violent polymerization during the prepolymerization stage. After the temperature was reduced to 80 °C, 0.504 g of hexamethylene diisocyanate (HDI) was weighed into a 10 mL centrifuge tube, and 6 mL of dimethyl sulfoxide (DMSO) solvent and a few drops of 2-ethylhexanoic acid stannous catalyst were added in turn, and then added dropwise into the three-necked flask after mixing well, and the prepolymerization reaction was carried out for 1 hour. The temperature was reduced to 40 °C, and 0.369 g of adipic acid dihydrazide (ADH) chain extender was dissolved in 20 mL of DMSO solvent, and was added dropwise into the three-necked flask in two batches, and the “climbing rod” phenomenon was observed during the chain extension. After that, it was stirred at 40 °C overnight. The entire reaction process from the beginning of the prepolymerization was always kept under a nitrogen atmosphere. Then the reaction was added to diethyl ether at a ratio of 1:20 for settlement, and the solid product obtained by settlement was suction filtered and washed with diethyl ether and pure water in turn for 3 times, and then placed in a vacuum drying oven for drying, and finally a dry white product was obtained, which was named PCU-MeSB. The same method was used to prepare a polyurethane control group with the same content of soft and hard segments, which was named PCU.

[0047] Example 3

[0048] Step 1, N-methyldiethanolamine (5.96 g, 50 mmol), 1,3-propane sultone (6.70 g, 55 mmol) and dichloromethane (200 mL) were added into a 500 mL round bottom flask. The mixture was stirred at 400 rpm in an oil bath at 40 °C for 24 h under nitrogen atmosphere. Then, the reaction solution was rotated to about 25 mL using a rotary evaporator. The remaining solution was precipitated with acetone at a ratio of 1:20, and then washed with acetone three times to obtain a white powder. Finally, the product was dried in a vacuum drying oven and weighed. 1 g of polyhexamethylene carbonate diol (Mn~2000) and 0.121 g of MeSB-diol were weighed into a 100 mL three-necked flask, and a mechanical stirring device was assembled and simply sealed with plastic wrap at the interface before being placed in a 120 °C oil bath for vacuum drying for 1 h to remove the moisture in the raw materials and avoid the occurrence of violent polymerization during the prepolymerization stage. After the temperature was reduced to 80 °C, 0.504 g of hexamethylene diisocyanate (HDI) was weighed into a 10 mL centrifuge tube, and 6 mL of dimethyl sulfoxide (DMSO) solvent and a few drops of 2-ethylhexanoic acid stannous catalyst were added in turn, and then added dropwise into the three-necked flask after mixing well, and the prepolymerization reaction was carried out for 1 h. The temperature was reduced to 40 °C, and 0.369 g of adipic acid dihydrazide (ADH) chain extender was dissolved in 20 mL of DMSO solvent, and was added dropwise into the three-necked flask in two batches, and the “climbing rod” phenomenon was observed during the chain extension. After that, the reaction was stirred at 40 °C overnight, and the entire reaction process from the beginning of the prepolymerization was always kept under a nitrogen atmosphere. Then, the reaction was precipitated in ether at a ratio of 1:20, and the solid product obtained by precipitation was filtered and washed with ether and pure water three times in turn, and then dried in a vacuum drying oven, and finally a dry white product was obtained, which was named PCU-MeSB. The same method was used to prepare a polyurethane control group with the same content of soft and hard segments, which was named PCU.

[0049] Reference: 3D printing of lubricative stiff supramolecular polymer hydrogels for meniscus replacement, Biomater. Sci., 2021, 9, 5116-5126.

[0050] Mechanical property test of 3D printed scaffold for meniscus replacement (PCU and PCU-MeSB): The 3D printed scaffold for meniscus replacement (PCU and PCU-MeSB) prepared was tested by using an electronic universal testing machine (Instron, USA): rectangular tensile samples (length: 20 mm, width: 5 mm, thickness: 0.5 mm) and cylindrical compression samples (diameter: 7 mm; height: 2 mm) were tested for tensile test at a strain rate of 50 mm / min, for compression test at a strain rate of 10 mm / min, the maximum strain of cyclic tensile loading-unloading experiment was fixed at 30%, the strain rate was 50 mm / min, the maximum strain of cyclic compression loading-unloading experiment was fixed at 30%, the strain rate was 10 mm / min (i.e. the maximum strain of all cycles was fixed at 30%), and the tear test used trouser-shaped samples and was performed at a strain rate of 50 mm / min. For the tear test, trouser-shaped PCU and PCU-MeSB printed samples were used, both legs were fixed on the tensile machine clamps, and stretching was performed at a loading rate of 50 mm / min. The tear energy of the sample was calculated as follows:

[0051]

[0052] wherein F is the average load during stable tearing process, in N, and d is the thickness of the sample, in mm.

[0053] (1) Figure 3 and Figure 4 The tensile mechanical properties of the printed structure were characterized. The 3D printed PCU-MeSB exhibited a tensile strength of 4.56 ± 0.78 MPa, a Young’s modulus of 14.92 ± 2.82 MPa, an elongation at break of 694.01 ± 209.61%, and a toughness of 23.48 ± 10.02 MJ·m -3 -2. In contrast, the 3D printed PCU exhibited relatively poor mechanical properties (tensile strength of 1.38 ± 0.83 MPa, Young’s modulus of 13.74 ± 5.38 MPa, elongation at break of 144.01 ± 87.35%, and toughness of 1.23 ± 0.92 MJ·m -3 -2) due to the uneven and defect-rich filaments formed during printing, thus intuitively indicating that the comprehensive mechanical properties of the 3D printed PCU-MeSB structure can better meet the requirements of load bearing.

[0054] (2) As shown in Figure 5 -8, in order to further evaluate the fatigue resistance of the 3D printed PCU-MeSB, cyclic tensile and compression loading-unloading tests were performed for 1000 cycles, and there was no rest between consecutive two cycles, Figure 6 and 8 the dissipated energy was Figure 5 and7 The area enclosed by the corresponding stress-strain curve and the coordinate axes. During cyclic tensile and compressive loading-unloading, the ultimate stress at 30% strain and dissipated energy first decreased and then gradually stabilized. In addition, the repeatability of the stress-strain curve of 3D printed PCU-MeSB was good with the increase of cycle number. In the cyclic tensile test, 3D printed PCU-MeSB achieved a maximum stress of 4.71 MPa and a maximum dissipated energy of 588.72 kJ·m -3 -2 in the first cycle, and had a larger stress-strain hysteresis loop. After 1000 cycles, the ultimate stress and dissipated energy decreased to a stable level of 4.04 MPa and 174.38 kJ·m -3 -2, respectively. The above results show that partial irreversible deformation occurs during the initial tensile process, in which energy dissipation dominates. In addition, similar results were observed in the cyclic compression test. The ultimate stress and dissipated energy of 3D printed PCU-MeSB reached 8.00 MPa and 266.97 kJ·m -3 -2, respectively, after 1000 cycles. It is particularly noteworthy that the ultimate stress of 3D printed PCU-MeSB exceeded the initial stress value after 1000 cycles, which may be due to the volatilization of water absorbed by the zwitterions in PCU-MeSB during constant compression, resulting in an increase in stress. It is well known that a significant stress-strain hysteresis curve helps to dissipate energy and resist changing loads; while low hysteresis and good fatigue resistance are more suitable for meniscus implants. (To illustrate that the printed PCU-MeSB exhibits excellent fatigue resistance).

[0055] (3) Poor tear resistance may limit the application of meniscus implants in vivo, because once a crack is formed during the implantation process, it will cause rapid tearing of the implant, ultimately leading to the failure of the meniscus replacement implantation surgery. As shown in Figure 9 , the tear energy of PCU-MeSB was as high as 12.51 ± 1.57 kJ·m -2 , about 7 times that of PCU (1.81 ± 0.2 kJ·m -2 ), indicating that the printed PCU-MeSB exhibits excellent tear resistance, indicating that the structure of the printed PCU-MeSB can withstand tearing and maintain its integrity after implantation.

[0056] Lubrication performance test of 3D printed scaffolds (PCU and PCU-MeSB) for meniscus replacement: The lubrication performance of 3D printed PCU-MeSB was tested by TRB ball-on-disc tribometer (CSM Tribometer) through the friction coefficient test. The stainless steel ball with a diameter of 6 mm was used as the friction pair, the sliding distance was 10 mm, the test frequency was 1 Hz. The test temperature was 25℃, the test time was 300 s, the lubricant was water, and the friction coefficient was calculated by the system software.

[0057] Before the in vivo implantation experiment, the lubrication performance of 3D printed PCU-MeSB was characterized by in vitro friction test. As shown in Figure 10 printed PCU-MeSB (COF = 0.13 ± 0.01) showed better lubricity than printed PCU (COF = 0.39 ± 0.03), indicating that the printed PCU-MeSB had certain surface lubrication performance, which indicated that the printed PCU-MeSB implant might have better cartilage protection in vivo. It should be noted that the COF of the printed PCU-MeSB was similar to that of the pig joint cartilage (0.11).

[0058] Animal experiment of 3D printed scaffolds (PCU and PCU-MeSB) for meniscus replacement implanted in rabbit model:

[0059] All procedures of animal experiments were performed in accordance with the guidelines of the Chinese Government Animal Experiment Management Committee. The animal experiments were approved by the Animal Ethics Committee of Tianjin Medical University General Hospital. Twenty-six male rabbits, weighing 2.5-3.0 kg, were used for experiments of 4 and 12 weeks, respectively. The experimental animals were randomly divided into 4 groups: sham operation group (2 rabbits per time point), meniscectomy group (3 rabbits per time point), PCU group (4 rabbits per time point), and PCU-MeSB group (4 rabbits per time point). The 75% ethanol soaked meniscal implants were sterilized with ultraviolet light for 1 hour before surgery, and all surgeries were performed under sterile conditions. Before surgery, the rabbits were anesthetized with intravenous injection of 10% chloral hydrate (3.5 mL / kg) into the ear vein, and local anesthesia was performed with xylazine hydrochloride (0.2 mL / kg) during the operation. Then, the rabbits were fixed on the operating table, and the leg hair was shaved and disinfected with iodophor. Then, the rabbits' bilateral medial meniscus was completely removed. First, the skin and fascia tissue was incised to form a 2 cm incision. Then, the medial collateral ligament was cut, and the posterior horn of the medial meniscus was exposed. Subsequently, the joint capsule tissue was opened, and the posterior horn and anterior horn of the meniscus were dissected along the junction of the medial meniscus and tibial plateau with a sharp scalpel, and then the entire meniscus was removed. Next, the appropriate size of the meniscal substitute was selected for implantation during the operation by gross observation. The PCU and PCU-MeSB meniscal implants were transplanted into the knee joint, and the anterior horn of the implant was sutured to the medial collateral ligament, and the posterior horn of the implant was sutured to the joint capsule tissue to horizontally fix the meniscus to the initial position of the medial meniscus of the knee joint. In comparison, only the meniscus was exposed in the sham operation group, and only the meniscectomy was performed in the meniscectomy group. Then, the medial collateral ligament was connected and the joint capsule was closed with non-absorbable surgical sutures, the fascia and skin tissues were sutured, and the surgical leg was cleaned with alcohol cotton pads. Finally, all rabbits were injected with penicillin three days after surgery to prevent infection. After surgery, the rabbits were returned to the cage to ensure that all rabbits were free to move around. In the following period of time, the rabbits' knee joint conditions and activities were observed and recorded. All rabbits were euthanized, and their femurs, tibias, and meniscal implants were collected at 4 and 12 weeks after surgery. The femoral condyles (FCs), tibial platforms (TPs), and meniscal substitutes of all rabbits were photographed and evaluated. The tissues were fixed in 4% paraformaldehyde for 3 days, and the corresponding distal femurs and proximal tibias were decalcified in ethylenediaminetetraacetic acid (EDTA) solution (10%) for 28 days. Then, all tissues were dehydrated using graded ethanol and embedded in paraffin. Finally, the obtained tissue sections were stained with hematoxylin-eosin (H&E) and safranin-fast green (SOFG) (thickness ~ 10 μm). The above-mentioned tissue sections were observed and photographed using an EVOS M500 inverted fluorescence microscope. The group with only exposed rabbit meniscus was set as the sham operation group, the group with only meniscectomy was set as the meniscectomy group, and the 3D printed meniscal scaffold (PCU and PCU-MeSB) groups were experimental groups, as shown inFigure 12 As shown in Fig. 14, the 3D printed meniscus support (PCU-MeSB) group is significantly better than other groups. According to the osteoarthritic cartilage histopathology evaluation system of the International Osteoarthritis Research Society (OARSI), four researchers independently evaluate the cartilage degeneration of FC and TP, and the higher the score, the worse the cartilage condition, as shown in Fig. 15. Figure 13 As shown in Fig. 14, the 3D printed meniscus support (PCU-MeSB) group is significantly better than other groups. Therefore, the PCU-MeSB of the present application is applied as a meniscus replacement material.

[0060] According to the adjustment of the process parameters according to the content of the present application, the preparation of modified polyurethane and 3D printed meniscus can be realized, and the test shows that the performance is basically consistent with the present application. The above has made an exemplary description of the present application, it should be explained that, without departing from the core of the present application, any simple transformation, modification or other equivalent replacement which can not cost the creative labor of the person skilled in the art falls into the protection scope of the present application.

Claims

1. A modified polyurethane, characterized in that, The modified polyurethane is prepared by the following steps: Step 1: Amphoteric glycol, polyhexamethylene carbonate glycol, and diisocyanate are uniformly dispersed in an organic solvent. A catalyst is added for prepolymerization. The mass ratio of polyhexamethylene carbonate glycol, amphoteric glycol, and diisocyanate is 1:(0.1-0.3):(0.5-0.8). The prepolymerization temperature is 70-90 degrees Celsius, and the time is 1-3 hours. The molecular formula of the amphoteric glycol is shown below: Step 2: After prepolymerization, the temperature of the reaction system is lowered to 40-50 degrees Celsius. The chain extender adipate dihydrazide is added dropwise to the reaction system and the reaction is carried out under the temperature to obtain modified polyurethane. Throughout the entire reaction process from the start of prepolymerization, the reaction is always carried out under an inert protective atmosphere. The mass ratio of polyhexamethylene carbonate diol to adipate dihydrazide is 1:(0.3-0.5).

2. The modified polyurethane according to claim 1, characterized in that, In step 1, the mass ratio of polyhexamethylene carbonate diol, zwitterionic diol, and diisocyanate is 1:(0.1-0.2):(0.5-0.6); the diisocyanate is the hard segment of hexamethylene diisocyanate, and the number average molecular weight of polyhexamethylene carbonate diol is 1k-3k; the catalyst is stannous 2-ethylhexanoate, the organic solvent is dimethyl sulfoxide, dimethylformamide, or dimethylacetamide, the prepolymerization temperature is 70-80 degrees Celsius, and the time is 1-2 hours.

3. The modified polyurethane according to claim 1, characterized in that, In step 2, the chain extender adipate dihydrazide is uniformly dispersed in the same solvent as in step 1, and then added dropwise to the reaction system in batches to carry out the chain extension reaction. The amount added in each batch is basically equal, and the "climbing rod" phenomenon is observed during the process. The inert protective atmosphere is nitrogen, helium or argon. The reaction temperature is 40-50 degrees Celsius, and the reaction time is 6-12 hours. The mass ratio of polyhexamethylene carbonate diol to adipate dihydrazide is 1:(0.3-0.4).

4. A method for preparing modified polyurethane, characterized in that, Follow these steps: Step 1: Amphoteric glycol, polyhexamethylene carbonate glycol, and diisocyanate are uniformly dispersed in an organic solvent. A catalyst is added for prepolymerization. The mass ratio of polyhexamethylene carbonate glycol, amphoteric glycol, and diisocyanate is 1:(0.1-0.3):(0.5-0.8). The prepolymerization temperature is 70-90 degrees Celsius, and the time is 1-3 hours. The molecular formula of the amphoteric glycol is shown below: Step 2: After prepolymerization, the temperature of the reaction system is lowered to 40-50 degrees Celsius. The chain extender adipate dihydrazide is added dropwise to the reaction system and the reaction is carried out under the temperature to obtain modified polyurethane. Throughout the entire reaction process from the start of prepolymerization, the reaction is always carried out under an inert protective atmosphere. The mass ratio of polyhexamethylene carbonate diol to adipate dihydrazide is 1:(0.3-0.5).

5. The method for preparing modified polyurethane according to claim 4, characterized in that, In step 1, the mass ratio of polyhexamethylene carbonate diol, zwitterionic diol, and diisocyanate is 1:(0.1-0.2):(0.5-0.6); the diisocyanate is the hard segment of hexamethylene diisocyanate, and the number average molecular weight of polyhexamethylene carbonate diol is 1k-3k; the catalyst is stannous 2-ethylhexanoate, the organic solvent is dimethyl sulfoxide, dimethylformamide, or dimethylacetamide, the prepolymerization temperature is 70-80 degrees Celsius, and the time is 1-2 hours.

6. The method for preparing modified polyurethane according to claim 4, characterized in that, In step 2, the inert protective atmosphere is nitrogen, helium, or argon; the reaction temperature is 40-50 degrees Celsius, and the reaction time is 6-12 hours; the mass ratio of polyhexamethylene carbonate diol to adipate dihydrazide is 1:(0.3-0.4); the same solvent as in step 1 is used to uniformly disperse the chain extender adipate dihydrazide, and then it is added dropwise to the reaction system in batches to carry out the chain extension reaction. The amount added in each batch is basically equal, and the "climbing rod" phenomenon is observed during the process.

7. The application of the modified polyurethane as described in any one of claims 1-3 in the preparation of meniscus substitute materials.

8. The application of the modified polyurethane according to claim 7 in the preparation of meniscus substitute materials, characterized in that, Modified polyurethane was 3D printed using a bio-3D printer. The modified polyurethane material was added to the high-temperature barrel of the printer, heated to 180-220℃, and held at that temperature for 10-20 minutes. Printing parameters were set, and continuous printing was performed. The extruded microfibers were deposited on a printing platform at 35-45℃. After printing was completed, the printed product was immersed in PBS buffer solution until swelling equilibrium was reached, resulting in a 3D-printed PCU-MeSB scaffold with a specific shape.

9. The application of the modified polyurethane according to claim 8 in the preparation of meniscus substitute materials, characterized in that, The printing mode is linear fill mode, the printing speed is 6-10mm / s, and the filament output speed is 0.05-0.15mm / s; the design architecture is constructed using a layer-by-layer stacking method.

10. The application of the modified polyurethane according to claim 7 in the preparation of meniscus substitute materials, characterized in that, The 3D-printed structure used for meniscus replacement has a Young's modulus of 14.92±2.82 MPa, a compressive modulus of 22.08±4.31 MPa, and a toughness of 23.48±10.02 MJ / m. 3 .

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