Decellularized artificial lymphatic fluid drainage tube and preparation method and application thereof

Abstract

The application discloses a kind of acellular artificial lymph fluid drainage tube and its preparation method and application, the preparation method of acellular artificial lymph fluid drainage tube includes the following steps: with animal blood vessel as raw material, the raw material is treated, washing treatment, enhancement treatment, pore preparation treatment and drying treatment are carried out.This application preparation method is prepared to the acellular artificial lymph fluid drainage tube of preparation, can solve the problem that current lymphatic vessel-venous anastomosis operation is long, operation is difficult, can drain the lymph of patient swelling site to venous system in time after implantation, can quickly relieve swelling symptom.The acellular artificial lymph fluid drainage tube of the application supports the growth, migration, proliferation of autologous cells, and has good biocompatibility, the degradation time can be adjusted to match the reconstruction time of autologous lymphatic return system, to ensure long-term patency.

Description

Technical Field

[0001] This application relates to the fields of medical devices and biomimetic materials technology, and in particular to a decellularized artificial lymphatic drainage tube, its preparation method, and its application. Background Technology

[0002] Lymphedema is a common chronic disease, often caused by cancer surgery, radiotherapy, or infection. It leads to obstructed lymphatic circulation and fluid accumulation in tissues, causing patients to experience limb swelling, functional impairment, and a decreased quality of life. Current treatments for lymphedema include conservative therapies (such as pressure therapy and physical therapy) and surgical treatments. Lymphaticovenous anastomosis (LVA), a minimally invasive surgical procedure, directly connects tiny lymphatic vessels (<0.5 mm in diameter) to veins to improve lymphatic drainage and is widely used for patients with early-stage lymphedema. The advantages of LVA include minimal trauma, rapid recovery, and significant reduction in swelling within days post-surgery. However, its drawbacks include a success rate limited by the functional state of the lymphatic vessels and very high requirements for the anastomosis equipment and techniques.

[0003] Inspired by LVA (Lymphatic Vascular Aqueduct) technology, an ideal artificial lymphatic drainage tube should terminate at an autologous vein to ensure timely and effective return of the continuously generated lymph. The autologous vein can be a primary branch of a large vein in the upper or lower limbs, typically containing valves to prevent backflow of blood into the drainage tube. Furthermore, selecting appropriate materials is crucial for the long-term patency, durable function, and reconstruction of the autologous lymphatic drainage system. Currently, some reports have described the use of synthetic polymer materials for lymphatic drainage (10.1089 / lrb.2018.0042). Polymer-based artificial lymphatic drainage tubes generally possess good bioinertness, helping to reduce immune stimulation. However, polymer materials are typically quite rigid, unable to mimic the contraction and relaxation behavior of the human vascular system, and difficult to anastomose with human blood vessels. In addition, these implanted polymer materials are generally non-degradable, unable to support the reconstruction of the lymphatic drainage system, and carry the risk of long-term inflammatory proliferative blockage. Summary of the Invention

[0004] Therefore, it is necessary to provide a decellularized artificial lymphatic drainage tube. The decellularized artificial lymphatic drainage tube of this application can solve the problems of long operation time and difficult operation of existing lymphatic vessel-venous anastomosis surgery.

[0005] One embodiment of this application provides a decellularized artificial lymphatic drainage tube.

[0006] A method for preparing a decellularized artificial lymphatic drainage tube includes the following steps:

[0007] Animal blood vessels are used as raw materials, and the raw materials are subjected to decellularization, cleaning, enhancement, pore-forming and drying processes.

[0008] In some embodiments, the animal blood vessels include one or more blood vessels from pigs, cattle, sheep, rabbits, rats, fish, and poultry.

[0009] In some embodiments, the animal blood vessels include one or a combination of arterial and venous vessels.

[0010] In some embodiments, the decellularization treatment includes one or a combination of alkali treatment, acid treatment, surfactant treatment, and enzyme treatment.

[0011] In some embodiments, the alkaline treatment includes the following steps: placing the raw material in an alkaline solution with an alkaline concentration of 0.5wt% to 5wt% and decellularizing it at room temperature or -4°C for 0.5h to 12h.

[0012] In some embodiments, the acid treatment includes the following steps: placing the raw material in an acid solution with an acid concentration of 0.1wt% to 0.5wt% and decellularizing it at room temperature or -4°C for 0.5h to 12h.

[0013] In some embodiments, the surfactant treatment includes the following steps: placing the raw material in a surfactant solution with a surfactant concentration of 1wt% to 5wt% and performing decellularization treatment at room temperature or -4°C for 0.5h to 12h.

[0014] In some embodiments, the enzyme treatment includes the following steps: placing the raw material in a pancreatic enzyme solution with a concentration of 0.25wt% to 1.0wt% and decellularizing it at a temperature of 30°C to 35°C for 4 to 24 hours.

[0015] In some embodiments, when the decellularization process includes at least two combinations of alkali treatment, acid treatment, surfactant treatment, and enzyme treatment, several washing steps are interspersed between the decellularization steps.

[0016] In some embodiments, the drying process includes the following step: freeze-drying the cleaned raw materials.

[0017] In some embodiments, the enhancement treatment includes one or a combination of crosslinking treatment and surface coating treatment.

[0018] In some embodiments, the crosslinking treatment includes one or a combination of thermal crosslinking and chemical crosslinking.

[0019] In some embodiments, the thermal crosslinking includes the following steps: freeze-drying the cleaned raw material, and then placing it in a vacuum environment at 60°C to 120°C for crosslinking treatment for 1 hour to 24 hours.

[0020] In some embodiments, the chemical crosslinking includes the following steps: placing the cleaned raw material in a chemical crosslinking agent solution with a concentration of 0.01wt% to 5wt%, and stirring at room temperature or 4°C for 1h to 72h for crosslinking.

[0021] In some embodiments, the chemical crosslinking agent includes one or more combinations of glutaraldehyde, formaldehyde, genipin, butylene glycol glycidyl ether, divinyl sulfone, oxaloyl diazid, carbodiimide, and glycidyl methacrylate.

[0022] In some embodiments, the surface coating treatment includes the following steps: placing the cleaned raw material in a biocompatible polymer solution with a concentration of 0.1wt% to 5wt% and immersing it for 30s to 10min, then removing the raw material and removing the solvent from the biocompatible polymer.

[0023] In some embodiments, the biocompatible polymer includes one or more combinations of chitosan, cellulose, polylactic acid, silk fibroin, collagen, polyvinyl alcohol, polylactic acid-glycolic acid copolymer, polymethyl methacrylate, polyamide, polyurethane, and polycaprolactone.

[0024] In some embodiments, the length of the animal blood vessel is 0.5cm to 50cm.

[0025] In some embodiments, the outer diameter of the animal blood vessel is 0.1 mm to 6 mm.

[0026] In some embodiments, the animal blood vessel includes one of a straight structure and a bifurcated structure.

[0027] One embodiment of this application also provides a decellularized artificial lymphatic drainage tube.

[0028] A decellularized artificial lymphatic drainage tube was prepared using the method described above.

[0029] One embodiment of this application also provides an application of a decellularized artificial lymphatic drainage tube.

[0030] An application of an acellular artificial lymphatic drainage tube in lymphovascular-venous anastomosis, wherein one end of the acellular artificial lymphatic drainage tube can be anastomosed with an autologous vein or its branch, or with another acellular artificial lymphatic drainage tube, and the other end of the acellular artificial lymphatic drainage tube can be anastomosed with an autologous lymphatic vessel, or with another acellular artificial lymphatic drainage tube, or have its opening left in the subcutaneous tissue.

[0031] The aforementioned decellularized artificial lymphatic drainage tube solves the problems of long operation time and difficult operation in existing lymphatic-venous anastomosis surgeries. It can drain lymph from the swollen area to the venous system in a timely manner after implantation, and can quickly relieve the symptoms of swelling. The decellularized artificial lymphatic drainage tube of this application supports the ingrowth, migration and proliferation of autologous cells, has good biocompatibility, and its degradation time can be adjusted to match the reconstruction time of the autologous lymphatic return system, ensuring long-term patency. Specifically, the artificial lymphatic drainage tube of this application with suitable degradation performance that simulates the elasticity of blood vessels is of great significance. The decellularized artificial lymphatic drainage tube based on animal blood vessels not only has excellent biocompatibility, but also retains the elasticity of the blood vessels themselves; its degradation rate can be adjusted according to the degree of enhancement process to match the reconstruction rate of the autologous lymphatic return system, thereby ensuring smooth lymphatic return in the long term. Attached Figure Description

[0032] To more clearly illustrate the technical solutions in the embodiments of this application, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the accompanying drawings described below are only some embodiments of this application. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0033] To gain a more complete understanding of this application and its beneficial effects, the following description will be provided in conjunction with the accompanying drawings. In the following description, the same reference numerals denote the same parts.

[0034] Figure 1 This is a digital photograph of a decellularized artificial lymphatic drainage tube derived from the porcine saphenous artery, as described in one embodiment of this application. Figure 1 It contains a ruler to indicate the length;

[0035] Figure 2 The microstructure of the outer surface of the decellularized artificial lymphatic drainage tube derived from the porcine saphenous artery as described in one embodiment of this application;

[0036] Figure 3 The microstructure of the inner surface of the decellularized artificial lymphatic drainage tube derived from the porcine saphenous artery as described in one embodiment of this application;

[0037] Figure 4This is an H&E stained section of a porcine saphenous artery before decellularization, according to an embodiment of this application.

[0038] Figure 5 for Figure 4 H&E stained section of porcine saphenous artery after decellularization. Detailed Implementation

[0039] To make the above-mentioned objects, features, and advantages of the present invention more apparent and understandable, specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings. Many specific details are set forth in the following description to provide a thorough understanding of the present invention. However, the present invention can be practiced in many other ways different from those described herein, and those skilled in the art can make similar modifications without departing from the spirit of the present invention. Therefore, the present invention is not limited to the specific embodiments disclosed below.

[0040] In the description of this invention, it should be understood that the terms "center," "longitudinal," "lateral," "length," "width," "thickness," "upper," "lower," "front," "rear," "left," "right," "vertical," "horizontal," "top," "bottom," "inner," "outer," "clockwise," "counterclockwise," "axial," "radial," and "circumferential" indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are used only for the convenience of describing this invention and simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on this invention.

[0041] In this invention, unless otherwise explicitly specified and limited, the terms "installation," "connection," "linking," and "fixing," etc., should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral part; they can refer to a mechanical connection or an electrical connection; they can refer to a direct connection or an indirect connection through an intermediate medium; they can refer to the internal communication of two components or the interaction between two components, unless otherwise explicitly limited. Those skilled in the art can understand the specific meaning of the above terms in this invention according to the specific circumstances.

[0042] In this invention, unless otherwise explicitly specified and limited, "above" or "below" the second feature can mean that the first feature is in direct contact with the second feature, or that the first feature is in indirect contact with the second feature through an intermediate medium. Furthermore, "above," "over," and "on top" of the second feature can mean that the first feature is directly above or diagonally above the second feature, or simply that the first feature is at a higher horizontal level than the second feature. "Below," "below," and "under" the second feature can mean that the first feature is directly below or diagonally below the second feature, or simply that the first feature is at a lower horizontal level than the second feature.

[0043] In the description of this invention, "several" means one or more, "multiple" means two or more, "greater than," "less than," and "exceeding" are understood to exclude the stated number, while "above," "below," and "within" are understood to include the stated number. The use of "first" and "second" in the description is merely for distinguishing technical features and should not be construed as indicating or implying relative importance, or implicitly indicating the number of indicated technical features, or implicitly indicating the order of the indicated technical features.

[0044] In this document, "optionally," "optionally," and "optional" mean that something is optional, that is, it is selected from either "with" or "without." If multiple "options" appear in a technical solution, unless otherwise specified and there are no contradictions or mutual constraints, each "option" is independent. In this application, descriptions such as "optionally contains" and "optionally includes" indicate "contains or does not contain."

[0045] In this invention, unless otherwise stated, the sum of the parts of each component in the composition may be 100 parts by weight. Unless otherwise specified, the percentages (including weight percentages) in this invention are based on the total weight of the composition, and "wt%" in this document refers to mass percentage.

[0046] In this application, when numerical intervals (i.e., numerical ranges) are mentioned, unless otherwise specified, the distribution of selectable numerical values ​​within the numerical interval is considered continuous, and includes the two endpoints of the numerical interval (i.e., the minimum and maximum values), as well as every numerical value between these two endpoints. Unless otherwise specified, when a numerical interval refers only to integers within that numerical interval, it includes the two endpoint integers of the numerical range, as well as every integer between the two endpoints, which is equivalent to directly listing every integer. When multiple numerical ranges are provided to describe features or characteristics, these numerical ranges can be merged. In other words, unless otherwise specified, the numerical ranges disclosed in this application should be understood to include any and all subranges included therein. The "numerical value" in the numerical interval can be any quantitative value, such as a number, percentage, ratio, etc. The term "numerical interval" can be broadly included to include percentage intervals, ratio intervals, proportion intervals, etc.

[0047] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The terminology used herein in the description of the invention is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items.

[0048] It should be noted that the room temperature in this article refers to 18℃~40℃, and preferably, the room temperature refers to 20℃~30℃.

[0049] This application provides a decellularized artificial lymphatic drainage tube to address the problem that some existing polymer drainage materials are typically too rigid and cannot simulate the contraction and dilation behavior of the human vascular system. Furthermore, these implanted polymer materials are generally non-degradable, cannot support the reconstruction of the lymphatic return system, and pose a risk of long-term inflammatory proliferative blockage. This application also further solves the problems of long operating times and difficult procedures in current lymphatic-venous anastomosis surgeries. The decellularized artificial lymphatic drainage tube will be described below with reference to the accompanying drawings.

[0050] The decellularized artificial lymphatic drainage tube provided in this application embodiment is exemplary; please refer to [link to example]. Figure 1 As shown, Figure 1 This is a schematic diagram of the structure of the decellularized artificial lymphatic drainage tube provided in an embodiment of this application. The decellularized artificial lymphatic drainage tube of this application can be used for medical surgical purposes, such as lymphatic vessel-vein anastomosis surgery.

[0051] To more clearly illustrate the structure of the decellularized artificial lymphatic drainage tube, the following description, in conjunction with the accompanying drawings, will be provided.

[0052] For example, please refer to Figure 1 As shown, Figure 1 This is a schematic diagram of the decellularized artificial lymphatic drainage tube provided in the embodiments of this application.

[0053] It should be noted that, unless otherwise stated, the reaction steps may be performed in the order described herein or not. For example, other steps may be included between reaction steps, and the order of reaction steps may be appropriately interchanged. This is something that those skilled in the art can determine based on conventional knowledge and experience. Preferably, the reaction methods described herein are performed sequentially.

[0054] A method for preparing a decellularized artificial lymphatic drainage tube includes the following steps:

[0055] Animal blood vessels are used as raw materials, and the raw materials are subjected to decellularization, cleaning, enhancement, pore-making and drying processes.

[0056] In some embodiments, the animal blood vessels include one or more blood vessels from pigs, cattle, sheep, rabbits, rats, fish, and poultry.

[0057] In some embodiments, the animal blood vessels include one or a combination of arterial vessels and venous vessels.

[0058] In some embodiments, the decellularization process includes one or a combination of alkali treatment, acid treatment, surfactant treatment, and enzyme treatment.

[0059] In some embodiments, the alkali treatment includes the following steps: placing the raw material in an alkali solution with an alkali concentration of 0.5wt% to 5wt% and decellularizing it at room temperature for 0.5h to 12h. Preferably, the alkali solution concentration is 0.8wt% to 4wt%. More preferably, the alkali solution concentration is 1.5wt% to 3.5wt%. The decellularization time during alkali treatment includes, but is not limited to, 0.5h, 1h, 2h, 3h, 4h, 4.5h, 5h, 6h, 7h, 8h, 8.5h, 9h, 10h, 11h, 12h, or any range between the foregoing.

[0060] In some embodiments, the alkaline solution includes sodium hydroxide, potassium hydroxide, etc.

[0061] In some embodiments, the acid treatment includes the following steps: placing the raw material in an acid solution with an acid concentration of 0.1wt% to 0.5wt%, and decellularizing it at room temperature or -4°C for 0.5h to 12h. Preferably, the acid solution concentration is 0.2wt% to 0.4wt%. The decellularization time during acid treatment includes, but is not limited to, 0.5h, 1h, 2h, 3h, 4h, 4.5h, 5h, 6h, 7h, 8h, 8.5h, 9h, 10h, 11h, 12h, or any range between the foregoing.

[0062] In some embodiments, the acid solution includes dilute hydrochloric acid, dilute acetic acid, dilute sulfuric acid, etc.

[0063] In some embodiments, the surfactant treatment includes the following steps: placing the raw material in a surfactant solution with a surfactant concentration of 1wt% to 5wt%, and decellularizing it at room temperature or -4°C for 0.5h to 12h. Preferably, the surfactant concentration is 0.8wt% to 4wt%. More preferably, the surfactant concentration is 1.5wt% to 3.5wt%. The decellularization time during surfactant treatment includes, but is not limited to, 0.5h, 1h, 2h, 3h, 4h, 4.5h, 5h, 6h, 7h, 8h, 8.5h, 9h, 10h, 11h, 12h, or any range between the foregoing.

[0064] In some embodiments, the surfactant includes one or more combinations of cetyltrimethylammonium bromide (CTAB), sodium dodecyl sulfate (SDS), Tween 20, Tween 80, Triton X-100, and glyceroglucoside.

[0065] In some embodiments, the enzyme treatment includes the following steps: placing the raw material in a trypsin solution with a concentration of 0.25wt% to 1wt%, and decellularizing it at a temperature of 30°C to 35°C for 4 to 24 hours. Preferably, the trypsin solution concentration is 0.5wt% to 0.8wt%. The decellularization time during enzyme treatment includes, but is not limited to, 4h, 5h, 6h, 7h, 8h, 8.5h, 9h, 10h, 11h, 12h, 14h, 15h, 18h, 20h, 22h, 24h, or any range between the foregoing.

[0066] In some embodiments, the washing process includes the following steps: placing the decellularized raw material in pure water or PBS buffer and washing for 5 to 30 minutes. The washing time may include, but is not limited to, 5 minutes, 6 minutes, 8 minutes, 10 minutes, 12 minutes, 15 minutes, 20 minutes, 25 minutes, 30 minutes, or any range between the two.

[0067] In some embodiments, the raw materials after decellularization are washed at least once during the cleaning process.

[0068] In some embodiments, the drying process includes the following step: freeze-drying the cleaned raw materials.

[0069] In some embodiments, the enhancement treatment includes one or a combination of crosslinking treatment and surface coating treatment.

[0070] In some embodiments, the crosslinking treatment includes one or a combination of thermal crosslinking and chemical crosslinking.

[0071] In some embodiments, thermal crosslinking includes the following steps: freeze-drying the cleaned raw material, and then placing it in a vacuum environment at 60°C to 120°C for crosslinking treatment for 1 hour to 24 hours. The value of the thermal crosslinking time includes, but is not limited to: 1 hour, 6 hours, 10 hours, 18 hours, 24 hours, or any range between the foregoing.

[0072] In some embodiments, chemical crosslinking includes the following steps: placing the cleaned raw material in a chemical crosslinking agent solution with a concentration of 0.01wt% to 0.5wt%, and stirring at room temperature or 4°C for 1h to 72h for crosslinking. The photocrosslinking time includes, but is not limited to, 1h, 5h, 10h, 24h, 36h, 48h, 72h or any range between the foregoing.

[0073] In some embodiments, the chemical crosslinking agent includes one or more combinations of genipin, butylene glycol glycidyl ether, divinyl sulfone, oxalohydrazide, carbodiimide, and glycidyl methacrylate.

[0074] In some embodiments, the surface coating treatment includes the following steps: placing the cleaned raw material in a biocompatible polymer solution with a concentration of 0.1wt% to 5wt% and immersing it for 30s to 10min, then removing the raw material and removing the solvent from the biocompatible polymer.

[0075] In some embodiments, the biocompatible polymer includes one or more combinations of chitosan, cellulose, polylactic acid, silk fibroin, collagen, polyvinyl alcohol, polylactic acid-glycolic acid copolymer, polymethyl methacrylate, polyamide, polyurethane, and polycaprolactone.

[0076] In some embodiments, the length of the animal blood vessel is 0.5cm to 50cm. The length of the animal blood vessel includes, but is not limited to, 0.5cm, 5cm, 10cm, 15cm, 25cm, 30cm, 35cm, 40cm, 50cm or any range between the foregoing.

[0077] In some embodiments, the outer diameter of the animal blood vessel is 0.1 mm to 6 mm. The outer diameter of the animal blood vessel includes, but is not limited to, 0.1 mm, 0.5 mm, 1 mm, 2 mm, 3 mm, 4 mm, 5 mm, 6 mm, or any range between the foregoing.

[0078] In some embodiments, the animal blood vessel includes one of a straight structure and a bifurcated structure.

[0079] One embodiment of this application also provides a decellularized artificial lymphatic drainage tube.

[0080] A decellularized artificial lymphatic drainage tube was prepared using the method described above.

[0081] One embodiment of this application also provides an application of a decellularized artificial lymphatic drainage tube.

[0082] An application of an acellular artificial lymphatic drainage tube in lymphovascular-venous anastomosis, wherein one end of the acellular artificial lymphatic drainage tube can be anastomosed with an autologous vein or its branch, or with another acellular artificial lymphatic drainage tube, and the other end of the acellular artificial lymphatic drainage tube can be anastomosed with an autologous lymphatic vessel, or with another acellular artificial lymphatic drainage tube, or have its opening left in the subcutaneous tissue.

[0083] Example 1

[0084] This embodiment provides a decellularized artificial lymphatic drainage tube.

[0085] The decellularized artificial lymphatic drainage tube in this embodiment was prepared using the following method:

[0086] (1) Obtain a whole piece of connective tissue from the subcutaneous tissue of the pig's hind limb and separate the pig saphenous artery with a length of about 10cm.

[0087] (2) Using porcine saphenous artery as raw material, the obtained porcine saphenous artery was placed in a 2wt% NaOH solution and decellularized at room temperature for 4 hours.

[0088] (3) Then rinse thoroughly with purified water for 5 minutes.

[0089] (4) The cleaned decellularized porcine saphenous artery was then placed in a 0.05wt% glutaraldehyde solution for cross-linking treatment for 6 hours.

[0090] (5) After thoroughly cleaning the cross-linked decellularized cryptal artery, use a needle with an outer diameter of about 0.5 mm to make evenly inserted holes at intervals of about 0.5 cm on the surface of the cross-linked decellularized abdominal aorta.

[0091] (6) Freeze-drying process was used to obtain decellularized artificial lymphatic drainage tubes derived from the porcine saphenous artery. See [link to relevant documentation]. Figure 1 As shown in the attached images. Scanning electron microscopy of the decellularized artificial lymphatic drainage tube derived from the porcine saphenous artery is shown in the appendix. Figure 2 and attached Figure 3 , Figure 2 This is the microstructure of the outer surface of the decellularized artificial lymphatic drainage tube derived from the porcine saphenous artery in this embodiment; Figure 3 The microstructure of the inner surface of the decellularized artificial lymphatic drainage tube derived from the porcine saphenous artery in this embodiment; Figure 4 Image of H&E stained section of material before decellularization; Figure 5 This is an image of an H&E-stained section of decellularized material.

[0092] Example 2

[0093] This embodiment provides a decellularized artificial lymphatic drainage tube.

[0094] The decellularized artificial lymphatic drainage tube in this embodiment was prepared using the following method:

[0095] (1) Euthanized rats were obtained and their abdominal aortic vessels, which were about 10 cm long, were isolated.

[0096] (2) Using abdominal aortic vessels as raw materials, the obtained abdominal aortic vessels were placed in a 0.25wt% pancreatic enzyme solution and decellularized at 33℃ for 4h.

[0097] (3) Then wash thoroughly with PBS buffer for 10 min.

[0098] (4) The cleaned decellularized abdominal aortic vessels were then placed in a 0.01wt% genipin solution and stirred at room temperature for 2 hours for cross-linking.

[0099] (5) After thoroughly rinsing the cross-linked decellularized abdominal aortic vessels with pure water, use a needle with an outer diameter of about 0.5 mm to make evenly spaced holes at intervals of about 0.5 cm on the surface of the cross-linked decellularized abdominal aortic vessels.

[0100] (6) Freeze-drying process to obtain decellularized artificial lymphatic drainage tubes derived from the rat abdominal aorta.

[0101] The decellularized artificial lymphatic drainage tubes in Examples 1 and 2 were subjected to performance tests, and the test results are shown in Table 1.

[0102] Table 1

[0103]

[0104] The in vitro degradation time was determined by immersing the material in a 60 U / mL collagenase solution and recording the time it took for the mass to decrease by more than 95%.

[0105] Cell viability was determined by immersing the material in MEM medium at a ratio of 0.2 g / mL for 72 h and then measuring the 24 h viability of mouse fibroblasts in the extract.

[0106] In summary, the aforementioned decellularized artificial lymphatic drainage tube solves the problems of long operation time and difficult operation in existing lymphatic vessel-venous anastomosis surgeries. It can drain lymph from the swollen area to the venous system in a timely manner after implantation, and can quickly relieve the symptoms of swelling. The decellularized artificial lymphatic drainage tube of this application supports the ingrowth, migration, and proliferation of autologous cells, has good biocompatibility, and its degradation time can be adjusted to match the reconstruction time of the autologous lymphatic return system, ensuring long-term patency. Specifically, the artificial lymphatic drainage tube of this application with suitable degradation performance that simulates the elasticity of blood vessels is of great significance. The decellularized artificial lymphatic drainage tube based on animal blood vessels not only has excellent biocompatibility, but also retains the elasticity of the blood vessels themselves; its degradation rate can be adjusted according to the degree of cross-linking process to match the reconstruction rate of the autologous lymphatic return system, thereby ensuring smooth lymphatic return in the long term.

[0107] In the above embodiments, the descriptions of each embodiment have different focuses. For parts not described in detail in a certain embodiment, please refer to the relevant descriptions in other embodiments.

[0108] The technical features of the above embodiments can be combined in any way. For the sake of brevity, not all possible combinations of the technical features in the above embodiments are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0109] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements all fall within the scope of protection of the present invention. Therefore, the scope of protection of this patent should be determined by the appended claims.

Claims

1. A method for preparing a decellularized artificial lymphatic drainage tube, characterized in that, Includes the following steps: Using animal blood vessels as raw materials, the raw materials are subjected to decellularization, cleaning, enhancement, pore-forming, and drying processes. The decellularization treatment includes at least two combinations of alkali treatment, acid treatment, and enzyme treatment, with several washing steps interspersed between each decellularization step. The alkali treatment includes the following steps: placing the raw material in an alkali solution with an alkali concentration of 0.5wt%-5wt% and decellularizing at room temperature or -4℃ for 0.5h-12h; the acid treatment includes the following steps: placing the raw material in an acid solution with an acid concentration of 0.1wt%-0.5wt% and decellularizing at room temperature or -4℃ for 0.5h-12h; the enzyme treatment includes the following steps: placing the raw material in a trypsin solution with a trypsin concentration of 0.25wt%-1.0wt% and decellularizing at 30℃-35℃ for 4h-24h; the enhancement treatment includes one or a combination of cross-linking treatment and surface coating treatment.

2. The method for preparing the decellularized artificial lymphatic drainage tube according to claim 1, characterized in that, The animal blood vessels include one or more of the blood vessels of pigs, cattle, sheep, rabbits, rats, fish, and poultry.

3. The method for preparing the decellularized artificial lymphatic drainage tube according to claim 1, characterized in that, The animal blood vessels include one or a combination of arterial and venous vessels.

4. The method for preparing the decellularized artificial lymphatic drainage tube according to claim 1, characterized in that, The crosslinking treatment includes one or a combination of thermal crosslinking and chemical crosslinking.

5. The method for preparing the decellularized artificial lymphatic drainage tube according to claim 4, characterized in that, The thermal crosslinking includes the following steps: freeze-drying the cleaned raw material, and then placing it in a vacuum environment at 60℃~120℃ for crosslinking treatment for 1h~24h.

6. The method for preparing the decellularized artificial lymphatic drainage tube according to claim 4, characterized in that, The chemical crosslinking includes the following steps: placing the cleaned raw material in a chemical crosslinking agent solution with a concentration of 0.01wt%~5wt%, and stirring at room temperature or 4°C for 1h~72h for crosslinking. And / or, the chemical crosslinking agent includes one or more combinations of glutaraldehyde, formaldehyde, genipin, butylene glycol glycidyl ether, divinyl sulfone, oxalic acid dihydrazide, carbodiimide, and glycidyl methacrylate.

7. The method for preparing the decellularized artificial lymphatic drainage tube according to claim 1, characterized in that, The surface coating treatment includes the following steps: placing the cleaned raw material in a biocompatible polymer solution with a concentration of 0.1wt% to 5wt% and soaking it for 30s to 10min, then removing the raw material and removing the solvent from the biocompatible polymer.

8. The method for preparing the decellularized artificial lymphatic drainage tube according to claim 7, characterized in that, The biocompatible polymers include one or more combinations of chitosan, cellulose, polylactic acid, silk fibroin, collagen, polyvinyl alcohol, polylactic acid-glycolic acid copolymer, polymethyl methacrylate, polyamide, polyurethane, and polycaprolactone.

9. The method for preparing the decellularized artificial lymphatic drainage tube according to any one of claims 1 to 8, characterized in that, The length of the animal blood vessels ranges from 0.5cm to 50cm; And / or, the outer diameter of the animal blood vessel is 0.1 mm to 6 mm; And / or, the animal blood vessels include one of the following structures: a straight structure and a bifurcated structure.

10. A decellularized artificial lymphatic drainage tube, characterized in that, It is prepared by the preparation method described in any one of claims 1 to 9.

Images