A biodegradable artificial bladder and its preparation method

Abstract

The present invention discloses a biodegradable artificial bladder and a preparation method thereof. The biodegradable artificial bladder comprises a bladder body, ureters located at both sides of the bladder body and a urethra located at one end of the bladder body. The bladder body, ureters and urethra are all three-layer structures, namely an inner layer, a middle layer and an outer layer. In the biodegradable artificial bladder and the preparation method thereof, the artificial bladder has a three-layer structure, and the inner layer and the outer layer are both made of polyvinyl alcohol hydrogel material or a mixed material of polyvinyl alcohol hydrogel and silk protein, and the middle layer is made of acellular matrix material. Since the degradation rate of polyvinyl alcohol hydrogel is relatively slow, after the artificial bladder is implanted into a human body, the duration of cell tissue regeneration can be longer, and the generated cell tissue can be more complete and reliable, which solves the problem that the animal-derived bladder in the prior art loses its structural integrity due to material aging or rapid biodegradation after long-term implantation into a human body.

Description

Technical Field

[0001] The invention belongs to the technical field of medical devices, and in particular relates to a degradable biological artificial bladder and a preparation method thereof. Background Art

[0002] Bladder cancer has become one of the most common malignant tumors in the urogenital system, and its increasing incidence has attracted worldwide attention. The treatment of bladder cancer usually involves total cystectomy, followed by urinary diversion to solve the problem of urine storage and discharge. Currently, the commonly used urinary diversion methods in clinical practice include ureterocutaneous ureterostomy and autologous intestinal bladder replacement.

[0003] Existing patent CN105792777A discloses a biodegradable in situ artificial bladder endoprosthesis, which is intended to replace the human bladder that has lost its normal function due to serious diseases. This technology forms a new bladder by using a shell made of a multilayer silicone membrane and covering it with a layer made of a biocompatible and biodegradable material. However, the preparation of the bladder is difficult, and the acquisition of the endoprosthesis is complex, time-consuming and costly; Existing patent CN110856669A discloses an in situ implantable medical artificial bladder, which consists of an artificial ureter, an artificial bladder body and an artificial urethra, and uses an animal bladder that has been chemically modified to remove the cell components in the original tissue and retain the framework for tissue regeneration. However, after long-term implantation in the human body, the animal-derived bladder loses its structural integrity due to material aging or rapid biodegradation, and there is still a risk of infection and immune rejection; Existing patent CN104717939A discloses an in situ artificial bladder endoprosthesis, which consists of a part that can be connected to the urethra and ureter, and a collapsible cover, and is made of a polymer copolymer. The artificial bladder surgery process may require precise surgical operations, long-term aging of mechanical properties, and high costs. Summary of the invention

[0004] The purpose of the present invention is to solve the problems raised in the background technology and to provide a degradable biological artificial bladder and a preparation method thereof.

[0005] To achieve the above object, the technical solution adopted by the present invention is:

[0006] The present invention proposes a biodegradable artificial bladder, comprising a bladder body, ureters located on both sides of the bladder body and a urethra located at one end of the bladder body. The bladder body, ureters and urethra are all three-layer structures, namely an inner layer, a middle layer and an outer layer, and the inner layer and the outer layer are both made of polyvinyl alcohol hydrogel material or a mixed material of polyvinyl alcohol hydrogel and silk fibroin, and the middle layer is made of a decellularized matrix material.

[0007] Preferably, the thickness of the inner layer is 0.5-3 mm, the thickness of the outer layer is 0-1 mm, the thickness of the middle layer is 0-1 mm, and the volume of the bladder body is 30-500 mL.

[0008] Preferably, the mixed material is 8-12% polyvinyl alcohol hydrogel and 6-15% silk fibroin mixed in a ratio of 1:4, 2:3, 1:1, 3:2 or 4:1.

[0009] Preferably, the port of the ureter away from the bladder body is a two-layer structure including an inner layer and a middle layer.

[0010] Preferably, the inner layer and the outer layer are both provided with regenerative cells, and the ports of the ureter and urethra away from the bladder body are coated with growth factors.

[0011] A method for preparing a biodegradable artificial bladder, comprising:

[0012] Injecting polymer material into the first mold in the shape of an artificial bladder, cooling it down in an environment of 2-8°C to form it, and demolding it to obtain an inner cavity support body;

[0013] The prepared inner cavity support body is placed in a second mold in the shape of an artificial bladder, and then a polyvinyl alcohol hydrogel solution or a mixed solution of the prepared polyvinyl alcohol hydrogel and silk fibroin is injected into the gap between the inner wall of the second mold and the inner cavity support body, and then frozen at <-40°C for 8-24 hours, and then warmed to room temperature for 8-12 hours, and repeated 3-6 times, and then the second mold is removed to obtain a first structure with an inner layer structure of polyvinyl alcohol hydrogel material or mixed material with an inner cavity support body;

[0014] Obtain a sheet of decellularized matrix and coat the surface of the inner layer structure in the first structure with distilled water or polyvinyl alcohol hydrogel to obtain a second structure with an inner layer structure of an inner cavity support body and a decellularized matrix middle layer structure;

[0015] Coating polyvinyl alcohol hydrogel on the surface of the acellular matrix middle layer structure in the second structure, then placing the coated second structure into a third mold in the shape of an artificial bladder, injecting a polyvinyl alcohol hydrogel solution or a prepared mixed solution of polyvinyl alcohol hydrogel and silk fibroin into the gap between the inner wall of the third mold and the second structure, freezing at <-40°C for 8-24 hours, then warming up at room temperature for 8-12 hours, repeating 3-6 times, and then removing the third mold to obtain an artificial bladder with an inner layer structure of an inner cavity support body and a decellularized matrix middle layer structure, and an outer layer structure of polyvinyl alcohol hydrogel material or mixed material;

[0016] The artificial bladder with the inner cavity support body is placed in a container and placed in a constant temperature water bath at 37±3°C until the inner cavity support body is completely dissolved to obtain the artificial bladder without the inner cavity support body;

[0017] Finally, the artificial bladder is sterilized by irradiation to complete the preparation of the artificial bladder.

[0018] Preferably, each of the molds includes two relatively clamped half shells, and the chamber formed by the two half shells includes a first chamber that is interconnected and in the shape of a bladder body, two second chambers in the shape of ureters, and a third chamber in the shape of a urethra, and the two second chambers are respectively located on both sides of the first chamber, and the third chamber is located at one end of the first chamber.

[0019] Preferably, before injecting the polymer material into the first mold in the shape of the artificial bladder, a detachable stent is placed into the first mold so that the stent is fixed in the inner cavity support body;

[0020] After removing the artificial bladder from the inner cavity support body, the stent is taken out from the artificial bladder;

[0021] Wherein, the bracket comprises a straight rod, a first branch rod and a second branch rod matched with each mold, the straight rod is provided with two placement grooves, each of the branch rods comprises two first cylindrical rods and a second cylindrical rod hinged to each other, one end of each of the first cylindrical rods away from the second cylindrical rod is provided with a latch corresponding to the placement groove, each of the second cylindrical rods is provided with a first limiting sleeve, and one end of each of the second cylindrical rods away from the first cylindrical rod and one end of the straight rod are both provided with a second limiting sleeve;

[0022] The second chamber is provided with a groove corresponding to the first limiting sleeve;

[0023] The straight rod extends from the third cavity of each mold into the first cavity, and is limited by the second limiting sleeve located outside the third cavity. The first branch and the second branch extend from the two second cavities of each mold into the first cavity respectively, and the two latches are inserted into the two placement grooves one by one, and the first branch and the second branch are both limited by the corresponding second limiting sleeve located outside the second cavity and the first limiting sleeve located in the groove.

[0024] The length of the second cavity in the third mold is smaller than the length of the second cavity in the first mold and the second mold.

[0025] Preferably, the preparation process of the mixed solution of polyvinyl alcohol hydrogel and silk fibroin comprises:

[0026] Add 6-15% silk fibroin to 8-12% polyvinyl alcohol hydrogel, and stir the two in a ratio of 1:4, 2:3, 1:1, 3:2 or 4:1 at 25°C±5, then place in a vacuum drying oven to remove bubbles to form a uniform and bubble-free mixed solution of polyvinyl alcohol hydrogel and silk fibroin.

[0027] Preferably, the prepared artificial bladder comprises a bladder body, ureters located on both sides of the bladder body, and a urethra located at one end of the bladder body;

[0028] Before the artificial bladder is sterilized by irradiation, growth factors are applied to the ports of the ureter and urethra in the artificial bladder that are away from the bladder body, and regenerative cells are seeded on the inner and outer structures of the artificial bladder.

[0029] The method for preparing the biodegradable artificial bladder further comprises: after obtaining the first structure, using glycerol or propylene glycol to harden and modify the polyvinyl alcohol hydrogel at the ureteral port and the urethral port in the inner layer structure of the first structure.

[0030] Compared with the prior art, the present invention has the following beneficial effects:

[0031] 1. The biodegradable biological artificial bladder and its preparation method have a three-layer structure, and the inner layer and the outer layer are both made of polyvinyl alcohol hydrogel or a mixed material of polyvinyl alcohol hydrogel and silk fibroin, and the middle layer is made of acellular matrix. Since the degradation rate of polyvinyl alcohol hydrogel is relatively slow, after the artificial bladder is implanted into the human body, the duration of cell tissue regeneration can be longer and faster, and the generated cell tissue can be more complete and reliable, which solves the problem that the animal-derived bladder in the prior art loses its structural integrity due to material aging or rapid biodegradation after long-term implantation in the human body; the polyvinyl alcohol hydrogel and the acellular matrix are both degradable, so that the material of the artificial bladder will not always be in the human body; at the same time, the silk fibroin can improve the tensile strength of polyvinyl alcohol hydrogel, which is more convenient for subsequent surgical suturing;

[0032] 2. In the process of preparing the artificial bladder, the artificial bladder is formed in one piece with the help of three molds, and the entire operation process is convenient and simple, thereby improving the preparation efficiency; the port of the ureter of the artificial bladder is set into a two-layer structure, and the acellular matrix material is used as the outer layer to directly contact the ureter of the human body, which better promotes cell differentiation and growth;

[0033] 3. By inoculating regenerative cells on the surface of the inner and outer layers, cell differentiation and growth are promoted, thereby better promoting the regeneration of the middle layer tissue, so that the artificial bladder can be connected and anastomosed with the human body. By applying growth factors at the ports of the two ureters and the ports of the ureters, cell regeneration is accelerated, so that the artificial bladder can be anastomosed with the corresponding parts of the human body. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 is a first right-angle schematic diagram of the artificial bladder of the present invention;

[0035] Figure 2 is a second right-angle schematic diagram of the artificial bladder of the present invention;

[0036] Figure 3 It is a schematic diagram of the first mold in the degradable bio-artificial bladder and the preparation method thereof of the present invention;

[0037] Figure 4 is a schematic diagram of a second mold of the present invention;

[0038] Figure 5 is a schematic diagram of a third mold of the present invention;

[0039] Figure 6 It is a side schematic diagram of each mold in the present invention;

[0040] Figure 7 It is a schematic diagram of the support and each mold structure of the present invention;

[0041] Figure 8 is a schematic diagram of the assembled support of the present invention;

[0042] Fig. 9 This is a schematic diagram of the bracket of the present invention after being disassembled. DETAILED DESCRIPTION

[0043] In order to make the purpose, technical solution and advantages of the present application more clearly understood, the present application is further described in detail below in conjunction with the accompanying drawings and embodiments. It should be understood that the specific embodiments described herein are only used to explain the present application and are not used to limit the present application.

[0044] In one embodiment, Figure 1-Figure 2 As shown, a degradable bio-artificial bladder is provided, including a bladder body, ureters located on both sides of the bladder body and a urethra located at one end of the bladder body. The bladder body, ureters and urethra are all three-layer structures, namely an inner layer, a middle layer and an outer layer, and the inner layer and the outer layer are both made of polyvinyl alcohol hydrogel material or a mixed material of polyvinyl alcohol hydrogel and silk fibroin, and the middle layer is made of acellular matrix material.

[0045] It should be noted that the acellular matrix is ​​derived from cattle or pigs, including but not limited to cattle tendons, pericardium or pig intestines, etc. In addition, one or more of the same concentration can be used in combination. Figure 1-Figure 2 In the figure, c represents the ureter, d represents the urethra, and e represents the bladder proper.

[0046] In this embodiment, the thickness of the inner layer is 0.5-3 mm (eg, 2 mm), the thickness of the outer layer is 0-1 mm (eg, 0.5 mm), the thickness of the middle layer is 0-1 mm (eg, 0.5 mm), and the volume of the bladder body is 30-500 mL (eg, 320 mL).

[0047] In this embodiment, when both the inner layer and the outer layer are made of polyvinyl alcohol hydrogel material, the polyvinyl alcohol hydrogel material is a polyvinyl alcohol hydrogel with a concentration of 8-12% (such as a concentration of 10% for the polyvinyl alcohol hydrogel);

[0048] When both the inner layer and the outer layer are a mixture of polyvinyl alcohol hydrogel and silk fibroin, the mixed material is 8-12% polyvinyl alcohol hydrogel and 6-15% silk fibroin mixed in a ratio of 1:4, 2:3, 1:1, 3:2 or 4:1; for example, if the concentration of the polyvinyl alcohol hydrogel is 10% and the concentration of the silk fibroin is 8%, and they are mixed in a ratio of 2:3, the silk fibroin can enhance the tensile strength of the polyvinyl alcohol hydrogel, making it easier to sew with human tissues.

[0049] In this embodiment, the port of the ureter away from the bladder body is a two-layer structure, including an inner layer and an intermediate layer. It should be noted that during the use of the artificial bladder, the two-layer structure of the two ureters of the artificial bladder is sleeved and sutured with the ureters of the human body (the ureters of the human body are sleeved on the outside of the port of the ureter of the artificial bladder, and then sutured), and the port of the ureter of the artificial bladder is set to a two-layer structure, and the decellularized matrix material is directly in contact with the ureters of the human body as the outer layer, which better promotes cell differentiation and growth.

[0050] In this embodiment, regenerative cells are provided on both the inner layer and the outer layer (regenerative cells are seeded on the surfaces of both the inner layer and the outer layer); it should be noted that by seeding the surfaces of the inner layer and the outer layer with regenerative cells, cell differentiation and growth are promoted, thereby better promoting the regeneration of the middle layer tissue, so that the artificial bladder is connected and matched with the human body.

[0051] In this embodiment, the ports of the ureter and urethra away from the bladder body are coated with growth factors; the regenerative cells may be stem cells, urothelial cells, etc., and the growth factors may be one or more of fibroblast growth factor, platelet-derived growth factor, and epidermal growth factor. It should be noted that during the use of the artificial bladder, the two-layer structure of the two ureters of the artificial bladder is sutured with the ureters of the human body through surgery, and the port of the urethra of the artificial bladder away from the bladder body is sutured with the urethra of the human body. By applying growth factors at the ports of the two ureters and the ports of the ureters, cell regeneration is accelerated, so that the artificial bladder is matched with the corresponding part of the human body. The traction of the polyvinyl alcohol hydrogel material is good and easy to sew. The traction of the polyvinyl alcohol hydrogel material is further enhanced by silk fibroin.

[0052] The artificial bladder has two layers of polyvinyl alcohol hydrogel, an inner layer and an outer layer. Since the polyvinyl alcohol hydrogel degrades slowly, after the artificial bladder is implanted in the human body, the cell tissue regeneration can last longer, making the generated cell tissue more complete and reliable. The polyvinyl alcohol hydrogel and the decellularized matrix are both degradable (the polyvinyl alcohol hydrogel degrades more slowly), so that the material of the artificial bladder will not always remain in the human body.

[0053] In another embodiment, Figure 1-Figure 9 As shown, a method for preparing a degradable bio-artificial bladder is disclosed based on the degradable bio-artificial bladder, comprising:

[0054] S1, placing a detachable bracket into a first mold;

[0055] Among them, since the bracket needs to be placed in three molds in the subsequent process, the way in which the bracket cooperates with each mold is described here. Figure 3-Figure 5 The figure shows the schematic diagram of each mold after being disassembled, where the first mold is represented by A, the second mold is represented by B, and the third mold is represented by C) and includes two relatively clamped half shells (such as Figure 3-Figure 5 As shown, one of the half shells is provided with a preset number of plugs a, such as three plugs, and the other half shell is provided with through holes b corresponding to the plugs, and the plugs are inserted into the through holes one by one for clamping, so as to realize the assembly between the two half shells, and as shown Figure 6As shown, the side of the half shell with the plug is an arc surface, and the side of the half shell with the through hole is a plane surface). The chambers formed by the two half shells include a first chamber that is interconnected and in the shape of a bladder body (as shown in Figure 1, one end of the bladder body is hemispherical and the other end is truncated, or it can be spherical, olive-shaped, etc. without limitation), two second chambers in the shape of ureters (ureters are tubular structures) and a third chamber in the shape of a urethra (urethra is tubular structure), and the two second chambers are respectively located on both sides of the first chamber, and the third chamber is located at one end of the first chamber (the two ureters are located on both sides of the truncated cone end of the bladder body, the urethra is located at the end of the truncated cone end of the bladder body, and the two ureters, the urethra and the bladder body are interconnected). The capacity of the three molds is determined according to the thickness of the layer actually prepared, and there is no limitation. The capacity of the first mold is smaller than that of the second mold, and the capacity of the second mold is smaller than that of the third mold.

[0056] Among them, Figure 8-Figure 9 As shown, the bracket includes a straight rod, a first branch rod and a second branch rod (the straight rod is represented by f, the first branch rod is represented by g, and the second branch rod is represented by h) that cooperate with each mold, and two placement grooves are provided on the straight rod (the placement groove is represented by i), and each branch rod includes two first cylindrical rods and a second cylindrical rod that are hinged to each other, and an end of each first cylindrical rod away from the second cylindrical rod is provided with a latch corresponding to the placement groove (the latch is integrally formed with the first cylindrical rod, and the latch is represented by j), and a first limiting sleeve is provided on each second cylindrical rod (the first limiting sleeve can be integrally formed with the second cylindrical rod, or a split design can be adopted, and the first limiting sleeve is represented by k), and a second limiting sleeve is provided on one end of each second cylindrical rod away from the first cylindrical rod and one end of the straight rod (the second limiting sleeve is integrally formed with the corresponding rod, or a split design can be adopted, and the second limiting sleeve is located outside the corresponding cavity to limit each rod to prevent each rod from completely entering the mold, and the second limiting sleeve is represented by l); as shown Figure 7 , which is a schematic diagram of the cooperation between the bracket and each mold, wherein l in the figure represents the second limiting sleeve, and the bracket pointing upward in the figure is the second limiting sleeve on the straight rod, the bracket pointing to the lower left is the second limiting sleeve on the first branch rod, and the bracket pointing to the lower right is the second limiting sleeve on the second branch rod;

[0057] The second chamber is provided with a groove corresponding to the first limiting sleeve (such as Figure 3-Figure 5 , the groove is represented by m);

[0058] The straight rod extends from the third cavity of each mold into the first cavity, and is limited by the second limiting sleeve located outside the third cavity. The first branch and the second branch extend from the two second cavities of each mold into the first cavity respectively, and the two pins are inserted into the two placement grooves one by one, and the first branch and the second branch are both limited by the corresponding second limiting sleeve located outside the second cavity and the first limiting sleeve located in the groove (the first limiting sleeve in each branch rod cooperates with the groove, so that the pin of each branch rod can be stably located in the placement groove and will not fall off).

[0059] It should be noted that, when placing the bracket into the first mold, the first mold can be disassembled first, and then the bracket is matched with one of the half shells, and then the other half shell is covered on the half shell to place the bracket in the first mold; after injecting the polymer material into the first mold, an inner cavity support body with the bracket is obtained (the bracket is fixed in the inner cavity support body); in the subsequent second mold, the inner cavity support body with the bracket is matched with one of the half shells of the second mold, and then the other half shell is covered on the half shell to place the inner cavity support body with the bracket in the second mold; in the third mold, the second structure (the second structure contains the bracket at this time) is matched with one of the half shells of the second mold, and then the other half shell is covered on the half shell to place the second structure with the bracket in the third mold. Each mold is made of transparent or translucent material, which can be a resin material.

[0060] S2, injecting polymer material into the first mold in the shape of an artificial bladder and into which the stent is placed (the injection can be performed using a syringe), cooling and molding in an environment of 2-8°C (such as cooling and molding in a refrigerator at 3°C), demolding to obtain an inner cavity support body, and fixing the stent in the inner cavity support body;

[0061] The polymer material injected into the first mold may be one or more of gelatin, liposomes (such as DPPC), polyethers (such as Pluronic F127), copolymers (such as poloxamer 407), etc., and the obtained inner cavity support body is a degradable or easily soluble material. The support makes the inner cavity support body more stable and prevents the polymer material from collapsing.

[0062] S3. Preparing a mixed solution of polyvinyl alcohol hydrogel and silk fibroin:

[0063] Add 6-15% silk fibroin to 8-12% polyvinyl alcohol hydrogel (e.g., the concentration of polyvinyl alcohol hydrogel is 10% and the concentration of silk fibroin is 8%), and stir the two at a ratio of 1:4, 2:3, 1:1, 3:2 or 4:1 at 25°C±5 (e.g., stir the two at a ratio of 2:3 at 25°C), then place in a vacuum drying oven to remove bubbles (until a uniform and bubble-free solution is formed), thereby forming a uniform and bubble-free mixed solution of polyvinyl alcohol hydrogel and silk fibroin.

[0064] S4. Place the prepared inner cavity support body into a second mold in the shape of an artificial bladder, and then inject a polyvinyl alcohol hydrogel solution (with a concentration of 8%-12%, such as 10%) or a mixed solution of the prepared polyvinyl alcohol hydrogel and silk fibroin into the gap between the inner wall of the second mold and the inner cavity support body, freeze at <-40°C for 8-24h, then warm up at room temperature for 8-12h, repeat 3-6 times (such as freezing at -40°C for 16h, then warming up at room temperature for 10h, repeat 5 times), and then remove the second mold to obtain a support body with an inner cavity. A first structure body with an inner layer structure of polyvinyl alcohol hydrogel material or a mixed material, and then the polyvinyl alcohol hydrogel at the ureteral port and urethral port in the inner layer structure of the first structure body is hardened and modified with glycerol (or propylene glycol, etc.) for standby use (the hardening modification makes it easier to connect and suture the ureteral port (or urethral port) of the artificial bladder with the human ureter (urethra) during subsequent surgery, and the human body's own ureter (or urethra) is more easily attached to the surface of the material, and the strength is increased after suturing, the toughness of the material is increased, and the operation is more convenient).

[0065] S5, obtaining a sheet of decellularized matrix and coating the surface of the inner layer structure in the first structure with distilled water or polyvinyl alcohol hydrogel (the concentration of the polyvinyl alcohol hydrogel may be 8-12%, such as 10%) to obtain a second structure having an inner layer structure with an inner cavity support body and a decellularized matrix middle layer structure;

[0066] Among them, the decellularized matrix in this embodiment adopts the lower mucosal matrix of the small intestine of pigs, and the lower mucosal matrix of the small intestine of pigs is divided into a preset number of sheets (of any size) with uniform thickness, and the sheet-like decellularized matrix is ​​coated on the surface of the inner layer structure in the first structure by distilled water or polyvinyl alcohol hydrogel. The specific number of coated sheet-like decellularized matrix is ​​set according to actual needs, and the specific number of coated layers can be one layer or multiple layers. The coated decellularized matrix constitutes a decellularized matrix intermediate layer structure.

[0067] S6. Coating polyvinyl alcohol hydrogel on the surface of the acellular matrix intermediate layer structure in the second structure (the concentration of the polyvinyl alcohol hydrogel can be 8-12%, such as 10%, to prevent the second structure from being dispersed by the subsequent injection of polyvinyl alcohol hydrogel solution or the prepared mixed solution of polyvinyl alcohol hydrogel and silk fibroin into the third mold, and to make the coated polyvinyl alcohol hydrogel and the polyvinyl alcohol hydrogel in the mixed solution injected later fit more firmly), and then put the coated second structure into the third mold in the shape of an artificial bladder, and align the inner wall of the third mold with the second structure. A polyvinyl alcohol hydrogel solution (such as a polyvinyl alcohol hydrogel concentration of 10% and a silk fibroin concentration of 8%) or a prepared mixed solution of polyvinyl alcohol hydrogel and silk fibroin is injected into the gap between the two molds, frozen at <-40°C for 8-24 hours, then warmed up at room temperature for 8-12 hours, and repeated 3-6 times (such as freezing at -40°C for 16 hours, then warming up at room temperature for 10 hours, and repeated 5 times), and then the third mold is removed to obtain an artificial bladder with an inner layer structure of an inner cavity support body, a middle layer structure of acellular matrix, and an outer layer structure of polyvinyl alcohol hydrogel material or a mixed material.

[0068] S7. Place the artificial bladder with the inner cavity support body into a container (such as a beaker), and place it in a constant temperature water bath at 37±3℃ (such as 37℃), and after the inner cavity support body is completely dissolved, obtain the artificial bladder without the inner cavity support body; after the artificial bladder without the inner cavity support body, remove the stent from the artificial bladder, and then obtain a separate artificial bladder (when removing the stent, directly pull out the two branches, and then pull out the straight rod, that is, remove the stent); the prepared artificial bladder includes the bladder body, the ureters located on both sides of the bladder body, and the urethra located at one end of the bladder body (such as Figure 1-Figure 2 FIG. 1 shows an artificial bladder, wherein c represents the ureter, d represents the urethra, and e represents the bladder body. Figure 2 As shown, one side of the bladder body is curved and the other side is flat).

[0069] S8. Growth factors are applied to the ports of the ureter and urethra in the artificial bladder that are respectively away from the bladder body (i.e., growth factors are applied to the ports of the ureter and urethra), and regenerative cells are inoculated on the inner and outer structures of the artificial bladder (i.e., regenerative cells are inoculated on the surface of the inner structure (i.e., the inner wall and end of the inner structure) and the surface of the outer structure (i.e., the outer wall and end of the outer structure)).

[0070] S9. Finally, the artificial bladder is sterilized by irradiation to complete the preparation of the artificial bladder (such as sterilization by electron beam irradiation and then stored at 0-5°C, such as stored at 0°C). The thickness of the inner layer structure of the artificial bladder is 0.5-3mm (such as 2mm), the thickness of the outer layer structure is 0-1mm (such as 0.5mm), the thickness of the middle layer structure of the decellularized matrix is ​​0-1mm (such as 0.5mm), and the volume of the bladder body in the artificial bladder is 30-500mL (such as 320mL).

[0071] In this embodiment, the length of the second cavity in the third mold is smaller than the length of the second cavity in the first mold and the second mold respectively; wherein the length of the second cavity in the first mold is substantially equal to the length of the second cavity in the second mold.

[0072] It should be noted that the length of the second chamber in the third mold is smaller than the length of the second chamber in the first mold, and the length of the second chamber in the second mold is smaller than the length of the second chamber in the first mold, so that the end of the ureter of the artificial bladder away from the bladder body has only two layers of structure (i.e., the inner layer structure and the decellularized matrix middle layer structure).

[0073] In the process of preparing the artificial bladder, the artificial bladder is formed in one piece with the help of three molds. The whole operation process is convenient and simple, thereby improving the preparation efficiency.

[0074] The process of implanting the artificial bladder of the present invention into the human body is as follows:

[0075] Under general anesthesia, the patient is subjected to total cystectomy. After complete hemostasis, the artificial bladder of the present invention is implanted in the original pelvic cavity. After adjusting the appropriate position, one end of the commonly used 5Fr ureteral stent in clinical practice is placed along the artificial bladder ureteral port, and then the artificial bladder ureteral port is anastomosed with the human ureteral stump. It should be noted that the normal ureteral mucosa is sleeved on the middle layer of the two-layer structure of the artificial ureter, and the human ureteral mucosa and the middle layer of the artificial ureter are sutured with absorbable sutures, and then the outer layer of the artificial bladder ureter (this outer layer is the outer layer of the three-layer structure of the ureter) and the human ureteral serosal layer are sutured with silk thread, and the contralateral artificial ureter is anastomosed in the same way. An 18Fr double-lumen catheter is inserted through the urethra and passes through the anastomotic opening of the human urethral stump and the artificial urethral port, and the artificial urethral port is anastomosed with the human urethral stump with silk thread. After the artificial bladder is implanted, the abdominal cavity is closed layer by layer to complete the operation.

[0076] The above-mentioned embodiments only express several implementation methods of the present application, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for a person of ordinary skill in the art, several variations and improvements can be made without departing from the concept of the present application, and these all belong to the protection scope of the present application. Therefore, the protection scope of the patent of the present application shall be subject to the attached claims.

Claims

1. A method for preparing a biodegradable artificial bladder, characterized in that: The preparation method of the degradable biological artificial bladder comprises: Injecting polymer material into the first mold in the shape of an artificial bladder, cooling it down in an environment of 2-8°C to form it, and demolding it to obtain an inner cavity support body; The prepared inner cavity support body is placed in a second mold in the shape of an artificial bladder, and then a polyvinyl alcohol hydrogel solution or a mixed solution of the prepared polyvinyl alcohol hydrogel and silk fibroin is injected into the gap between the inner wall of the second mold and the inner cavity support body, and then frozen at <-40°C for 8-24 hours, and then warmed to room temperature for 8-12 hours, and repeated 3-6 times, and then the second mold is removed to obtain a first structure with an inner layer structure of polyvinyl alcohol hydrogel material or mixed material with an inner cavity support body; Obtain a sheet of decellularized matrix and coat the surface of the inner layer structure in the first structure with distilled water or polyvinyl alcohol hydrogel to obtain a second structure with an inner layer structure of an inner cavity support body and a decellularized matrix middle layer structure; Coating polyvinyl alcohol hydrogel on the surface of the acellular matrix middle layer structure in the second structure, then placing the coated second structure into a third mold in the shape of an artificial bladder, injecting a polyvinyl alcohol hydrogel solution or a prepared mixed solution of polyvinyl alcohol hydrogel and silk fibroin into the gap between the inner wall of the third mold and the second structure, freezing at <-40°C for 8-24 hours, then warming up at room temperature for 8-12 hours, repeating 3-6 times, and then removing the third mold to obtain an artificial bladder with an inner layer structure of an inner cavity support body and a decellularized matrix middle layer structure, and an outer layer structure of polyvinyl alcohol hydrogel material or mixed material; The artificial bladder with the inner cavity support body is placed in a container and placed in a constant temperature water bath at 37±3°C until the inner cavity support body is completely dissolved to obtain the artificial bladder without the inner cavity support body; Finally, the artificial bladder is sterilized by irradiation to complete the preparation of the artificial bladder.

2. The method for preparing a degradable bio-artificial bladder according to claim 1, characterized in that: Each of the molds includes two relatively clamped half shells, and the chamber formed by the two half shells includes a first chamber that is interconnected and in the shape of a bladder body, two second chambers in the shape of ureters, and a third chamber in the shape of a urethra, and the two second chambers are respectively located on both sides of the first chamber, and the third chamber is located at one end of the first chamber.

3. The method for preparing the degradable bio-artificial bladder according to claim 2, characterized in that: Before injecting the polymer material into the first mold in the shape of an artificial bladder, placing a detachable stent into the first mold so that the stent is fixed in the inner cavity support body; After removing the artificial bladder from the inner cavity support body, the stent is taken out from the artificial bladder; Wherein, the bracket comprises a straight rod, a first branch rod and a second branch rod matched with each mold, the straight rod is provided with two placement grooves, each of the branch rods comprises two first cylindrical rods and a second cylindrical rod hinged to each other, one end of each of the first cylindrical rods away from the second cylindrical rod is provided with a latch corresponding to the placement groove, each of the second cylindrical rods is provided with a first limiting sleeve, and one end of each of the second cylindrical rods away from the first cylindrical rod and one end of the straight rod are both provided with a second limiting sleeve; The second chamber is provided with a groove corresponding to the first limiting sleeve; The straight rod extends from the third cavity of each mold into the first cavity, and is limited by the second limiting sleeve located outside the third cavity. The first branch and the second branch extend from the two second cavities of each mold into the first cavity respectively, and the two latches are inserted into the two placement grooves one by one, and the first branch and the second branch are both limited by the corresponding second limiting sleeve located outside the second cavity and the first limiting sleeve located in the groove. The length of the second cavity in the third mold is smaller than the length of the second cavity in the first mold and the second mold.

4. The method for preparing a degradable bio-artificial bladder according to claim 1, characterized in that: The concentration of the polyvinyl alcohol hydrogel is 8-12%; The preparation process of the mixed solution of polyvinyl alcohol hydrogel and silk fibroin comprises: Add 6-15% silk fibroin to 8-12% polyvinyl alcohol hydrogel, and stir the two in a ratio of 1:4, 2:3, 1:1, 3:2 or 4:1 at 25°C±5, then place in a vacuum drying oven to remove bubbles to form a uniform and bubble-free mixed solution of polyvinyl alcohol hydrogel and silk fibroin.

5. The method for preparing the degradable bio-artificial bladder according to claim 1, characterized in that: The prepared artificial bladder includes a bladder body, ureters located on both sides of the bladder body, and a urethra located at one end of the bladder body; Before the artificial bladder is sterilized by irradiation, growth factors are applied to the ports of the ureter and urethra in the artificial bladder that are away from the bladder body, and regenerative cells are seeded on the inner and outer structures of the artificial bladder. The method for preparing the biodegradable artificial bladder further comprises: after obtaining the first structure, using glycerol or propylene glycol to harden and modify the polyvinyl alcohol hydrogel at the ureteral port and the urethral port in the inner layer structure of the first structure.

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