Biological adhesive patch for layered induced tissue regeneration of digestive tract

Abstract

The present invention provides a biological adhesive patch for layered induced tissue regeneration of the digestive tract, comprising a bottom layer and a top layer wherein the bottom layer comprises a core portion in direct contact with a wound surface and a peripheral portion surrounding the core portion to close a wound; the size and shape of the core part of the bottom layer correspond to the size and shape of a wound, the core part of the bottom layer induces tissue regeneration and is prepared from a chiral hydrogel composition, and the core part of the bottom layer comprises a first layer formed on the top layer and a second layer formed on the first layer, the first layer comprises gastric primary smooth muscle cells and the second layer comprises gastric epithelial organoid. The patch has the effect of dual induction of tissue regeneration, can realize inflammatory reaction inhibition, tissue filling growth and blood vessel induction growth at the tension wound defect part, and completes three-dimensional multi-layer repair of the digestive tract tension wound.

Description

Technical Field

[0001] The present disclosure relates to a biological patch, and more particularly, to a bioadhesive patch for layered induction of tissue regeneration of the digestive tract and capable of being degraded in vivo. Background Art

[0002] The clinical incidence of tension wounds in the digestive tract is relatively high, mostly caused by full-thickness defects of the digestive tract. Tension wounds are subjected to high stress that opens to both sides, making wound healing difficult and prone to infection, especially in the esophageal area. Considering the complex anatomical structures of the surrounding chest cavity, trachea, mediastinum, etc., traditional surgical treatment is difficult and has many complications.

[0003] The causes of tension wounds include trauma, iatrogenic, and pathological. Traumatic wounds are irregular in shape and often contaminated; iatrogenic wounds are mainly surgical incisions and perforations, such as local lesion excision, scar release, etc.; pathological wounds are mostly caused by ulcers caused by various reasons that gradually worsen, and their occurrence time is relatively slow.

[0004] Clinically, the methods for treating tension wounds in the digestive tract are quite limited. For wounds with larger defects, tension-reducing sutures are generally used, but this method is very traumatic and changes the geometric structure of the digestive tract, which is closely related to functions such as digestive tract motility and nerve conduction. For wounds with smaller defects, endoscopic stenting or endoscopic high-tension OTSC (over the scope clipping) closure is generally used, but endoscopic stenting is highly dependent on the spreading and healing ability of the digestive tract tissue itself and is prone to displacement. Endoscopic OTSC can be used to treat digestive tract fistulas with tension characteristics. After release, the entire layer of the digestive tract can be clamped and can be used to close fistulas of 1-2 cm within 7 days, but OTSC is expensive, cumbersome to operate, and has a short treatment window.

[0005] The healing characteristics of tension wounds determine the high clinical standards. First, infection factors run through the entire process of wound healing. Second, the true three-dimensional repair of tension wounds must include not only the repair of the defect structure level, but also the repair of functional levels such as microvascular regeneration and layered induced tissue regeneration.

[0006] Wound healing is a complex pathophysiological process that requires the coordinated action of multiple cell types to maintain tissue integrity and homeostasis. The disordered cell behavior caused by trauma can lead to healing impairment and the formation of chronic non-healing wounds. The main pathological change after trauma is inflammation, which causes the wound to persist through continuous damage to the wound surface. There are a large number of Langerhans cells, neutrophils, proinflammatory macrophages and proteases in the wound surface, which are related to the severity of clinical ulcers. With the increase in the infiltration of specific immune cell subsets, the function of pathological immune cells is disturbed, and these factors work together to lead to poor healing. Excessive activation of neutrophils produces neutrophil extracellular traps, which in turn leads to cytotoxicity and delayed wound healing.

[0007] The wound healing process can be divided into the coagulation phase, the inflammatory phase, the repair phase, and the maturation phase. A key activity in the proliferation phase is the formation of new blood vessels. In an intact tissue, the microvascular system maintains homeostasis, allowing sufficient nutrients and oxygen to be delivered to the tissue. Trauma causes the destruction of the microvascular system, tissue fluid accumulation, inflammation, and hypoxia. Hypoxia and inflammatory cytokines activate endothelial cells, leading to immune cytotoxicity. Neutrophils are recruited, followed by macrophages. Macrophages are essential for coordinating angiogenic responses during wound healing and can produce a variety of factors that regulate cardiovascular growth. Endogenous growth factors can induce angiogenesis and play a key role in the wound healing process. In addition, there are gene-based, stem cell-based, and drug-based therapies for angiogenesis in wounds.

[0008] Current problems in wound repair research at home and abroad are as follows: the research subjects are mainly concentrated on the surface skin, and there are few studies on wound repair of the digestive tract, especially the esophagus. The main reason is that the digestive tract is difficult to observe because it is deep in the human body, and the digestive tract of experimental animals is more difficult to observe than other digestive tract structures. Among them, research specifically on tension-related digestive tract wound repair is even rarer.

[0009] Therefore, there is an urgent clinical need for a method that can maintain the geometric shape of the digestive tract, induce tissue regeneration with low tension control, and achieve structural and functional reconstruction of tension lesions in the digestive tract.

[0010] From the perspective of biomechanics, the present invention will study the mechanical properties of the tension wounds of the digestive tract. On the premise of anti-inflammation, biomechanical research will be carried out from three perspectives: basic structure repair, nutritional function repair, and motor function repair. A combination of multiple repair and reconstruction methods will be used to achieve the inhibition of inflammatory response at the tension wound defect, tissue filling growth, vascular induced growth, and layered induced tissue regeneration, thereby completing the three-dimensional and multi-level repair of the tension wounds of the digestive tract. Summary of the invention

[0011] In view of this, the present invention provides a bioadhesive patch for layered induced tissue regeneration of the digestive tract, which can integrate anti-inflammatory repair, tissue repair, vascular repair, layered induced tissue regeneration and other aspects to perform multi-dimensional repair and reconstruction.

[0012] The present invention provides a bioadhesive patch for layered induced tissue regeneration of the digestive tract, which comprises a bottom layer and a top layer, wherein the bottom layer is a wound contact surface, and the bottom layer comprises a core portion directly in contact with the wound surface and a peripheral portion surrounding the core portion, wherein the peripheral portion of the bottom layer provides adhesion, so that the bioadhesive patch for layered induced tissue regeneration can be attached to the outer wall of the digestive tract to seal the wound; the size and shape of the core portion of the bottom layer correspond to the size and shape of the wound, the core portion of the bottom layer induces tissue regeneration, and is prepared from a chiral hydrogel composition, and the core portion of the bottom layer comprises a first layer formed on the top layer and a second layer formed on the first layer, the first layer comprises primary gastric smooth muscle cells, and the second layer comprises gastric epithelial organoids.

[0013] The bioadhesive patch for layered induced tissue regeneration of the digestive tract according to the present invention is used to be attached to the outer wall of the digestive tract, and can form double induced tissue regeneration. Compared with the non-chiral hydrogel, the chiral hydrogel in the core part of the bottom layer can induce rapid tissue growth. In addition, the two layers of the core part of the bottom layer respectively contain cells corresponding to the composition structure of the digestive tract, which can further induce tissue regeneration, form layered induced tissue regeneration, and make the wound heal faster.

[0014] Preferably, the top layer is a hydrophilic polyurethane material.

[0015] The top layer has excellent mechanical properties, can provide mechanical strength for the bioadhesive patch, and after a period of time, the top layer can be degraded and absorbed.

[0016] Preferably, the thickness of the top layer is 100-300 μm.

[0017] Preferably, the thickness of the peripheral portion of the bottom layer is 100 to 300 μm.

[0018] Preferably, the total thickness of the core portion of the bottom layer is 100-400 μm, and the thickness ratio of the first layer to the second layer is 1:0.6-1.3, more preferably 1:1.

[0019] Preferably, in the first layer, the content of primary gastric smooth muscle cells is 1*10 5 ~3*10 6 Pieces / ml.

[0020] Preferably, in the second layer, the content of gastric epithelial organoids is 01*10 5 ~3*106 Pieces / ml.

[0021] Preferably, the chiral hydrogel composition comprises chiral hydrogel, PLGA microspheres encapsulating VEGF and bFGF, and antibacterial drugs.

[0022] VEGF is the most critical vascular stimulator in the process of angiogenesis, while bFGF is a multifunctional growth factor and angiogenesis factor. VEGF and bFGF are encapsulated in PLGA microspheres and then loaded onto the bioadhesive patch together with free quinolone antibiotics. After bonding, the antibacterial drugs in the chiral hydrogel composition are rapidly released to fight inflammation and inhibit infection, while the PLGA microspheres remain in the wound and slowly degrade, and VEGF and bFGF are slowly released to promote tissue growth and vascular reconstruction in the wound.

[0023] Preferably, based on the total weight of the chiral hydrogel composition, the content of VEGF is 1 to 100 ppm, and the content of bFGF is 1 to 100 ppm.

[0024] Preferably, the antibacterial drug is a quinolone, for example, selected from norfloxacin, levofloxacin and / or moxifloxacin.

[0025] Preferably, based on the total weight of the chiral hydrogel composition, the content of the antibacterial drug is 0.5% to 1%.

[0026] Preferably, the chiral hydrogel is a hydrogel comprising a chiral molecular structure, for example, a hydrogel formed by a cross-linking reaction between sodium alginate and L-lysine.

[0027] Preferably, the chiral hydrogel is prepared by the following method:

[0028] 1) preparing a sodium alginate aqueous solution with a concentration of 1.5 wt% to 3 wt%, adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide and N-hydroxysuccinimide thereto, wherein the amount of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide is 0.8 wt% to 1.5 wt%, and the amount of N-hydroxysuccinimide is 0.8 wt% to 1.5 wt% relative to the total weight of the sodium alginate aqueous solution;

[0029] 2) Add L-lysine to the mixture obtained in step 1), with the amount of L-lysine being 0.4 wt% to 1 wt% relative to the total weight of the sodium alginate aqueous solution, and stir to react uniformly to obtain a chiral hydrogel.

[0030] Preferably, the PLGA microspheres encapsulated with VEGF and bFGF are prepared by the following method:

[0031] 1) adding PLGA (polylactic acid-co-glycolic acid) powder and Tween-80 into dichloromethane to completely dissolve them, wherein the concentration of PLGA is 2wt% to 8wt%, and the concentration of Tween is 15wt% to 30wt%;

[0032] 2) adding an aqueous solution of VEGF and bFGF to the solution in step 1), and ultrasonically forming colostrum, wherein in the aqueous solution of VEGF and bFGF, the concentration of VEGF is 0.01wt% to 0.1wt%, the concentration of bFGF is 0.01wt% to 0.1wt%, the weight ratio of PLGA to VEGF is 1000:0.5 to 1.5, and the weight ratio of PLGA to VEGF is 1000:0.5 to 1.5;

[0033] 3) adding polyvinyl alcohol to water, heating and dissolving, and then cooling, wherein the concentration of polyvinyl alcohol is 0.5wt% to 2wt%;

[0034] 4) adding the colostrum obtained in step 2) to the polyvinyl alcohol solution obtained in step 3), and ultrasonicating to form a double emulsion, wherein the content ratio of polyvinyl alcohol to VEGF is 2000:0.5-2;

[0035] 5) stirring the emulsion overnight and evaporating the dichloromethane;

[0036] 6) centrifuging the product of step 5), removing the supernatant, and washing with pure water for multiple times, centrifuging after each washing;

[0037] 7) The product obtained by centrifugation in step 6) is dispersed in pure water to obtain PLGA microspheres encapsulating VEGF and bFGF.

[0038] Preferably, the molar ratio of the lactic acid structural unit to the glycolic acid structural unit in PLGA is 2-4:1.

[0039] Preferably, the bioadhesive of the peripheral portion of the bottom layer is an interpenetrating network of covalently cross-linked poly(acrylic acid-N-hydroxysuccinimide) ester (PAA-NHS) and physically cross-linked poly(vinyl alcohol) (PVA).

[0040] Beneficial Effects

[0041] 1. The bioadhesive patch for layered induced tissue regeneration of the digestive tract according to the present invention is biodegradable and does not need to be removed after the repair is completed, thus avoiding secondary surgery.

[0042] 2. The bioadhesive patch for layered induced tissue regeneration of the digestive tract according to the present invention has a dual induction tissue repair effect. First, the chiral hydrogel advantageously induces tissue repair of the wound surface, and secondly, the different layers of the core part of the bottom surface contain cells corresponding to the tissue structure, which can use layered induced tissue regeneration to achieve tissue repair.

[0043] 3. The bioadhesive patch for layered induced tissue regeneration of the digestive tract according to the present invention contains antibacterial drugs and vascular stimulating factors, etc., which can inhibit inflammatory response at tension wound defects, fill tissue growth, and induce blood vessel regeneration, thereby completing three-dimensional multi-level repair of tension wounds in the digestive tract. DETAILED DESCRIPTION

[0044] The embodiments of the present disclosure will be described in more detail below. It should be understood that the present disclosure can be implemented in various forms and should not be construed as being limited to the embodiments set forth herein. On the contrary, these embodiments are provided to provide a more thorough and complete understanding of the present disclosure. It should be understood that the embodiments of the present disclosure are only for exemplary purposes and are not intended to limit the scope of protection of the present disclosure.

[0045] It should be noted that the titles of any sections / subsections provided herein are not restrictive. Various embodiments are described throughout this article, and any type of embodiment may be included under any section / subsection. In addition, the embodiments described in any section / subsection may be combined in any manner with any other embodiments described in the same section / subsection and / or different sections / subsections.

[0046] In the description of the embodiments of the present disclosure, the term "including" and similar terms should be understood as open inclusion, that is, "including but not limited to". The term "based on" should be understood as "based at least in part on". The term "one embodiment" or "the embodiment" should be understood as "at least one embodiment". The term "some embodiments" should be understood as "at least some embodiments". Other explicit and implicit definitions may be included below. The terms "first", "second", etc. may refer to different or the same objects. Other explicit and implicit definitions may be included below.

[0047] The embodiments of the present disclosure may involve user data, data acquisition and / or use, etc. These aspects are subject to the corresponding laws, regulations and relevant provisions. In the embodiments of the present disclosure, all data collection, acquisition, processing, processing, forwarding, use, etc. are carried out on the premise that the user knows and confirms. Accordingly, when implementing each embodiment of the present disclosure, the type, scope of use, usage scenario, etc. of the data or information that may be involved should be informed to the user and the user's authorization should be obtained in an appropriate manner in accordance with the relevant laws and regulations. The specific notification and / or authorization method can vary according to the actual situation and application scenario, and the scope of the present disclosure is not limited in this respect.

[0048] The starting materials used in the examples, unless otherwise stated, were commercially available.

[0049] Example 1

[0050] Preparation of PLGA microspheres encapsulated with VEGF and bFGF

[0051] 1) Weigh 100 mg of PLGA powder (lactic acid / glycolic acid copolymer (75:25), Mw=20000), add 2 mL of dichloromethane and 0.5 g of Tween-80, and sonicate until completely dissolved;

[0052] 2) adding 0.2 mL of ultrapure water (containing 100 μg VEGF + 100 μg bFGF growth factor) to the solution obtained in step 1), and ultrasonicating for 5 minutes to form colostrum;

[0053] 3) Weigh 1 g of polyvinyl alcohol (Mw = 13000, hydrolysis rate above 99%), add 100 mL of ultrapure water, stir at 90°C until completely dissolved, and then cool to room temperature for later use;

[0054] 4) adding the colostrum in step 2) into 20 mL of the polyvinyl alcohol solution prepared in step 3), and performing high power ultrasound for 10 minutes to form a double emulsion;

[0055] 5) The emulsion was stirred at 900 rpm overnight to fully remove dichloromethane;

[0056] 6) Centrifuge at 6000 rpm for 10 min, remove the supernatant, and wash the precipitate six times by centrifugation in this way;

[0057] 7) Disperse the precipitate in 10 mL of ultrapure water. The preparation is complete.

[0058] Example 2

[0059] Preparation of a bioadhesive patch for layered induced tissue regeneration of the digestive tract

[0060] 1) Preparation of the top layer

[0061] 8 g of hydrophilic polyurethane (HydroMed D3) was dissolved in 92 g of ethanol aqueous solution (ethanol: water = 93:7 v / v), dried, and formed into a hydrophilic polyurethane film with a thickness of 200 μm;

[0062] 2) Preparation of the peripheral portion of the bottom layer

[0063] Acrylic acid, PVA (Mw = 150000, 99% or more hydrolyzed), α-ketoglutaric acid and polyethylene dimethacrylate (Mw = 550) are added to the dispersion of pure water and stirred thoroughly to make the concentration of acrylic acid 35wt%, PVA 8%, α-ketoglutaric acid 0.2wt%, and polyethylene dimethacrylate 0.05wt%. After stirring evenly, 30mg / 1ml of acrylic acid-N-hydroxysuccinimide ester is added thereto to prepare a bioadhesive precursor solution. The area corresponding to the core part is reserved, and the precursor solution is coated on the top layer prepared in step 1) to form the peripheral part of the bottom layer, wherein the size and shape of the bottom layer, the core part and the peripheral part are determined by the wound size. Irradiate under ultraviolet light (350nm, 10W power) for 30 minutes and dry, and the peripheral part of the bottom layer formed has a thickness of 200μm after drying.

[0064] 3) Preparation of the core part of the bottom layer

[0065] Preparation of the first layer: 0.15 g of norfloxacin, 5*10 6 0.20 g of sodium alginate, 0.12 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), and 0.12 g of N-hydroxysuccinimide (NHS) were added after stirring evenly, and 0.08 g of L-lysine was added after stirring to obtain the first layer of precursor solution of the core part, and the precursor solution was coated on the reserved area of ​​the top layer (i.e., formed on the top layer and surrounded by the outer part), and the thickness after drying was 150 μm.

[0066] Preparation of the second layer: 0.15 g of norfloxacin, 5*10 6Gastric epithelial organoids were prepared, and after stirring evenly, 0.20 g of sodium alginate, 0.12 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), and 0.12 g of N-hydroxysuccinimide (NHS) were added, and after continuing to stir evenly, 0.08 g of L-lysine was added, and the second layer of precursor solution of the core part was obtained by stirring, and the precursor solution was coated on the dried first layer, and the thickness after drying was 150 μm.

[0067] Thus, a bioadhesive patch for layered induced tissue regeneration of the digestive tract was prepared.

[0068] Example 3

[0069] Preparation of a bioadhesive patch for layered induced tissue regeneration of the digestive tract

[0070] 1) Preparation of the top layer

[0071] 8 g of hydrophilic polyurethane (HydroMed D3) was dissolved in 92 g of ethanol aqueous solution (ethanol: water = 93:7 v / v), dried, and formed into a hydrophilic polyurethane film with a thickness of 200 μm;

[0072] 2) Preparation of the peripheral portion of the bottom layer

[0073] Acrylic acid, PVA (Mw = 150000, 99% or more hydrolyzed), α-ketoglutaric acid and polyethylene dimethacrylate (Mw = 550) are added to the dispersion of pure water and stirred thoroughly to make the concentration of acrylic acid 35wt%, PVA 8%, α-ketoglutaric acid 0.2wt%, and polyethylene dimethacrylate 0.05wt%. After stirring evenly, 30mg / 1ml of acrylic acid-N-hydroxysuccinimide ester is added thereto to prepare a bioadhesive precursor solution. The area corresponding to the core part is reserved, and the precursor solution is coated on the top layer prepared in step 1) to form the peripheral part of the bottom layer, wherein the size and shape of the bottom layer, the core part and the peripheral part are determined by the wound size. Irradiate under ultraviolet light (350nm, 10W power) for 30 minutes and dry, and the peripheral part of the bottom layer formed has a thickness of 200μm after drying.

[0074] 3) Preparation of the core part of the bottom layer

[0075] Preparation of the first layer: 0.15 g of norfloxacin, 5*10 60.20 g of sodium alginate, 0.12 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), and 0.12 g of N-hydroxysuccinimide (NHS) were added after stirring evenly, and 0.06 g of L-lysine was added after stirring to obtain a first layer of precursor solution, which was coated on the reserved area of ​​the top layer (i.e., formed on the top layer and surrounded by the outer part), and the thickness after drying was 150 μm.

[0076] Preparation of the second layer: 0.15 g of norfloxacin, 5*10 6 Gastric epithelial organoids were prepared, and after stirring evenly, 0.20 g of sodium alginate, 0.12 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), and 0.12 g of N-hydroxysuccinimide (NHS) were added, and after continuing to stir evenly, 0.06 g of L-lysine was added, and the second layer of precursor solution was obtained by stirring, and the precursor solution was coated on the dried first layer, and the thickness after drying was 150 μm.

[0077] Thus, a bioadhesive patch for layered induced tissue regeneration of the digestive tract was prepared.

[0078] Example 4

[0079] Preparation of a bioadhesive patch for layered induced tissue regeneration of the digestive tract

[0080] 1) Preparation of the top layer

[0081] 8 g of hydrophilic polyurethane (HydroMed D3) was dissolved in 92 g of ethanol aqueous solution (ethanol: water = 93:7 v / v), dried, and formed into a hydrophilic polyurethane film with a thickness of 200 μm;

[0082] 2) Preparation of the peripheral portion of the bottom layer

[0083] Acrylic acid, PVA (Mw = 150000, 99% or more hydrolyzed), α-ketoglutaric acid and polyethylene dimethacrylate (Mw = 550) are added to the dispersion of pure water and stirred thoroughly to make the concentration of acrylic acid 35wt%, PVA 8%, α-ketoglutaric acid 0.2wt%, and polyethylene dimethacrylate 0.05wt%. After stirring evenly, 30mg / 1ml of acrylic acid-N-hydroxysuccinimide ester is added thereto to prepare a bioadhesive precursor solution. The area corresponding to the core part is reserved, and the precursor solution is coated on the top layer prepared in step 1) to form the peripheral part of the bottom layer, wherein the size and shape of the bottom layer, the core part and the peripheral part are determined by the wound size. Irradiate under ultraviolet radiation (350nm, 10W power) for 30 minutes and dry to form the peripheral part of the bottom layer with a thickness of 200μm.

[0084] 3) Preparation of the core part of the bottom layer

[0085] Preparation of the first layer: 0.15 g of norfloxacin, 5*10 6 0.20 g of sodium alginate, 0.12 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), and 0.12 g of N-hydroxysuccinimide (NHS) were added after stirring evenly, and 0.12 g of L-lysine was added after stirring evenly, and the first layer of precursor solution was obtained by stirring. The precursor solution was coated on the reserved area of ​​the top layer (i.e., formed on the top layer and surrounded by the outer part), and dried. The thickness after drying was 150 μm.

[0086] Preparation of the second layer: 0.15 g of norfloxacin, 5*10 6 Gastric epithelial organoids were prepared, and after stirring evenly, 0.20 g of sodium alginate, 0.12 g of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC), and 0.12 g of N-hydroxysuccinimide (NHS) were added, and after continuing to stir evenly, 0.12 g of L-lysine was added, and stirred to obtain a second layer of precursor solution, which was coated on the dried first layer and dried to a thickness of 150 μm after drying.

[0087] Thus, a bioadhesive patch for layered induced tissue regeneration of the digestive tract was prepared.

[0088] Comparative Example 1

[0089] Preparation of bioadhesive patches

[0090] 1) Preparation of the top layer

[0091] 8 g of hydrophilic polyurethane (HydroMed D3) was dissolved in 92 g of ethanol aqueous solution (ethanol: water = 93:7 v / v), dried, and formed into a hydrophilic polyurethane film with a thickness of 200 μm;

[0092] 2) Preparation of the bottom layer

[0093] Add acrylic acid, PVA (Mw = 150000, more than 99% hydrolyzed), α-ketoglutaric acid and polyethylene dimethacrylate (Mw = 550) to the dispersion of pure water and stir well to make the concentration of acrylic acid 35wt%, PVA 8%, α-ketoglutaric acid 0.2wt%, and polyethylene dimethacrylate 0.05wt%. After stirring evenly, add acrylic acid-N-hydroxysuccinimide ester in an amount of 30mg / 1ml to prepare a bioadhesive precursor solution. Apply the precursor solution on the top layer prepared in step 1) to form a bottom layer. Irradiate under ultraviolet light (350nm, 10W power) for 30 minutes. The outer part of the formed bottom layer has a thickness of 200μm.

[0094] Thus, a bioadhesive patch was prepared.

[0095] Test Example

[0096] Female healthy domestic pigs (25 to 30 kg) fasted for 24 hours before surgery. The animals were placed in a dorsal recumbent position, the abdomen was cut open and aseptic preparation was performed. An incision was made in the midline of the abdomen with a blade, and electrocautery was used to extend it if necessary to expose the stomach, which was first bandaged with moist sterile gauze, and then a defect was made to prevent contamination of the abdominal cavity. A biopsy puncture with a diameter of 5 mm was used to cause damage to the stomach wall. Then, the bioadhesive patch obtained in the embodiment and the comparative example was affixed to the damaged area of ​​the outer wall of the stomach to form a seal (2 animals per group), and finally the abdominal incision was closed. The animals were humanely euthanized on the 14th day after surgery, the wound area sealed by the patch was removed, and fixed in 10% formalin for 24 hours, and pathological sections were made for observation and analysis under a microscope.

[0097] Observation and analysis of pathological sections revealed that the wounds in the injured area of ​​the animals using the bioadhesive patch for layered induced tissue regeneration of the digestive tract prepared in Examples 2 to 4 were basically healed. In the slice specimens, repaired gastric epithelial cells were found near the inner wall of the stomach (corresponding to the position of the second layer of the core area of ​​the bottom layer of the bioadhesive patch for layered induced tissue regeneration of the digestive tract), and smooth muscle cells were found near the outer wall of the stomach (corresponding to the position of the first layer of the core area of ​​the bottom layer of the bioadhesive patch for layered induced tissue regeneration of the digestive tract). It can be seen that the bioadhesive patch for layered induced tissue regeneration of the digestive tract prepared in Examples 2 to 4 achieves tissue layered induced regeneration, so that the gastric injury area is repaired in layers according to the tissue structure. However, only a large number of collagen fibers and blood vessels were observed in the slices of the injured area of ​​the animals using the bioadhesive patch of Comparative Example 1, and no similar layered repair structure was observed.

[0098] It should be understood that the various embodiments in this specification are described in a progressive manner, and the same or similar parts between the various embodiments can be referred to each other, and each embodiment focuses on the differences from other embodiments. In particular, for the method embodiment, since it is basically similar to the method described in the device and system embodiments, the description is relatively simple, and the relevant parts can be referred to the partial description of other embodiments.

[0099] It should be understood that the above description is of a specific embodiment of the present specification. Other embodiments are within the scope of the claims. In some cases, the actions or steps recorded in the claims can be performed in an order different from that in the embodiments and still achieve the desired results.

[0100] It should be understood that the singular description of an element shown herein does not limit the number of the element to one. In addition, modules or elements described or shown as separate herein may be combined into a single module or element, and modules or elements described or shown as single herein may be split into multiple modules or elements.

[0101] It should also be understood that the terms and expressions used herein are for description only, and one or more embodiments of this specification should not be limited to these terms and expressions. The use of these terms and expressions does not mean to exclude any equivalent features of the illustrations and descriptions (or parts thereof), and it should be recognized that various modifications that may exist should also be included in the scope of the claims. Other modifications, changes and substitutions may also exist. Accordingly, the claims should be deemed to cover all such equivalents.

Claims

1. A bioadhesive patch for layered induction of tissue regeneration in the digestive tract, characterized in that: It includes a bottom layer and a top layer, wherein the bottom layer is a wound contact surface, and the bottom layer includes a core part that directly contacts the wound surface and a peripheral part that surrounds the core part, wherein the peripheral part of the bottom layer provides adhesion, so that the bioadhesive patch that induces tissue regeneration can be attached to the outer wall of the digestive tract to seal the wound; the size and shape of the core part of the bottom layer correspond to the size and shape of the wound, the core part of the bottom layer induces tissue regeneration, and is prepared from a chiral hydrogel composition, and the core part of the bottom layer includes a first layer formed on the top layer and a second layer formed on the first layer, the first layer includes primary gastric smooth muscle cells, and the second layer includes gastric epithelial organoids.

2. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 1, characterized in that: The top layer is a hydrophilic polyurethane material; And / or, the thickness of the top layer is 100-300 μm.

3. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 1, characterized in that: The thickness of the peripheral portion of the bottom layer is 100 to 300 μm; And / or, the total thickness of the core part of the bottom layer is 100-400 μm, and the thickness ratio of the first layer to the second layer is 1:0.6-1.

3.

4. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 1, characterized in that: In the first layer, the content of primary gastric smooth muscle cells is 1*10 5 ~3*10 6 Pieces / ml; And / or, in the second layer, the content of gastric epithelial organoids is 01*10 5 ~3*10 6 Pieces / ml.

5. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 1, characterized in that: The chiral hydrogel composition comprises chiral hydrogel, PLGA microspheres encapsulating VEGF and bFGF, and antibacterial drugs.

6. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 5, characterized in that: Based on the total weight of the chiral hydrogel composition, the content of VEGF is 1 to 100 ppm, and the content of bFGF is 1 to 100 ppm; And / or, the antibacterial drug is a quinolone.

7. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 5, characterized in that: The antibacterial drug is norfloxacin, levofloxacin and / or moxifloxacin; and / or, based on the total weight of the chiral hydrogel composition, the content of the antibacterial drug is 0.5% to 1%; And / or, the chiral hydrogel is a hydrogel comprising a chiral molecular structure.

8. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 5, characterized in that: The chiral hydrogel is a hydrogel formed by a cross-linking reaction between sodium alginate and L-lysine.

9. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 5, characterized in that: The chiral hydrogel is prepared by the following method: 1) preparing a sodium alginate aqueous solution with a concentration of 1.5 wt% to 3 wt%, adding 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide and N-hydroxysuccinimide thereto, wherein the amount of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide is 0.8 wt% to 1.5 wt%, and the amount of N-hydroxysuccinimide is 0.8 wt% to 1.5 wt% relative to the total weight of the sodium alginate aqueous solution; 2) Add L-lysine to the mixture obtained in step 1), with the amount of L-lysine being 0.4 wt% to 1 wt% relative to the total weight of the sodium alginate aqueous solution, and stir to react uniformly to obtain a chiral hydrogel.

10. The bioadhesive patch for layered induction of tissue regeneration of the digestive tract according to claim 1, characterized in that: The bioadhesive of the peripheral portion of the bottom layer is an interpenetrating network of covalently cross-linked poly(acrylic acid-N-hydroxysuccinimide) ester (PAA-NHS) and physically cross-linked poly(vinyl alcohol) (PVA).