Hydrogel plugging umbrella for gastric perforation and preparation method thereof

Abstract

The invention discloses a hydrogel plugging umbrella for gastric perforation and a preparation method thereof. The hydrogel plugging umbrella comprises a plugging part, and fixing parts are integrally formed at the two ends of the plugging part. The excellent plugging effect is achieved through the triple synergistic effect, the space mechanical interlocking effect brought by the double-umbrella-shaped design provides the immediate plugging and continuous self-expansion compaction effect, and physical and chemical adhesion of a tissue-gel interface ensures that under the corrosive gastric juice environment and the dynamic mechanical load, the plugging effect is good, and the plugging effect is good. A safe and lasting gastric perforation sealing effect can still be provided; the caffeic acid modified chitosan component has unique proton adsorption and biomolecule capture characteristics and can effectively block permeation of digestive enzymes such as gastric acid and pepsase, the protection effect not only maintains the activity of endogenous growth factors in a wound microenvironment, but also creates conditions beneficial to tissue regeneration, and the wound healing effect is improved. Therefore, healing and repairing of gastric perforation are remarkably promoted.

Description

Technical Field

[0001] The invention relates to the field of medical technology, in particular to a hydrogel occluding umbrella for gastric perforation and a preparation method thereof. Background Art

[0002] Gastric perforation refers to a full-thickness defect of the gastric wall caused by a variety of causes, including severe peptic ulcers, trauma, malignant tumors, iatrogenic injuries, and active perforation during endoscopic surgery. If medical intervention is not performed in time, gastric contents can penetrate into the abdominal cavity through the perforation, causing acute and severe abdominal pain and systemic inflammation, and may even lead to fatal sepsis. Traditional surgical operations are often more traumatic and are accompanied by the risk of serious complications such as anastomotic leakage, wound infection, and postoperative bleeding. In contrast, endoscopic metal clip treatment is a less invasive alternative, but it is difficult to operate and requires doctors to have extremely high professional skills. It is also prone to failure when dealing with inflamed or edematous tissues.

[0003] Over the past few decades, hydrogel-based bioadhesives have become potential materials for gastric perforation repair due to their excellent biocompatibility, tissue adhesion and biodegradability. These hydrogel formulations include injectable, powder assembly and preformed types, which are designed to repair gastric perforation by adhering to the wound site. However, in clinical applications, the mechanical and chemical digestive environment of the stomach poses a major challenge to the performance of hydrogels. Mechanical digestion in the stomach requires that the hydrogel has structural stability and mechanical fatigue resistance under the condition of frequent contraction and relaxation of the circular and longitudinal muscles of the stomach wall. Chemical digestion in the stomach requires that the hydrogel can still maintain adhesion and strength after long-term contact with gastric acid and digestive enzymes. At the same time, gastric acid, as a strong acid, irritates the wound surface and aggravates inflammation. Digestive enzymes can decompose endogenous growth factors and delay the healing process. At present, most hydrogels are difficult to effectively protect the wound surface and promote wound healing in the harsh gastric environment.

[0004] The atrial septum occluder commonly used in cardiology has been innovatively used in the treatment of gastrointestinal perforation and fistula in recent years due to its unique structural design. Although its metal structure can effectively resist mechanical and chemical digestion in the stomach, its non-degradability requires a secondary surgery to remove it later, which inevitably causes secondary damage. We have tried to develop a new type of degradable hydrogel occluder that can not only be accurately delivered to the lesion site through endoscopy, but also has good biocompatibility and controllable degradability, which can effectively occlude the defect and promote tissue healing. Summary of the invention

[0005] The purpose of the present invention is to provide a hydrogel occluding umbrella for gastric perforation and a preparation method thereof, which solves the problems raised in the background technology.

[0006] To achieve the above-mentioned purpose, the present invention provides the following technical solution: a hydrogel occluding umbrella for gastric perforation, comprising an occluding part, both ends of which are integrally formed with fixing parts, the diameter and height of the occluding part are both 5 mm, and the diameter and height of the fixing part are 10 mm and 3 mm respectively.

[0007] The present invention also relates to a method for preparing a hydrogel occluding umbrella for gastric perforation, comprising the following steps:

[0008] Step 1: dissolving chitosan in deionized water at room temperature and stirring continuously, then dissolving caffeic acid in anhydrous ethanol by ultrasonic treatment to obtain a uniform solution, dissolving 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride and N-hydroxysuccinimide in a small amount of deionized water, and mixing with the caffeic acid ethanol solution, then dropping the activated caffeic acid solution into the chitosan solution, stirring at room temperature, dialyzing the obtained solution in deionized water for three days for purification after the stirring is completed, and then freeze-drying to obtain a caffeic acid grafted chitosan product, that is, obtaining caffeic acid modified chitosan;

[0009] Step 2: Add polyacrylamide, caffeic acid-modified chitosan, photoinitiator and N,N'-cystamine bisacrylamide solution into deionized water, and then heat to obtain a mixture, and oscillate the mixture on an oscillator to obtain a transparent caffeic acid-modified chitosan / polyacrylamide solution:

[0010] Step 3: Inject the prepared caffeic acid modified chitosan / polyacrylamide solution into the mold, cure it under ultraviolet light overnight, and finally take out the prepared hydrogel occluder and store it in a sterile sealed container.

[0011] As a preferred embodiment of the present invention, the stirring time in step 1 is 48 hours.

[0012] As a preferred embodiment of the present invention, the heating temperature in step 2 is 60° C. and the heating time is 20 min.

[0013] As a preferred embodiment of the present invention, the aseptic sealed storage temperature condition in step three is 10°C-20°C.

[0014] As a preferred embodiment of the present invention, the photoinitiator is Irgacure2959.

[0015] Compared with the prior art, the present invention has the following beneficial effects:

[0016] 1. The hydrogel occluder adopts a unique double umbrella design, which achieves excellent occlusion effect through triple synergistic effects: the rapid spatial mechanical interlocking effect brought by the double umbrella design, the continuous self-expansion and compaction effect, and the physical and chemical adhesion of the tissue-gel interface. This multiple mechanism ensures that it can still provide a fast, safe and long-lasting gastric perforation sealing effect under the corrosive gastric fluid environment and dynamic mechanical load.

[0017] 2. The caffeic acid-modified chitosan / polyacrylamide component has unique proton adsorption and biomolecule capture properties, which can effectively block the penetration of digestive enzymes such as gastric acid and pepsin. This protective effect not only maintains the activity of endogenous growth factors in the wound microenvironment, but also creates conditions conducive to tissue regeneration, thereby significantly promoting the healing and repair of gastric perforation.

[0018] 3. The mold casting technology can accurately and customarily prepare hydrogel occluders of various shapes and sizes with good biocompatibility and biodegradability, and is suitable for endoscopic delivery, which helps to repair gastric perforation in various clinical applications. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Other features, objects and advantages of the present invention will become more apparent from the detailed description of non-limiting embodiments made with reference to the following drawings:

[0020] Figure 1 This is a schematic structural diagram of a hydrogel occluding umbrella for gastric perforation according to the present invention;

[0021] Figure 2 It is a cross-sectional view of a mold of a hydrogel occluding umbrella for gastric perforation according to the present invention;

[0022] Figure 3 is the hydrogen nuclear magnetic resonance spectrum of the present invention;

[0023] Figure 4 It is the Fourier transform infrared spectrum of the present invention;

[0024] Figure 5 It is the mechanical performance diagram of the present invention;

[0025] Figure 6 It is the cyclic stretching and cyclic compression diagram of the present invention;

[0026] Figure 7 It is the electron microscope structure diagram of the present invention;

[0027] Figure 8 This is a swelling performance diagram of the present invention;

[0028] Fig. 9 It is the in vitro barrier protection and cell migration diagram of the present invention;

[0029] Fig.10is the in vitro biocompatibility diagram of the present invention;

[0030] Fig.11 The in vivo degradability and blood biochemical profile of the present invention;

[0031] Fig.12 This is a surgical treatment and endoscopic tracking diagram of rabbit gastric perforation according to the present invention;

[0032] Fig.13 This is a general image of the rabbit stomach wall tissue after 28 days of treatment with the present invention

[0033] Fig.14 It is the in vivo X-ray tracing diagram of the present invention;

[0034] Fig.15 This is an immunofluorescence staining image of rabbit stomach wall tissue after 7 days of treatment with the present invention;

[0035] Fig.16 This is an immunohistochemical staining image of rabbit stomach wall tissue after 7 days of treatment with the present invention;

[0036] Fig.17 This is an endoscopic tracking image of the wound healing of the pig stomach of the present invention;

[0037] Fig.18 This is the gross tissue and HE section of pig stomach after 28 days of treatment by the present invention;

[0038] Fig.19 This is the biochemical analysis diagram of pig blood and HE slice diagram of important organs of the present invention.

[0039] In the figure: 1, blocking part; 11, fixing part. DETAILED DESCRIPTION

[0040] In order to make the technical means, creative features, objectives and effects achieved by the present invention easy to understand, the present invention is further explained below in conjunction with specific implementation methods.

[0041] A hydrogel occluding umbrella for gastric perforation comprises an occluding part 1, both ends of which are integrally formed with fixing parts 11.

[0042] It should be noted that the diameter and height of the blocking portion 1 are both 5 mm, and the diameter and height of the fixing portion 11 are respectively 10 mm and 3 mm.

[0043] A preparation method of a hydrogel occluding umbrella for gastric perforation is as follows:

[0044] 5 g of chitosan (CS) was dissolved in deionized water at room temperature and stirred continuously. Then, 1.22 g of caffeic acid (CA) was dissolved in anhydrous ethanol by ultrasonic treatment to obtain a uniform solution. To activate the carboxyl groups on the caffeic acid, 1.30 g of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (hereinafter referred to as EDCI) and 0.78 g of N-hydroxysuccinimide were dissolved in a small amount of deionized water and mixed with the caffeic acid ethanol solution. Subsequently, the activated CA solution was added dropwise to the CS solution and stirred at room temperature for 48 hours. The resulting solution was dialyzed in deionized water for three days for purification and then freeze-dried to obtain the caffeic acid modified chitosan (CA-CS) product.

[0045] Polyacrylamide PAM (1.8 g), CA-CS (200 mg), photoinitiator Irgacure 2959 (22.4 mg) and N,N'-cystamine bisacrylamide solution (46 μL, 10 mg / mL) were added to 8 mL of deionized water, and after heating at 60°C for about 20 min, the mixture was shaken on a shaker to obtain a transparent CA-CS / PAM solution.

[0046] The polymethyl methacrylate (PMMA) cavity mold was made by injection molding. Its design was inspired by the atrial septum / ventricular septum occlusion device of the heart. The design scheme included two umbrella-shaped fixing parts and a central occluding part, wherein the diameter and height of the central part were both 5 mm, and the diameter and height of the end part were 10 mm and 3 mm respectively. The prepared CA-CS / PAM solution was injected into the mold and cured overnight under ultraviolet light (150W). Finally, the prepared hydrogel occluding umbrella was taken out and stored in a sterile sealed container at 10℃-20℃.

[0047] What needs to be further said is:

[0048] 1. Design and characterization of CA-CS / PAM composite hydrogels

[0049] like Figure 3 As shown, the 1H-NMR spectrum suggests that CA was successfully grafted onto CS;

[0050] like Figure 4 As shown, Fourier transform infrared spectroscopy (FTIR) spectrum shows that CA-CS / PAM hydrogel has a new characteristic absorption peak at 1407 cm-1, which is attributed to the aromatic C=C peak of the catechol group rich in phenolic hydroxyl groups in CA-CS. The CA-CS skeleton can effectively dissipate energy, while the soft and ductile PAM network maintains the integrity of the hydrogel during deformation, giving it excellent mechanical properties.

[0051] like Figure 5As shown in Figure 2, CA-CS / PAM hydrogel exhibits significant tensile properties and can be stretched to 20 times its original length, with a breaking strength of 77.18 kPa and a toughness of 0.62 MJ / m 3 The highest value, compressive strength 287.80kPa.

[0052] like Figure 6 As shown in the data, CA-CS / PAM hydrogel exhibited excellent mechanical stability and fatigue resistance in cyclic tensile and cyclic compression tests, which laid a solid foundation for CA-CS / PAM hydrogel to resist fracture or external deformation during the dynamic movement of gastric tissue.

[0053] like Figure 7 As shown, it can be seen under an electron microscope that the present invention has a dense microstructure.

[0054] like Figure 8 As shown, CA-CS / PAM hydrogel exhibited rapid swelling behavior, enabling it to quickly fill gastric perforations and exert a self-expansion compaction effect to achieve effective and durable closure. In PBS and SGF solutions, CA-CS / PAM hydrogel swelled rapidly within the first hour, and the swelling ratios after 24 hours reached 500.14% and 580.60%, respectively.

[0055] 2. Wound protection function, biocompatibility and degradability of CA-CS / PAM composite hydrogel

[0056] Gastric acid and digestive proteases can hinder the gastric wound healing process by aggravating tissue damage and degrading factors that promote regeneration, especially basic fibroblast growth factor and vascular endothelial growth factor, which are essential for angiogenesis and tissue repair. CS has proton adsorption capacity, and the catechol groups in CA have a molecular retention effect on biomacromolecules. CA-CS / PAM hydrogel can protect wounds from gastric juice penetration, thereby retaining endogenous growth factors in the wound bed to promote perforation repair.

[0057] like Fig. 9 As shown, a simple transwell system was developed, in which the hydrogel in the upper chamber blocked the simulated gastric fluid to evaluate the wound protection effect of CA-CS / PAM hydrogel. The cell migration results in the lower chamber showed that after 48 hours, the CA-CS / PAM hydrogel group showed more significant cell migration (67.3±3.4%), compared with the PAM hydrogel group (28.2±2.3%), confirming its ability to effectively prevent gastric acid and pepsin penetration and promote wound healing. These results indicate that CA-CS / PAM hydrogel has unique H+ ion adsorption and biomacromolecule chelating capabilities, which are beneficial for preventing wounds from being exposed to acidic conditions and external decomposition factors (such as pepsin), increasing local growth factor accumulation, and promoting gastric wound healing.

[0058] To further evaluate the long-term protective effect of CA-CS / PAM hydrogel in simulated gastric fluid, a transwell device was used to monitor the permeation of H+ ions and pepsin through the hydrogel barrier. Specifically, pure PAM hydrogel and CA-CS / PAM hydrogel were placed at the bottom of the upper chamber, and an acidic solution (pH = 2.0) and a pepsin solution (2 mg / mL) were added. The pH value and pepsin concentration of the lower chamber were systematically detected every 24 hours. The results showed that as H+ ions freely permeated into the lower chamber, the pH value in the PAM hydrogel steadily decreased, reaching 4.98±0.4 on the third day. In contrast, the CA-CS / PAM hydrogel effectively prevented PBS acidification for at least 7 days, and the acid resistance was proportional to the CS content. Similarly, the CA-CS / PAM hydrogel significantly reduced the diffusion of pepsin, with only 14.32% permeability after 7 days, while the PAM hydrogel group was 83.33%. The permeability of pepsin decreased with the increase of modified caffeic acid content. These results collectively indicate that the CA-CS / PAM hydrogel is able to maintain its chemical properties for a long time and withstand long-term exposure to acidic conditions and decomposing enzymes, thereby providing a protective barrier for gastric wounds.

[0059] like Fig.10 As shown, in vitro biocompatibility studies using live / dead cell staining and cell counting kit-8 (CCK-8) assays indicated that CA-CS / PAM hydrogels had no significant effect on cell viability compared with the control group.

[0060] like Fig.11 As shown, in vivo degradability and biocompatibility are crucial for effective tissue repair, minimizing complications, and avoiding secondary surgical removal. To this end, we implanted CA-CS / PAM hydrogels (diameter: 1 cm) subcutaneously into rats and collected samples for further evaluation to explore the biodegradability of the hydrogel. The remaining weight of the CA-CS / PAM hydrogel after 12 weeks was significantly reduced to one-fifth of that at week 1. In addition, hematological and biochemical analyses showed that there were no significant differences in complete blood cell counts (CBCs) and biochemical indices between hydrogel-implanted rats and healthy controls for up to 12 weeks, indicating that it has excellent in vivo biocompatibility and that the hydrogel degradation products have no potential systemic toxicity.

[0061] Embodiment 1

[0062] A full-thickness perforation with a diameter of 5 mm was created in the anterior wall of the rabbit stomach. Fig.12 A), and then the perforation is sealed with a hydrogel occluder;

[0063] During the wound healing process, endoscopic examinations were performed on the wounds of the two groups on the 3rd, 7th, 14th and 28th days. After the operation, the hydrogel occluder expanded rapidly and was stably fixed at the perforation site for at least three days, effectively blocking the rabbit gastric perforation. Endoscopic follow-up on the 14th day showed that the healing effect of the occluder group was better than that of the traditional suture group, which still had mild inflammation ( Fig.12 B), on day 28, there were no obvious unhealed wounds in both groups;

[0064] Examination of gastric tissues collected on day 28 confirmed that the mucosal and serosal surfaces of gastric perforations in both groups had completely healed, and H&E staining further showed that the perforated area was completely bridged, indicating that the gastric wall in both groups had been fully repaired on day 28. Fig.13 As shown;

[0065] In order to track the shedding of the hydrogel occluder umbrella, we added three metal beads to each of the double umbrella-shaped ends of the hydrogel occluder umbrella and observed the position of the hydrogel occluder umbrella by X-ray. The hydrogel occluder umbrella group remained at the perforation site during the first 5 days and gradually fell off on the 7th day. The gastric part was discharged through the gastrointestinal tract, while the abdominal part remained firmly attached to the original site, as shown in Figure 2. Fig.14 As shown;

[0066] Immunofluorescence staining of the wound site was performed on the 7th day, and the results of CD86 (M1 macrophages) and CD206 (M2 macrophages) immunofluorescence staining showed that compared with the suture group, the hydrogel occlusion umbrella group had the lowest level of M1 macrophages and the highest level of M2 macrophages in the wound on the 7th day, indicating that the wound shifted to an anti-inflammatory phenotype, such as Fig.15 As shown;

[0067] On the 7th day, the wound site was subjected to immunohistochemistry (IHC) staining to study its efficacy in retaining endogenous growth factors in vivo. The results showed that compared with the suture group, the hydrogel occluder had significantly enhanced growth factor expression of bFGF and angiogenic VEGF on the 7th day, and the number of new blood vessels also increased. This was due to the proton adsorption capacity of chitosan in the component and the sealing effect of the catechol group in caffeic acid on biomacromolecules through nucleophilic reaction, which gave the hydrogel occluder a barrier protection effect on the wound surface, protecting it from the invasion of gastric acid and digestive enzymes, and promoting the repair of gastric perforation. Fig.16 shown.

[0068] Embodiment 2

[0069] A 10 mm diameter perforation was made in the anterior wall of the pig stomach using a Dual knife. After modeling, pneumoperitoneum was formed, and then the perforation was immediately blocked with a hydrogel occluder. After using the hydrogel occluder, the artificial abdominal gas caused by gastric perforation surgery was effectively treated, confirming the excellent blocking effect of the occluder.

[0070] During the wound healing process, endoscopic examinations were performed on the wounds of the two groups on days 3, 7, 14, and 28. The hydrogel occluder effectively blocked the pig stomach perforation. On day 28, there were no obvious unhealed wounds in both groups. Fig.17 As shown;

[0071] Examination of gastric tissues collected on day 28 confirmed that the mucosal and serosal surfaces of gastric perforations in both groups had completely healed, and H&E staining further showed that the perforated area was completely bridged, indicating that the gastric wall had been fully repaired in both groups on day 28. Fig.18 As shown;

[0072] Blood analysis results on day 28 after treatment were unremarkable, and H&E staining of major organs (heart, liver, spleen, lung, and kidney) showed normal tissues, with no evidence of inflammation or systemic inflammatory injury secondary to gastric perforation, e.g. Fig.19 shown.

[0073] The above shows and describes the basic principles and main features of the present invention and the advantages of the present invention. It is obvious to those skilled in the art that the present invention is not limited to the details of the above exemplary embodiments, and the present invention can be implemented in other specific forms without departing from the spirit or basic features of the present invention. Therefore, no matter from which point of view, the embodiments should be regarded as exemplary and non-restrictive. The scope of the present invention is defined by the appended claims rather than the above description, and it is intended that all changes falling within the meaning and scope of the equivalent elements of the claims are included in the present invention.

[0074] In addition, it should be understood that although the present specification is described according to implementation modes, not every implementation mode contains only one independent technical solution. This description of the specification is only for the sake of clarity. Those skilled in the art should regard the specification as a whole. The technical solutions in each embodiment may also be appropriately combined to form other implementation modes that can be understood by those skilled in the art.

Claims

1. A hydrogel occluding umbrella for gastric perforation, characterized in that: It comprises a blocking portion (1), and both ends of the blocking portion (1) are integrally formed with fixing portions (11).

2. The hydrogel occluder for gastric perforation according to claim 1, characterized in that: The diameter and height of the blocking portion (1) are both 5 mm, and the diameter and height of the fixing portion (11) are respectively 10 mm and 3 mm.

3. A method for preparing a hydrogel occluding umbrella for gastric perforation, applicable to the hydrogel occluding umbrella according to claims 1-2, characterized in that: The method comprises the following steps: Step 1: dissolving chitosan in deionized water at room temperature and stirring continuously, then dissolving caffeic acid in anhydrous ethanol by ultrasonic treatment to obtain a uniform solution, dissolving 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride and N-hydroxysuccinimide in a small amount of deionized water, and mixing with the caffeic acid ethanol solution, then dropping the activated caffeic acid solution into the chitosan solution, stirring at room temperature, dialyzing the obtained solution in deionized water for three days for purification after the stirring is completed, and then freeze-drying to obtain a caffeic acid grafted chitosan product, that is, obtaining caffeic acid modified chitosan; Step 2: Add polyacrylamide, caffeic acid-modified chitosan, photoinitiator and N,N'-cystamine bisacrylamide solution into deionized water, and then heat to obtain a mixture, and oscillate the mixture on an oscillator to obtain a transparent caffeic acid-modified chitosan / polyacrylamide solution: Step 3: Inject the prepared caffeic acid modified chitosan / polyacrylamide solution into the mold, cure it under ultraviolet light overnight, and finally take out the prepared hydrogel occluding umbrella and store it in a sterile sealed container.

4. The method for preparing a hydrogel occluding umbrella for gastric perforation according to claim 3, characterized in that: The stirring time described in step 1 is 48 hours.

5. The method for preparing a hydrogel occluding umbrella for gastric perforation according to claim 3, characterized in that: The heating temperature in step 2 is 60° C. and the heating time is 20 min.

6. The method for preparing a hydrogel occluding umbrella for gastric perforation according to claim 3, characterized in that: The aseptic sealed storage temperature condition described in step 3 is 10°C-20°C.

7. The method for preparing a hydrogel occluding umbrella for gastric perforation according to claim 3, characterized in that: The photoinitiator is Irgacure 2959.

8. The method for preparing a hydrogel occluding umbrella for gastric perforation according to claim 3, characterized in that: The mold described in step three is a polymethyl methacrylate cavity mold made by injection molding.

9. The method for preparing a hydrogel occluding umbrella for gastric perforation according to claim 3, characterized in that: The power of the ultraviolet lamp is 150W.

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