Degradable biological membrane, preparation method thereof and application of degradable biological membrane in precutting line marking of gastric resection operation
By preparing dye-labeled degradable biofilms and combining 3D printing technology, the problem of inaccurate resection range during gastrectomy surgery is solved, and accurate gastrectomy and postoperative recovery are achieved.
Patent Information
- Application Number
- CN202510685859.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-05-26
- Publication Date
- 2025-08-22
AI Technical Summary
In gastrectomy, it is difficult for the prior art to accurately locate the resection range and retain the volume of the stomach, resulting in inconsistent surgical results, affecting the patient's nutrition and postoperative recovery.
Degradable biofilms containing dye labeling function were prepared, stomach models were constructed through 3D printing technology, dyes were embedded using PLGA microcapsules and chitosan-sodium alginate nanoparticles, cross-linked gelatin to form biofilms, and precise pretangents were used to form during surgery using the effects of thrombin and dopamine.
Accurate resection of gastrectomy surgery is achieved, the quality and efficiency of the surgery are improved, and the accuracy of the resection range and the patient's postoperative recovery is ensured.
Smart Images
Figure CN120514934A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of degradable biomedical materials, in particular to a degradable biofilm, a preparation method thereof and an application thereof in pre-cutting line marking in gastrectomy surgery. Background Art
[0002] Currently, when performing gastric surgery, gastrointestinal surgeons mostly make cuts based on experience and simple anatomical landmarks, without a clear positioning of the gastric resection range and the volume of the stomach to be retained. This sometimes results in insufficient resection range and sometimes too little gastric volume to be retained. Especially when performing sleeve gastrectomy for metabolic weight loss, there is no precise positioning of the cutting line for the volume of the retained stomach (generally required to be 60-100 ml to achieve a good weight loss effect). Instead, the surgeons rely solely on the temporary insertion of a correction gastric tube to estimate the volume of the retained stomach. This results in different gastric resection and retained volumes, which has a certain impact on the patient's postoperative nutrition and leads to differences in the effectiveness of the surgery.
[0003] Biomedical materials are used to diagnose, treat, repair, and replace damaged tissues and organs, or enhance their function. Because they are used directly on the human body or are closely related to human health, they are subject to strict standards and requirements for their use. Biodegradable biomaterials, as new materials in the medical field, are receiving significant attention and are experiencing rapid development.
[0004] The use of degradable biomedical materials to prepare degradable biofilms with dye marking functions is expected to form pre-cut lines in surgical operations such as gastrectomy, providing convenience for the operation and solving the defects of existing methods such as inaccurate resection range and affecting surgical results. Summary of the Invention
[0005] The purpose of the present invention is to provide a degradable biofilm, a preparation method thereof and its application in pre-cutting line marking for gastrectomy surgery. By preparing a degradable biofilm containing a dye marking function, it is applied during gastrectomy surgery to self-degrade to form an accurate pre-cutting line, thereby improving the accuracy of resection.
[0006] In view of this, the solution of the present invention is:
[0007] The first object of the present invention is to provide a method for preparing a degradable biofilm, comprising the following steps:
[0008] PLGA microcapsules loaded with thrombin were prepared by solvent method;
[0009] The dye is embedded in chitosan-sodium alginate nanoparticles to obtain dye nanoparticles;
[0010] PLGA microcapsules and dye nanoparticles are added to gelatin solution, uniformly dispersed, and then cross-linked to form a solid after casting, thereby obtaining a degradable biofilm.
[0011] Furthermore, the PLGA microcapsules containing thrombin are obtained by emulsifying a PLGA oil phase and a thrombin aqueous phase; and / or the particle size of the PLGA microcapsules containing thrombin is 1-5 μm.
[0012] Furthermore, the preparation method of the dye nanoparticles includes embedding the dye in chitosan by an ion gel method, and then adding sodium alginate to obtain the dye nanoparticles; and / or, the dye loading amount in the dye nanoparticles is 0.1-0.5wt%, and the dye is selected from methylene blue, methyl violet or indocyanine green.
[0013] Furthermore, transglutaminase is added during the cross-linking process, and the enzyme is inactivated below 50°C after cross-linking for 5-10 minutes at room temperature; and / or, the gelatin solution concentration is 5-10% (w / v), and the PLGA microcapsules account for 5-10% of the solid matter obtained by cross-linking.
[0014] Furthermore, the sensing peptide GPLGIAGQ is added during the cross-linking process.
[0015] The second object of the present invention is to propose the use of a biofilm in the preparation of a surgical marking material, wherein the biofilm is prepared by the preparation method described in the first aspect.
[0016] Furthermore, the surgical operation is gastrectomy.
[0017] The third object of the present invention is to provide a biofilm marking method for pre-cutting lines of gastrectomy surgery based on 3D printing, the steps comprising:
[0018] S1. Constructing a 3D stomach model based on stomach feature data;
[0019] S2. Constructing a pre-cut portion based on a 3D model of the stomach, and constructing a biofilm attached to the pre-cut portion, the biofilm being prepared by the preparation method described in the first aspect;
[0020] S3. Place the biofilm obtained in step S2 into the abdominal cavity to fit the biofilm to the stomach, and form a pre-cut line for gastrectomy surgery after the biofilm is degraded.
[0021] Furthermore, the stomach 3D model is obtained by 3D printing based on stomach characteristic data; and / or the pre-cut part model includes the gastric body and gastric fundus of the greater curvature of the stomach.
[0022] Furthermore, in step S3, the inner side of the biofilm is coated with dopamine.
[0023] Compared with the prior art, the present invention has the following beneficial effects:
[0024] The degradable biofilm provided by the present invention is obtained by encapsulating thrombin microcapsules and dye nanoparticles into gelatin and cross-linking them. The biofilm can dye the target area and release thrombin under surgical conditions, and can be used as a surgical line marker. The preparation method is simple and feasible, and the degradation performance is good.
[0025] The degradable biofilm described in the present invention can be inserted into the patient's abdominal cavity through a laparoscopic troca. After physiological saline activates the adhesion layer, it directly adheres to the stomach surface. Body temperature (37°C) triggers dopamine oxidative adhesion and thrombin release. The material degrades within 5-8 minutes to avoid interfering with subsequent surgical steps. Methylene blue falls off on the stomach wall to form a pre-cut line for resection positioning, thereby achieving precise gastric resection and improving surgical quality and efficiency. BRIEF DESCRIPTION OF THE DRAWINGS
[0026] Figure 1 This is a schematic diagram of the degradable biofilm obtained in the preparation example of the present invention.
[0027] Figure 2 This is a flow chart of the animal experiment simulating gastrectomy surgery process in Example 2 of the present invention. Figure 2 Middle: ① Free the pre-resected stomach; ② Place a correction gastric tube as an indicator for gastric resection; ③ Cut the stomach along the correction gastric tube (the dotted line represents the intended part of the stomach, which can be covered with a membrane); ④ Prepare a sleeve stomach; ⑤ Prepare a 3D gastric model based on imaging data; ⑥ Prepare a pre-cutting line biofilm based on the 3D model; ⑦ Cover the pre-resected stomach with a biofilm as an indicator, and cut the stomach along the pre-cutting line. DETAILED DESCRIPTION
[0028] The technical solutions of the present invention will be clearly and completely described below in conjunction with preferred embodiments. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. All other embodiments obtained by ordinary technicians in this field based on the embodiments of the present invention without making any creative efforts shall fall within the scope of protection of the present invention.
[0029] Preparation Example Preparation of degradable biofilm
[0030] Material composition: gelatin concentration 5-10%, TGase 10U / g, gelatin buffer phosphate buffer (pH 7.0), thrombin microencapsulation and methylene blue nanoparticles, polylactic acid-glycolic acid copolymer, chitosan, sodium alginate
[0031] 1) Preparation of thrombin microcapsules
[0032] Poly(lactic acid-co-glycolic acid) copolymer was used as the microcapsule shell material, and thrombin microcapsules were prepared by emulsification-solvent evaporation method. The specific steps were as follows: thrombin (100 IU / cm 2) is dissolved in the aqueous phase, PLGA is dissolved in dichloromethane solvent, the aqueous phase and the oil phase are mixed and emulsified to form a W / O / W emulsion, and the solvent is evaporated to obtain PLGA microcapsules (particle size 1-5 μm), that is, PLGA microcapsules encapsulating thrombin are obtained.
[0033] 2) Methylene blue nanoparticle embedding
[0034] Chitosan-sodium alginate nanoparticles are used to encapsulate methylene blue. The specific steps are: chitosan (positively charged) and methylene blue are mixed, sodium tripolyphosphate is added, and nanoparticles (50-200 nm in diameter) are formed through ion gelation. The nanoparticles are then dispersed into a sodium alginate solution and stirred for 10 minutes. The solution is then centrifuged to obtain chitosan-sodium alginate nanoparticles encapsulated with methylene blue. The negatively charged sodium alginate coating enhances the stability of the nanoparticles.
[0035] 3) Penetration of microcapsules and nanoparticles into gelatin matrix
[0036] During the preparation of a gelatin solution (5-10% w / v, phosphate buffer, pH 7.0), the PLGA microcapsules and chitosan-alginate nanoparticles prepared in the above steps were added and sonicated at 40kHz for 30 seconds to ensure uniform dispersion. Microcapsules were loaded at 5-10% (w / w) of the total solids to avoid excessive loading that would affect the mechanical properties of the material. Nanoparticles were loaded at 0.1-0.5% (w / w) of methylene blue to ensure color development without affecting the degradation rate.
[0037] 4) Cross-linking of gelatin
[0038] To the gelatin solution containing PLGA microcapsules and chitosan-alginate nanoparticles, a collagenase-sensitive peptide chain (GPLGIAGQ) and a TG enzyme solution were added. The mixture was stirred for 5 minutes, incubated at 37°C for 10 minutes (low cross-linking conditions), and cooled to form a degradable gel. TG enzyme cross-linking conditions: an enzyme concentration of 10 U / g gelatin, reaction at room temperature for 10 minutes to avoid high temperatures damaging the particle structure. After cross-linking, the mixture was heated to 45°C to inactivate the enzyme and avoid damaging the integrity of the PLGA microcapsules.
[0039] 5) Performance test of degradable gel
[0040] After cross-linking by casting according to the method of step 4), the film was immersed in a dopamine solution with a concentration of 1-5 wt% for 30 minutes, and then washed with water until no obvious elution impurities were found, to obtain a gel film with an average thickness of 1 mm. Figure 1 shown.
[0041] 6) Characterization of the gel film and testing of its physical properties and degradation performance.
[0042] a. Determine the maximum absorption wavelength (664nm) by preparing dye solutions of different concentrations;
[0043] b. Thrombin catalyzes the hydrolysis of the chromogenic substrate to release a yellow product (p-nitroaniline, pNA). The absorbance at 405 nm is measured and the enzyme activity is converted using a standard curve.
[0044] c. Dopamine undergoes redox reaction on the electrode surface, generating a characteristic current peak. Dopamine has an oxidation peak at approximately 0.25V, and the peak current is proportional to the concentration.
[0045] d. Physical performance tests and standards:
[0046] ①Tested by ISO37 tensile test.
[0047] ② Simulated peritoneal environment standards: temperature 37±0.5°C, pH 7.3-7.4, ion concentration 0.9% NaCl, and addition of matrix metalloproteinase (MMP-2).
[0048] ③ Adhesion strength: select shear-tensile composite test according to the biomimetic wet adhesion material, at least 5 parallel samples: deviation ≤ 20%.
[0049] The specific preparation examples 1-3 and the related components and performance detection results of the degradable composite biofilms prepared therefrom are shown in Table 1 and Table 2, respectively.
[0050]
[0051] Table 2:
[0052] Test results Preparation Example 1 Preparation Example 2 Preparation Example 3 Gel film color blue transparent blue transparent blue transparent Thrombin Checkout Checkout Checkout Collagenase-sensitive peptide chain Checkout Checkout Checkout Degradation time / min 6.0 5.5 5.0 Elastic modulus / kPa 15.6 15.3 15.8 <![CDATA[Adhesion strength / N / cm 2 > 0.42 0.37 0.49
[0053] The test results in Table 2 above show that thrombin and collagenase sensitive peptide chains are successfully encapsulated in the biodegradable biofilm; the degradation time meets the requirement of 5-8 minutes; the elastic modulus of the biofilm is ≥15kPa, and the elastic modulus of human stomach wall tissue is between 10-50kPa, indicating that the material has good biocompatibility and mechanical matching. The 15kPa gel can avoid compression damage caused by being too hard or failure caused by being too soft. Adhesion strength ≥0.3N / cm 2 , meeting the adhesion strength requirements.
[0054] In addition, the degradable biofilm is vacuum-packed and stored away from light under freeze-drying conditions (-80°C, 24 hours). It has been proven that its shelf life at room temperature is 3 to 6 months. Soaking in saline for 1 minute before use allows the material to restore its elasticity and adhesion.
[0055] Example 1 Biofilm production based on 3D model
[0056] Using a 3D model obtained from preoperative CT images, the three-dimensional surface of the stomach is constructed using surface meshing technology. Next, a virtual "membrane" is assumed to cover the surface of the stomach to estimate its volume. The coverage area of the membrane can be calculated through geometric calculations of the three-dimensional mesh, and the surface area of the stomach can be deduced. Combined with the preoperative expansion of the stomach wall, the volume of the stomach can be calculated. This volume measurement can be used as a reference to estimate the amount of stomach that needs to be retained during surgery. A specific algorithm is used to correlate the different coverage areas of the membrane with the actual stomach volume, and the volume of the stomach that should be retained during surgery can be deduced.
[0057] 1) Import DICOM format CT images into medical image processing software (3D Slicer, Mimics), and have a radiologist outline the stomach (from the cardia to the pylorus). Alternatively, use AI tools (such as nnUNet and TotalSegmentator) for preliminary segmentation. Manually correct the results to distinguish between the gastric cavity and the gastric wall, ensuring that the segmentation results include the entire gastric structure (from the cardia to the pylorus).
[0058] 2) Meshing algorithm to calculate stomach volume. The segmented stomach mask was converted into a triangular mesh (STL format) using the Marching Cubes algorithm. The membrane was assumed to be a continuous surface that adhered closely to the stomach surface and whose geometry was consistent with the 3D stomach mesh. The membrane's coverage area was mapped to the stomach volume, establishing an area-to-volume conversion relationship.
[0059] Triangle patch method: Calculate the area of each triangle patch Ai = 1 / 2 (v1 × v2) where v1 and v2 are the edge vectors of the patch, and the total surface area S = ∑Ai;
[0060] The expansion of the stomach is uniform and isotropic, and the relationship between volume V and surface area S approximately satisfies V∝S 3 / 2 ;
[0061] The gastric surface is divided into N sub-regions (such as cardia, fundus, body, and pylorus), with an area of Si for each region;
[0062] The contribution weights wi of each area and volume are fitted by preoperative CT data to satisfy V = ∑wi Si;
[0063] Define gastric wall expansion δ = V 术前 / V 术中 , δ was adjusted by intraoperative traction status.
[0064] Intraoperative gastric status correction:
[0065] If the preoperative CT shows a drinking state (volume V CT ), intraoperative fasting state volume Vfasting = kV CT, k is a correction factor. A deformation coefficient γ (γ = 0.7 represents a 30% volume reduction) is applied according to the surgical procedure (eg, traction strength, anatomical range of mobilization).
[0066] The retention volume is calculated as: V 保留 =δ·γ·∑wi Si
[0067] In one embodiment, a patient's preoperative CT volume V CT =1200mL, k=0.1, γ=0.7, then V 保留 =0.1×0.7×1200=84mL.
[0068] 3) Biofilm production
[0069] a. By extracting the characteristic parameters of the patient's stomach and its related anatomical structures (such as the greater and lesser curvatures of the stomach, pylorus, cardia, and tumor), an accurate 3D model is obtained. A three-dimensional digital model (STL format) is constructed and then a physical 3D model is obtained through 3D printing technology.
[0070] b. Create a biofilm based on the extent of the stomach to be resected: Sleeve gastrectomy requires resection of the gastric body and fundus along the greater curvature, retaining a volume of 60-100 ml (approximately 1 / 10). For tumors, the biofilm is removed 5 cm from the tumor edge, avoiding important blood vessels. A pre-cut line parallel to the greater curvature of the stomach (or parallel to the cardia / pylorus for tumors) is obtained. Based on the biomechanical deformation modeling in steps 1) and 2), the pre-cut line is marked on the 3D model. An additional 1 cm is added to the pre-cut line to allow for precise biofilm coverage of the resected portion of the stomach and for hemostatic material penetration, achieving adhesion, positioning, and hemostasis. In a cleanroom, a solution of collagenase-sensitive peptide (GPLGIAGQ) and TGase was added to the gelatin solution containing PLGA microcapsules and chitosan-alginate nanoparticles described in Preparation Example 1. The 3D model marked with the pre-cut line was immersed in this solution and incubated at 37°C for 10 minutes. After cross-linking, the temperature was raised to 45°C to inactivate the enzyme, resulting in a cross-linked biofilm attached to the 3D model. The biofilm was then trimmed along the pre-cut line to precisely cover the excision site and sterilized using ethanol or ethylene oxide (to avoid structural damage caused by high temperature and high pressure).
[0071] Example 2 Application of Biofilm in Forming Pre-cut Line Marking
[0072] This embodiment simulates the process of obtaining the pre-cutting line and performing the resection during gastrectomy surgery through animal experiments. Figure 2 The specific steps are as follows:
[0073] 1) Preoperative preparation
[0074] a. Obtain a scan image of the stomach using CT scan;
[0075] b. Extract characteristic parameters of the stomach and its related anatomical structures to obtain an accurate 3D model and the pre-resection site (resection target 100ml±5ml) ( Figure 2 Steps ⑥-⑦);
[0076] c. A pre-cut biofilm was prepared according to Example 1, and a 5 wt % dopamine solution was sprayed on the inside of the biofilm and then washed after 30 min.
[0077] 2) Intraoperative implementation:
[0078] a. Tissue mobilization according to surgical needs ( Figure 2 Steps ①-⑤);
[0079] b. The biofilm is rolled up and passed through the troca into the abdominal cavity, unfolded and covered with the target area (the side coated with dopamine is in contact with the stomach), with the boundary position of the biofilm-covered area at -1 cm as the pre-cut line; for the convenience of illustrating the formation of the pre-cut line, Figure 1 Step ⑧ shows a schematic diagram of the free stomach covered with biofilm, and the boundary of the blue biofilm can be clearly seen.
[0080] c. Precisely cut the gastric tissue along the pre-cut line. During the cutting process, the biofilm is completely degraded within 8 minutes, without affecting the surgical process.
[0081] 3) Postoperative verification
[0082] The actual volume of the resected gastric specimen was measured by injecting water into the specimen, and the actual volume was 98 ml, with an error rate of 3.9%. The resection wound healed rapidly, significantly faster than the surgical method without the biofilm labeling of thrombin.
[0083] It can be seen that the above method of forming a pre-cut line through the biofilm has high accuracy, which can effectively solve the inaccurate resection range of existing methods and improve the quality of surgery.
[0084] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions, and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.
Claims
1. A method for preparing a degradable biofilm, characterized in that the steps include: PLGA microcapsules loaded with thrombin were prepared by solvent method; The dye is embedded in chitosan-sodium alginate nanoparticles to obtain dye nanoparticles; PLGA microcapsules and dye nanoparticles are added to gelatin solution, uniformly dispersed, and then cross-linked to form a solid after casting, thereby obtaining a degradable biofilm.
2. The preparation method according to claim 1, characterized in that The PLGA microcapsules containing thrombin are obtained by emulsifying a PLGA oil phase and a thrombin aqueous phase; And / or, the particle size of the PLGA microcapsules encapsulating thrombin is 1-5 μm.
3. The preparation method according to claim 1, characterized in that The preparation method of the dye nanoparticles comprises embedding the dye in chitosan by an ion gel method, and then adding sodium alginate to obtain the dye nanoparticles; And / or, the dye loading amount in the dye nanoparticles is 0.1-0.5 wt %, and the dye is selected from methylene blue, methyl violet or indocyanine green.
4. The preparation method according to claim 1, characterized in that Transglutaminase is added during the cross-linking process, and the enzyme is inactivated below 50°C after cross-linking for 5-10 minutes at room temperature; And / or, the gelatin solution has a concentration of 5-10% (w / v), and the PLGA microcapsules account for 5-10% of the solid matter obtained by cross-linking.
5. The preparation method according to claim 1, characterized in that The sensor peptide GPLGIAGQ was added during the cross-linking process.
6. Use of a biofilm in preparing a surgical marking material, wherein the biofilm is prepared by the preparation method according to any one of claims 1 to 5.
7. The use according to claim 6, characterized in that The surgical operation is gastrectomy.
8. A biofilm marking method for pre-cutting lines of gastrectomy surgery based on 3D printing, characterized in that: Steps include; S1. Constructing a 3D stomach model based on stomach feature data; S2. constructing a pre-cut portion based on a 3D model of the stomach, and constructing a biofilm attached to the pre-cut portion, wherein the biofilm is prepared by the preparation method according to any one of claims 1 to 5; S3. Place the biofilm obtained in step S2 into the abdominal cavity and adhere it to the stomach. After the biofilm degrades, a dye boundary is formed, which is the pre-cut line for the resection surgery.
9. The biofilm marking method according to claim 1, characterized in that The stomach 3D model is obtained by 3D printing based on stomach characteristic data; And / or, the pre-cut part model includes the gastric body and gastric fundus of the greater curvature of the stomach.
10. The biofilm marking method according to claim 1, characterized in that In step S3, the inner side of the biofilm is coated with dopamine.