Thoracic surgical biological patch and preparation method therefor

Abstract

Disclosed is a thoracic surgical biological patch. The biological patch has a maximum tensile elongation greater than 17.2%, an elastic deformation rate greater than 18.3%, and a single-thread suture pulling force greater than 15.5 N. Further disclosed is a preparation method for the thoracic surgical biological patch. The thoracic surgical biological patch provided by the present invention can be used for lining and sealing incisal margins of lung and trachea tissues, promote the healing and repair of the tissues at the incisal margins, and effectively prevent various complications caused by postoperative air leakage.

Description

A thoracic surgery biological patch and a preparation method thereof TECHNICAL FIELD

[0001] The present application relates to a thoracic surgery biological patch and a preparation method thereof. BACKGROUND

[0002] Lung volume reduction surgery is an effective means of treating lung or bronchial diseases, including lung wedge and local resection, lower esophageal cancer resection, lung pleural resection, total lung resection, lung lobe and lung segment resection, etc. Postoperative lung air leakage is the key to the success of lung volume reduction surgery and is the main cause of lung function and other complications. The "National Expert Consensus on Perioperative Continuous Lung Air Leakage Management Strategy for Thoracic Surgery (2023)" recommends using appropriate surgical techniques to minimize or avoid visceral pleural rupture. Patients with high-risk factors such as chronic obstructive pulmonary disease or emphysema undergoing lung surgery can standardize the use of thoracic surgery biological patches with related indications to prevent lung air leakage after preoperative evaluation. (Recommendation level: 1B level; Expert agreement rate: 97%)

[0003] At present, although the thoracic surgery biological patch on the market can reduce lung air leakage to a certain extent, the problem of lung air leakage still exists. Therefore, the performance of the thoracic surgery biological patch in clinical practice is required to be higher, and the biological patch needs to have better compliance, not easy to slip from the tissue, good fit, etc., in order to reduce the difficulty of surgical operation and postoperative complications, and to reduce the occurrence of intraoperative and postoperative air leakage with better technology and products. SUMMARY

[0004] The present application has found that by controlling the four key parameters of the maximum tensile elongation rate, the elastic deformation rate, the single line suture pulling force, and the value of the elastic deformation rate accounting for the maximum tensile elongation rate of the biological patch, the obtained biological patch has good resilience and good compliance, which can effectively prevent lung air leakage in thoracic surgery.

[0005] In the present application, the determination of the maximum tensile elongation rate and the single line suture pulling force refers to the existing literature (Li Chongchong, Liu Li, Wang Shuo, et al. Comparison of mechanical properties of allogeneic and animal-derived patches [J]. Beijing biomedical engineering, 2021.). The determination of the elastic deformation rate is using the conventional determination method in the art. The prepared biological patch is cut into 4 cm long and 1 cm wide, and the biological patch is clamped on the tensile testing machine along its length direction, and a tensile load is applied at a speed of 100 mm / min to stretch the biological patch until it is pulled apart, and a tensile curve is drawn. Based on the elastic deformation segment of the tensile curve, the elastic deformation rate of the biological patch is calculated.

[0006] As an aspect of the present application, it relates to a thoracic surgery biological patch, the maximum tensile elongation of which is greater than 17.2%, the elastic deformation rate is greater than 18.3%, and the single thread suture pull is greater than 15.5N.

[0007] As a preference, the maximum tensile elongation of the biological patch is 17.2-51.9%.

[0008] As a preference, the elastic deformation rate of the biological patch ranges from 18.3-39.4%.

[0009] As a preference, the single thread suture pull of the biological patch is 15.5-33.4N.

[0010] As a preference, the elastic deformation rate of the biological patch accounts for 49-93% of the value of the maximum tensile elongation.

[0011] As a preference, the maximum tensile elongation of the biological patch is 25.6-51.9%, the single thread suture pull ranges from 17.2-33.4N, the elastic deformation rate ranges from 18.3-39.4%, and the elastic deformation rate accounts for 49-93% of the value of the maximum tensile elongation.

[0012] As a preference, the thoracic surgery biological patch is provided with two perforations at opposite ends of the biological patch.

[0013] As another aspect of the present application, it relates to an anastomat set comprising the above thoracic surgery biological patch. Since the current lung volume reduction surgery cutting is mostly in the form of cutting anastomat, the application of the patch needs to be combined with the pre-installation of the anastomat staple clip, i.e. the patch is stretched and fixed at both ends of the staple clip before the operation.

[0014] As another aspect of the present application, it relates to a preparation method of the above thoracic surgery biological patch, comprising:

[0015] (1) Pretreatment:

[0016] ① The healthy bovine pericardium sheet is immersed in low-osmotic Hank's solution, and the low-osmotic Hank's solution is repeatedly replaced after multiple rinsing to remove the swollen and broken cell fragments, cell nuclei and organelles after decellularization treatment;

[0017] ② The tissue sheet after the above treatment is repeatedly rinsed with physiological saline, each time for 90-150min, and the physiological saline is replaced each time. The total rinsing times are determined according to the fact that no visible cells or cell components and cell fragments can be seen under a microscope, and the protein and nucleic acid quantitative determination is performed until no soluble protein and nucleic acid can be detected.

[0018] (3) removing phospholipids and non-structural proteins and part of the tissue matrix in the tissue piece with a surfactant solution; the surfactant solution can be Tween 80, sodium dodecyl sulfate or Triton X-100;

[0019] (4) soaking in a glutaraldehyde solution with a concentration of 2.5-4% for 3-3.5h;

[0020] (2) chemical modification:

[0021] placing the pretreated tissue material in a Cr 3+ The ion concentration of the hydroxyl chromium solution is 0.0625 mol / dm 3 , OH / Cr is 0.5, and the first water bath oscillation is performed at 20-28 DEG C for 2-4h, the pH of the material treatment solution is detected and is increased by 0.3-0.6 pH units with 10% NaHCO3, and then the second water bath oscillation is performed at 32-40 DEG C for 3-5h to obtain the biological patch in a single-layer sheet shape.

[0022] The present application obtains the thoracic surgery biological patch which can be fused with the lung tissue of a patient, has good biocompatibility, can meet the requirements of treatment effect, and has the anti-calcification and biomechanical properties by decellularizing, removing immunogenicity and chemical modification of the bovine pericardium tissue.

[0023] When the tissue is clamped, the nail hole and suture formed after the titanium nail of the anastomat penetrates through the biological patch can be rapidly contracted due to the elastic retraction deformation of the biological patch to prevent air leakage, and the elastic deformation of the patch also makes the tissue clamping more stable and reliable.

[0024] In the lung volume reduction surgery, the thoracic surgery biological patch can be used for padding and sealing of the lung and tracheal tissue incisal edge, can ensure stable adhesion with the lung tissue, does not slip off, does not cause postoperative air leakage, can promote healing and repair of the incisal edge tissue, reduces postoperative adhesion and pain, greatly improves the use comfort of the patient, reduces the foreign body sensation, greatly improves the postoperative life quality of the patient, and effectively prevents the occurrence of complications such as large-area subcutaneous emphysema, dyspnea, lung infection, incision infection, empyema and the like caused by postoperative air leakage of the lung volume reduction surgery.

[0025] The thoracic surgery biological patch provided by the present application has one rough surface and one smooth surface, the rough surface has a large friction force after contacting with the nail cartridge or nail anvil of the anastomat, so that the thoracic surgery biological patch is not easy to slip off during cutting and anastomosis of the anastomat, after installation, the smooth surface is in close contact with the lung tissue, the adhesion degree with the lung tissue is good, the use comfort of the patient is improved, and the foreign body sensation is reduced. Through surgical implantation of the thoracic surgery biological patch provided by the present application, the patient obtains a lung tissue incisal edge which is reinforced as a whole through tissue fusion and reconstruction, and the whole reconstructed lung tissue has normal blood transportation.

[0026] The thoracic surgery biological patch provided by the application has excellent mechanical properties, and can effectively prevent air leakage of patients (especially for patients with poor lung quality such as chronic obstructive pulmonary disease, long-term smoking, radiotherapy and chemotherapy) during and after operation. Lung air leakage is caused by the fact that the height of the formed anastomosis nail cannot perfectly match the thickness of the tissue. The height of the formed anastomosis nail is fixed, but the thickness of the lung tissue of different patients and different parts is different. When the lung tissue is too thick or too thin relative to the height of the anastomosis nail, air leakage of the lung tissue after cutting and suturing is likely to occur. The thoracic surgery biological patch provided by the application has good resilience and toughness, and can meet the biomechanical properties required for the treatment of patients of different ages and different conditions. After clamping the tissue, the nail hole formed after the nail penetrates the biological patch and the suture site will rapidly shrink due to the elastic retraction deformation of the patch to prevent leakage. At the same time, the elastic deformation of the patch also makes the tissue clamping more stable and reliable, preventing postoperative air leakage.

[0027] Compared with the biological patch obtained by using high molecular synthetic material polyglycolic acid, after implanting the thoracic surgery biological patch obtained by the application, the patient's own tissue cells continuously grow into the thoracic surgery biological patch. The growing tissue cells begin to divide and differentiate, and the mature fibroblasts begin to secrete collagen. The proliferation of collagen tissue thickens the weak lung tissue, and small blood vessels are generated in the thoracic surgery biological patch. With the fusion and reconstruction of the thoracic surgery biological patch with the patient's own lung tissue, the lung tissue at the cutting and suturing site can be strengthened more quickly and more durably.

[0028] The thoracic surgery biological patch obtained by the process of the embodiment removes the immunogenicity of the tissue piece itself, retains the skeleton and part of the tissue matrix of the natural biological tissue, has good biocompatibility in fusion and reconstruction with the patient's lung tissue, is easy for host cells to grow into, and is suitable for repair of various soft tissues. BRIEF DESCRIPTION OF DRAWINGS

[0029] Figure 1 is a schematic view of prepositioning the biological patch on the anastomat; wherein A is a biological patch with different perforation distances; B is an anastomat without prepositioning of the biological patch; C is a prepositioned anastomat biological patch; D is a local enlarged view of E in figure C.

[0030] Reference signs: biological patch 1, perforation 2, anastomat 3, clamping piece 4, nail cartridge or titanium nail anvil 5. DETAILED DESCRIPTION

[0031] The following will be further described in detail through specific embodiments:

[0032] In the process of completing the present application, the inventors first refer to the existing literature (Li Chongchong, Liu Li, Wang Shuo, et al. Comparison of mechanical properties of allogeneic and animal-derived patches [J]. Beijing biomedical engineering, 2021.) for the problem of lung air leakage prone to occur in thoracic surgery, and take the tensile strength, maximum tensile elongation and single line suture pulling force of the biological patch as the parameters for screening the biological patch. However, it is found after determination that the tensile strength of the obtained biological patch fluctuates greatly and the value is unstable, and it is found through animal experiments that the correlation between the tensile strength of the biological patch and the lung air leakage is unstable, and the tensile strength of the biological patch cannot be used as a parameter for screening the biological patch. Therefore, the inventors determine the screening and evaluation parameters of the biological patch as the maximum tensile elongation, elastic deformation rate and single line suture pulling force, and determine the preferred range of the three parameters through clinical trials as the maximum tensile elongation range of 17.2-51.9%, the elastic deformation rate range of 18.3-39.4%, and the single line suture pulling force of 15.5-33.4N. However, in this process, the inventors accidentally found that the value of the elastic deformation rate accounted for the maximum tensile elongation also affects the lung air leakage.

[0033] The inventors have verified this finding through clinical trials, and confirmed that when the maximum tensile elongation, elastic deformation rate and single line suture pulling force are within a certain range, and the value of the elastic deformation rate accounted for the maximum tensile elongation is not less than 49%, it will be beneficial to prevent lung air leakage in thoracic surgery.

[0034] I. Preparation method of biological patch

[0035] (1) Pretreatment

[0036] In the present application, the pretreatment specifically refers to the operation steps of decellularization and removal of immunogenicity of biological tissue, which is a conventional technical means in the art, and the present application does not limit the specific operation process for the purpose of removing cellular tissue components and removing immunogenicity. The specific operation process can be referred to as follows:

[0037] ①Immerse the healthy bovine pericardium sheet tissue in hypotonic Hank's solution, and repeatedly replace the hypotonic Hank's solution after multiple rinsing to fully swell and break various cells in the tissue, so as to remove the swollen and broken cell fragments, cell nuclei and organelles after decellularization treatment.

[0038] ②Rinse the tissue sheet after the above treatment with physiological saline repeatedly, each time for 90-150min, and replace the physiological saline each time. The total rinsing times are determined according to that there are no visible cells or cell components and cell fragments under a microscope, and the protein and nucleic acid quantitative determination is performed until no soluble protein and nucleic acid are detected; for example, the physiological saline rinsing time in a specific embodiment of the present application is preferably 90min.

[0039] ③ Use a surfactant solution to remove phospholipids and non-structural proteins, as well as some tissue matrix immunogenic molecules such as hyaluronic acid, various chondroitin sulfates, and mucopolysaccharides from the tissue slices; the surfactant solution can be Tween 80, sodium dodecyl sulfate (SDS), or Triton X-100. For example, in a specific embodiment of the present invention, Tween 80 is preferred.

[0040] ④ Soak in a 2.5-4% glutaraldehyde solution for 3-3.5 hours. For example, in a specific embodiment of the present invention, the preferred soaking conditions are: 3 hours in a 2.5% glutaraldehyde solution.

[0041] (2) Chemical modification

[0042] Composite crosslinking modification is performed on free carboxyl groups within and / or between tissue matrix collagen molecules and between tissue matrix collagen and tissue matrix. The crosslinking agent is a polymer of a coordination compound of hydroxychromium, enabling the modified tissue sheet to acquire corresponding mechanical properties and anti-calcification properties. For specific procedures, please refer to: placing the pretreated tissue material in Cr... 3+ The ion concentration is 0.0625 mol / dm³. 3 The first water bath shaking is performed in a hydroxychromium solution with an OH / Cr ratio of 0.5, under the following conditions: shaking in a water bath at 20-28°C for 2-4 hours. For example, in a specific embodiment of the present invention, the preferred conditions for the first water bath shaking are shaking in a water bath at 20°C for 2 hours.

[0043] The pH of the material treatment solution was measured and increased by 0.3-0.6 pH units using 10% NaHCO3. A second water bath shaking was then performed at 32-40°C for 3-5 hours to obtain a monolayer sheet-like biological patch. For example, in a preferred embodiment of the present invention, the second water bath shaking conditions were: increasing the pH by 0.3 units using 10% NaHCO3, followed by shaking at 32°C for 3 hours, ultimately yielding a monolayer sheet-like biological patch.

[0044] The obtained biological patch has one rough surface and one smooth surface. As shown in FIG. 1, two or more through holes 2 are formed on the obtained biological patch 1, the through holes 2 are consistent in direction and slightly smaller in size than the width of the clamping sheet 4, the free end of the clamping sheet 4 is sequentially inserted into two through holes 2 (the distance between the two through holes 2 is selected according to the operation requirement), and the position of the biological patch 1 is further adjusted manually to make the biological patch 1 lay flat on the inner side of the clamping sheet 4, so that the rough surface contacts the anastomat nail bin or nail anvil 5, and after contact, there is a large friction force, so that the biological patch 1 is not easy to slip off during cutting and anastomosis by the anastomat 3; the smooth surface is in contact with the lung tissue to ensure the adhesion with the lung tissue. The size of the through hole 2 is slightly smaller than the width of the clamping sheet 4, which can ensure that the biological patch 1 remains in a stretched state when hanging on the clamping sheet 4, and does not slip or fall off from the clamping sheet 4.

[0045] Examples 1-24

[0046] In Examples 1-24, the main differences in the preparation process of the biological patch are shown in Table 1:

[0047] Table 1: Main differences in the preparation method of Examples 1-24

[0048] II. Determination of tensile strength, maximum tensile elongation and single thread suture pull force

[0049] Good mechanical properties can reduce or even avoid lung air leakage, reduce postoperative complications, improve the compliance of the biological patch, and improve the comfort of the patient. In order to explore the relationship between the mechanical parameters and the air leakage, compliance and comfort of the thoracic surgical biological patch, the inventors first consulted the literature and determined the tensile strength, maximum tensile elongation and single thread suture pull force of the biological patch obtained in Examples 1-24 (10 biological patches were obtained in each example, and the parameters were determined according to the literature, and the average value was taken) based on the existing literature (Li Chongchong, Liu Li, Wang Shuo, et al. Comparison of mechanical properties of allogeneic and animal-derived patches [J]. Beijing biomedical engineering, 2021.) The determination results are shown in Table 2:

[0050] Table 2: Maximum tensile elongation, tensile strength and single thread suture pull force of Examples 1-24

[0051] From Table 2 above, it can be concluded that the tensile strength of the 10 biological patches obtained in each of Examples 1-24 fluctuates greatly and the values are unstable, which may be because there are other influencing factors of the tensile strength of the biological patch that the inventors have not found, so that the tensile strength of the biological patch cannot be accurately controlled, which is not suitable for use as a surgical material.

[0052] The maximum tensile elongation of the biological patches obtained in Examples 1-24 ranges from 15.6-51.9%, the tensile strength is 16.3-39.5 MPa, and the single-thread suture pull force ranges from 5.1-33.4 N.

[0053] Ten biological patches were obtained for each example, and each was cut into a length of 4 cm and a width of 1 cm to test whether it could be preloaded onto the stapler.

[0054] The preloading of the biological patch onto the stapler requires appropriate stretching, which is related to the maximum tensile elongation. In theory, the larger the maximum tensile elongation, the better. If the maximum tensile elongation is too small, the biological patch will break after a small distance of stretching. Specifically, ten biological patches obtained from Example 21 (maximum tensile elongation of 15.6±0.1%) were assembled into a stapler, and it was found that nine were torn apart and one had obvious cracks. Ten biological patches obtained from Example 14 (maximum tensile elongation of 16.3±0.5%) were assembled into a stapler, and it was found that seven were torn apart and three had obvious cracks. This reflects the toughness of the biological patch, which is extremely difficult to deform, and after deformation, it quickly enters permanent deformation and rapidly breaks down.

[0055] It was found that biological patches with a maximum tensile elongation of 15.6% and 16.3% cannot meet the requirements for installation onto a stapler. Based on the biological patches obtained from other examples meeting the stretching requirements, they can be preloaded onto a stapler, so the maximum tensile elongation of the biological patch obtained from Example 3 is 17.2±0.2%, which is the minimum value of the maximum tensile elongation that meets the installation of the biological patch onto the stapler.

[0056] Animal experiments were conducted on examples that met the preloading requirements of the stapler (Examples 1-13, Examples 15-20, and Examples 22-24).

[0057] III. Animal Experiments

[0058] White pigs are ideal animal models for studying human lung diseases and injuries due to many similarities in anatomical structure and physiological function between white pigs and humans. White pigs are of moderate size, making them easy to handle and manage. Compared to small animals such as mice and rats, white pigs provide more operating space, making it easier for researchers to perform surgical procedures. At the same time, white pigs are easy to raise and manage under laboratory conditions, are less likely to develop infections after surgery, are easier to control in the long term, and have a high long-term survival rate.

[0059] The healthy experimental animals were purchased according to the SOP-5 Experimental Animal and Receiving Operation Regulations, and the experimental animals were provided by Jiangxi Yinsha Biotechnology Co., Ltd. (License No. SCXK (Jiangxi) 2023-0002). All animals were used for animal experiments for the first time at the beginning of the experiment. All test animals were quarantined before surgery. Only test animals that passed the quarantine were included in the study.

[0060] 1. Inclusion criteria

[0061] Comply with the requirements of the National Animal Quarantine Management Measures; body weight is 50-60 kg; physiological indicators are normal; no obvious serious animal diseases.

[0062] 2. Feeding

[0063] The experimental animals were tracked and managed according to the SMP-5 Experimental Animal Care and Management Regulations. The experimental animals were given an appropriate amount of feed twice a day. The animals could drink water freely through an automatic water supply system. The feed and drinking water were clean and safe.

[0064] 3. Grouping and number

[0065] 1-month experimental group: 110 pigs in total; 3-month experimental group: 110 pigs; gender is not limited, and random grouping. Examples 1-13, Examples 15-20, and Examples 22-24 (a total of 22 examples), each example uses a total of 10 pigs (5 pigs for the 1-month experimental group and 5 pigs for the 3-month experimental group) for experiments. The 10 biological patches obtained from each example correspond to one pig.

[0066] Animal information was recorded during the experiment, including body weight, operation time, survival days, biological patch information, etc.

[0067] Preoperative measurement of blood routine, blood biochemistry, coagulation function, and blood gas.

[0068] Postoperative measurement of blood routine, blood biochemistry, coagulation function, and blood gas at 1 month and 3 months after surgery.

[0069] 4. Experimental steps

[0070] ①Preoperative preparation; ②Preoperative blood test according to routine examination requirements and record preoperative data; ③During operation, the test pig is fully anesthetized, intubated, and assisted with a respirator; ④The pig is placed in a supine position, the abdomen is exposed by laparotomy, the lung tissue is separated, and an appropriate position is selected; the incision is cut; ⑤A 4cm-long and 1cm-wide biological patch is installed on a disposable anastomat, the anastomat with the pre-installed biological patch is placed at the predetermined position, and the angle and depth are adjusted to ensure close fit with the surrounding tissue; ⑥The anastomotic stoma and the biological patch are carefully examined to observe whether the biological patch is fixed stably, whether the patch and the lung tissue are anastomosed well, whether the resilience of the biological patch meets the requirements, whether there is air leakage at the staple holes and suture lines, and whether the abdomen is closed after confirming that there is no air leakage.

[0071] After the operation, the test animals are sent to the observation room for observation, and the respirator is supported until the animals wake up. After the animals can stand and move, they are sent to the animal house for feeding. The animals are alternately illuminated for 12h / 12h per day, and are given an appropriate amount of animal feed once per day in the morning and afternoon, and can drink water freely. The mental state, appetite, respiration, wound complications, and other adverse symptoms are observed daily.

[0072] After the operation, the animals are given intramuscular injection of ceftriaxone sodium 2g for 1 week for anti-infection. The mental state, appetite, respiration, wound healing state, and whether the animals have the symptoms of vomiting, subcutaneous emphysema, dyspnea, lung infection, wound infection, empyema, and the like are observed every day. The surgical wound is disinfected with iodophor until the wound heals. The veterinarian conducts regular physical examination, blood test, and takes anti-infection prevention and treatment measures according to the results. The postoperative medication and adverse events (including death, infection, etc.) within 24h are recorded.

[0073] At the end of the test, whether the patch is complete, has defects, and leaks air is observed macroscopically, whether there is a thrombus on the surface, and whether there are changes such as bleeding and necrosis in the surrounding tissue.

[0074] 5. Evaluation criteria for animal experiments

[0075] ①During the operation, whether there is air leakage at the anastomotic stoma and the surrounding tissue after the operation. ②Whether the biological patch can be closely fitted at the anastomotic stoma. ③The incidence of postoperative complications such as infection and rejection during feeding. ④Biological patch gross specimen observation: the test animals are sacrificed at 1 month and 3 months after the operation, respectively, to observe whether the anastomotic stoma is restored well, and whether the biological patch promotes tissue regeneration and repair. ⑤Histological examination: the surgical biological patch and tissue specimens are taken at 1 month and 3 months after the operation for histological examination to observe whether there are thromboembolism, inflammation, necrosis, and the like.

[0076] 6. Results of animal experiments

[0077] (1) The maximum tensile elongation range is 17.2-51.9%, which can meet the requirements of installation on the anastomat.

[0078] None of the pigs with the biological patch of the 13 embodiments appeared lung air leakage, which were example 1, example 4-6, example 8-13, example 18, example 23-24, respectively.

[0079] The pigs without lung air leakage successfully completed the operation, the biological patch was closely attached to the lung tissue, there was no air leakage at the resection edge, and no instrument failure of the anastomat and components occurred during the operation. The lung resection edge was smooth during the operation, with no air leakage and exudate. At the follow-up endpoints of 1 month and 3 months, scar tissue formed at the lung resection edge, which was not decomposed, which indicated that the biological patch could act as a stent to promote the repair and healing of the lung resection edge, and embodied the good biocompatibility of the thoracic surgery biological patch, which could form good combination with the surrounding tissue and promote the repair and healing of the lung resection edge.

[0080] During the operation, the vital signs of the anesthetized animals were normal, and no adverse events occurred.

[0081] After the operation, the number, shape and quality of the whole blood cells of the animals were normal, and the detection results of white blood cells, red blood cells, hemoglobin and platelets were stable, and no abnormality of important blood routine indexes was found. The liver and kidney function values were almost within the normal range, and no abnormality of liver and kidney function was found.

[0082] During the survival and feeding of the animals, the general condition was good, the body temperature, diet and excretion were normal, the autonomous activity was good, and no obvious weight loss, fever, anorexia, mania and other abnormal performances occurred, and the animals successfully survived to the endpoint.

[0083] (2) The single thread suture pulling force reflects the ability of the biological patch to resist shear force. If the single thread suture pulling force is too small, tearing is likely to occur at the nail hole and the suture line. The biological patches obtained by example 3 (single thread suture pulling force 5.1±0.3N), example 2 (single thread suture pulling force 12.5±0.1N) and example 16 (single thread suture pulling force 14.9±0.3N) had obvious tearing at the nail hole, and the tearing intensified over time, while the biological patch of example 19 (single thread suture pulling force 15.5±0.5N) had no obvious tearing, which indicated that in order to use titanium nails to fix the biological patch without tearing at the nail hole and the suture line, the single thread suture pulling force of the biological patch should be no less than 15.5N, i.e. the single thread suture pulling force of the biological patch should be in the range of 15.5-33.4N.

[0084] (3) In theory, the biological patch needs to have a certain tensile strength when pre-installed in the anastomat, which helps to ensure the compliance of the biological patch and prevent lung air leakage. However, in the animal experiment, it was found that:

[0085] The biological patches obtained by example 2, example 3, example 7, example 15-17, example 19, example 20 and example 22 had lung air leakage.

[0086] Specifically, the experimental groups of the biological patches obtained by Example 2 (tensile strength 16.3 ± 2.2 MPa), Example 19 (tensile strength 19.5 ± 2.3 MPa), Example 3 (tensile strength 20.2 ± 2.5 MPa), Example 17 (tensile strength 20.4 ± 5.3 MPa), Example 20 (tensile strength 22.9 ± 1.4 MPa), Example 15 (tensile strength 25.4 ± 1.9 MPa), Example 16 (tensile strength 25.5 ± 3.3 MPa), Example 22 (tensile strength 29.5 ± 1.6 MPa), and Example 7 (tensile strength 39.5 ± 3.1 MPa) appeared lung air leakage.

[0087] However, the examples with similar tensile strength as these examples did not appear lung air leakage, for example, Example 12 (tensile strength 19.1 ± 4.9 MPa) and Example 18 (tensile strength 21.6 ± 3.4 MPa) with similar tensile strength as Example 19 and Example 3 did not appear lung air leakage. Example 4 (tensile strength 24.3 ± 3.1 MPa) and Example 8 (tensile strength 26.2 ± 4.7 MPa) with similar tensile strength as Example 15 and Example 16 did not appear lung air leakage. Example 6 (tensile strength 31.8 ± 2.1 MPa), Example 9 (tensile strength 27.8 ± 1.5 MPa), Example 11 (tensile strength 30.7 ± 2.6 MPa), Example 23 (tensile strength 33.2 ± 4.1 MPa), and Example 24 (tensile strength 33.7 ± 4.2 MPa) with similar tensile strength as Example 22 and Example 7 did not appear lung air leakage.

[0088] Since it is unstable to explore whether the biological patch will appear lung air leakage based on the tensile strength, it is necessary to reconsider the mechanical parameters that can indicate lung air leakage.

[0089] Based on the research of the inventors of the present application on the application of animal-derived biological materials in thoracic surgery implantation for many years, the inventors found that the imperfect fit between the staple height of the anastomotic staple and the tissue thickness is an important reason leading to lung air leakage. The staple height of the anastomotic staple is fixed, but the lung tissue thickness of different patients and different parts is different, when the lung tissue is too thick or too thin relative to the anastomotic staple height, it is possible to cause lung incision edge tissue air leakage. Therefore, the biological patch needs to have good resilience, the inventors want to introduce “elasticity” into the biological patch of biological origin, and replace “tensile strength” with “elastic deformation rate”, and continue to explore the thoracic surgical biological patch for preventing lung air leakage by comprehensively considering the three mechanical parameters of elastic deformation rate, maximum tensile elongation rate and single line suture pulling force.

[0090] Four, determination of elastic deformation rate

[0091] 1. Measurement method

[0092] A 4 cm long and 1 cm wide biological patch was clamped on a tensile testing machine in the length direction, and a tensile load was applied to stretch the biological patch at a speed of 100 mm / min until it was pulled apart. A tensile curve was drawn, and the elastic deformation rate of the biological patch was calculated based on the elastic deformation section of the tensile curve during the test.

[0093] 2. Measurement results

[0094] Table 3: Elastic deformation rates of Examples 1-24

[0095] As can be seen from Table 3, the elastic deformation rates of the biological patches obtained in Examples 1-24 are relatively stable, and the elastic deformation rate range is 8.9-39.4%.

[0096] In addition to the reasons for the occurrence of lung air leakage in the experimental groups corresponding to Examples 2, 3, and 16, in which tears occurred at the staple holes and suture lines, based on the experimental results of the animal experiments, the inventors unexpectedly found that the biological patches obtained using Example 20 (elastic deformation rate 12.7±0.1%) and Example 22 (elastic deformation rate 14.9±0.4%) both had 7 pigs that experienced lung air leakage; the biological patches obtained using Example 19 (elastic deformation rate 15.4±0.7%) and Example 7 (elastic deformation rate 16.2±1.1%) both had 4 pigs that experienced lung air leakage; the biological patch obtained using Example 15 (elastic deformation rate 17.2±1.9%) had only 3 pigs that experienced lung air leakage; and the biological patch obtained using Example 17 (elastic deformation rate 17.5±0.6%) had only 1 pig that experienced lung air leakage. That is, as the elastic deformation rate increased, the number of pigs that experienced lung air leakage decreased, and the elastic deformation rates of the biological patches obtained in these examples were all lower than those of the examples in which lung air leakage did not occur.

[0097] Therefore, the inventors speculate that the greater the elastic deformation rate, the less likely it is that lung air leakage will occur. Based on the experimental results of the animal experiments, it can be preliminarily proven that the inventors' introduction of the "elastic deformation rate" is correct, and it is speculated that the reason for the occurrence of lung air leakage is that the biological patch has poor resilience, a slightly poor degree of fit with the pig's tissue, and insufficient contraction elasticity at the staple holes and suture lines, thereby causing lung air leakage.

[0098] Based on the animal experiments, it can be preliminarily determined that the range of the elastic deformation rate of the biological patch should be controlled to be 18.3-39.4%.

[0099] Five, clinical trial one

[0100] 1. Inclusion criteria, exclusion criteria, and collection of clinical data

[0101] (1) Inclusion criteria

[0102] ① Age: 18-75 years old, gender unlimited; ② Meet the clinical surgical indications, including various types of lung volume reduction surgery (surgical approach is not required, traditional open, laparoscopic, robotic); ③ Can understand the purpose of the test, voluntarily participate and sign the informed consent, willing to accept relevant examination and clinical follow-up.

[0103] (2) Exclusion criteria

[0104] ① Patients who need to be transferred to the intensive care unit for ventilator-assisted breathing after surgery; ② Patients who are converted to open chest surgery due to bleeding or severe chest adhesion during surgery; ③ Patients diagnosed with bronchopleural fistula after surgery; Patients who died or had incomplete clinical data during hospitalization.

[0105] (3) Clinical data collection

[0106] Collect patient data that meets the conditions: ① Demographic data: gender, age, body mass index (BMI), smoking history; ② Preoperative assessment data: lung function (forced expiratory volume in one second (FEV1%) and forced expiratory volume in one second to forced vital capacity ratio (FEV1% / FVC%)), peripheral blood lymphocyte count, albumin, prognostic nutritional index (PNI); ③ Surgical data: surgical site, surgical approach (lobectomy or segmentectomy), whether lymph node dissection was performed, whether there was pleural adhesion, pathological result was benign or malignant, whether there was air leakage in the water test lung expansion method; ④ Postoperative stage: daily air leakage assessment, postoperative 12h pleural cavity pressure difference.

[0107] 2. Test grouping

[0108] Test group: divided into 13 groups, respectively using the 13 examples of animal experiments to obtain biological patches for clinical trials, 10 patients in each group, respectively using the biological patch obtained by the example to perform lung volume reduction surgery. Before surgery, the biological patch is pre-installed on the anastomat.

[0109] Control group: select patients who only use anastomat to close / anastomose to perform surgery.

[0110] 3 Test procedure

[0111] 1. Test procedure

[0112] The specific procedure of the test includes: (1) signing the informed consent form; (2) case screening, confirming the clinical inclusion and exclusion criteria, and synchronously completing the baseline data collection; (3) determining the operation time; (4) grouping into the test group or the control group according to the random grouping; (5) performing the operation; (6) observing the air leakage and recording it into the operation report; (7) evaluating the postoperative condition of the subject; (8) completing the pre-discharge, 1-month postoperative, and 3-month postoperative follow-up of the subject according to the relevant inspection requirements in the scheme.

[0113] All patients underwent surgery under general anesthesia. The diseased lung was closed and resected using a stapler during the operation, and the bronchus was disconnected using a stapler. After complete hemostasis, an appropriate amount of warm normal saline was poured into the chest cavity for chest irrigation, and lung ventilation was performed at the same time. The lung tissue and bronchial stump in the chest cavity were observed for air leakage. The lung wound or bronchial stump with air leakage was reinforced and sutured with absorbable thread. Warm normal saline was poured into the chest cavity again to observe the air leakage. After confirming that there was no air leakage, the chest was closed. One or two chest closed drainage tubes were placed in the patient after the operation, which reached the top of the pleura, and the drainage tubes were connected to the chest drainage bottle containing 500 ml of normal saline. The air leakage of the chest drainage bottle was monitored after the operation, and symptomatic treatment was given, such as oxygen inhalation, atomization, expectorant, asthma, nutrition support, and anti-infection, etc.

[0114] 4. Test method

[0115] Prospective, multi-center, randomized controlled, open, and superiority.

[0116] 5. Test results

[0117] (1) Comparison of general data

[0118] First, 150 patients who met the conditions were included in the clinical trial study, and the occurrence of lung air leakage is shown in Table 4.

[0119] Table 4: Test results of clinical trial one Note: Other lung volume reduction surgery includes total lung resection, lung pleurodesis, etc. The number of cases with lung air leakage is before the “ / ”; the total number of patients is after the “ / ”.

[0120] From the results of Table 4, it can be found that:

[0121] (1) On the basis of the maximum tensile elongation range (17.2-51.9%) determined in the previous experiments, the elastic deformation rate range (18.3-39.4%), and the single suture tension range (15.5-33.4N), the test group of the biological patch obtained by Example 6 (elastic deformation rate 19.9±1.1%) showed lung air leakage. In theory, the greater the elastic deformation rate, the better. The inventors speculate that the elasticity of the biological patch needs to be further improved to ensure that the biological patch can further seal the nail hole and the suture to meet the requirements of frequent expansion and contraction of the lung.

[0122] Further comparison, the elastic deformation rate of the biological patch obtained by Example 6 (elastic deformation rate 19.9±1.1%), Example 11 (elastic deformation rate 18.3±0.2%) and Example 24 (elastic deformation rate 18.4±0.3%) was compared. The inventors unexpectedly found that the value of the elastic deformation rate of the biological patch obtained by Example 6 accounted for the maximum tensile elongation (41%) was smaller. Therefore, the inventors speculate that there is some correlation between the value of the elastic deformation rate and the clinical results, and further verify it in clinical experiments.

[0123] Table 5: Value of elastic deformation rate of Examples 1-24 accounted for the maximum tensile elongation

[0124] As can be seen from Table 5, the value of the elastic deformation rate of the biological patch obtained by Examples 1-24 accounted for the maximum tensile elongation is in the range of 31-93%.

[0125] Six, clinical test two

[0126] 1. Based on the preparation method of the biological patch, the biological patch with the following parameters is obtained by adjusting the preparation method.

[0127] Table 6: Biomechanical parameters of Examples 25-26

[0128] Table 7: Main differences in preparation methods of Examples 25-26

[0129] On the basis of clinical test one, 20 patients undergoing lung volume reduction surgery were further enrolled, and Examples 25 and 26 (10 patients for each example, without limiting the lung resection site) were used to further verify the effect of the value of the elastic deformation rate accounted for the maximum tensile elongation on the clinical application effect. The test results are shown in Table 8.

[0130] Table 8: Test results of clinical test two

[0131] Compared with the lung air leakage in the clinical trial I, the maximum tensile elongation, the elastic deformation rate and the single thread suture pull force of the biological patch obtained in Example 25 and Example 26 meet the application requirements mentioned in the clinical trial I, except that the value of the elastic deformation rate accounting for the maximum tensile elongation is lower than 49% (Example 24: the value of the elastic deformation rate accounting for the maximum tensile elongation is 49%). It can be proved again from Table 8 that the value of the elastic deformation rate accounting for the maximum tensile elongation is indeed related to the lung air leakage. Although the air leakage of Example 25 and Example 26 is improved, it is still unacceptable in the medical field.

[0132] The mechanical properties of the biological patch need to meet the requirements of the operation of a specific site. Compared with the digestive surgery biological patch, the digestive surgery biological patch is used to prevent liquid leakage and bleeding, while the thoracic surgery biological patch is used to prevent air leakage. Since the viscosity of liquid is greater than that of gas, the biological patch used to prevent lung air leakage needs to have better elasticity. According to the animal experiment, the clinical trial I and the clinical trial II, the biomechanical parameters of the biological patch are as follows: the maximum tensile elongation ranges from 17.2% to 51.9%, the elastic deformation rate ranges from 18.3% to 39.4%, the single thread suture pull force ranges from 15.5N to 33.4N, and the value of the elastic deformation rate accounting for the maximum tensile elongation ranges from 49% to 93%. The biomechanical characteristics of the biological patch with the above parameters are excellent, which can ensure the adhesion of the biological patch to the lung tissue and prevent lung air leakage.

[0133] In addition, in addition to the examples that cannot be installed on the anastomat and the examples with lung air leakage in the animal experiment and the clinical trial I, the biological patches obtained in Example 1, Example 4-5, Example 8-13, Example 16, Example 18 and Example 23-24 meet the requirements of application to thoracic surgery, and the specific parameter ranges are as follows: the maximum tensile elongation ranges from 25.6% to 51.9%, the single thread suture pull force ranges from 17.2N to 33.4N, the elastic deformation rate ranges from 18.3% to 39.4%, and the value of the elastic deformation rate accounting for the maximum tensile elongation ranges from 49% to 93%.

[0134] The preferred embodiments of the present application are described in detail above. It should be understood that those skilled in the art can make many modifications and changes without creative labor based on the concept of the present application. Therefore, any technical solutions obtained by logical analysis, reasoning or limited experiments based on the prior art according to the concept of the present application shall be within the protection scope defined by the claims.

Claims

1. A thoracic surgical biological patch, characterized by: The maximum tensile elongation of the biological patch is greater than 17.2%, the elastic deformation is greater than 18.3%, and the single-thread suture pull is greater than 15.5N.

2. The thoracic surgical biological patch of claim 1, wherein: The maximum tensile elongation of the biological patch is 17.2-51.9%.

3. The thoracic surgical biological patch of claim 2, wherein: The elastic deformation of the biological patch ranges from 18.3-39.4%.

4. The thoracic surgical biological patch of claim 3, wherein: The single-thread suture pull of the biological patch is 15.5-33.4N.

5. The thoracic surgical biological patch of claim 4, wherein: The elastic deformation of the biological patch accounts for 49-93% of the maximum tensile elongation.

6. The thoracic surgical biological patch according to any one of claims 1-5, wherein: The maximum tensile elongation of the biological patch is 25.6-51.9%, the single-thread suture pull ranges from 17.2-33.4N, the elastic deformation ranges from 18.3-39.4%, and the elastic deformation accounts for 49-93% of the maximum tensile elongation.

7. The thoracic surgical biological patch according to any one of claims 1-6, wherein: The thoracic surgery biological patch is made of bovine pericardium.

8. The thoracic surgical biological patch of any one of claims 1-7, wherein, The thoracic surgery biological patch is provided with two perforations located at opposite ends of the biological patch.

9. An anastomat kit comprising the thoracic surgery biological patch of any one of claims 1-8.

10. A method of making a thoracic surgical bioprosthetic patch according to any one of claims 1-7, characterized in that, Comprising: (1) Pretreatment: ① Soak the healthy bovine pericardium sheet tissue in hypotonic Hank's solution, and repeatedly replace the hypotonic Hank's solution after multiple rinsing to remove the swollen and broken cell fragments, cell nuclei and organelles after decellularization treatment; ② Rinely the tissue sheet after the above treatment with physiological saline repeatedly, 90-150 min each time, replace the physiological saline each time, and the total rinsing times are determined according to that there are no visible cells or cell components and cell fragments under a microscope, and no soluble protein and nucleic acid are detected by protein and nucleic acid quantitative determination; ③ Remove the phospholipids and non-structural proteins and part of the tissue matrix in the tissue sheet with a surfactant solution; the surfactant solution can be Tween 80, sodium dodecyl sulfate or triton X-100; ④ Soak in a 2.5-4% glutaraldehyde solution for 3-3.5h; (2) Chemical modification: The pretreated tissue material was placed in a Cr 3+ The ion concentration was 0.0625 mol / dm 3 The first water bath oscillation was performed in a hydroxyl chromium solution with OH / Cr of 0.5, under the conditions of 20-28°C water bath oscillation for 2-4h, the pH of the material treatment solution was detected and increased by 0.3-0.6 pH units with 10% NaHCO3, and then the second water bath oscillation was performed, under the conditions of 32-40°C water bath oscillation for 3-5h, to obtain a single-layer sheet-shaped biological patch.

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