Method for constructing a myocardial ischemia-reperfusion injury model

Abstract

The application relates to a method for constructing a myocardial ischemia-reperfusion injury model, inserting a blood vessel blocking body into a blood vessel of a target mouse and sending the blood vessel blocking body to a position of a coronary artery to be blocked, blocking the position of the coronary artery to be blocked by using the blood vessel blocking body, and making part or all of the blood vessel blocking body made of biodegradable material, so as to simulate myocardial ischemia-reperfusion injury through degradation of the blood vessel blocking body in the target mouse. The blood vessel blocking body is made of biodegradable material, the biodegradable material can be degraded in the animal body, myocardial ischemia-reperfusion can be simulated without additional operation, the different thicknesses of the degradable parts of the blood vessel blocking body can be set, the coronary artery ischemia-reperfusion time can be accurately controlled, the controllability of the reperfusion time is improved, and the disease process and reperfusion injury of human myocardial ischemia / acute myocardial infarction can be accurately simulated.

Description

TECHNICAL FIELD

[0001] The application relates to a method for constructing a myocardial ischemia-reperfusion injury model and belongs to the technical field of animal model construction. BACKGROUND

[0002] After acute myocardial infarction, early recovery of myocardial perfusion through thrombolysis or percutaneous coronary intervention (PCI) is the most effective method to reduce the myocardial infarction area and improve the clinical prognosis. However, the recovery of blood flow in ischemic myocardium may cause irreversible myocardial ischemia-reperfusion injury (MIR), and due to the lack of effective clinical intervention means, reducing reperfusion injury has become the focus of the prevention and treatment of ischemic heart disease.

[0003] In the basic research of myocardial ischemia-reperfusion injury, a myocardial ischemia-reperfusion injury model is usually used to simulate myocardial ischemia / acute myocardial infarction in the human body. For example, the patent application for invention with publication number CN115777629A discloses a method for establishing and evaluating a myocardial ischemia-reperfusion model, which includes the following steps: preoperative weighing and anesthesia fixation, tracheal intubation, double-needle ligation of the left anterior descending branch of the coronary artery, postoperative suturing, and removal of the tracheal tube after the animal recovers from spontaneous breathing. The modeling success rate and model stability of the myocardial ischemia-reperfusion model are improved, the required sample size is significantly reduced, and the model evaluation indicators are more comprehensive and accurate. For another example, the patent application for invention with publication number CN114758726A discloses a myocardial ischemia-reperfusion influencing factor data detection method, which includes the following steps: step one: myocardial ischemia influencing factor acquisition: through experiments and data queries, a series of reactions that are prone to occur in myocardial ischemia-reperfusion are obtained, and it is concluded that the essential cause is cell pyroptosis during myocardial ischemia-reperfusion; step two: cell pyroptosis influencing factor: a mouse myocardial ischemia-reperfusion model is established, and a control group, an ischemia group, and an experimental group are set up. In the control group, only the mice are subjected to surgical operation without ligation of the coronary artery. After analysis, it is concluded that injection of MCC950 for inhibiting NLPR3 inflammasome and Caspase-1 inhibitor within 5 minutes of myocardial ischemia reduces cell pyroptosis, thereby reducing myocardial ischemia-reperfusion injury. Injection of MCC950 and Caspase-1 inhibitor before myocardial ischemia can reduce myocardial necrosis caused by myocardial ischemia. The above-mentioned technologies each have their own characteristics. However, the myocardial ischemia-reperfusion models in the above-mentioned technologies are all reperfused when myocardial ischemia does not exceed 1 hour (a small animal model is prepared by ligation of the coronary artery for no more than 1 hour followed by reperfusion), which cannot simulate the long disease process (e.g., 4-6 hours or more) from myocardial ischemia to stent surgery in a clinical patient. In addition, the above-mentioned technologies all need to perform a thoracotomy on a small animal when establishing the myocardial ischemia-reperfusion model, which itself causes a large trauma and an immune inflammatory response. On the one hand, it is easy to cause model construction failure (death of the small animal), and on the other hand, it affects the body functions of the small animal (additional trauma and response compared to the treatment of myocardial ischemia / acute myocardial infarction in the human body), thereby interfering with the research and evaluation of myocardial ischemia-reperfusion injury. The defects of the above-mentioned myocardial ischemia-reperfusion models greatly limit the research of the pathological mechanism of myocardial ischemia-reperfusion injury, especially the short ischemia period (≤1 hour) hinders the research and development of drugs for prophylactic administration during the ischemia period, and limits the preclinical evaluation of the treatment effect of drug and non-drug therapies. SUMMARY

[0004] To overcome the above-mentioned defects of the prior art, the present application provides a method for constructing a myocardial ischemia-reperfusion injury model, which can accurately simulate the disease process and reperfusion injury of myocardial ischemia / acute myocardial infarction in the human body.

[0005] The technical scheme adopted by the present application is: a method for constructing a myocardial ischemia-reperfusion injury model, a blood vessel blocking body is inserted into a blood vessel of a target mouse and sent to (or guided to) a position of a coronary artery to be blocked, the position of the coronary artery to be blocked is blocked by the blood vessel blocking body, part or all of the blood vessel blocking body is made of a biodegradable material (polymer), and the blood vessel blocking body is degraded in the target mouse to simulate myocardial ischemia-reperfusion injury (establish a myocardial ischemia-reperfusion injury model).

[0006] Preferably, the position of the coronary artery to be blocked is the position of the left anterior descending branch of the coronary artery.

[0007] Preferably, the blood vessel blocking body is inserted into the coronary artery (the position of the coronary artery to be blocked) from the radial artery of the upper limb or the femoral artery of the lower limb of the target mouse (minimally invasive).

[0008] Preferably, the blood vessel blocking body is inserted into the blood vessel of the target mouse under the guidance of photoacoustic imaging technology.

[0009] Preferably, the position of the coronary artery to be blocked is completely blocked by the blood vessel blocking body, the blood flow is blocked, and reperfusion is achieved by controlling the biodegradability of the degradable part of the blood vessel blocking body.

[0010] Preferably, the biodegradable material is polylactic acid (or polylactide).

[0011] Preferably, the blood vessel blocking body is umbrella-shaped, including a collapsible and expandable umbrella surface and an intervention guide wire, one end of the intervention guide wire is connected to the center of the umbrella surface by a connection method that can be disconnected when the tension reaches a set threshold.

[0012] Further, one end of the intervention guide wire passes through the center of the umbrella surface, the center of the umbrella surface is provided with a through hole for the intervention guide wire to pass through, the end of the intervention guide wire passing through the center of the umbrella surface (the end outside the umbrella surface) is provided with an outer plug that can deform towards the inside of the umbrella surface under external force, the outer plug abuts against the outside of the center of the umbrella surface, and the intervention guide wire in the inside direction of the umbrella surface is provided with an inner plug that does not deform under external force, the inner plug abuts against the inside of the center of the umbrella surface; or, one end of the intervention guide wire is interference-fitted or bonded with the center of the umbrella surface.

[0013] Preferably, the thickness of the middle part of the umbrella surface is smaller than the thickness of the edge of the umbrella surface.

[0014] Preferably, at least the middle part of the umbrella surface is made of the biodegradable material.

[0015] Further, the middle part of the umbrella surface is made of the biodegradable material, and the edge of the umbrella surface is made of a hard medical instrument material for coronary intervention surgery, such as nickel-titanium alloy, cobalt-chromium alloy, or stainless steel.

[0016] Preferably, the blood vessel obstruction body is made of 3D printing technology.

[0017] Preferably, by setting different thicknesses of the degradable part of the umbrella surface, the degradation time of the degradable part of the umbrella surface is changed (the time of achieving ischemia-reperfusion in the animal body), so as to accurately control the coronary artery ischemia-reperfusion time and simulate the disease process of different time lengths from myocardial ischemia to surgical treatment of the clinical patient.

[0018] The beneficial effects of the present application are:

[0019] 1. The biodegradable material is used to prepare the blood vessel obstruction body, which can be degraded in the animal body without additional operation, so as to simulate myocardial ischemia-reperfusion, and the different thicknesses of the degradable part (the part made of biodegradable material) of the blood vessel obstruction body can be set to accurately control the coronary artery ischemia-reperfusion time, improve the controllability of the reperfusion time, accurately simulate the disease process and reperfusion injury of human myocardial ischemia / acute myocardial infarction (clinical myocardial ischemia / acute myocardial infarction patients), provide a powerful model support for the mechanism research of myocardial ischemia-reperfusion injury, help the in-depth research of the pathological mechanism of myocardial ischemia-reperfusion injury, facilitate the more objective preclinical evaluation of drug and non-drug therapy for myocardial ischemia-reperfusion injury, and promote the effective research and development of related clinical treatment schemes.

[0020] 2. The present application adopts a minimally invasive method to construct a myocardial ischemia-reperfusion injury model, without the need for thoracotomy on the target mouse, and the trauma of the target mouse is small, almost no immune inflammatory reaction, the success rate and stability of the model construction are high, the sample quantity is small, and the impact of the model construction on the physical function of the target mouse (compared with the additional trauma and reaction of human myocardial ischemia / acute myocardial infarction treatment) is small, which does not interfere with the research and evaluation of myocardial ischemia-reperfusion injury.

[0021] 3. Since the thickness of the edge of the umbrella of the vascular occlusion body is greater than that of the middle portion, on the one hand, the weight of the edge of the umbrella is greater than that of the middle portion, so that the vascular occlusion body can always maintain balance during the process of being inserted into the blood vessel and delivered to the position where the coronary artery is to be blocked, and can be delivered smoothly to the position where the blood vessel is to be blocked, thereby avoiding deflection under the impact of blood flow, resulting in inability to move smoothly, damaging the blood vessel, and preventing smooth deployment after reaching the position where the blood vessel is to be blocked, or failing to completely block the blood vessel after deployment. On the other hand, during reperfusion (the middle portion of the umbrella degrades or the middle portion degrades first), the blood vessel can be effectively supported, reperfusion can be achieved within a specific time, and the efficiency and quality of reperfusion can be guaranteed. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Fig. 1 is a schematic diagram of the process of delivering the vascular occluding body to the location in the coronary artery where vascular occlusion is to be formed;

[0023] Fig. 2 Schematic diagram of the process of the umbrella of the vascular occluding body unfolding to block the coronary artery;

[0024] Fig. 3 is a schematic diagram of a state in which the umbrella of the vascular occluding body is unfolded and the coronary artery is completely blocked;

[0025] Figs. 1-3 In the figure, 1-coronary artery; 2-umbrella; 3-interventional guidewire. DETAILED DESCRIPTION

[0026] See also Figs. 1-3 The present invention discloses a method for constructing a myocardial ischemia-reperfusion injury model, wherein a vascular occluding body is inserted into a blood vessel of a target mouse and delivered (or guided) to a location in a coronary artery 1 where vascular occlusion is to be formed. The location in the coronary artery where vascular occlusion is to be formed is blocked by the vascular occluding body. Part or all of the vascular occluding body is made of a biodegradable material (polymer) (having deformation properties). Myocardial ischemia-reperfusion injury is simulated by degradation and penetration of the vascular occluding body in the target mouse (establishing a myocardial ischemia-reperfusion injury model).

[0027] The target mice are SD rats or mice.

[0028] The location of the coronary artery where vascular occlusion is to be formed is preferably the left anterior descending coronary artery.

[0029] The vascular occlusion body is preferably inserted into the radial artery of the upper limb or the femoral artery of the lower limb of the target mouse in a minimally invasive manner and delivered to the coronary artery (the location of the coronary artery where vascular occlusion is to be formed), so that during the construction of the myocardial ischemia-reperfusion injury model, the target mouse suffers little trauma and has almost no immune inflammatory response.

[0030] The process of inserting the blood vessel blocking body into the blood vessel of the target mouse is preferably completed under the guidance of photoacoustic imaging technology, which can effectively improve the accuracy of the blood vessel blocking body to the position and improve the accuracy of the myocardial ischemia-reperfusion injury model.

[0031] When the blood vessel blocking body is used to block the position of the coronary artery to be formed into a blood vessel, the blood vessel blocking body preferably completely blocks the position of the coronary artery to be formed into a blood vessel, blocks the blood flow, and helps the subsequent simulation of the reperfusion process and accurate research.

[0032] The biodegradable material is preferably polylactic acid (or polylactide), which can be degraded in the animal body without additional operation to simulate myocardial ischemia-reperfusion.

[0033] The blood vessel blocking body preferably has an umbrella shape, including a collapsible and expandable umbrella surface 2 and an interventional guide wire 3, one end of the interventional guide wire is connected to the center of the umbrella surface by a connection method that can be disconnected when the tension reaches a set threshold. Part or all of the umbrella surface is made of the biodegradable material so that the corresponding part or all can be degraded in the target mouse's body. The interventional guide wire is preferably made of nickel-titanium alloy, platinum-tungsten alloy, cobalt-chromium alloy, or stainless steel, so that it has a certain hardness, making it easy to push the umbrella surface to travel in the blood vessel. During the process of inserting the blood vessel blocking body into the blood vessel of the target mouse and sending it to the position of the coronary artery to be formed into a blood vessel, the umbrella surface is in a collapsed state and is pushed to travel along the direction of the blood flow in the blood vessel (the tip of the collapsed umbrella surface faces the direction of the blood flow). Because the tip of the umbrella surface faces the direction of the blood flow in the blood vessel and the direction of travel is the same as the direction of the blood flow, the umbrella surface does not expand during travel. When the umbrella surface is sent to the position of the coronary artery to be formed into a blood vessel, the interventional guide wire is pulled in the opposite direction. Under the combined action of the tension of the interventional guide wire and the pushing force of the blood flow in the blood vessel on the inner side of the umbrella surface, the umbrella surface expands until the edges of the umbrella surface are stably supported on the inner wall of the blood vessel, achieving complete blocking of the blood vessel. Continue to pull the interventional guide wire until the tension reaches a set threshold (the set threshold of the tension can be flexibly set according to the actual situation of the support strength of the umbrella surface on the inner wall of the blood vessel after expansion). The interventional guide wire and the umbrella surface are disconnected, the umbrella surface remains in the position to be formed into a blood vessel and is stably supported on the inner wall of the blood vessel, and the interventional guide wire is removed from the blood vessel.

[0034] The connection between the interventional guide wire and the umbrella surface can adopt any of the following connection methods:

[0035] (1) the intervention guide wire passes through the center of the umbrella surface, the center of the umbrella surface is provided with a through hole for the intervention guide wire to pass through, the intervention guide wire passes through the through hole, the end of the intervention guide wire (the end outside the umbrella surface) is provided with an outer plug which can be deformed towards the inside of the umbrella surface under the action of external force, the outer plug abuts against the outside of the center of the umbrella surface, the outer plug can be circular or hollow conical (the tip is towards the intervention guide wire and is fixedly connected with the intervention guide wire), and the diameter (the diameter of the circular or the diameter of the bottom surface of the conical) is greater than the hole diameter of the through hole, the outer plug can be made of elastic material, such as medical-grade elastomer TPE material or polycarbonate urethane (PCU), the intervention guide wire in the inside direction of the umbrella surface is provided with an inner plug which does not deform under the action of external force, the inner plug abuts against the inside of the center of the umbrella surface, the inner plug can be circular or cylindrical, and the diameter (the diameter of the circular or the outer diameter of the cylindrical) is greater than the hole diameter of the through hole, the inner plug can be made of the same material as the intervention guide wire, the inner plug can be integrally formed with the intervention guide wire, when the umbrella surface is pushed to travel in the blood vessel through the intervention guide wire, since the inner plug abuts against the inside of the center of the umbrella surface, the intervention guide wire will not slide in the through hole, which can ensure that the umbrella surface smoothly travels in the blood vessel, when the intervention guide wire is pulled in the reverse direction, since the outer plug abuts against the outside of the center of the umbrella surface, the intervention guide wire will not slide in the through hole, which can ensure that the umbrella surface is unfolded under the joint action of the pulling force and the pushing force of the blood flow, and is stably supported on the inner wall of the blood vessel, the pulling force continues to be applied to the intervention guide wire, when the pulling force reaches the set threshold value, the outer plug is deformed towards the inside of the umbrella surface, passes through the through hole, the intervention guide wire is separated from the umbrella surface (the connection is disconnected), and the elasticity of the outer plug can be flexibly set according to the actual situation of the support strength of the umbrella surface on the inner wall of the blood vessel after unfolding.

[0036] (2) the one end of the intervention guide wire is in interference fit with the center of the umbrella face, the inner side of the center of the umbrella face is provided with a groove conforming to the end of the intervention guide wire, the corresponding end of the intervention guide wire is in interference fit with the groove, when the umbrella face is pushed to travel in the blood vessel through the intervention guide wire, the umbrella face can travel smoothly in the blood vessel because the corresponding end of the intervention guide wire is inserted in the groove and cannot be separated, when the intervention guide wire is pulled reversely, the umbrella face cannot be separated from the intervention guide wire because the corresponding end of the intervention guide wire is in interference fit with the groove, the umbrella face can be unfolded and stably supported on the inner wall of the blood vessel under the joint action of the pulling force and the pushing force of the blood flow, the corresponding end of the intervention guide wire is pulled out of the groove to separate the intervention guide wire from the umbrella face (the connection is disconnected) when the pulling force reaches the set threshold, the degree of interference fit between the corresponding end of the intervention guide wire and the umbrella face can be flexibly set according to the actual situation of the support strength of the umbrella face on the inner wall of the blood vessel after unfolding.

[0037] (3) the one end of the intervention guide wire is bonded with the center (referring to the inner side of the center of the umbrella face) of the umbrella face, the soft tissue medical glue, matrix glue or degradable surgical adhesive can be used for bonding, when the umbrella face is pushed to travel in the blood vessel through the intervention guide wire, the umbrella face can travel smoothly in the blood vessel because the corresponding end of the intervention guide wire is bonded with the umbrella face and cannot be separated, when the intervention guide wire is pulled reversely, the umbrella face cannot be separated from the intervention guide wire because the corresponding end of the intervention guide wire is bonded with the umbrella face, the umbrella face can be unfolded and stably supported on the inner wall of the blood vessel under the joint action of the pulling force and the pushing force of the blood flow, the corresponding end of the intervention guide wire is separated from the umbrella face (the connection is disconnected) when the pulling force reaches a certain value, the corresponding end of the intervention guide wire overcomes the bonding force between the intervention guide wire and the umbrella face, the bonding strength between the corresponding end of the intervention guide wire and the umbrella face can be flexibly set according to the actual situation of the support strength of the umbrella face on the inner wall of the blood vessel after unfolding.

[0038] The intervention guide wire and the umbrella face can also be connected by other suitable connection modes in which the two connecting members can be disconnected when the external force reaches a certain value.

[0039] The thickness of the middle part of the umbrella surface is preferably less than the thickness of the edge of the umbrella surface, and at least the middle part of the umbrella surface is made of the biodegradable material, so that the middle part of the umbrella surface can be degraded first, or only the middle part of the umbrella surface can be degraded, so as to realize reperfusion of the coronary artery through degradation of the biodegradable material. In a preferred embodiment, the middle part of the umbrella surface is made of the biodegradable material, and the edge of the umbrella surface is made of a hard medical device material used in coronary intervention surgery, such as nickel-titanium alloy, cobalt-chromium alloy or stainless steel, etc. In this way, on the one hand, the weight of the edge of the umbrella surface is greater than that of the middle part, so that the balance can be maintained during the process of inserting the blood vessel occlusion body into the blood vessel and sending it to the position of the coronary artery to be occluded, and the blood vessel occlusion body can be smoothly sent to the position to be occluded, so as to avoid the situation that the blood vessel occlusion body is deflected under the impact of blood flow, cannot smoothly travel, damages the blood vessel, cannot be smoothly deployed after reaching the position to be occluded, or cannot completely occlude the blood vessel after being deployed. On the other hand, the edge of the umbrella surface can provide effective support to the blood vessel during reperfusion, so as to realize reperfusion within a certain time and ensure the efficiency and quality of reperfusion.

[0040] The blood vessel occlusion body can be made by 3D printing technology.

[0041] The outer diameter of the edge of the umbrella surface after deployment can be appropriately designed according to the inner diameter of the position of the coronary artery to be occluded in the target mouse, and is usually not greater than 1000 μm.

[0042] The myocardial ischemia-reperfusion injury model can realize precise control of the ischemia-reperfusion time of the coronary artery by setting different thicknesses of the degradable part of the umbrella surface and changing the time required for the degradable part of the umbrella surface to be degraded (the time of realizing ischemia-reperfusion in the animal body), so as to improve the controllability of the reperfusion time, simulate the disease process of different time lengths from myocardial ischemia to surgical treatment of clinical patients, provide strong model support for the mechanism research of myocardial ischemia-reperfusion injury, facilitate the in-depth research of the pathological mechanism of myocardial ischemia-reperfusion injury, facilitate more objective preclinical evaluation of drug and non-drug therapies for myocardial ischemia-reperfusion injury, and promote the effective research and development of related clinical treatment schemes.

[0043] Unless otherwise specified, the various preferred and optional technical means disclosed in the present application can be combined in any manner, forming several different technical solutions.

Claims

1. A method for constructing a model of myocardial ischemia-reperfusion injury, characterized by Inserting a blood vessel blocking body into a blood vessel of a target mouse and sending it to a position of a coronary artery to be blocked, blocking the position of the coronary artery to be blocked by the blood vessel blocking body, part or all of the blood vessel blocking body being made of a biodegradable material, simulating myocardial ischemia-reperfusion injury by degradation of the blood vessel blocking body in the target mouse; The blood vessel blocking body is umbrella-shaped and includes a collapsible and expandable umbrella surface and an intervention guide wire, one end of the intervention guide wire being connected to the center of the umbrella surface by a connection mode that can be disconnected when the tension reaches a set threshold, the thickness of the middle part of the umbrella surface being smaller than the thickness of the edge of the umbrella surface, at least the middle part of the umbrella surface being made of the biodegradable material, the degradation of the degradable part of the umbrella surface being changed by setting different thicknesses of the degradable part of the umbrella surface, the time required for the degradation of the degradable part of the umbrella surface being changed to achieve precise control of the coronary artery ischemia-reperfusion time, and the disease process of a clinical patient from myocardial ischemia to surgical treatment being simulated.

2. The method of claim 1, wherein the method further comprises The position of the coronary artery to be blocked is the left anterior descending coronary artery position.

3. The method of claim 1, wherein the method further comprises The blood vessel blocking body is inserted from the radial artery of the upper limb or the femoral artery of the lower limb of the target mouse and sent to the coronary artery.

4. The method of claim 1, wherein the method further comprises The blood vessel blocking body is inserted into the blood vessel of the target mouse under the guidance of photoacoustic imaging technology.

5. The method of claim 1, wherein the method further comprises The blood vessel blocking body completely blocks the position of the coronary artery to be blocked, blocks the blood flow, and realizes reperfusion by controlling the biodegradability of the degradable part of the blood vessel blocking body.

6. The method of claim 1, wherein the method further comprises The biodegradable material is polylactic acid.

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