Establishment of a noninvasive ischemic stroke model induced by magnetic field combined with photochemical therapy

Abstract

The present application discloses a method for establishing a non-invasive ischemic stroke model induced by a magnetic field combined with photochemical induction, comprising injecting photosensitive magnetic nanoparticles into the tail vein of a mouse; controlling the magnetic field at a preset position of the mouse to guide at least a portion of the photosensitive magnetic nanoparticles to form microthrombi with flowing blood at the preset position; wherein the mouse is exposed to light of a first preset wavelength so that the photosensitive magnetic nanoparticles emit a first color of light; and the mouse is exposed to light of a second preset wavelength so that the photosensitive magnetic nanoparticles emit a second color of light to induce a photochemical reaction, stimulate the coagulation process, and obtain a non-invasive ischemic stroke model. The present application applies magnetic fields of different sizes, and through the first color of light positioning mark, the photosensitive magnetic nanoparticles are accurately attracted to a preset position in the internal carotid artery or the brain, and the second color of light is used to induce a photochemical reaction, which has the advantages of being non-invasive, simple and convenient to operate, and precise.

Description

Technical Field

[0001] The present application relates to the technical field of animal models, and in particular to a method for establishing a non-invasive ischemic stroke model induced by a magnetic field combined with photochemistry. Background Art

[0002] Animal disease models refer to animals created in various medical and scientific research settings to mimic human disease manifestations. They are primarily used in experimental physiology, experimental pathology, and experimental therapeutics (including new drug screening). The development of human disease is extremely complex, and using humans as experimental subjects to deeply explore the mechanisms of disease and promote the development of medicine has been slow. Accumulated clinical experience is not only limited in time and space, but many experiments are also ethically and methodologically constrained. Indirect research using animal models, however, allows for the deliberate modification of factors that are impossible or difficult to eliminate under natural conditions, allowing for more accurate observation of the model's experimental results and comparative studies with human diseases. This facilitates a more convenient and effective understanding of the patterns of human disease development and the study of preventive and therapeutic measures.

[0003] Ischemic stroke, as a serious neurological disease, has become the leading cause of death and disability in my country. There are also many clinical studies on ischemic stroke. However, the stroke models in the existing technology all require invasive surgery, in which an incision is made in the animal's neck to expose the internal carotid artery. The surgery itself is difficult to operate and may have a certain impact on the animal's physiology, thereby affecting the accuracy and reliability of the animal model, causing the established animal model to lose its reference value. In addition, in the existing technology, when a thrombus forms in the animal's neck, it is impossible to determine the exact location of the thrombus formation. Only the approximate location of the thrombus can be known, which also causes the established animal model to be inaccurate and lose its reference value.

[0004] Therefore, there is a need for a method for establishing a non-invasive ischemic stroke model induced by a magnetic field combined with photochemistry, which can achieve the establishment of a non-invasive ischemic stroke model with simple and convenient operation. At the same time, it can accurately form a thrombus at a specific location, thereby improving the accuracy and reliability of the model. Summary of the Invention

[0005] In response to the problems existing in the above-mentioned prior art, the present application provides a method for establishing a non-invasive ischemic stroke model induced by a magnetic field combined with photochemical induction. This method can achieve the establishment of a non-invasive ischemic stroke model with simple and convenient operation. At the same time, it can accurately form a thrombus at a specific location, improving the accuracy and reliability of the model. The technical solution is as follows:

[0006] The present application provides a method for establishing a non-invasive ischemic stroke model induced by a magnetic field combined with photochemistry, comprising:

[0007] Photosensitive magnetic nanoparticles were injected into the tail vein of mice;

[0008] controlling a magnetic field at a preset position of the mouse to guide at least a portion of the photosensitive magnetic nanoparticles to form microthrombi with flowing blood at the preset position; wherein the mouse is exposed to light of a first preset wavelength so that the photosensitive magnetic nanoparticles emit a first color of light;

[0009] The mice are exposed to light of a second preset wavelength so that the photosensitive magnetic nanoparticles emit a second color of light to induce a photochemical reaction, stimulate the coagulation process, and obtain a non-invasive ischemic stroke model.

[0010] Furthermore, the mice are healthy adult male C57 / B mice aged 3 to 4 months.

[0011] Furthermore, the particle size of the photosensitive nanomagnetic particles is 80 to 150 nm.

[0012] Furthermore, the tail vein injection of magnetic nanoparticles into mice comprises:

[0013] Injecting different concentrations of the magnetic nanoparticles into the tail veins of multiple groups of mice; or

[0014] The same concentration of the magnetic nanoparticles was injected into the tail veins of multiple groups of mice.

[0015] Furthermore, the volume of the photosensitive nanomagnetic particles injected into the tail vein is 130 to 150 μL; the concentration of the injected photosensitive magnetic nanoparticles is one of a first concentration, a second concentration, and a third concentration; the first concentration is greater than 0 mg / mL and not greater than 2 mg / mL, the second concentration is greater than 2 mg / mL and not greater than 4 mg / mL, and the third concentration is greater than 4 mg / mL and not greater than 6 mg / mL.

[0016] Furthermore, before controlling the magnetic field at a preset position of the mouse to guide at least a portion of the photosensitive magnetic nanoparticles to form microthrombi with flowing blood at the preset position, the method further comprises:

[0017] The mice were anesthetized and placed on a constant temperature blanket;

[0018] The head and neck hair of the mice were removed, and the positions of the common carotid artery, internal carotid artery, and external carotid artery of the mice were marked.

[0019] Furthermore, the preset adjustment range of the magnetic field is 0 to 1T.

[0020] Furthermore, the first color light is red light; and the second color light is green light.

[0021] Furthermore, after exposing the mouse to light of a second preset wavelength so that the photosensitive magnetic nanoparticles emit a second color of light to induce a photochemical reaction and stimulate the coagulation process to obtain a non-invasive ischemic stroke model, the method further includes:

[0022] The mice were returned to housing and subjected to neurobehavioral assessments.

[0023] Furthermore, the neurobehavioral assessment is:

[0024] In a single-blind condition, the mice were subjected to a grid walking test, a cylinder test, and a spaghetti grasping test; wherein the neurobehavioral assessment was conducted within 1 to 15 weeks after the injection of the photosensitive magnetic nanoparticles.

[0025] The implementation of this application has the following beneficial effects:

[0026] The present application utilizes the characteristic that nanomagnetic particles flow with hemodynamics to apply magnetic fields of different sizes at specific locations, forcibly attracting photosensitive magnetic nanoparticles to a preset location to form microthrombi with the flowing blood. At the same time, the present application utilizes light of a first preset wavelength to mark the location of the photosensitive magnetic nanoparticles, which is beneficial for accurately guiding the photosensitive magnetic nanoparticles to the preset location, playing a role in precise positioning with high accuracy. The light of a second preset wavelength can induce a photochemical reaction, causing platelets in the blood to adhere to the surface of the vascular endothelium and undergo a release reaction, stimulating the coagulation process, making it easier for microthrombi to form at the preset location. The magnetic field and the light of the second preset wavelength are combined to control the degree of coagulation, thereby obtaining a series of non-invasive ischemic stroke models of different degrees formed at different preset locations. Compared with the surgical methods of traditional stroke models, the present application has the advantages of being non-invasive, simple and convenient to operate, and precise. The model has high accuracy and reliability, which is convenient for simulating and observing the characteristics of different degrees of damage in human ischemic stroke, laying a solid foundation for the pathogenesis of cerebral ischemia and drug screening. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] In order to more clearly illustrate the technical solution of the present invention, the following is a brief introduction to the drawings used in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0028] Figure 1 The figure is a logic flow chart of a method for establishing a non-invasive ischemic stroke model induced by a magnetic field combined with photochemistry in a possible embodiment of the present invention. DETAILED DESCRIPTION

[0029] The following will be combined with the accompanying drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present application. Obviously, the embodiments described are only part of the embodiments of the present application, not all of the embodiments, and therefore should not be understood as limiting the present application. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of this application.

[0030] It should be noted that the terms "including" and "having" and any variations thereof in the specification and claims of this application are intended to cover non-exclusive inclusions. For example, a process, method, system, product or server that includes a series of steps or units is not necessarily limited to those steps or units clearly listed, but may include other steps or units that are not clearly listed or are inherent to these processes, methods, products or devices.

[0031] Example 1

[0032] Based on the current situation that the mouse ischemic stroke model cannot be established non-invasively and has low accuracy, the present application provides a method for establishing a non-invasive ischemic stroke model induced by a magnetic field combined with photochemical induction, as shown in the attached manual. Figure 1 As shown, including:

[0033] S1, Photosensitive magnetic nanoparticles were injected into the tail vein of mice.

[0034] S2, controlling the magnetic field at a preset position of the mouse to guide at least a portion of the photosensitive magnetic nanoparticles to form microthrombi with the flowing blood at the preset position; wherein, the mouse is exposed to light of a first preset wavelength so that the photosensitive magnetic nanoparticles emit a first color of light.

[0035] S3, exposing the mouse to light of a second preset wavelength, so that the photosensitive magnetic nanoparticles emit a second color of light to induce a photochemical reaction, stimulate the coagulation process, and obtain a non-invasive ischemic stroke model.

[0036] Specifically, in one possible embodiment of the present specification, the mice used in step S1 are 3-4 month old adult healthy male C57 / B mice.

[0037] In this embodiment, the magnetic field at the preset position of the mouse can be controlled by a magnetic field controller. The magnetic field controller is placed on the neck of the mouse and above, and then the magnetic field controller is moved to the preset position, thereby guiding the photosensitive magnetic nanoparticles to move to the preset position, and precisely forming microthrombi at the preset position.

[0038] Exposing mice to light of a first preset wavelength or a second preset wavelength can be controlled by a laser emitter, and the laser emitter is placed at a preset position of the mouse; when the photosensitive magnetic nanoparticles are required to continuously display the first color light, the laser emitter is used to emit light of the first preset wavelength for excitation; similarly, the laser emitter is used to emit light of the second preset wavelength, thereby exciting the photosensitive magnetic nanoparticles to display the second color light and inducing a photochemical reaction.

[0039] Optionally, the preset position is located in the internal carotid artery or in the brain; when the preset position is located in the internal carotid artery, the magnetic field controller is placed in the internal carotid artery of the mouse to guide the photosensitive magnetic nanoparticles to form microthrombi at the preset position in the internal carotid artery; and when the preset position is located in the brain, the magnetic field controller is placed on the head of the mouse, and then the magnetic field controller is moved to the corresponding preset position in the brain to guide the photosensitive magnetic nanoparticles to form microthrombi at the preset position in the brain.

[0040] Specifically, before step S2, the establishment method further includes:

[0041] The mice were anesthetized and placed on a constant temperature blanket;

[0042] The head and neck hair of the mice were removed, and the positions of the common carotid artery, internal carotid artery, and external carotid artery of the mice were marked.

[0043] Among them, the temperature of the constant temperature blanket is set to 37±0.2℃, which is as close to the body temperature of the mice as possible to avoid the abnormal environment affecting the physiology of the mice, thereby affecting the accuracy of the non-invasive ischemic stroke model; and removing the mouse hair can expose the mouse's skin, which is convenient for observing and marking the mouse's brain, common carotid artery, internal carotid artery and external carotid artery, and then facilitating the placement of the magnetic field controller and laser emitter at the preset positions, thereby improving the accuracy of magnetic field control and light control, so that microthrombi can be accurately formed at the preset position, improving operational convenience and model accuracy.

[0044] Specifically, in one possible embodiment of the present specification, depending on the level of sophistication of the non-invasive ischemic stroke model to be established, the injection of photosensitive magnetic nanoparticles into the tail vein of the mouse in step S1 may include:

[0045] S101, injecting different concentrations of the photosensitive magnetic nanoparticles into the tail veins of multiple groups of mice respectively; or

[0046] S102, injecting the photosensitive magnetic nanoparticles of the same concentration into the tail veins of multiple groups of mice.

[0047] Among them, when the requirement for the model's precision is low, step S102 is executed. At the same injection concentration, a series of non-invasive ischemic stroke models of different degrees are established only by adjusting the intensity of the magnetic field in step S2. When the photosensitive magnetic nanoparticles injected into the mouse body cannot meet the establishment of a large number of non-invasive ischemic stroke models of different degrees at a single concentration, step S101 is executed. By jointly adjusting different concentrations of photosensitive magnetic nanoparticles and different sizes of magnetic fields, the degree range of the simulated non-invasive ischemic stroke model is expanded, the non-invasive ischemic stroke model is further enriched, and the accuracy and reliability are improved.

[0048] Specifically, in one possible embodiment of the present specification, the photosensitive magnetic nanoparticles can be selected as Fe3O4, which has moderate magnetism, can be stably attracted by the magnetic field, and can form microthrombi quickly and conveniently; and, in another possible embodiment of the present specification, the particle size of the photosensitive magnetic nanoparticles can be selected as 80-150 nm, which is a moderate particle size; optionally, the particle size of the photosensitive magnetic nanoparticles is 100-120 nm, which facilitates the movement of the photosensitive magnetic nanoparticles in the blood vessels.

[0049] Specifically, in one possible embodiment of the present specification, in step S1, a certain volume of photosensitive magnetic nanoparticles is drawn and injected into the tail vein of a mouse, and the volume of the drawn and injected photosensitive magnetic nanoparticle reagent can be selected to be 130-150 μL, and the concentration of the multiple groups of photosensitive magnetic nanoparticles includes one of a first concentration c1, a second concentration c2, and a third concentration c3; wherein, 0 < c1 ≤ 2 mg / mL, 2 mg / mL < c2 ≤ 4 mg / mL, and 4 mg / mL < c3 ≤ 6 mg / mL, then the total number of photosensitive magnetic nanoparticles injected from the tail vein of the mouse can be adjusted by adjusting the specific volume and specific concentration of the photosensitive magnetic nanoparticles to achieve more and more detailed changes, thereby coordinating with the subsequent S The magnetic fields of different sizes in step 2 simulate more non-invasive ischemic stroke models of different degrees; and when the injection concentration is low, the total number of its photosensitive magnetic nanoparticles has a highest threshold. When the magnetic field increases, its ability to attract photosensitive magnetic nanoparticles is likely to be far greater than the threshold of the photosensitive magnetic nanoparticles actually injected, resulting in the inability to form a non-invasive ischemic stroke model of the corresponding degree and model distortion; while the injection of high concentrations of photosensitive magnetic nanoparticles can match the larger magnetic field, that is, the total number of injected photosensitive magnetic nanoparticles is always higher than the number of photosensitive magnetic nanoparticles that can be attracted by the magnetic field of the corresponding group, and the non-invasive ischemic stroke model of the corresponding degree (severe) can be accurately formed with high reliability.

[0050] For example, in this embodiment, the volume of the injected photosensitive magnetic nanoparticles can be selected to be 150 μL, and the mice in step S1 are divided into three groups, and the injection concentrations of the three groups are set to 1 mg / ml, 3 mg / ml and 5 mg / ml, respectively. Then, in step S2, different magnetic field intensities are applied to the three groups of mice, the group with an injection concentration of 1 mg / ml is applied with a smaller magnetic field intensity, the group with an injection concentration of 3 mg / ml is applied with a medium magnetic field intensity, and the group with an injection concentration of 5 mg / ml is applied with a larger magnetic field intensity, so as to form non-invasive ischemic stroke models of different degrees, which is simple and convenient to operate; and multiple mice can also be set within each group, and multiple mice in the same group are injected with the same concentration of photosensitive magnetic nanoparticles and applied with the same magnetic field intensity, so that the establishment method of the present application is reproducible and more reliable.

[0051] In another possible embodiment of the present specification, different magnetic field intensities can also be applied to multiple mice in each group (all mice in the group have been injected with the same concentration of photosensitive magnetic nanoparticles); due to the limitation of the injection concentration itself, the number of photosensitive magnetic nanoparticles in the mouse body is limited. Assuming that the injection of photosensitive magnetic nanoparticles at a concentration of 1 mg / ml can meet the regulation requirements of a smaller magnetic field intensity range (set as interval T1), then within the smaller magnetic field intensity range (interval T1), different smaller magnetic field intensities of different sizes can be further applied to the multiple mice in the group, and these different smaller magnetic field intensities are all within the range of interval T1, thereby obtaining mice with different mild microthrombi. The mice in this group all show different degrees of mildness, that is, different degrees of non-invasive ischemic stroke model, further refining the model; In this case, the mice are actually grouped into the first level according to the concentration, and then grouped into the second level according to the magnetic field strength. The final total number of groups is equal to the number of groups in the first level multiplied by the number of groups in the second level. It should be noted that the concentration-based grouping in step S1 is not limited to three groups. Four, five or even more groups can be set according to actual needs, and the concentration interval between adjacent groups is smaller, thereby simulating a more refined non-invasive ischemic stroke model. In addition, the grouping of mice in step S1 is only for the convenience of description and is not limited to grouping based on concentration. Grouping can also be based on different magnetic field strengths. Then, multiple mice in the group (the same magnetic field strength will be applied in the subsequent S2 step in the same group) are injected with different concentrations of photosensitive magnetic nanoparticles, and the establishment method is flexible and changeable.

[0052] In the pathogenesis of stroke, microthrombi can form in the internal carotid artery or in the brain. After the magnetic field setting in step S2 is completed, moving the magnetic field controller upward along the internal carotid artery or moving it on the head can be more conducive to guiding the photosensitive magnetic nanoparticles to the preset position, so that the microthrombi can be accurately formed at the preset position, with precise positioning, thereby improving the reliability of the model.

[0053] Specifically, in a possible implementation manner of the present specification, the preset adjustment range of the magnetic field is set to 0-1T.

[0054] The preset adjustment range has a wide adjustment range, which can allow the formation of stroke models within a large range; and in this embodiment, according to the increase in injection concentration, when the injection concentrations of the three groups of mice are 1 mg / ml, 3 mg / ml and 5 mg / ml, respectively, the magnetic field applied to the three groups of mice can be set in the range of 0-10mT (excluding 0), 20-30mT and 31-40mT, respectively, to form mild, moderate and severe non-invasive ischemic stroke models, respectively; similarly, in other possible implementations of the present specification, the magnetic field range of each group can also be narrowed according to the number of groups, and the concentration of the photosensitive nanomagnetic particles to be injected can be adjusted according to the narrowed magnetic field range to add more groups of non-invasive ischemic stroke models and further enrich the non-invasive ischemic stroke models.

[0055] It should be noted that the photosensitive magnetic nanoparticles used in the present application can emit fluorescence of different colors under irradiation with light of different wavelengths, thereby exerting different effects. In one possible embodiment of the present specification, before injecting the photosensitive magnetic nanoparticles into the tail vein of the mouse, the method further includes:

[0056] The magnetic nanoparticles are dyed with a photosensitivity dye to obtain the photosensitive magnetic nanoparticles.

[0057] The photosensitivity dye can emit different colors of light under different specific wavelengths of light, so that the dyed photosensitive magnetic nanoparticles show the same color of light as the photosensitivity dye. In this embodiment, the photosensitivity dye can be rose red.

[0058] Specifically, before adjusting the magnetic field, that is, before step S2, it is necessary to use a laser emitter to excite the fluorescence of the photosensitive magnetic nanoparticles, so that the photosensitive magnetic nanoparticles continue to emit the first color light, which plays a role of marking and precise positioning, and identifies the position of the photosensitive magnetic nanoparticles in real time, so as to facilitate intuitive observation of the movement path and arrival position of the photosensitive magnetic nanoparticles in the internal carotid artery or the brain, so that the microthrombus is accurately formed at the preset position, thereby improving the accuracy and reliability of each model.

[0059] In a possible implementation of the present specification, the first color light is red light, so that the photosensitive magnetic nanoparticles emit the first color light of a first preset wavelength, and the first preset wavelength can be selected to be 590-620 nm.

[0060] Specifically, in step S3, the second preset wavelength is used to excite the photosensitive magnetic nanoparticles to emit a second color light, which can be selected as green light; under green light, a photochemical reaction can be induced, wherein the photosensitivity dye on the photosensitive magnetic nanoparticles can generate free radicals, causing vascular endothelial damage, producing lipid peroxides, and then platelet aggregation occurs. The platelets adhere to the surface of the vascular endothelium and release reaction occurs, stimulating the coagulation process; that is, in the present application, on the one hand, through magnetic field induction, and on the other hand, through the laser emitter to make the photosensitive magnetic nanoparticles emit green light for photochemical reaction, the two cooperate to form microthrombi together, making it easier to form microthrombi in blood vessels and shortening the formation time of microthrombi; in a possible embodiment of the present specification, the second preset wavelength can be selected as 460-550nm.

[0061] Specifically, in this application, photosensitive magnetic nanoparticles are attracted by a magnetic field, and a photochemical reaction is induced by light of a second preset wavelength, so that the formation time of microthrombi at a preset position is less than 4 min. The time required is short, and a large number of non-invasive ischemic stroke models can be obtained quickly and conveniently, saving time.

[0062] Specifically, after step S3, that is, after obtaining the non-invasive ischemic stroke model, the method further includes:

[0063] The mice were returned to housing and subjected to neurobehavioral assessments.

[0064] Neurobehavioral assessments may include:

[0065] The mice were subjected to a grid walking test, a cylinder test, and a spaghetti grasping test under single-blind conditions.

[0066] Among them, the evaluation time for neurobehavioral assessment includes 1 to 15 weeks after the injection of photosensitive magnetic nanoparticles, and neurobehavioral assessment can also be performed one week before surgery. The evaluation results one week before surgery can be used as a control group to facilitate the comparison of normal mice with non-invasive ischemic stroke models of different degrees; in this embodiment, the evaluation time can be further set to one week before surgery to 1, 3, 5, 7, 11, and 15 weeks after surgery.

[0067] The grid walking test is an evaluation method for detecting whether an animal has the ability to accurately control the placement of its hind paws after brain or spinal cord injury. It is suitable for small animals such as cats and mice. Depending on the angle of the grid placement, it can be divided into a horizontal grid test and an inclined grid test. During the evaluation, the mouse is placed on a horizontal or inclined grid (with a distance of 2.5 cm between the two ends), and behavioral data such as the number of times the mouse's hind paws miss the grid, the number of footsteps, and the time it takes to cover this distance are recorded over a certain period of time.

[0068] The cylinder test is a research tool for asymmetric central nervous system diseases. It mainly studies the asymmetric use of limbs in rodents during wall standing. It is also called the spontaneous forelimb task assessment test. During the assessment, mice are placed in a cylinder with a diameter of 20 cm and a height of 40 cm. The camera in the automatic recording system records the number, duration and proportion of times the mouse stands against the wall with its left forelimb, right forelimb, and both forelimbs simultaneously within a certain period of time. The automatic recording system records and analyzes the data, which can ensure the objectivity of the assessment results while saving assessment time and manpower.

[0069] The spaghetti grasping test uses video to record the changes in the animal's forepaw movements when grasping and eating 2.5 cm long uncooked spaghetti noodles. During this process, the position and angle of the camera can be adjusted to obtain the best view of the forepaw movement, and slow-motion playback can clearly observe the adjustments of the mouse's forepaws and determine the clear movement and replacement of any toes of the forepaw grasping the noodles after the mouse starts eating. The number of times the mouse's forepaws adjust on each piece of rice noodles and the time it takes to finish eating each piece of rice noodles are recorded in each trial. This is repeated at least four times to ensure quantitative analysis of changes in forepaw sensitivity.

[0070] The present application utilizes light of a first preset wavelength to cause photosensitive magnetic nanoparticles to emit red light, making it easy to observe the position of the photosensitive magnetic nanoparticles, so that microthrombi are precisely formed at the preset position, thereby achieving precise positioning. Furthermore, the present application utilizes a magnetic field controller to adjust magnetic fields of different sizes based on the relationship that the number of aggregated photosensitive magnetic nanoparticles is proportional to the magnetic field size of the magnetic field controller, so as to control the number of photosensitive magnetic nanoparticles attracted to the preset position, so that different numbers of photosensitive magnetic nanoparticles form microthrombi of different sizes with the flowing blood. Simultaneously, light of a second preset wavelength is used to induce a photochemical reaction, causing platelets in the blood to adhere to the surface of the vascular endothelium and undergo a release reaction, thereby stimulating the coagulation process, making it easier to form microthrombi, and also being able to control the degree of coagulation, thereby quickly, conveniently, and accurately obtaining a series of non-invasive ischemic stroke models of varying degrees formed at different preset positions. Taking three groups of mice as an example, in this embodiment, the establishment method can be specifically as follows:

[0071] Several mice were divided into three groups. 1 mg / mL of photosensitive nanomagnetic particles were injected into the tail vein of the first group of mice, 3 mg / mL of photosensitive nanomagnetic particles were injected into the tail vein of the second group of mice, and 5 mg / mL of photosensitive nanomagnetic particles were injected into the tail vein of the third group of mice. The volume of photosensitive magnetic nanoparticles injected in the above three groups was the same, all 150 μL.

[0072] Mice were anesthetized with isoflurane and placed on a constant temperature blanket. The hair on the neck of the mice was removed, and the positions of the common carotid artery, internal carotid artery, and external carotid artery of the mice were marked for subsequent operation and observation.

[0073] The magnetic field controller and laser emitter were placed on the internal carotid artery of the mouse, and the laser emitter was adjusted to emit light so that the photosensitive magnetic nanoparticles emitted red light.

[0074] The setting value of the magnetic field controller was adjusted so that the magnetic field intensity applied to the necks of the first group of mice was in the range of 0 to 10 mT (excluding 0), the magnetic field intensity applied to the necks of the second group of mice was in the range of 20 to 30 mT, and the magnetic field intensity applied to the necks of the third group of mice was in the range of 31 to 40 mT.

[0075] The adjusted magnetic field controller is moved upward along the internal carotid artery to guide the photosensitive nanomagnetic particles to flow into the preset position in the internal carotid artery, forming microthrombi with the flowing blood. The light emitted by the laser emitter is then adjusted to make the photosensitive magnetic nanoparticles emit green light, inducing a photochemical reaction, further accelerating the formation of microthrombi. Three groups of non-invasive ischemic stroke models are obtained, and the corresponding relationship between their stroke severity should be as shown in Table 1.

[0076] Table 1 Corresponding relationship between injection concentration, magnetic field intensity and stroke severity in this embodiment

[0077]

[0078]

[0079] The treated mice were returned to their homes and subjected to neurobehavioral assessments, including a grid walking test, a cylinder test, and a spaghetti grasping test. The stroke severity indicated by the assessment results was consistent with the predicted results in Table 1. This indicates that the method for establishing a noninvasive ischemic stroke model induced by a magnetic field combined with photochemical induction is simple, convenient, and accurate. It utilizes a combination of light of a second preset wavelength and magnetic field intensity to induce microthrombus formation without the need for surgical intervention. The established noninvasive ischemic stroke model is also highly accurate and reliable, with high reference value, laying a solid foundation for the pathogenesis of cerebral ischemia and drug screening.

[0080] Example 2

[0081] The difference between this embodiment and embodiment 1 is that the mice are divided into nine groups, and different magnetic field intensities are applied to the nine groups of mice, and the corresponding relationships are shown in Table 2. The rest is the same as embodiment 1.

[0082] Table 2 Corresponding relationship between injection concentration, magnetic field intensity and stroke severity in Example 2

[0083]

[0084] It should be noted that for the case of an injection concentration of 1 mg / mL and a magnetic field strength of 21 mT or above in Table 2, due to the limitation of the total number of photosensitive magnetic nanoparticles in the mouse body, even if a higher magnetic field strength is applied, microthrombi corresponding to the magnetic field strength cannot be formed. Instead, only microthrombi corresponding to the magnetic field threshold (i.e., the magnetic field strength corresponding to the complete attraction of 1 mg / mL of photosensitive magnetic nanoparticles) can be formed. This is actually meaningless and is therefore not listed in this example.

[0085] What is described above are only some embodiments of the present application and are not intended to limit the present application. Those skilled in the art should understand that the present application may be subject to various changes and improvements, and any modifications, equivalent substitutions, and improvements made in accordance with the present application shall fall within the scope of protection required by the present application.

Claims

1. A method for establishing a non-invasive ischemic stroke model induced by magnetic field combined with photochemical therapy, characterized in that: include: Photosensitive magnetic nanoparticles were injected into the tail vein of mice; controlling a magnetic field at a preset position of the mouse to guide at least a portion of the photosensitive magnetic nanoparticles to form microthrombi with flowing blood at the preset position; wherein the mouse is exposed to light of a first preset wavelength so that the photosensitive magnetic nanoparticles emit a first color of light; The mice are exposed to light of a second preset wavelength so that the photosensitive magnetic nanoparticles emit a second color of light to induce a photochemical reaction, stimulate the coagulation process, and obtain a non-invasive ischemic stroke model.

2. The establishment method according to claim 1, characterized in that The mice were healthy adult male C57 / B mice aged 3 to 4 months.

3. The establishment method according to claim 1, characterized in that The particle size of the photosensitive magnetic nanoparticles is 80-150 nm.

4. The establishment method according to claim 1, characterized in that The method of injecting photosensitive magnetic nanoparticles into the tail vein of mice comprises: injecting different concentrations of the photosensitive magnetic nanoparticles into the tail veins of multiple groups of mice; or The same concentration of the photosensitive magnetic nanoparticles was injected into the tail veins of multiple groups of mice.

5. The establishment method according to claim 1, characterized in that: The volume of the photosensitive magnetic nanoparticles injected into the tail vein is 130~150 μL; the concentration of the injected photosensitive magnetic nanoparticles is one of a first concentration, a second concentration, and a third concentration; the first concentration is greater than 0 mg / mL and not greater than 2 mg / mL, the second concentration is greater than 2 mg / mL and not greater than 4 mg / mL, and the third concentration is greater than 4 mg / mL and not greater than 6 mg / mL.

6. The establishment method according to claim 1, characterized in that: Before controlling the magnetic field at a preset position of the mouse to guide at least a portion of the photosensitive magnetic nanoparticles to form microthrombi with flowing blood at the preset position, the method further comprises: The mice were anesthetized and placed on a constant temperature blanket; The head and neck hair of the mice were removed, and the positions of the common carotid artery, internal carotid artery, and external carotid artery of the mice were marked.

7. The establishment method according to claim 1, characterized in that: The preset adjustment range of the magnetic field is 0~1T.

8. The establishment method according to claim 1, characterized in that: The first color light is red light; the second color light is green light.

9. The establishment method according to claim 1, characterized in that: After exposing the mouse to light of a second preset wavelength so that the photosensitive magnetic nanoparticles emit a second color of light to induce a photochemical reaction and stimulate a coagulation process to obtain a non-invasive ischemic stroke model, the method further includes: The mice were returned to housing and subjected to neurobehavioral assessments.

10. The establishment method according to claim 9, characterized in that: The neurobehavioral assessments are: In a single-blind condition, the mice were subjected to a grid walking test, a cylinder test, and a spaghetti grasping test; wherein the neurobehavioral assessment was conducted within 1 to 15 weeks after the injection of the photosensitive magnetic nanoparticles.

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