Method for constructing a human kidney organoid xenotransplantation model

Abstract

The present invention provides a method for constructing a humanized kidney organoid xenotransplantation model, which relates to the technical field of xenotransplantation model construction, and comprises the following steps: acquiring physical sign data of a number of immunodeficient mice to obtain a physical sign data set; matching each physical sign data in the physical sign data set with reference physical sign data respectively; after successful matching, using the corresponding immunodeficient mouse as an experimental carrier; pre-processing the experimental carrier to acquire a first image in real time; determining whether the renal capsule of the experimental carrier is exposed by matching the first image with a first reference image; after successful matching, generating a transplantation instruction; after receiving the transplantation instruction, a pipette sucks the kidney organoid and implants the kidney organoid into the renal capsule of the experimental carrier, and after implanting the kidney organoid, suturing the incision; acquiring a second image in real time; determining whether the incision of the experimental carrier is sutured by matching the second image with a second reference image; after successful matching, generating a monitoring instruction.

Description

Technical Field

[0001] The present invention relates to the technical field of xenotransplantation model construction, and in particular to a method for constructing a human-derived kidney organoid xenotransplantation model. Background Art

[0002] Human kidney organoid xenotransplantation refers to the establishment of human kidney organoids in animals. The animals can provide an in vivo experimental platform for studying human kidney organoids, which is of great significance in drug trials, kidney development, regenerative medicine and other aspects.

[0003] The steps for constructing the existing humanized kidney organoid xenotransplantation model are as follows: (1) prepare agarose rods of a specific diameter and soak them in a culture medium containing vascular endothelial growth factor (VEGF) to promote angiogenesis; (2) make an incision on the mouse renal capsule and insert the agarose rods in a V-shaped arrangement to create a free space and reduce tension; (3) transplant spheres of iPS cell-derived renal progenitor cells mixed with mouse embryonic spinal cord, and spheres of human umbilical vein endothelial cells (HUVECs) mixed with mesenchymal stem cells (MSCs) under the mouse renal capsule through the incision to complete the construction of the model. The preoperative preparation of the existing method requires the preparation and soaking of agarose rods, which is time-consuming and has many influencing factors. During the operation, the kidney organoids need to be mixed with other types of cells or tissues and then transplanted under the mouse renal capsule together, making it difficult to complete the entire construction process through a computational program; due to the introduction of too many confounding factors, it is not conducive to the separate analysis of the effect of the mouse renal capsule on the kidney organoids. Summary of the invention

[0004] The purpose of the present invention is to provide a method for constructing a humanized kidney organoid xenotransplantation model. The technical problem to be solved is how to complete the model construction process through a computational program, thereby reducing the influence of other types of cells or tissues on the kidney organoids transplanted into the mouse renal capsule.

[0005] The present invention is achieved through the following technical solutions:

[0006] The method for constructing a human kidney organoid xenotransplantation model comprises the following steps:

[0007] S100, obtaining physical sign data of a number of immunodeficient mice to obtain a physical sign data set;

[0008] S200, calling reference physical sign data, and matching each physical sign data in the physical sign data set with the reference physical sign data respectively;

[0009] If the match is successful, the immunodeficient mice corresponding to the physical sign data are used as experimental carriers;

[0010] If the matching fails, the process returns to S100 to continue execution and add new vital sign data;

[0011] S300, after obtaining the above experimental carrier, pre-treating the experimental carrier;

[0012] During preprocessing, image information of the experimental carrier is acquired in real time to obtain a first image;

[0013] S400, calling the first reference image, and determining whether the renal capsule of the experimental carrier is exposed by matching the first image with the first reference image;

[0014] If the match is successful, the renal capsule of the experimental carrier is exposed and a transplantation instruction is generated;

[0015] If the match fails, continue to pre-process the above experimental vector;

[0016] S500, after receiving the above transplantation instruction, the pipette sucks up the kidney organoid, implants the kidney organoid into the renal capsule of the experimental carrier, and sutures the incision after implanting the kidney organoid;

[0017] When executing the above transplantation instruction, image information of the experimental carrier is acquired in real time to obtain a second image;

[0018] S600, calling a second reference image, and determining whether the incision of the experimental carrier is sutured by matching the second image with the second reference image;

[0019] If the match is successful, the incision suture of the experimental carrier is completed and the monitoring instruction is generated;

[0020] If the match fails, continue to execute the transplantation instruction on the above experimental vector.

[0021] The vital sign data of immunodeficient mice were collected and analyzed by computer programs, compared with the preset reference vital sign data, and qualified experimental carriers were screened out, ensuring that only the most qualified mice were used as experimental carriers, thereby reducing unnecessary experimental interference caused by individual differences in mice; during the pretreatment process, real-time image acquisition and comparison were used to enable the experimental carrier to be detected instantly and accurately when the renal capsule was exposed, ensuring that the transplantation operation was performed when the renal capsule was exposed, reducing other tissue damage or contamination caused by improper operation timing, and thus reducing the potential impact of other cells or tissues on the transplanted kidney organoids; the transplantation instructions were completed by a pipette, reducing errors introduced by human factors, and the implantation was completed by injection, reducing physical damage to the kidney organoids and surrounding tissues during the transplantation process, reducing postoperative inflammatory reactions and unnecessary activation of other types of cells, which was beneficial to the survival and functional recovery of transplanted kidney organoids; by real-time monitoring of the incision suturing situation and immediate feedback on the suturing effect, the quality of the operation was ensured, which not only avoided problems such as incision infection and leakage caused by improper suturing, but also reduced possible inflammatory reactions and other tissue damage, indirectly protecting the transplanted kidney organoids from the negative effects of other cells or tissues.

[0022] The model construction of the present invention does not require the preparation of agarose rods, and the kidney organoids are directly implanted under the renal capsule of the experimental carrier, which shortens the model construction time and simplifies the construction process; when implanting the kidney organoids, there is no need to introduce other types of cells or tissues to mix with the kidney organoids, which reduces the mixed use with other types of cells or tissues, and reduces the influence of other types of cells or tissues on the kidney organoids transplanted in the mouse renal capsule, which helps to separately analyze the influence of the renal capsule on the kidney organoids, and provides a more reliable and efficient experimental platform for kidney organoid xenotransplantation research.

[0023] Furthermore, the reference sign types in the above reference sign data include the age, fasting duration and water deprivation duration of immunodeficient mice; the reference sign type data corresponding to the above reference sign types are: the age of immunodeficient mice is 8 to 12 weeks old, the fasting duration is 3.5 to 4.5 hours, and the water deprivation duration is 3.5 to 4.5 hours.

[0024] Furthermore, the specific steps of S200 include:

[0025] S210, extracting a physical sign data from the physical sign data set, and extracting a type corresponding to a reference physical sign type from the physical sign data to obtain a physical sign type set;

[0026] S220, extracting data corresponding to each physical sign type in the physical sign type set from the physical sign data to obtain physical sign type data;

[0027] S230, matching the above physical sign type data with reference physical sign type data of corresponding reference physical sign types;

[0028] S240: If the physical sign type data belongs to the reference physical sign type data corresponding to the reference physical sign type, the physical sign type data is matched successfully; otherwise, the physical sign type data is matched unsuccessfully;

[0029] S250, traversing the above-mentioned physical sign type set, if any physical sign type data belongs to the reference physical sign type data of the corresponding reference physical sign type, the physical sign data matching is successful; otherwise, the physical sign data matching fails.

[0030] Select one physical sign data from the physical sign data set, and compare the physical sign data with the reference physical sign data; extract the type and specific data corresponding to the reference physical sign type (such as age, fasting time, water deprivation time) from the physical sign data, and for each physical sign type data, determine whether it belongs to the reference physical sign type data range of the corresponding type, ensuring that each physical sign type is strictly screened; traverse the physical sign type set, check the matching results of all physical sign type data, if any physical sign type data is matched successfully, that is, all physical signs meet the requirements of the reference physical sign type data, then the physical sign data is matched successfully, and the immunodeficient mouse is selected as the experimental carrier; if any physical sign type data fails to match, then the physical sign data fails to match, and it is necessary to reselect one physical sign data from the physical sign data set and continue screening until an immunodeficient mouse that meets the requirements is screened out.

[0031] Through the above screening method, it is ensured that only immunodeficient mice that fully meet the preset conditions are selected for the experiment, thereby reducing the experimental errors and uncertainties caused by individual differences in experimental vectors, and improving the success rate and reliability of constructing human kidney organoid xenotransplantation models.

[0032] Further, when all the physical sign data in the above physical sign data set fail to match, new physical sign data is added;

[0033] The newly added physical sign data are added to the physical sign data set, the reference physical sign data are called, and the newly added physical sign data are matched with the reference physical sign data.

[0034] In the process of constructing a humanized kidney organoid xenotransplantation model, when all the physical sign data in the physical sign data set do not meet the reference physical sign data requirements, their physical sign data will be added to the physical sign data set and matched, ensuring that the screening process of the experimental vector is a dynamic and continuous process until immunodeficient mice that meet the preset conditions are screened out.

[0035] Furthermore, the specific steps of S300 include:

[0036] S310, after obtaining the above-mentioned experimental carrier, generating an anesthesia instruction; injecting anesthetic into the experimental carrier according to the above-mentioned anesthesia instruction, and detecting whether the experimental carrier has a pain response;

[0037] S320, if the experimental carrier has no pain response, a fixation and disinfection instruction is generated; the experimental carrier is fixed in a prone position according to the fixation and disinfection instruction, and after the fixation is completed, the left back area of ​​the experimental carrier is disinfected;

[0038] If the experimental carrier has a pain response, the process returns to S310 to continue the process and injects an anesthetic into the experimental carrier.

[0039] S330, after disinfection, generating a first separation instruction; using the first separation instruction, making a longitudinal incision in the left back area of ​​the experimental carrier to separate the skin layer and the muscle layer; acquiring image information of the experimental carrier to obtain a first image;

[0040] S340, calling the third reference image and the fourth reference image, respectively matching the first image with the third reference image and the first image with the fourth reference image, to determine whether the experimental carrier is separated correctly;

[0041] By matching the first image and the third reference image, determining whether the fat tissue of the experimental carrier is exposed; by matching the first image and the fourth reference image, determining whether the left kidney of the experimental carrier is exposed;

[0042] If the adipose tissue and the left kidney of the above experimental vector are exposed in sequence, the experimental vector is separated correctly; otherwise, the experimental vector is separated incorrectly;

[0043] S350, after the above-mentioned experimental vector is separated correctly, a renal capsule incision instruction is generated; through the above-mentioned renal capsule incision instruction, an incision is made in the left kidney of the experimental vector.

[0044] The above-mentioned pretreatment includes anesthesia, fixation and disinfection, separation of the skin layer and the muscle layer, and confirmation of the exposure of the anatomical structure (such as adipose tissue and the left kidney); specifically: in order to avoid the risk of surgical failure caused by the movement of the experimental carrier, an appropriate amount of anesthetic is injected into the experimental carrier, and it is detected whether the experimental carrier has completely lost consciousness and pain. If the experimental carrier still has a pain response, the anesthetic is injected again until the experimental carrier is completely pain-free; after confirming that the experimental carrier has no pain, the experimental carrier is fixed in a prone position to facilitate surgical operations; after fixation is completed, the left back area of ​​the experimental carrier (i.e., the surgical area) is disinfected to reduce the risk of infection; in the left back area A longitudinal incision is made in the domain to separate the skin layer and the muscle layer. At the same time, a first image is acquired, and whether the fat tissue of the experimental carrier has been correctly exposed is determined by matching the first image with the third reference image; whether the left kidney of the experimental carrier has been correctly exposed is determined by matching the first image with the fourth reference image; only when the fat tissue and the left kidney are exposed in sequence, the experimental carrier is considered to be correctly separated; if the experimental carrier is separated incorrectly (i.e., the fat tissue or the left kidney is not correctly exposed), it is necessary to re-perform the separation operation or adjust the surgical strategy; after confirming that the experimental carrier is separated correctly, an incision is made on the left kidney of the experimental carrier to prepare for kidney organoid transplantation.

[0045] Furthermore, before receiving the transplantation instruction, the pipette needs to be disinfected and equipped with cell culture medium.

[0046] Although kidney organoids are kept in cell culture medium before transplantation, in order to avoid the kidney organoids being away from the cell culture medium for too long during the transplantation process and affecting their activity and function, the kidney organoids are also kept in cell culture medium during the transplantation process; when implanting kidney organoids, the cell culture medium must be removed before the implantation operation is performed, reducing the immune response caused by the cell culture medium entering the renal capsule.

[0047] Furthermore, S500 further includes the following steps:

[0048] S510, implanting the kidney organoid into the renal capsule of the experimental carrier, acquiring image information of the experimental carrier, and obtaining a second image;

[0049] S520, calling the fifth reference image, and determining whether there is bleeding in the renal capsule of the experimental carrier after the kidney organoid is implanted by matching the second image with the fifth reference image;

[0050] If the match is successful, there is no bleeding in the renal capsule of the experimental carrier;

[0051] If the match failed, the renal capsule of the experimental carrier was bleeding.

[0052] In order to improve the success rate of transplantation and the safety of the experimental carrier, the renal capsule of the experimental carrier is judged in time to react intuitively. After the implantation is completed, the image information of the experimental carrier is obtained by using imaging technology (such as ultrasound, CT, MRI or optical imaging, etc.), which is called the "second image"; the pre-stored "fifth reference image" is called, and the fifth reference image is the image information of the experimental carrier under similar operation or experimental conditions without bleeding or other complications. By comparing and matching the second image (i.e., the image of the experimental carrier after implantation of the kidney organoid) with the fifth reference image, the state of the renal capsule is evaluated; if the match is successful, it means that the second image and the fifth reference image are highly similar in key features and show no signs of renal capsule bleeding. Therefore, it can be judged that the renal capsule of the experimental carrier has no bleeding after implantation of the kidney organoid; if the match fails, it indicates that there are significant differences between the second image and the fifth reference image, which may indicate renal capsule bleeding or other abnormalities. In this case, emergency measures need to be taken to deal with the bleeding.

[0053] Furthermore, when executing the above monitoring instructions, the experimental carrier is placed in an environment with a temperature of 20 to 26°C and a relative humidity of 40 to 70%.

[0054] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0055] The vital sign data of immunodeficient mice were collected and analyzed by computer programs, compared with the preset reference vital sign data, and qualified experimental carriers were screened out, ensuring that only the most qualified mice were used as experimental carriers, thereby reducing unnecessary experimental interference caused by individual differences in mice; during the pretreatment process, real-time image acquisition and comparison were used to enable the experimental carrier to be detected instantly and accurately when the renal capsule was exposed, ensuring that the transplantation operation was performed when the renal capsule was exposed, reducing other tissue damage or contamination caused by improper operation timing, and thus reducing the potential impact of other cells or tissues on the transplanted kidney organoids; the transplantation instructions were completed by a pipette, reducing errors introduced by human factors, and the implantation was completed by injection, reducing physical damage to the kidney organoids and surrounding tissues during the transplantation process, reducing postoperative inflammatory reactions and unnecessary activation of other types of cells, which was beneficial to the survival and functional recovery of transplanted kidney organoids; by real-time monitoring of the incision suturing situation and immediate feedback on the suturing effect, the quality of the operation was ensured, which not only avoided problems such as incision infection and leakage caused by improper suturing, but also reduced possible inflammatory reactions and other tissue damage, indirectly protecting the transplanted kidney organoids from the negative effects of other cells or tissues.

[0056] Compared with the prior art, the model construction of the present invention does not require the preparation of agarose rods, and the kidney organoids are directly implanted under the renal capsule of the experimental carrier, which shortens the model construction time and simplifies the construction process; when implanting the kidney organoids, there is no need to introduce other types of cells or tissues to mix with the kidney organoids, which reduces the mixed use with other types of cells or tissues, and reduces the influence of other types of cells or tissues on the kidney organoids transplanted in the mouse renal capsule, which helps to separately analyze the influence of the renal capsule on the kidney organoids, and provides a more reliable and efficient experimental platform for kidney organoid xenotransplantation research. BRIEF DESCRIPTION OF THE DRAWINGS

[0057] In order to more clearly illustrate the technical solutions of the exemplary embodiments of the present invention, the following briefly introduces the drawings required for use in the embodiments. It should be understood that the following drawings only illustrate certain embodiments of the present invention and should not be regarded as limiting the scope. For those of ordinary skill in the art, other relevant drawings can be obtained based on these drawings without creative work. In the drawings:

[0058] Figure 1 The main flow chart of the construction method;

[0059] Figure 2 This is a schematic diagram of the left kidney after implantation of kidney organoids. The rectangular box shows the implanted kidney organoids. DETAILED DESCRIPTION

[0060] In order to make the objectives, technical solutions and advantages of the present invention more clearly understood, the present invention is further described in detail below in conjunction with embodiments and drawings. The exemplary embodiments of the present invention and their description are only used to explain the present invention and are not intended to limit the present invention.

[0061] Example 1

[0062] Combination Figure 1 The method for constructing a human kidney organoid xenotransplantation model comprises the following steps:

[0063] S100, obtaining physical sign data of a number of immunodeficient mice to obtain a physical sign data set, wherein the immunodeficient mice may be NPSC immunodeficient mice;

[0064] S200, calling reference physical sign data, and matching each physical sign data in the physical sign data set with the reference physical sign data respectively;

[0065] If the match is successful, the immunodeficient mice corresponding to the physical sign data are used as experimental carriers;

[0066] If the matching fails, the process returns to S100 to continue execution and add new vital sign data;

[0067] S300, after obtaining the above experimental carrier, pre-treating the experimental carrier;

[0068] During preprocessing, image information of the experimental carrier is acquired in real time to obtain a first image;

[0069] S400, calling a first reference image, wherein the first reference image is image information of the experimental carrier exposing the renal capsule under similar operation or experimental conditions, and the image has an incision under the left kidney of the experimental carrier to form a pocket; determining whether the renal capsule of the experimental carrier is exposed by matching the first image with the first reference image;

[0070] If the match is successful, the renal capsule of the experimental carrier is exposed and a transplantation instruction is generated;

[0071] If the match fails, continue to pre-process the above experimental vector;

[0072] S500, after receiving the above transplantation instruction, the pipette sucks up the kidney organoid and implants the kidney organoid into the renal capsule of the experimental carrier. After implanting the kidney organoid, the incisions of the skin layer and the muscle layer are sutured;

[0073] When executing the above transplantation instruction, image information of the experimental carrier is acquired in real time to obtain a second image;

[0074] S600, calling a second reference image, wherein the second reference image is image information when the experimental carrier is sutured under similar operation or experimental conditions; determining whether the incision of the experimental carrier is sutured by matching the second image with the second reference image;

[0075] If the match is successful, the incision suture of the experimental carrier is completed and the monitoring instruction is generated;

[0076] If the match fails, continue to execute the transplantation instruction on the above experimental vector.

[0077] The vital sign data of immunodeficient mice were collected and analyzed by computer programs, compared with the preset reference vital sign data, and qualified experimental carriers were screened out, ensuring that only the most qualified mice were used as experimental carriers, thereby reducing unnecessary experimental interference caused by individual differences in mice; during the pretreatment process, real-time image acquisition and comparison were used to enable the experimental carrier to be detected instantly and accurately when the renal capsule was exposed, ensuring that the transplantation operation was performed when the renal capsule was exposed, reducing other tissue damage or contamination caused by improper operation timing, and thus reducing the potential impact of other cells or tissues on the transplanted kidney organoids; the transplantation instructions were completed by a pipette, reducing errors introduced by human factors, and the implantation was completed by injection, reducing physical damage to the kidney organoids and surrounding tissues during the transplantation process, reducing postoperative inflammatory reactions and unnecessary activation of other types of cells, which was beneficial to the survival and functional recovery of transplanted kidney organoids; by real-time monitoring of the incision suturing situation and immediate feedback on the suturing effect, the quality of the operation was ensured, which not only avoided problems such as incision infection and leakage caused by improper suturing, but also reduced possible inflammatory reactions and other tissue damage, indirectly protecting the transplanted kidney organoids from the negative effects of other cells or tissues.

[0078] The model construction of the present invention does not require the preparation of agarose rods, and the kidney organoids are directly implanted under the renal capsule of the experimental carrier, which shortens the model construction time and simplifies the construction process; when implanting the kidney organoids, there is no need to introduce other types of cells or tissues to mix with the kidney organoids, which reduces the mixed use with other types of cells or tissues, and reduces the influence of other types of cells or tissues on the kidney organoids transplanted in the mouse renal capsule, which helps to separately analyze the influence of the renal capsule on the kidney organoids, and provides a more reliable and efficient experimental platform for kidney organoid xenotransplantation research.

[0079] Example 2

[0080] Based on Example 1, the reference sign types in the above reference sign data include the age, fasting duration and water deprivation duration of immunodeficient mice; the reference sign type data corresponding to the above reference sign types are: the age of immunodeficient mice is 8 to 12 weeks old, the fasting duration is 3.5 to 4.5 hours, and the water deprivation duration is 3.5 to 4.5 hours.

[0081] The immune system of mice aged 8 to 12 weeks is relatively mature but not yet fully adult. Mice in this age group have better tolerance to surgery and transplantation, which is conducive to the smooth progress of the experiment. Based on considerations of the metabolic rate and water balance of mice, ensure that the mice are in a suitable physiological state before surgery. Fasting and water deprivation for 3.5 to 4.5 hours can help reduce the risk of vomiting and aspiration during anesthesia and surgery, and maintain the physiological stability of mice during surgery.

[0082] By controlling the age, fasting and water deprivation duration of the experimental carriers, the risks during the operation, such as vomiting and aspiration, are reduced, thereby reducing damage to other cells or tissues caused by surgical complications; the appropriate physiological state helps the experimental carriers to better accept the transplantation operation, reduce postoperative rejection and inflammatory reactions, and thus protect the transplanted kidney organoids from attacks by other cells or tissues; standardized reference vital signs data ensures consistency between experimental carriers, making the experimental results more reliable and repeatable, helping researchers to more accurately evaluate the transplantation effect and identify key factors that may affect the success of the transplantation; by reducing stress reactions and injuries during the operation, the experimental carriers can recover their health faster, provide a better growth environment for transplanted kidney organoids, and help transplanted kidney organoids achieve better survival and functional recovery in xenogeneic bodies.

[0083] Example 3

[0084] Based on Example 2, the specific steps of S200 include:

[0085] S210, extracting a physical sign data from the physical sign data set, and extracting a type corresponding to a reference physical sign type from the physical sign data to obtain a physical sign type set;

[0086] S220, extracting data corresponding to each physical sign type in the physical sign type set from the physical sign data to obtain physical sign type data;

[0087] S230, matching the above physical sign type data with reference physical sign type data of corresponding reference physical sign types;

[0088] S240: If the physical sign type data belongs to the reference physical sign type data corresponding to the reference physical sign type, the physical sign type data is matched successfully; otherwise, the physical sign type data is matched unsuccessfully;

[0089] S250, traversing the above-mentioned physical sign type set, if any physical sign type data belongs to the reference physical sign type data of the corresponding reference physical sign type, the physical sign data matching is successful; otherwise, the physical sign data matching fails.

[0090] Select one physical sign data from the physical sign data set, and compare the physical sign data with the reference physical sign data; extract the type and specific data corresponding to the reference physical sign type (such as age, fasting time, water deprivation time) from the physical sign data, and for each physical sign type data, determine whether it belongs to the reference physical sign type data range of the corresponding type, ensuring that each physical sign type is strictly screened; traverse the physical sign type set, check the matching results of all physical sign type data, if any physical sign type data is matched successfully, that is, all physical signs meet the requirements of the reference physical sign type data, then the physical sign data is matched successfully, and the immunodeficient mouse is selected as the experimental carrier; if any physical sign type data fails to match, then the physical sign data fails to match, and it is necessary to reselect one physical sign data from the physical sign data set and continue screening until an immunodeficient mouse that meets the requirements is screened out.

[0091] Through the above screening method, it is ensured that only immunodeficient mice that fully meet the preset conditions are selected for the experiment, thereby reducing the experimental errors and uncertainties caused by individual differences in experimental vectors, and improving the success rate and reliability of constructing human kidney organoid xenotransplantation models.

[0092] A specific example for reference is given a physical sign data set, which consists of physical sign data one to physical sign data three, and the type of each physical sign data includes age, fasting time, water deprivation time, body size and weight. The physical sign data set is shown in Table 1.

[0093] Table 1

[0094] Age (days) Fasting duration (hours) Water fasting duration (hours) Body size (cm) Weight (g) Vital signs data 1 70 3 4 10 28 Vital signs data 2 63 4 4 9 27 Vital signs data 3 49 4 4 5 23

[0095] When screening experimental vectors, match the physical sign data one to the physical sign data three with the reference physical sign data in turn; here, taking the matching of physical sign data one with the reference physical sign data as an example, the specific steps are as follows: extract data corresponding to age, fasting duration and water deprivation duration from physical sign data one to obtain age data, fasting duration data and water deprivation duration data (i.e., physical sign type data); match the age data (70 days) with 8 to 12 weeks of age, the age data belongs to 8 to 12 weeks of age, and the age data is successfully matched; match the fasting duration data (3 hours) with 3.5 to 4.5 hours, the fasting duration data Since the water fasting time data does not belong to 3.5-4.5 hours, the fasting time data matching fails. The above steps can be executed sequentially or simultaneously. When executed sequentially, the vital sign data one is directly identified as a matching failure, and the vital sign data two is matched. When processed in parallel, the water fasting time data (4 hours) is matched with 3.5-4.5 hours at the same time. Since the water fasting time data belongs to 3.5-4.5 hours, the water fasting time data is matched successfully. Then, whether the vital sign data one is matched successfully is determined based on the matching results of all vital sign type data. Since the fasting time data matching fails, the vital sign data one matching fails.

[0096] Example 4

[0097] Based on any of the above embodiments, when all the physical sign data in the above physical sign data set fail to match, new physical sign data is added; the newly added physical sign data is added to the physical sign data set, the reference physical sign data is called, and the newly added physical sign data is matched with the reference physical sign data.

[0098] In the process of constructing a humanized kidney organoid xenotransplantation model, when all the physical sign data in the physical sign data set do not meet the reference physical sign data requirements, their physical sign data will be added to the physical sign data set and matched, ensuring that the screening process of the experimental vector is a dynamic and continuous process until immunodeficient mice that meet the preset conditions are screened out.

[0099] Example 5

[0100] Based on any of the above embodiments, the specific steps of S300 include:

[0101] S310, after obtaining the above-mentioned experimental carrier, generating an anesthesia instruction; injecting anesthetic into the experimental carrier according to the above-mentioned anesthesia instruction, and detecting whether the experimental carrier has a pain response;

[0102] S320, if the experimental carrier has no pain response, a fixation and disinfection instruction is generated; the experimental carrier is fixed in a prone position according to the fixation and disinfection instruction, and after fixation, the left back area of ​​the experimental carrier is disinfected, and 70% alcohol or iodine can be used for disinfection;

[0103] If the experimental carrier has a pain response, the process returns to S310 to continue the process and injects an anesthetic into the experimental carrier.

[0104] S330, after disinfection, generating a first separation instruction; using the first separation instruction, making a longitudinal incision in the left back area of ​​the experimental carrier to separate the skin layer and the muscle layer; acquiring image information of the experimental carrier to obtain a first image;

[0105] S340, calling a third reference image and a fourth reference image, wherein the third reference image is image information when the experimental carrier exposes fat tissue under similar operation or experimental conditions; and the fourth reference image is image information when the experimental carrier exposes the left kidney under similar operation or experimental conditions;

[0106] Matching the first image with the third reference image and the first image with the fourth reference image respectively to determine whether the experimental carrier is separated correctly;

[0107] By matching the first image and the third reference image, determining whether the fat tissue of the experimental carrier is exposed; by matching the first image and the fourth reference image, determining whether the left kidney of the experimental carrier is exposed;

[0108] If the adipose tissue and the left kidney of the above experimental vector are exposed in sequence, the experimental vector is separated correctly; otherwise, the experimental vector is separated incorrectly;

[0109] S350, after the above-mentioned experimental vector is separated correctly, a renal capsule incision instruction is generated; through the above-mentioned renal capsule incision instruction, an incision is made in the left kidney of the experimental vector.

[0110] The above-mentioned pretreatment includes anesthesia, fixation and disinfection, separation of the skin layer and the muscle layer, and confirmation of the exposure of the anatomical structure (such as adipose tissue and the left kidney); specifically: in order to avoid the risk of surgical failure caused by the movement of the experimental carrier, an appropriate amount of anesthetic is injected into the experimental carrier, and it is detected whether the experimental carrier has completely lost consciousness and pain. If the experimental carrier still has a pain response, the anesthetic is injected again until the experimental carrier is completely pain-free; after confirming that the experimental carrier has no pain, the experimental carrier is fixed in a prone position to facilitate surgical operations; after fixation is completed, the left back area of ​​the experimental carrier (i.e., the surgical area) is disinfected to reduce the risk of infection; in the left back area A longitudinal incision is made in the domain to separate the skin layer and the muscle layer. At the same time, a first image is acquired, and whether the fat tissue of the experimental carrier has been correctly exposed is determined by matching the first image with the third reference image; whether the left kidney of the experimental carrier has been correctly exposed is determined by matching the first image with the fourth reference image; only when the fat tissue and the left kidney are exposed in sequence, the experimental carrier is considered to be correctly separated; if the experimental carrier is separated incorrectly (i.e., the fat tissue or the left kidney is not correctly exposed), it is necessary to re-perform the separation operation or adjust the surgical strategy; after confirming that the experimental carrier is separated correctly, an incision is made on the left kidney of the experimental carrier to prepare for kidney organoid transplantation.

[0111] A specific example for reference is to use anesthetics to perform general anesthesia on the experimental carrier. After confirming that the experimental carrier has completely lost consciousness and has no pain response, the experimental carrier is fixed on the operating table in a prone position, and 70% alcohol or iodine is used to disinfect the surgical area, especially the left back area; starting from the lower edge of the middle part of the 11th rib on the left back of the experimental carrier, a longitudinal incision of about 1 to 2 cm is made on the side close to the abdomen with a sterile surgical blade. The longitudinal incision is parallel to the spine of the experimental carrier, and sterile blunt-tipped scissors or a scalpel are used to peel off the tissue along the anatomical plane (longitudinal incision), separate the skin layer and the muscle layer, and expose the retroperitoneal fat tissue. The appearance of the adipose tissue is loose, yellow fat masses, which are clearly distinguished from the surrounding muscle tissue and other structures; continue to separate the adipose tissue and muscle layer until the left kidney is exposed. The appearance of the left kidney is oval or slightly curved oval, with a smooth surface, clear contours, and usually dark red or reddish brown, which is significantly different from the surrounding adipose tissue and muscle tissue in color; use sterile tweezers and scissors to separate and remove the adipose tissue covering the surface of the left kidney. During the separation process, find the sunken area of ​​the kidney (i.e., the renal hilum), and use sterile tweezers to tear a small hole in the renal capsule about 2 mm to the left of the renal hilum to form a "pocket" to complete the pretreatment.

[0112] Afterwards, use a sterilized 20ul range pipette with the corresponding gun tip, cut off the tip area of ​​about 2mm at the front end of the gun tip to obtain a pipette; use negative pressure to suck the kidney organoids and cell culture fluid into the pipette. At this time, due to the presence of cell culture fluid, the kidney organ tube is located at the rear end of the pipette. In order to reduce the impact of the cell culture fluid on the experimental carrier, it is necessary to slowly discharge the cell culture fluid at the front end of the pipette until the kidney organoid reaches the front end of the pipette, and then bounce the kidney organoid into the incision "pocket" under the left kidney capsule to complete the implantation of the kidney organoid. The left kidney after implantation is as follows Figure 2 As shown, from Figure 2 It can be seen that the kidney organoids transplanted by this method have no obvious subrenal capsule bleeding, and do not require the assistance of a microscope, nor are there any other biological agents or cell tissues mixed in. Use sterile surgical sutures (such as 4-0 or 5-0 silk) to suture the longitudinal incisions of the skin layer and muscle layer, ensuring that the sutures are tight to prevent postoperative infection and incision dehiscence. Place the mice in a warm, dry recovery environment and closely monitor their recovery.

[0113] Example 6

[0114] Based on any of the above embodiments, before receiving the transplantation instruction, the pipette needs to be disinfected and equipped with cell culture medium in the pipette, and the cell culture medium can be Advanced RPMI 1640 culture medium.

[0115] Although kidney organoids are kept in cell culture medium before transplantation, in order to avoid the kidney organoids being away from the cell culture medium for too long during the transplantation process and affecting their activity and function, the kidney organoids are also kept in cell culture medium during the transplantation process; when implanting kidney organoids, the cell culture medium must be removed before the implantation operation is performed, reducing the immune response caused by the cell culture medium entering the renal capsule.

[0116] Example 7

[0117] Based on any of the above embodiments, S500 further includes the following steps:

[0118] S510, implanting the kidney organoid into the renal capsule of the experimental carrier, acquiring image information of the experimental carrier, and obtaining a second image;

[0119] S520, calling the fifth reference image, and determining whether there is bleeding in the renal capsule of the experimental carrier after the kidney organoid is implanted by matching the second image with the fifth reference image;

[0120] If the match is successful, there is no bleeding in the renal capsule of the experimental carrier;

[0121] If the match failed, the renal capsule of the experimental carrier was bleeding.

[0122] In order to improve the success rate of transplantation and the safety of the experimental carrier, the renal capsule of the experimental carrier is judged in time to react intuitively. After the implantation is completed, the image information of the experimental carrier is obtained by using imaging technology (such as ultrasound, CT, MRI or optical imaging, etc.), which is called the "second image"; the pre-stored "fifth reference image" is called, and the fifth reference image is the image information of the experimental carrier under similar operation or experimental conditions without bleeding or other complications. By comparing and matching the second image (i.e., the image of the experimental carrier after implantation of the kidney organoid) with the fifth reference image, the state of the renal capsule is evaluated; if the match is successful, it means that the second image and the fifth reference image are highly similar in key features and show no signs of renal capsule bleeding. Therefore, it can be judged that the renal capsule of the experimental carrier has no bleeding after implantation of the kidney organoid; if the match fails, it indicates that there are significant differences between the second image and the fifth reference image, which may indicate renal capsule bleeding or other abnormalities. In this case, emergency measures need to be taken to deal with the bleeding.

[0123] In a specific embodiment, when executing the above monitoring instructions, the experimental carrier is placed in an environment with a temperature of 20-26° C. and a relative humidity of 40-70%.

[0124] The specific implementation methods described above further illustrate the objectives, technical solutions and beneficial effects of the present invention in detail. It should be understood that the above description is only a specific implementation method of the present invention and is not intended to limit the scope of protection of the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A method for constructing a human kidney organoid xenotransplantation model, characterized in that: The following steps are involved: S100, obtaining physical sign data of a number of immunodeficient mice to obtain a physical sign data set; S200, calling reference physical sign data, and matching each physical sign data in the physical sign data set with the reference physical sign data respectively; If the match is successful, the immunodeficient mice corresponding to the physical sign data are used as experimental carriers; If the matching fails, the process returns to S100 to continue execution and add new vital sign data; S300, after obtaining the experimental carrier, pre-treating the experimental carrier; During preprocessing, image information of the experimental carrier is acquired in real time to obtain a first image; S400, calling a first reference image, and determining whether the renal capsule of the experimental carrier is exposed by matching the first image with the first reference image; If the match is successful, the renal capsule of the experimental carrier is exposed and a transplantation instruction is generated; If the match fails, continue to pre-process the experimental vector; S500, after receiving the transplantation instruction, the pipette sucks up the kidney organoid, implants the kidney organoid into the renal capsule of the experimental carrier, and sutures the incision after implanting the kidney organoid; When executing the transplantation instruction, image information of the experimental carrier is acquired in real time to obtain a second image; S600, calling a second reference image, and determining whether the incision of the experimental carrier is sutured by matching the second image with the second reference image; If the match is successful, the incision suture of the experimental carrier is completed and the monitoring instruction is generated; If the match fails, the transplantation instruction continues to be executed on the experimental vector.

2. The method for constructing a human kidney organoid xenotransplantation model according to claim 1, characterized in that: The reference sign types in the reference sign data include the age, fasting duration and water deprivation duration of the immunodeficient mice; the reference sign type data corresponding to the reference sign types are: the age of the immunodeficient mice is 8 to 12 weeks old, the fasting duration is 3.5 to 4.5 hours, and the water deprivation duration is 3.5 to 4.5 hours.

3. The method for constructing a human kidney organoid xenotransplantation model according to claim 2, characterized in that: The specific steps of S200 include: S210, extracting a physical sign data from the physical sign data set, and extracting a type corresponding to a reference physical sign type from the physical sign data to obtain a physical sign type set; S220, extracting data corresponding to each physical sign type in the physical sign type set from the physical sign data to obtain physical sign type data; S230, matching the physical sign type data with reference physical sign type data of corresponding reference physical sign types; S240: If the physical sign type data belongs to the reference physical sign type data corresponding to the reference physical sign type, the physical sign type data is matched successfully; otherwise, the physical sign type data is matched unsuccessfully; S250, traversing the physical sign type set, if any physical sign type data belongs to the reference physical sign type data of the corresponding reference physical sign type, the physical sign data matching is successful; otherwise, the physical sign data matching fails.

4. The method for constructing a human kidney organoid xenotransplantation model according to claim 1, characterized in that: When all the physical sign data in the physical sign data set fail to match, adding physical sign data; The newly added physical sign data is added to the physical sign data set, the reference physical sign data is called, and the newly added physical sign data is matched with the reference physical sign data.

5. The method for constructing a human kidney organoid xenotransplantation model according to claim 1, characterized in that: The specific steps of S300 include: S310, after obtaining the experimental carrier, generating an anesthesia instruction; injecting anesthetic into the experimental carrier according to the anesthesia instruction, and detecting whether the experimental carrier has a pain response; S320, if the experimental carrier has no pain response, a fixation and disinfection instruction is generated; the experimental carrier is fixed in a prone position according to the fixation and disinfection instruction, and after the fixation is completed, the left back area of ​​the experimental carrier is disinfected; If the experimental carrier has a pain response, the process returns to S310 to continue the process and inject an anesthetic into the experimental carrier. S330, after disinfection, generating a first separation instruction; using the first separation instruction, making a longitudinal incision in the left back area of ​​the experimental carrier to separate the skin layer and the muscle layer; acquiring image information of the experimental carrier to obtain a first image; S340, calling the third reference image and the fourth reference image, respectively matching the first image with the third reference image and the first image with the fourth reference image, to determine whether the experimental carrier is separated correctly; By matching the first image and the third reference image, determining whether the fat tissue of the experimental carrier is exposed; by matching the first image and the fourth reference image, determining whether the left kidney of the experimental carrier is exposed; If the adipose tissue and the left kidney of the experimental vector are exposed in sequence, the experimental vector is separated correctly; otherwise, the experimental vector is separated incorrectly; S350, after the experimental carrier is correctly separated, a renal capsule incision instruction is generated; an incision is made in the left kidney of the experimental carrier according to the renal capsule incision instruction.

6. The method for constructing a human kidney organoid xenotransplantation model according to claim 1, characterized in that: Before receiving the transplantation instruction, the pipette needs to be disinfected and equipped with cell culture fluid.

7. The method for constructing a human kidney organoid xenotransplantation model according to claim 1, characterized in that: S500 also includes the following steps: S510, implanting the kidney organoid into the renal capsule of the experimental carrier, acquiring image information of the experimental carrier, and obtaining a second image; S520, calling the fifth reference image, and determining whether there is bleeding in the renal capsule of the experimental carrier after the kidney organoid is implanted by matching the second image with the fifth reference image; If the match is successful, there is no bleeding in the renal capsule of the experimental carrier; If the match failed, the renal capsule of the experimental carrier was bleeding.

8. The method for constructing a human kidney organoid xenotransplantation model according to claim 1, characterized in that: When executing the monitoring instructions, the experimental carrier is placed in an environment with a temperature of 20 to 26° C. and a relative humidity of 40 to 70%.

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