CN109682586B

CN109682586B - Method for evaluating and testing in vitro intravascular stent based on microfluidic chip and application

Info

Publication number: CN109682586B
Application number: CN201910032581.XA
Authority: CN
Inventors: 徐紫宸; 王贵学
Original assignee: Chongqing University
Current assignee: Chongqing University
Priority date: 2019-01-14
Filing date: 2019-01-14
Publication date: 2021-01-29
Anticipated expiration: 2039-01-14
Also published as: CN109682586A

Abstract

The invention belongs to the technical field of microfluidic chips, and particularly relates to a method for performing an in vitro intravascular stent evaluation experiment based on a microfluidic chip and application of the method. The method comprises the following steps: the micro-fluidic chip in-vitro intravascular stent evaluation system comprises a sample adding valve 1, a circulating pulsation micro pump 6 and a cell culture pool 4, is used for sampling through a sampling plug 13, collects experimental data and evaluates experimental results. The method reduces the time cost and the consumption of raw material cost for evaluating the experiment requirements, ensures the identity of the experiment under multiple conditions, and ensures the comparability of the experiment results.

Description

Method for evaluating and testing in vitro intravascular stent based on microfluidic chip and application

Technical Field

The invention belongs to the technical field of microfluidic chips, and particularly relates to a method for performing an in vitro intravascular stent evaluation experiment based on a microfluidic chip and application of the method.

Background

The prevalence rate of cardiovascular diseases is rising year by year, and according to the aging situation and population growth rate of the population in China, the prevalence rate of cardiovascular diseases in China is predicted to increase to 50% in 2030, and the severity and universality of cardiovascular diseases are not neglected. One of the most important treatment modalities for cardiovascular diseases is intervention, in which the implantation of vascular stents is the most common means in intervention. The research and development of new vascular stents more suitable for the complex internal environment of human bodies have great significance for relieving the incidence of the increasingly severe cardiovascular diseases.

For the relevant study of vascular stents,evaluating the distance between the stent and the blood flow and the vascular tissue after implantation in a blood vessel The interaction effect is a very critical ring, which is related to the incidence of clinical adverse events after stent implantation and the late heart after surgery The recurrence rate of vascular disease ensures that the stent can perform the therapeutic action as expected and does not produce other fatal later-period side effects The application is as follows.In addition, the research on the force-biology mechanism is also an important thrust for detecting the occurrence and the development of cardiovascular diseases, and the optimal intervention scheme can be constructed by carrying out specific modification and modification on the stent or a stent coating through the related mechanism.

Currently, relevant evaluation studies are often limited to animal model testing. The animal experiment has many limiting factors, such as the control of the physiological state and the interventional operation condition of the animal,The cost for raising experimental animals and constructing animal models, etc., also has the disadvantages of long preparation period, complicated preparation procedure, etc. However, the evaluation test of the stent is indispensable.

Therefore, in order to enable researchers to complete quickly and convenientlyAn evaluation experiment of the vascular stent is also designed by utilizing the microfluidic technology to accelerate the research process of the vascular stent and solve the treatment problem of the closed coronary artery diseaseMethod for carrying out related evaluation experiment in vitroIs very necessary.

Disclosure of Invention

In view of the above, an object of the present invention is to provide a method for performing an in vitro intravascular stent evaluation experiment based on a microfluidic chip, which can conveniently and effectively evaluate the intravascular stent, and overcomes the disadvantages of the feeding cost of experimental animals, the construction cost of animal models, long preparation period, complicated preparation procedures, and the like.

In order to achieve the purpose, the invention adopts the following scheme:

a method for carrying out in-vitro intravascular stent evaluation experiments based on a microfluidic chip comprises the following steps:

1) setting a micro-fluidic chip in-vitro vascular stent evaluation system comprising a sample adding valve 1, a circulating pulsation micro-pump 6 and a cell culture pool 4;

2) closing all the stop valves 11 and the sample adding valve 1, sleeving the blood vessel support 12 on the matched support experiment tube 7, adding a culture medium from the sample port 8, flowing into the cell culture tank 4, inoculating the living cell suspension on the blood vessel support 12, and opening the stop valves 11 after the cells adhere to the wall;

3) sampling is performed through the sampling plug 13, and experimental data is collected to evaluate the experimental results.

The chip mainly comprises a sample adding valve 1, a circulating pulsation micropump 6 and a cell culture pool 4; the chip is circular or rectangular, and 2 side-by-side cylindrical sample adding valves 1 are processed at the center of the chip; each cylindrical sample adding valve 1 is provided with 2 opposite openings on the same section diameter line, and each opening is respectively connected with 1 sample adding pipe 2; each sampling pipe 2 is respectively led to one respective

cell culture pond

4, and 4 cell culture ponds 4 are formed in total and are communicated with a disc-shaped channel 3 processed at the bottom in each cell culture pond 4;

the 4 cell culture ponds 4 are uniformly distributed at four corners of the rectangular chip or at four symmetrical edges of the circular chip; 1 circulating pulsation micropump 6 is arranged between a pair of cell culture ponds 4, the circulating pulsation micropump 6 is communicated in series between 2 circulating pipes 5, and the 2 circulating pipes 5 are respectively communicated with the disc-shaped channels 3 in the cell culture ponds 4 at two sides of the circulating pulsation micropump 6; the circulating pipe 5 and the sample injection pipe 2 are communicated with the disc-shaped channel 3 at a T-shaped interface 17 at the side of the cell culture pond 4; the middle of the disc-shaped channel 3 is provided with a pipeline partition wall 19, and the disc-shaped channel 3 is divided into 2 disc-shaped double small pipes 18 from the T-shaped interface 17; the disc-shaped channel 3 is coiled to the center of the cell culture pool 4 along the bottom surface of the cell culture pool 4 and is communicated with a bracket experiment tube 7 which is processed and fixed at the center; the bracket experiment tube 7 is a hollow tube which is made of elastic materials and is higher than the cell culture pool 4, and a sampling plug 13 is arranged at the opening of the top end of the bracket experiment tube; the sampling plug 13 is internally provided with a pressure sensing probe.

2 sample ports 8 are processed in the middle of the chip between 2 side-by-side circular sample adding valves 1, sample channels 9 are processed between the 2 sample ports 8 and are communicated with each other, and stop valves 11 are processed on the communicated sample channels 9; the 2 sample ports 8 are also respectively provided with 2 sample channels 9 which are respectively communicated with the 2 cell culture pools 4.

The sampling tube 2 is respectively provided with 1 one-way circulating tube 10 beside 2 sample adding valves 1 and is communicated with the sampling tubes 2 at two ends of the sample adding valves 1; each one-way circulation pipe 10 is provided with a stop valve 11.

All the sample inlet pipes 2, the disc-shaped channels 3, the circulating pipe 5, the sample channels 9, the one-way circulating pipe 10 and the bracket experiment pipe 7 of the chip are sealed pipelines; the disc-shaped channel 3 is sealed by an ion exchange membrane, so that the ion solution in the disc-shaped channel 3 and the solution in the cell culture pool 4 can not be mixed; the cylindrical interior of the sample adding valve 1 is a valve body 16, an upper opening is processed in the valve body 16, and an inner valve channel 14 with the size of an end head matched with the sample inlet pipe 2 is formed in the valve body 16; the valve body 16 is provided with a valve knob 15 at the top.

Further, when the cells are evaluated as variables, step 2) comprises closing all the stop valves 11 and the sample adding valve 1, sleeving the same blood vessel support 12 on the matched support experiment tube 7, adding the same culture medium from the sample port 8 and flowing into the cell culture tank 4, inoculating different living cell suspensions on the blood vessel support 12, opening the stop valves 11 after the cells adhere to the wall, keeping the stop valve between the two sample ports 8 closed, and adjusting the circulating pulsation micropump 6 to start the evaluation experiment.

The function of the stop valve is mainly to isolate two different culture environments.

Further, the medium is a mixed reagent of a nonionic reagent and a cell culture solution.

The mixing of the non-ionic agent with the cell culture fluid usually requires agitation to achieve a uniform state, and in this device, the purpose of uniform agitation can be achieved by the action of the circulating pulsating micro-pump 6.

Further, when the culture environment is evaluated as a variable, step 2) comprises closing all the stop valves 11 and the sample adding valves 1, sleeving the same blood vessel support 12 on the matched support experiment tube 7, adding the same culture medium from the sample port 8 to flow into the cell culture tank 4, inoculating the same living cell suspension on the blood vessel support 12, opening the stop valves 11 after the cells adhere to the wall, keeping the stop valve between the two sample ports 8 closed, opening the sample adding valves 1, adding different additives, adjusting the circulating pulsation micropump 6, and then starting the evaluation experiment.

Further, when the vascular stent 12 is evaluated as a variable, step 2) is to close all the stop valves 11 and the sample adding valve 1, to sleeve different vascular stents 12 on the matched stent experiment tube 7, to add the same culture medium from the sample port 8 to flow into the cell culture tank 4, to inoculate the same living cell suspension on the vascular stent 12, to open the stop valves 11 after the cells adhere to the wall, to keep the stop valve between the two sample ports 8 open, to adjust the circulating pulsation micropump 6, to start the evaluation experiment.

The diameter size of the bracket experiment tube 7 has different specifications, and the blood vessel brackets 12 with different pipe diameters can be attached to the sleeve.

Further, sampling cock 13 can penetrate the sample syringe needle sample, and sampling cock 13 is inside to be equipped with pressure sensing probe and can exports the liquid pressure in the support experiment pipe 7 to the display and show.

Further, the disk-shaped channel 3 of the chip is sealed by an ion exchange membrane, and the ionic solution in the disk-shaped channel 3 and the solution in the cell culture pond 4 are isolated.

The ion exchange membrane has ion groups, can selectively permeate ions in a solution, and can controllably treat and adjust liquid in the device.

Further, the chip is a closed circulation system, and liquid in the chip can flow back.

Further, the temperature of the evaluation experiment was 37 ± 0.5 ℃, and was selected as 37 ℃.

And finally, collecting experimental data according to the degradation or deformation degree of the stent before and after the experiment, the adhesion and growth degree of cells on the surface of the stent and the change of components in the culture solution, and evaluating the experimental result.

The second purpose of the present invention is to provide an application of the method for the in vitro vascular stent evaluation test, specifically an application in the in vitro vascular stent evaluation.

The method for carrying out in-vitro intravascular stent evaluation experiment based on the microfluidic chip is the combination of a circulatory system and cell culture, and is suitable for evaluation experiment work and scientific research work of intravascular stents.

In certain embodiments, in vitro stenting experiments are designed following the idea of controlling unique variables.

When cells were evaluated as variables: in the study of blood vessels and problems associated with vascular stents, it is common practice to study cell types. Including but not limited to: human Umbilical Vein Endothelial Cells (HUVEC), Human Umbilical Artery Smooth Muscle Cells (HUASMC), rat thoracic aorta smooth muscle cells (A7r 5).

When the culture environment was evaluated as a variable: the culture medium is different from RPMI-1640 culture medium, DMEM culture medium (containing high sugar, medium sugar and low sugar formula); the chip can realize the separation of the compartments through the control of the valve, and on the premise that the planted cells are the same and the specifications of the added brackets are the same, additional cell action factors such as inflammatory factors, cell growth factors and the like are added into different compartments. In theory, substances that can act on cells can be used as tools for constructing different culture environments as long as they are soluble in the culture medium or can be uniformly dispersed in the culture medium.

When vascular stents were evaluated as variables: the difference of the stent can be embodied not only in the specification difference, but also in the material, the surface pattern and the surface coating. Even if the stent has the same specification, the coating composition carried on the stent surface or the way of carrying the coating is different, and the comparison test can be carried out as the only variable.

The invention has the beneficial effects that:

1) the method for carrying out the in vitro intravascular stent evaluation experiment based on the microfluidic chip overcomes the defects of feeding of experimental animals, construction cost of animal models and the like, long preparation period, complicated preparation procedures and the like, and reduces the time cost and the consumption of raw material cost required by the evaluation experiment;

2) the method for carrying out the in-vitro intravascular stent evaluation experiment based on the microfluidic chip can ensure the identity of the experiment under multiple conditions and ensure the comparability of the experiment result;

3) the ion exchange membrane adopted by the method for carrying out the in vitro intravascular stent evaluation experiment based on the microfluidic chip effectively shields the interference of other ions while ensuring the concentration of cations in a cell culture area to be unchanged, realizes the tightness in the experiment process, integrates the experiment reaction and the result detection, and is convenient and quick.

Drawings

Fig. 1 is a schematic front view of a rectangular chip.

Fig. 2 is a schematic front view of a circular chip.

FIG. 3 is a schematic partial cross-sectional view of a cell culture tank.

Fig. 4 is a schematic view of a front view structure of a sample injection valve in an open state.

Fig. 5 is a schematic top view of a sample injection valve in an open state.

Fig. 6 is a schematic view of a front view of a sample adding valve in a closed state.

Fig. 7 is a schematic top view of a sample injection valve in a closed state.

FIG. 8 is a partial schematic view of a T-port of a cell culture well.

Fig. 9 is an SEM scan of the surface of different coatings.

FIG. 10 shows the results of experiments on smooth muscle cell proliferation on the surface of each set of coated stents.

In the figure: 1. a sample adding valve; 2. a sample inlet pipe; 3. a disk-shaped channel; 4. a cell culture pond; 5. a circulation pipe; 6. a circulating pulsation micropump; 7. a bracket test tube; 8. a sample port; 9. a sample channel; 10. a one-way circulation pipe; 11. a stop valve; 12. a vascular stent; 13. a sampling plug; 14. an in-valve passage; 15. a valve knob; 16. a valve body; a T-type interface; 18. a disc-shaped double small tube; 19. pipeline partition wall.

Detailed Description

The examples are given for the purpose of better illustration of the invention, but the invention is not limited to the examples. Therefore, those skilled in the art should make insubstantial modifications and adaptations to the embodiments of the present invention in light of the above teachings and remain within the scope of the invention.

Example 1 method for performing in vitro intravascular stent evaluation experiment based on microfluidic chip under different cell effects

The micro-fluidic chip for the rectangular blood vessel stent experiment shown in the figure 1 is adopted, and the chip mainly comprises a sample adding valve 1, a circulating pulsation micro pump 6, a cell culture pool 4 and the like. The chip is circular or rectangular, and 2 side-by-side cylindrical sample adding valves 1 are processed at the center of the chip; each cylindrical sample adding valve 1 is provided with 2 opposite openings on the same section diameter line, and each opening is respectively connected with 1 sample adding pipe 2; each sample inlet pipe 2 is respectively communicated with one

cell culture pond

4, and 4 cell culture ponds 4 are formed in total and are communicated with a disc-shaped channel 3 formed at the bottom in each cell culture pond 4.

All shut-off valves 11 and sample addition valves 1 are closed. The same blood vessel stent 12 is sleeved on the matched stent experiment tube 7. The same medium is added from the sample port 8 and flows into the cell culture tank 4. Different living cell (such as human umbilical vein endothelial cells, human umbilical artery smooth muscle cells and rat thoracic aorta smooth muscle cells) suspensions are inoculated on the vascular stent 12, after the cells adhere to the wall, the stop valve 11 is opened, wherein the stop valve between the two sample ports 8 needs to be kept closed, and the circulating pulsation micropump 6 is adjusted, so that the related experiment can be started. In the experimental process, the sample adding valve 1 is in a closed state, a cell culture medium is added through the sample port 8 (or a nonionic reagent is added firstly and a cell culture solution is added) according to the state of cells in the cell culture pool 4, sampling analysis can be carried out through the sampling plug 13, experimental data is collected according to the degradation or deformation degree of the bracket before and after the experiment, the adhesion and growth degree of cells on the surface of the bracket and the change of components in the culture solution, and the experimental result is evaluated.

Example 2 method for performing in vitro intravascular stent evaluation experiment based on microfluidic chip under different culture environments

The round intravascular stent experimental microfluidic chip shown in fig. 2 is adopted, and the chip structure is consistent with the rectangular chip structure used in example 1. All shut-off valves 11 and sample addition valves 1 are closed. The same blood vessel stent 12 is sleeved on the matched stent experiment tube 7. The same medium is added from the sample port 8 and flows into the cell culture tank 4. The same suspension of living cells is seeded onto the vascular stent 12. After the cells adhere to the wall, the stop valve 11 is opened, wherein the stop valve between the two sample ports 8 needs to be kept closed, the sample adding valve 1 is opened, different experimental reagents (such as inflammatory factors and cell growth factors) are added, and the circulating pulsation micropump 6 is adjusted, so that the related experiment can be started. During the experiment, the sample adding valve 1 is in a closed state. Depending on the state of the cells in the cell culture chamber 4, cell culture medium is added through the sample port 8. Sampling analysis can be carried out through the sampling plug 13, experimental data is collected according to the degradation or deformation degree of the stent before and after the experiment, the adhesion and growth degree of cells on the surface of the stent and the change of components in the culture solution, and the experimental result is evaluated.

Example 3 method for performing in vitro different intravascular stent evaluation experiments based on microfluidic chip

All the stop valves 11 and the sample adding valves 1 are closed by adopting the microfluidic chip for the rectangular intravascular stent experiment described in example 1. Different blood vessel stents 12 are sleeved on the matched stent experiment tube 7. The same medium is added from the sample port 8 and flows into the cell culture tank 4. Inoculating the same living cell suspension on a blood vessel bracket 12, opening a stop valve 11 after the cells adhere to the wall, keeping the stop valve between two sample ports 8 open, and adjusting a circulating pulsation micropump 6 to start a related experiment. During the experiment, the sample adding valve 1 is in a closed state, and cell culture medium is added through the sample port 8 according to the state of the cells in the cell culture pool 4. Sampling analysis can be carried out through the sampling plug 13, experimental data is collected according to the degradation or deformation degree of the stent before and after the experiment, the adhesion and growth degree of cells on the surface of the stent and the change of components in the culture solution, and the experimental result is evaluated.

Example 4 platelet adhesion assay

All the stop valves 11 and the sample adding valves 1 are closed by adopting the microfluidic chip for the rectangular intravascular stent experiment described in example 1. The blood vessel stent 12 with different coating components (the coating components can be dopamine, PEI and drugs) processed on the surface is sleeved on the matched stent experiment tube 7. The same medium is added from the sample port 8 and flows into the cell culture tank 4. Platelet-rich plasma with the same concentration is added into the device, so that the plasma can flow through the surface of the stent at a constant speed. The experiment temperature is 37 ℃, the experiment time is 1h, the same living cell suspension is inoculated on the blood vessel support 12, after the cells adhere to the wall, the stop valve 11 is opened, and the circulating pulsation micropump 6 is adjusted, so that the related experiment can be started. Evaluation of the results of the subsequent experiments was performed using Scanning Electron Microscopy (SEM). Six fields of magnification 3000 were randomly selected for each concentration sample in this assay, as shown in FIG. 9, where A is a 2mg/ml dopamine +5mg/ml PEI group; b is a group of 2mg/ml dopamine +10mg/ml PEI; c is a group of 2mg/ml dopamine +15mg/ml PEI; d is a blank control group, namely a 316L stainless steel bracket group without a coating, 1 represents an unloaded medicine group, and 2 is a loaded medicine group.

As is clear from the analysis of the platelet morphology and the number of platelets in the visual field frame, in the method for producing the hybrid membrane, the number of platelets in the visual field frame was smaller in the drug-loaded composition than in the drug-unloaded composition, and more importantly, the morphology of the adhered platelets was not changed much and no false feet were protruding. Among the three drug-carrying groups, the B-2 group, namely the drug-carrying combination of 2mg/ml dopamine and 10mg/ml PEI, has the least adhesion quantity, most adhered platelets are in a round normal state, the activation degree is low, and the effect in a platelet adhesion test is relatively good.

Example 5 vascular smooth muscle cell proliferation assay

1. 4 groups of brackets (316LSS groups, 5mg/ml PEI + DA + GO + DTX coating groups, 10mg/ml PEI + DA + GO + DTX coating groups and 15mg/ml PEI + DA + GO + DTX coating groups) of different experimental types are sterilized by ultraviolet front and back irradiation in a clean bench for 12 hours;

2. placing the sterilized stent in a pore plate, and adding a smooth muscle cell suspension to allow cells to adhere to the surface of the stent;

3. sterilizing the chip, placing the bracket in the chip, adding a cell culture medium, and culturing in a 5% CO2 incubator at 37 ℃;

4. taking out the chip at the specified time (1d, 3d and 5d), taking out the bracket, putting the bracket into a new pore plate, adding 500 mu L of culture medium and 50 mu L of LMTS detection solution into each pore, and putting the pore plate back to the cell culture box at 37 ℃ for wet incubation for 2 h;

5. the well plate is taken out, the reaction solution is shaken up and 100 microlitres of the reaction solution is absorbed and added into a 96 well plate, the well plate is placed at the 490nm wavelength of an enzyme labeling instrument, and the OD value of absorbance is measured.

As shown in fig. 10, there was a significant difference (p < 0.05) between the OD values of the three drug-loaded scaffolds and the 316LSS bare chip scaffold at all 1, 3, 5d, with a very significant difference (p < 0.01) between the three drug-loaded scaffolds and the 316LSS bare chip scaffold at 5 d. The OD value of the drug-loaded stent is far lower than that of a non-drug-loaded stent and a bare-die stent, which shows that DTX (drug) carrying of the group of coatings is successful and the coating has a remarkable inhibiting effect on proliferation of smooth muscle cells.

Finally, the above embodiments are only for illustrating the technical solutions of the present invention and not for limiting, although the present invention has been described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that modifications or equivalent substitutions may be made to the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, and all of them should be covered in the claims of the present invention.

Claims

1. A method for carrying out in-vitro intravascular stent evaluation experiments based on a microfluidic chip comprises the following steps:

1) the method comprises the following steps of arranging a micro-fluidic chip in-vitro intravascular stent evaluation system comprising a sample adding valve (1), a circulating pulsation micro-pump (6) and a cell culture pool (4);

2) closing all the stop valves (11) and the sample adding valve (1), sleeving a blood vessel support (12) on a matched support experiment tube (7), adding a culture medium from a sample port (8) to flow into a cell culture pond (4), inoculating a living cell suspension on the blood vessel support (12), and opening the stop valves (11) after the cells adhere to the wall;

3) sampling is carried out through a sampling plug head (13), experimental data are collected, and experimental results are evaluated;

the chip mainly comprises a sample adding valve (1), a circulating pulsation micropump (6) and a cell culture pool (4); the chip is circular or rectangular, and 2 side-by-side cylindrical sample adding valves (1) are processed at the center of the chip; each cylindrical sample adding valve (1) is provided with 2 opposite openings on the same section diameter line, and each opening is respectively connected with 1 sample adding pipe (2); each sample inlet pipe (2) is respectively communicated with one respective cell culture pond (4), and 4 cell culture ponds (4) are communicated with a disc-shaped channel (3) processed at the bottom in each cell culture pond (4);

the 4 cell culture ponds (4) are uniformly distributed at four corners of the rectangular chip or at four symmetrical edges of the circular chip; 1 circulating pulsation micropump (6) is arranged between the pair of cell culture ponds (4), the circulating pulsation micropump (6) is communicated in series between 2 circulating pipes (5), and the 2 circulating pipes (5) are respectively communicated with the disc-shaped channels (3) in the cell culture ponds (4) at two sides of the circulating pulsation micropump (6); the circulating pipe (5) and the sample injection pipe (2) are communicated with the disc-shaped channel (3) at a T-shaped interface (17) at the side of the cell culture pool (4); a pipeline partition wall (19) is arranged in the middle of the disc-shaped channel (3), and the disc-shaped channel (3) is divided into 2 disc-shaped double small pipes (18) from the T-shaped interface (17); the disc-shaped channel (3) is coiled to the center of the cell culture pool (4) along the bottom surface of the cell culture pool (4) and is communicated with a bracket experiment tube (7) which is processed and fixed at the center; the bracket experiment tube (7) is a hollow tube which is made of elastic materials and is higher than the cell culture pool (4), and a sampling plug (13) is arranged at the opening of the top end of the bracket experiment tube; a pressure sensing probe is arranged in the sampling plug head (13);

2 sample ports (8) are processed between 2 side-by-side circular sample adding valves (1) in the middle of the chip, sample channels (9) are processed between the 2 sample ports (8) and are communicated with each other, and stop valves (11) are processed on the communicated sample channels (9); 2 sample channels (9) are respectively processed on the 2 sample ports (8) and are respectively communicated with the 2 cell culture pools (4);

the sampling tube (2) is respectively provided with 1 one-way circulating tube (10) beside 2 sample adding valves (1), and the sampling tube (2) is communicated with two ends of the sample adding valves (1); each one-way circulation pipe (10) is provided with a stop valve (11).

2. The method according to claim 1, characterized in that when the cells are evaluated as variables, the evaluation experiment can be started by closing all the stop valves (11) and the sample adding valve (1) in step 2), sleeving the same blood vessel bracket (12) on the matched bracket experiment tube (7), adding the same culture medium from the sample port (8) into the cell culture tank (4), inoculating different living cell suspensions on the blood vessel bracket (12), opening the stop valves (11) after the cells adhere to the wall, keeping the stop valve between the two sample ports (8) closed, and adjusting the circulating pulsation micropump (6).

3. The method according to claim 1, wherein when the culture environment is evaluated as a variable, the evaluation experiment can be started by closing all the stop valves (11) and the sample adding valves (1) in step 2), sleeving the same blood vessel stent (12) on the matched stent experiment tube (7), adding the same culture medium from the sample port (8) into the cell culture pond (4), inoculating the same living cell suspension on the blood vessel stent (12), opening the stop valves (11) after the cells adhere to the wall, keeping the stop valve between the two sample ports (8) closed, opening the sample adding valves (1), adding different additives, and adjusting the circulating pulsation micropump (6).

4. The method according to claim 1, characterized in that, when the blood vessel support (12) is evaluated as a variable, the evaluating experiment can be started by closing all the stop valves (11) and the sample adding valves (1) in step 2), sleeving different blood vessel supports (12) on the matched support experiment tube (7), adding the same culture medium from the sample port (8) into the cell culture tank (4), inoculating the same living cell suspension on the blood vessel support (12), opening the stop valves (11) after the cells are attached to the wall, keeping the stop valve between the two sample ports (8) open, and adjusting the circulating pulsation micropump (6).

5. The method according to claim 1, characterized in that the sampling plug (13) can penetrate a sampling needle to sample, and a pressure sensing probe is arranged inside the sampling plug (13) and can output the liquid pressure in the stent test tube (7) to a display to display.

6. The method of claim 2, wherein the culture medium is a mixed reagent of a non-ionic reagent and a cell culture fluid.

7. The method according to claim 1, wherein the disk-shaped channel (3) of the chip is closed by an ion exchange membrane, and the ionic solution in the disk-shaped channel (3) is separated from the solution in the cell culture pond (4).

8. The method of claim 1, wherein the chip is a closed loop system, and the liquid in the chip is recirculated.

9. The method according to any one of claims 1 to 8, wherein the temperature of the evaluation experiment is 37 ± 0.5 ℃.