A method for evaluating the cell migration ability of a stent microbridge - microfluidic chip and a bioactive material

Abstract

The present invention provides a stent microbridge - microfluidic chip and a method for evaluating the cell migration ability of bioactive materials. The chip includes: a substrate, an intermediate interlayer on the substrate, and a cover glass covering the intermediate interlayer; wherein, a culture flow channel and migration flow channels located on both sides of the culture flow channel are provided on one side of the substrate close to the intermediate interlayer; a plurality of through - microbridge regions are reserved between the culture flow channel and each migration flow channel; a plurality of microbridge regions are embedded with stent microbridges made of bioactive materials for reconstructing the microfluidic chip structure. The plurality of microbridge regions can be embedded with different or the same types of stent microbridges for studying or evaluating the cell migration ability caused by the stent microbridges. The bioactive materials are used as a part of the microfluidic chip structure for reconstructing the structure of the microfluidic chip. This chip can be used to study the cell migration ability of various bioactive materials, providing a reliable basis for the research and evaluation of bioactive materials.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedical experimental instruments, and particularly relates to a stent microbridge-microfluidic chip and a method for evaluating the cell migration ability of bioactive materials. Background Art

[0002] Bioactive ceramics, as medical devices, can meet specific clinical needs. For example, research has produced ceramic materials with excellent osteoinduction, osteoconductivity, and biocompatibility. The extended tissue induction in regenerative medicine has expanded the boundaries of the repair concept in clinical practice. The introduction of technologies represented by three-dimensional (3D) printing technology has created conditions for solving problems such as complex / composite materials, multi-tissue organ coordination, and personalized repair of specific parts. Bioactive ceramics have become an essential category in medical devices.

[0003] Cell migration plays a crucial role in the realization of the bone tissue repair function of bioactive materials. According to ISO19090:2018 and the Chinese national standard YY / T 1744-2020, there are three standard methods for testing the cell migration ability of bioactive materials: one is to drop a cell suspension onto the material surface and test the penetration ability of the cell suspension under the action of gravity. Another is to add the material to the cell suspension and drive the cells to adsorb on the material by shaking. The third is to cover the cell layer with the material and let the cells migrate onto the material and proliferate. These three methods lack the dynamic circulation of body fluids and cannot simulate cell migration under the in vivo microenvironment. Body fluids are sufficient to carry cells through the small gap between the implant material and the host bone tissue.

[0004] Therefore, the current methods for evaluating cell migration ability have great limitations. It is crucial to establish an in vitro cell migration model that can simulate the in vivo microenvironment and accurately control the dynamic circulation of liquids. Summary of the Invention

[0005] Aiming at the above problems existing in the prior art, the present invention provides a stent microbridge-microfluidic chip and a method for evaluating the cell migration ability of bioactive materials. This chip can provide a simulated in vivo microenvironment for bioactive materials, so as to accurately study and evaluate bioactive materials in vitro. The specific content of the invention is as follows:

[0006] In the first aspect, the present invention provides a stent microbridge-microfluidic chip, including: a substrate 1, an intermediate interlayer 2 located above the substrate, and a cover glass 3 covering the intermediate interlayer;

[0007] Wherein, a culture flow channel 101 and migration flow channels 102 located on both sides of the culture flow channel 101 are provided on one side of the substrate 1 close to the intermediate interlayer 2;

[0008] There are multiple through micro-bridge regions 103 reserved between the culture flow channel 101 and each of the migration flow channels 102;

[0009] Multiple of the micro-bridge regions 103 are embedded with scaffold micro-bridges for reconstructing the microfluidic chip structure, including:

[0010] Multiple of the micro-bridge regions 103 are embedded with different types of the scaffold micro-bridges for studying or evaluating the cell behaviors caused by different types of the scaffold micro-bridges; or

[0011] Multiple of the micro-bridge regions 103 are used to place the same type of the scaffold micro-bridges for studying or evaluating the cell behaviors caused by the same type of the scaffold micro-bridges;

[0012] The scaffold micro-bridges are made of bioactive materials;

[0013] The regions of the scaffold micro-bridges in contact with the micro-bridge regions 103 are sealed.

[0014] Optionally, the culture flow channel 101 and the migration flow channels 102 located on both sides of the culture flow channel 101 are arranged in an array;

[0015] Multiple of the micro-bridge regions 103 include a first micro-bridge 1031 and a second micro-bridge 1032;

[0016] The first micro-bridge 1031 and the second micro-bridge 1032 are symmetrically arranged.

[0017] Optionally, liquid inlets 106 and liquid outlets 107 are respectively arranged at both ends of the culture flow channel 101 and each of the migration flow channels 102;

[0018] First through holes 1061 corresponding to the liquid inlets 106 of each flow channel and second through holes 1071 corresponding to the liquid outlets 107 of each flow channel are respectively formed in the cover plate 3 and the middle interlayer 2, so that each flow channel is communicated with an external perfusion system.

[0019] Optionally, the middle interlayer 2 is made of a flexible organic polymer material, and the base 1 and the cover sheet 3 are made of a rigid organic polymer material; the base 1, the middle interlayer 2 and the cover sheet 3 are connected by fastening members to facilitate replacement of the scaffold micro-bridges.

[0020] Optionally, the scaffold micro-bridges are composed of a bioactive material and a hydrogel; the bioactive material includes any one of bioactive ceramic materials, metal materials, polymer materials and composite materials.

[0021] Optionally, the pore diameter of the scaffold micro-bridges is 500 μm, and the porosity of the scaffold micro-bridges is 50%.

[0022] Optionally, the scaffold microbridge is prepared by a foaming method, 3D printing or femtosecond laser etching process.

[0023] Optionally, the size of the scaffold microbridge is adapted to the size of the microbridge region 103 to achieve sealing and plugging of the microbridge region 103.

[0024] In a second aspect, the present invention provides a method for evaluating the cell migration ability of a bioactive material. The method is applicable to the scaffold microbridge-microfluidic chip described in the first aspect above and includes:

[0025] Embed scaffold microbridges into a plurality of microbridge regions 103 to complete the reconstruction of the microfluidic chip structure;

[0026] Inoculate cells in the culture channel 101 and inject growth factors into the migration channel 102;

[0027] Connect the microfluidic chip to a perfusion system, and the perfusion system continuously or intermittently supplies culture medium to the culture channel and the migration channel to reproduce the local microenvironment in vivo;

[0028] Analyze the number of live cells that enter the migration channel through the scaffold microbridge and adhere to the inner wall;

[0029] Based on the number of the live cells, determine the cell migration ability value of the bioactive material.

[0030] Optionally, when different types of scaffold microbridges are embedded in the plurality of microbridge regions 103, the same type of cells is inoculated in the culture channel 101;

[0031] When the same type of scaffold microbridges are embedded in the plurality of microbridge regions 103, different types of cells are inoculated in the culture channel 101.

[0032] The present invention provides a scaffold microbridge - microfluidic chip, comprising: a substrate 1, an intermediate interlayer 2 located above the substrate, and a cover plate 3 covering the intermediate interlayer; wherein, on one side of the substrate 1 close to the intermediate interlayer 2, a culture flow channel 101 and migration flow channels 102 located on both sides of the culture flow channel 101 are provided; a plurality of through - microbridge regions 103 are reserved between the culture flow channel 101 and each of the migration flow channels 102; a plurality of the microbridge regions 103 are embedded with scaffold microbridges for reconstructing the microfluidic chip structure, including: a plurality of the microbridge regions 103 are embedded with different types of the scaffold microbridges for studying or evaluating the cell behaviors (cell migration ability) caused by different types of the scaffold microbridges; or, a plurality of the microbridge regions 103 are used to place the same type of the scaffold microbridges for studying or evaluating the cell behaviors (cell migration ability evaluation) caused by the same type of the scaffold microbridges; the scaffold microbridges are made of bioactive materials. By reconstructing the structure of the microfluidic chip, the scaffold microbridges made of bioactive materials (such as calcium phosphate ceramics as bioactive materials) are directly used as a part of the chip structure, and an in - vitro chip system for simulating the human microenvironment can be obtained after connecting an external perfusion device. This chip system can be used to study the cell migration ability of various bioactive materials, providing a reliable basis for the research and evaluation of bioactive materials. Brief Description of the Drawings

[0033] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the following will briefly introduce the drawings required for use in the description of the embodiments or the prior art. Obviously, the following drawings are only some embodiments of the present application. For those of ordinary skill in the art, other drawings can be obtained based on these drawings without creative efforts.

[0034] Figure 1 Shows the schematic diagram of the hierarchical structure of the microfluidic chip provided by the embodiment of the present invention;

[0035] Figure 2 Shows the schematic diagram of the structure of the substrate in the microfluidic chip provided by the embodiment of the present invention;

[0036] Figure 3 Shows the surface topography diagram of the scaffold microbridge provided by the embodiment of the present invention;

[0037] Figure 4 Shows the flowchart of the method for evaluating the cell migration ability of the bioactive material provided by the embodiment of the present invention. Detailed Description of the Embodiments

[0038] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. The following description of at least one exemplary embodiment is actually only illustrative and in no way limits the present invention and its application or use. Based on the embodiments of the present invention, any product identical or similar to the present invention obtained by anyone under the inspiration of the present invention or by combining the features of the present invention with those of other existing technologies falls within the protection scope of the present invention. In addition, all other embodiments obtained by those of ordinary skill in the art without creative work also belong to the protection scope of the present invention.

[0039] For the specific experimental steps or conditions not specified in the embodiments, the operations or conditions of the conventional experimental steps described in the existing technologies in this field can be followed. The reagents and other instruments not indicating the manufacturer can be obtained as conventional reagent products through commercial purchase. In addition, the accompanying drawings are only schematic diagrams of the embodiments of the present invention and are not necessarily drawn to scale. The same reference numerals in the drawings represent the same or similar parts, and thus the repeated description of them will be omitted. Some of the block diagrams shown in the drawings are functional entities and do not necessarily correspond to physically or logically independent entities.

[0040] For the technologies, methods, and equipment known to those of ordinary skill in the relevant fields, detailed discussions may not be made, but under appropriate circumstances, the said technologies, methods, and equipment should be regarded as part of the authorization specification.

[0041] In the description of the present invention, it should be understood that the use of words such as "first" and "second" to limit the components is only for the convenience of distinguishing the corresponding components. Without further statement, the above words have no special meaning, and thus cannot be understood as a limitation on the protection scope of the present invention.

[0042] In addition, the technical features involved in different embodiments of the present invention described below can be combined with each other as long as they do not conflict with each other.

[0043] Before the detailed description of a microfluidic chip and a method for evaluating the cell migration ability of a bioactive material provided by the present invention, it is necessary to make the following description of the related technologies:

[0044] Since bioactive materials are not completely inert biologically or chemically, complex changes will occur in their interactions with human cells and tissues after implantation. Therefore, the research on such materials requires a model close to the human microenvironment. However, the existing standard methods for testing the cell migration ability of bioactive materials all have the problem of lacking the dynamic circulation of body fluids and cannot simulate cell migration in the in vivo microenvironment.

[0045] In the related art, as an alternative method to traditional cell culture and animal experiments, microfluidic chips have been used to construct in vitro organ models, including the liver, kidney, lung, intestine, heart, vascularized bone tissue, and cancer bone metastasis. That is, the current design of microfluidic chips mainly focuses on the study of specific tissue functions and is mainly used to study drug toxicity, physiology, and pathology. However, no microfluidic chip has been developed that can be used to study the interaction between tangible materials and cells and tissues.

[0046] Given that microfluidic chip technology can precisely control dynamic fluid circulation and maintain the characteristics of miniaturization, integration, and high throughput. Microfluidic chip technology has the potential to provide a reliable in vitro model for studying the interaction between tangible materials and cells and tissues. Based on these considerations, we redesigned and fabricated a microfluidic chip to evaluate the effects of different bioactive materials on cell migration. The specific implementation is as follows:

[0047] In a first aspect, the present invention provides a microfluidic chip, Figure 1 shows a schematic diagram of the layered structure of the microfluidic chip provided by an embodiment of the present invention, as Figure 1 shown, including: a substrate 1, an intermediate interlayer 2 located above the substrate, and a cover sheet 3 covering the intermediate interlayer 2; Figure 2 shows a schematic diagram of the structure of one side of the substrate in an embodiment of the present invention, as Figure 2 shown, a culture flow channel 101 is provided on one side of the substrate 1 close to the intermediate interlayer 2 and migration flow channels 102 are located on both sides of the culture flow channel 101; a plurality of through micro-bridge regions 103 are reserved between the culture flow channel and each of the migration flow channels, and a plurality of the micro-bridge regions 103 are embedded with scaffold micro-bridges for reconstructing the microfluidic chip structure, including: a plurality of the micro-bridge regions 103 are embedded with different types of the scaffold micro-bridges for studying or evaluating the cell migration ability caused by different types of the scaffold micro-bridges; or

[0048] a plurality of the micro-bridge regions 103 are used to place the same type of the scaffold micro-bridges for studying or evaluating the cell migration ability caused by the same type of the scaffold micro-bridges;

[0049] The scaffold micro-bridge is made of a bioactive material; the area where the scaffold micro-bridge contacts the micro-bridge region 103 is sealed.

[0050] See Figure 1 and Figure 2, In specific implementation, in order to combine the bioactive material to be evaluated with the microfluidic chip, the present invention reconstructs the structure of the microfluidic chip, and reserves an embedding position for the bioactive material on the chip substrate 1. Specifically, the microfluidic chip provided by the present invention is a detachable structure composed of a substrate 1, an intermediate sandwich layer 2 and a cover plate 3. The existence of the intermediate sandwich layer 2 well ensures the overall sealing performance of the microfluidic chip, preventing the liquid in the flow channel from leaking and affecting the results of the cell migration experiment. The culture channel 101 is the channel for inoculating the original cells in the cell migration experiment. Two identical migration channels 102 are respectively arranged on both sides of it, and a microbridge area 103 communicating the culture channel 101 and the migration channel 102 is arranged between the culture channel 101 and each adjacent migration channel 102. The microbridge area 103 is used to carry the scaffold microbridge made of the bioactive material. The scaffold microbridge is a movable element and can be conveniently placed into or taken out of the microbridge area 103, so as to analyze or evaluate the migration ability of different cells by the same bioactive material, or analyze or evaluate the migration ability of the same cells by different bioactive materials. Moreover, the periphery of the scaffold microbridge is in full contact with the microbridge area 103 to ensure that there is no gap between the scaffold microbridge and the microbridge area 103. The migration of cells in the culture channel 101 to the migration channel 102 can only occur through the scaffold microbridge, ensuring the accuracy of the evaluation results of the cell migration ability of the bioactive material.

[0051] In some embodiments, the scaffold microbridge can be prepared by the foaming method, 3D printing or femtosecond laser etching process to control the shape and size of the bioactive material, so as to realize the precise assembly of the microbridge area 103 and the microbridge scaffold. The pore diameter of the scaffold microbridge is controlled to be 200 - 900 μm, and the porosity of the scaffold microbridge is controlled to be 30 - 80%. In this way, the complete structure of the microfluidic chip is reconstructed together with the scaffold microbridge and the above-mentioned substrate 1, intermediate sandwich layer 2 and cover plate 3. This scaffold microbridge - microfluidic chip can provide a simulated in vivo microenvironment for evaluating the cell migration ability of the scaffold microbridge prepared from the bioactive material, and provide accurate and reliable evaluation results for the practical application of the bioactive material.

[0052] In specific implementation, the scaffold microbridge is preferably prepared by 3D printing method, or can also be prepared by the foaming method. When prepared by the foaming method, cutting and polishing means need to be used to make the material into a shape structure adapted to the size of the microbridge. Figure 3 The surface topography of the scaffold microbridge provided by the embodiment of the present invention is shown, as Figure 3As shown, through-holes are distributed on the surface of the scaffold micro-bridge. Its constituent materials include Matrigel matrix gel and specific bioactive materials to be evaluated. Specifically, the bioactive materials to be evaluated can be any one of bioactive ceramic materials, metal materials, and ceramic-metal composite materials. Such as biphasic calcium phosphate ceramics (BCP), hydroxyapatite (HA), β-tricalcium phosphate (TCP), etc. The Matrigel matrix gel has thermosensitive properties and can well encapsulate bioactive materials. In some embodiments, the bioactive materials are uniformly dispersed in the Matrigel matrix gel, and the pore size, shape, and size are precisely controlled by 3D printing technology to prepare a scaffold micro-bridge that can be precisely adapted to the size of the micro-bridge 103, realizing the sealing of the micro-bridge 103. Specifically, the pore size of the scaffold micro-bridge is controlled at 500 μm, and the porosity is controlled at 50%.

[0053] Continue to refer to Figure 1 , the culture channels 101 and the migration channels 102 located on both sides of the culture channels 101 are arranged in an array; there are through first micro-bridges 1031 and second micro-bridges 1032 between the culture channels and each migration channel; the first micro-bridges 1031 and the second micro-bridges 1032 are symmetrically arranged.

[0054] In specific implementation, the microfluidic chip provided by the present invention can verify the migration ability of the same bioactive material to different cells, or the difference in the migration ability of different bioactive materials to the same cell. When verifying the difference in the migration ability of different bioactive materials to the same cell, to ensure that the in vivo microenvironment simulated by the microfluidic chip has the same influence on different bioactive materials (scaffold micro-bridges) to be evaluated, the present invention symmetrically arranges two identical migration channels 102 on both sides of the culture channel 101, so that the culture channel 101 and the migration channels 102 located on both sides of the culture channel 101 are arranged in an array. At the same time, the through micro-bridge region 103 is arranged between the culture channel 101 and each migration channel 102. That is, there are a first micro-bridge 1031 and a second micro-bridge 1032 that communicate with the migration channel 102 on both sides of the culture channel 101.

[0055] During specific settings, considering that the position of the micro-bridge determines the position of the scaffold micro-bridge, and to ensure that different bioactive materials are evaluated for the cell migration ability of the same cells in the same simulated microenvironment, the first micro-bridge 1031 and the second micro-bridge 1032 are symmetrically arranged. That is, the first micro-bridge 1031 and the second micro-bridge 1032 are paired and symmetrically arranged on both sides of the culture channel 101. The specific number of micro-bridge regions 103 is determined according to the actual situation, but at least one set (one first micro-bridge 1031 and one second micro-bridge 1032 arranged symmetrically) is set. In this way, the migration path and the force condition of the cells in the middle culture channel 101 migrating to the migration channel 102 are the same. The only difference is that the materials of the scaffold micro-bridges arranged in the first micro-bridge 1031 and the second micro-bridge 1032 are different, that is, the bioactive materials are different. To verify the difference in the migration ability of the same type of cells for two different materials.

[0056] During specific implementation, the microfluidic chip is provided with an interface connected to an external circulating perfusion device. Figure 1 and Figure 2 It can be seen that the microfluidic chip provided by the present invention is provided with a liquid inlet 106 and a liquid outlet 107 at both ends of each culture channel 101 and migration channel 102. The cover plate 3 and the intermediate sandwich layer 2 are also respectively provided with a first through hole 1061 corresponding to the liquid inlet 106 of each channel, and a second through hole 1071 corresponding to the liquid outlet 107 of each channel, so that each channel is connected to an external perfusion system.

[0057] In some embodiments, the substrate 1 and the cover glass 3 of the microfluidic chip provided by the present invention are made of a rigid organic polymer material, and the intermediate sandwich layer 2 is made of a flexible organic polymer material, and the substrate 1, the intermediate sandwich layer 2 and the cover glass 3 are fixed together by fastening members. Among them, the flexible intermediate sandwich layer 2 located between the substrate 1 and the cover glass 3 can well ensure the overall sealing of the microfluidic chip. Specifically, the intermediate sandwich layer 2 can be a polydimethylsiloxane flexible material; the constituent materials of the substrate 1 and the cover glass 3 can be a polymethyl methacrylate rigid material.

[0058] In a second aspect, the present invention also provides a method for evaluating the cell migration ability of a bioactive material, which is applied to the scaffold micro-bridge - microfluidic chip described in the first aspect above. Figure 4 The flowchart of the method for evaluating the cell migration ability of the bioactive material provided by the embodiment of the present invention is shown. As Figure 4 shown, the method includes:

[0059] S1. Embedding scaffold micro-bridges into multiple micro-bridge regions 103 to complete the reconstruction of the microfluidic chip structure;

[0060] S2. Inoculating cells in the culture channel 101 and injecting growth factors into the migration channel 102;

[0061] S3. Connect the microfluidic chip to the perfusion system, and the perfusion system supplies the culture medium to the culture channel and the migration channel continuously or intermittently to reproduce the local microenvironment in vivo;

[0062] S4. Analyze the number of living cells that enter the migration channel through the scaffold microbridge and adhere to the inner wall;

[0063] S5. Based on the number of the living cells, determine the cell migration ability value of the bioactive material.

[0064] During specific implementation, when different types of scaffold microbridges are embedded in the multiple microbridge regions 103, the same type of cells are inoculated in the culture channel 101; when the same type of scaffold microbridges are embedded in the multiple microbridge regions 103, different types of cells are inoculated in the culture channel 101. The present invention takes the microfluidic chip including the Figure 2 shown substrate structure as an example, and combines the Figure 1 to illustrate how to evaluate the difference in the migration ability of the same bioactive material to different cells. As Figure 1 、 2 shown, the substrate structure includes two groups of symmetrically arranged microbridge regions 103. The microbridge region 103 specifically includes two first microbridges 1031 on one side of the culture channel 101 and two second microbridges 1032 on the other side of the culture channel 101. During the experiment, the scaffold microbridge made of the bioactive material to be evaluated, such as PBCP, is placed at any position of the first microbridge 1031, the negative control PMMA scaffold microbridge is placed at the position of the other microbridge 1031, and the position of the second microbridge 1032; the MSCs and HUVEC cells are inoculated in the culture channel 101 at a concentration of 1×10 5 / mL. After the cells adhere, connect the external circulation perfusion device to the microfluidic chip through the liquid inlet 106 and the liquid outlet 107, and continuously or intermittently perfuse the normal culture medium into the culture channel 101 and the migration channel 102 at a preset rate of 6 mm / s; after culturing for 1, 3, 5, and 7 days respectively, observe the migration of the cells to the migration channel 102 under an inverted fluorescence microscope after FDA / PI staining. The number of cells in each migration channel 102 was semi-quantitatively analyzed using ImageJ (NIH, USA) to determine the number of migrated cells, so as to determine the evaluation of the difference in the migration ability of the PBCP bioactive material to MSCs and HUVEC cells according to the number of migrated cells in each migration channel 102.

[0065] The present invention continues to take the microfluidic chip including the Figure 2 shown substrate structure as an example, and combines the Figure 1 to illustrate how to evaluate the migration ability of different bioactive materials to the same cells.

[0066] Continue to refer to Figure 1 . Place the porous microbridge scaffold made of the bioactive material to be evaluated, such as PBCP, into the first microbridge 1031, and place the porous microbridge scaffold made of another bioactive material, such as PTCP material, into the second microbridge 1032; Inoculate MSCs or HUVEC cells into the culture channel 101 at a concentration of 1×10 5 / mL. After the cells adhere, connect an external circulating perfusion device to the microfluidic chip through the inlet 106 and the outlet 107, and continuously or intermittently perfuse the normal culture medium into the culture channel 101 and the migration channel 102 at a preset rate of 6 mm / s; After culturing for 1, 3, 5, and 7 days respectively, observe the migration of cells to the migration channel 102 by FDA / PI staining under an inverted fluorescence microscope. Use ImageJ (NIH, USA) to semi-quantitatively analyze the number of cells in each migration channel 102 to determine the number of migrating cells, so as to determine the cell migration ability values and the differences in cell migration ability of the PBCP bioactive material and the PTCP bioactive material according to the number of migrating cells in each migration channel 102.

[0067] The method for evaluating the cell migration ability of the bioactive material provided by the embodiment of the present invention directly uses the scaffold microbridge made of the bioactive material (the bioactive material such as calcium phosphate ceramic) as a part of the chip structure to form a scaffold microbridge - microfluidic chip. After connecting the external perfusion system, the perfusion system continuously or intermittently supplies the culture medium solution to the culture channel 101 and the migration channel 102 of the scaffold microbridge - microfluidic chip to reproduce the local microenvironment in vivo. An in vitro chip system for simulating the human microenvironment is obtained. It is used to study or evaluate the cell behavior (cell migration ability evaluation) caused by different types of scaffold microbridges; or, to study or evaluate the cell behavior (cell migration ability evaluation) caused by the same type of the scaffold microbridges; to realize comparing the differences in the migration ability of different bioactive materials on the same cells, as well as the differences in the migration ability of the same bioactive material on different cells, and to provide a reliable basis for the research and evaluation of bioactive materials.

[0068] In the description of this specification, the description referring to terms such as "one embodiment", "some embodiments", "example", "specific example", or "some examples" means that the specific features, structures, materials, or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials, or characteristics described can be combined in a suitable manner in any one or more embodiments or examples. In addition, those skilled in the art can combine and combine the different embodiments or examples described in this specification.

[0069] For the method embodiments, for the sake of simple description, they are all expressed as a series of combinations of actions. However, those skilled in the art should be aware that the present invention is not limited by the described order of actions, because according to the present invention, certain steps can be performed in other orders or simultaneously. Secondly, those skilled in the art should also be aware that the embodiments described in the specification are all preferred embodiments, and the actions and components involved are not necessarily essential to the present invention.

[0070] The above has introduced in detail a method for evaluating the cell migration ability of a stent microbridge - microfluidic chip and a bioactive material provided by the present invention. Specific examples are used herein to elaborate on the principle and implementation manner of the present invention. The description of the above embodiments is only used to help understand the method and its core idea of the present invention; at the same time, for those of ordinary skill in the art, according to the idea of the present invention, there will be changes in the specific implementation manner and application scope. In summary, the content of this specification should not be construed as a limitation to the present invention.

Claims

1. A stent microbridge - microfluidic chip, characterized in that, Comprising: a substrate (1), an intermediate interlayer (2) located above the substrate, and a cover sheet (3) covering the intermediate interlayer; wherein, on one side of the substrate (1) close to the intermediate interlayer (2), a culture flow channel (101) and migration flow channels (102) located on both sides of the culture flow channel (101) are provided; a plurality of through micro-bridge regions (103) are reserved between the culture flow channel (101) and each of the migration flow channels (102); a plurality of the micro-bridge regions (103) are embedded with scaffold micro-bridges for reconstructing a microfluidic chip structure, including: a plurality of the micro-bridge regions (103) are embedded with different types of the scaffold micro-bridges for studying or evaluating cell behaviors caused by different types of the scaffold micro-bridges; or a plurality of the micro-bridge regions (103) are used to place the same type of the scaffold micro-bridges for studying or evaluating cell behaviors caused by the same type of the scaffold micro-bridges; the scaffold micro-bridges are made of a bioactive material; the regions of the scaffold micro-bridges in contact with the micro-bridge regions (103) are sealed; the substrate (1), the intermediate interlayer (2) and the cover sheet (3) are connected by fastening members to facilitate replacement of the scaffold micro-bridges; the bioactive material includes any one of bioactive ceramic materials, metal materials, polymer materials and composite materials.

2. The stent microbridge - microfluidic chip according to claim 1, wherein the culture flow channel (101) and the migration flow channels (102) located on both sides of the culture flow channel (101) are arranged in an array; a plurality of the micro-bridge regions (103) include a first micro-bridge (1031) and a second micro-bridge (1032); the first micro-bridge (1031) and the second micro-bridge (1032) are symmetrically arranged.

3. The stent microbridge - microfluidic chip according to claim 1, characterized in that, liquid inlets (106) and liquid outlets (107) are respectively arranged at both ends of the culture flow channel (101) and each of the migration flow channels (102); first through holes (1061) corresponding to the liquid inlets (106) of each flow channel and second through holes (1071) corresponding to the liquid outlets (107) of each flow channel are respectively formed in the cover sheet (3) and the intermediate interlayer (2) so that each flow channel is communicated with an external perfusion system.

4. The stent microbridge-microfluidic chip according to claim 1, wherein the intermediate interlayer (2) is made of a flexible organic polymer material, and the substrate (1) and the cover sheet (3) are made of a rigid organic polymer material.

5. The stent microbridge-microfluidic chip according to claim 1, characterized in that, the scaffold micro-bridges are composed of a bioactive material and a hydrogel.

6. The stent microbridge - microfluidic chip according to claim 1, characterized in that, the pore diameter of the scaffold micro-bridges is 200 - 900 μm, and the porosity of the scaffold micro-bridges is 30 - 80%.

7. The stent microbridge - microfluidic chip according to claim 1, characterized in that, the scaffold micro-bridges are prepared by a foaming method, 3D printing or a femtosecond laser etching process.

8. The stent microbridge-microfluidic chip according to claim 1, wherein, the size of the scaffold micro-bridges is adapted to the size of the micro-bridge regions (103) to achieve sealing and plugging of the micro-bridge regions (103).

9. A method for evaluating the cell migration ability of a bioactive material, characterized in that, The method is applicable to the scaffold micro-bridge - microfluidic chip according to any one of claims 1 - 8 above, including: embedding scaffold micro-bridges into a plurality of micro-bridge regions (103) to complete the reconstruction of the microfluidic chip structure; seeding cells in the culture flow channel (101) and injecting growth factors into the migration flow channels (102); Connect the microfluidic chip to the perfusion system, and the perfusion system supplies culture medium to the culture channel and the migration channel continuously or intermittently to reproduce the local microenvironment in vivo; Analyze the number of live cells that enter the migration channel through the scaffold microbridges and adhere to the inner wall; Based on the number of the live cells, determine the cell migration ability value of the bioactive material.

10. The method for evaluating the cell migration ability of the bioactive material according to claim 9, wherein, When different types of scaffold microbridges are embedded in the multiple microbridge regions (103), the same type of cells are inoculated in the culture channel (101); When the same type of scaffold microbridges are embedded in the multiple microbridge regions (103), different types of cells are inoculated in the culture channel (101).

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