A universal organ chip module and a three-dimensional multi-organ chip

Abstract

This invention discloses a universal organ-on-a-chip module, comprising a base plate, a lower substrate, a middle substrate, and a top plate. The lower substrate is stacked on the base plate and has through-holes forming a tissue culture chamber with the base plate. This tissue culture chamber is used to culture micro-tissues to simulate organs. The middle substrate is stacked on top of the lower substrate, and its lower side surface is etched with horizontal microchannels. The top plate is disposed on the middle substrate and is used to press the middle and lower substrates together. A fluid interface communicating with the horizontal microchannels is provided on the top plate. The sidewall of the lower substrate has longitudinal microchannels communicating with the horizontal microchannels on the middle substrate. This invention also discloses a three-dimensional multi-organ chip composed of multiple universal organ-on-a-chip modules and its applications. The universal organ-on-a-chip module of this invention enables modularization of organ chips, making the construction of three-dimensional multi-organ chips possible.

Description

Technical Field

[0001] The present invention relates to the technical field of organ chips, and in particular to a universal organ chip module and a three-dimensional multi-organ chip. Background Art

[0002] An organ chip is an organ physiological microsystem built on a glass slide-sized chip, which contains key elements of the organ microenvironment, such as living cells, tissue interfaces, biofluids, and mechanical forces. It can simulate the main structural and functional characteristics of different tissues and organs of the human body and the complex connections between organs in vitro, and is used to predict the human body's response to drugs or different external stimuli. It has broad application prospects in life science and medical research, new drug development, personalized medicine, toxicity prediction, and biodefense.

[0003] Multi-organ chips are the jewel of organ chips. They can simulate the human body and examine the overall effects of drug action. However, the common problems with multi-organ chips are: first, the number of organs integrated into multi-organ chips is not large enough, making it difficult to simulate the human body; second, each multi-organ chip has its own design, which makes it difficult to standardize and industrialize production. Summary of the invention

[0004] The technical problem to be solved by the present invention is to provide a universal multi-organ chip building module. By freely combining these modules, a multi-organ chip can be constructed, and the multi-organ chip can be three-dimensional.

[0005] In order to solve the above technical problems, the present invention provides the following technical solutions:

[0006] The present invention provides a universal organ chip module, which is composed of a bottom plate, a lower substrate, a middle substrate and a top plate;

[0007] The lower substrate is stacked on the bottom plate, and a through hole is formed on the lower substrate. The through hole and the bottom plate form a tissue culture chamber. The tissue culture chamber is used to culture micro-tissues to simulate organs. The micro-tissues include primary mammalian tissues, cell spheres and organoids.

[0008] The middle substrate is stacked on top of the lower substrate, a horizontal microchannel is etched on the lower side of the middle substrate in the horizontal direction, and the micro-tissues in the tissue culture chamber communicate through the fluid in the horizontal microchannel;

[0009] The top plate is arranged on the middle substrate and is used for pressing the middle substrate and the lower substrate together; the top plate is provided with a fluid interface communicating with the horizontal microchannel;

[0010] A longitudinal microchannel is provided on the side wall of the lower substrate, and the longitudinal microchannel is communicated with the horizontal microchannel on the middle substrate.

[0011] Furthermore, the bottom surface of the tissue culture chamber is a super-hydrophobic surface with micro-nano morphology, and its contact angle with the aqueous solution is greater than 120°;

[0012] Or the bottom surface of the tissue culture chamber is modified with a polymer coating with micro-nano morphology, and the contact angle between the polymer coating and the aqueous solution is less than 90°.

[0013] In the present invention, the bottom surface of the tissue culture chamber is a modified low-adhesion surface, so that when the micro-tissue is cultured, the micro-tissue will not adhere to the bottom of the well and disintegrate.

[0014] In one embodiment, the method for forming the super-hydrophobic surface is: mixing nano-silica particles, n-hexane and chloroform and then performing ultrasonic treatment to disperse the nano-silica particles in the mixed solution; then, immersing the PMMA plate in the mixed solution, taking it out and drying it, thereby obtaining a super-hydrophobic surface with a micro-nano structure.

[0015] Preferably, the method for forming the super-hydrophobic surface is as follows: weigh 0.25-2g of hydrophobic nano-silicon dioxide particles with a diameter of 2-200nm, measure 20-500ml of n-hexane and 0.5-50ml of chloroform. Mix the above materials and perform ultrasonic treatment to completely disperse the nano-silicon dioxide particles in the mixed solution. Immerse the PMMA plate in the above solution for about 5-300 seconds, take it out and dry it, and a PMMA surface with a surface super-hydrophobic micro-nano structure is obtained.

[0016] In another embodiment, the method for forming the super-hydrophobic surface is: immersing the PDMS plate in tetraethyl orthosilicate to make it swell, then taking out the swollen PDMS plate and placing it in an ethylenediamine aqueous solution; then taking out the PDMS plate, rinsing it and performing a heat treatment to obtain a super-hydrophobic surface with a micro-nano structure.

[0017] Preferably, the method for forming the super hydrophobic surface is as follows: immerse the PDMS sheet in tetraethyl orthosilicate at 30-70° C. for 10-200 minutes, then take out the swollen PDMS sheet from the tetraethyl orthosilicate solution and immediately float it in a 5%-40% ethylenediamine aqueous solution. After 3-24 hours, take it out and rinse it with deionized water for 3-5 times, and heat treat it in an oven for 0.5-5 hours to obtain a PDMS sheet with a surface super hydrophobic micro-nano structure.

[0018] In another embodiment, the method of forming the super hydrophobic surface is:

[0019] a. pouring a prepolymer of an elastic resin onto a template having a surface super-hydrophobic micro-nano structure, polymerizing the elastic resin prepolymer into a first elastic solid, and then peeling the first elastic solid from the template;

[0020] b. The surface of the first elastic solid is silanized and modified, and the silanized first elastic solid is used as a template, and then a prepolymer of an elastic resin is poured to polymerize the elastic resin prepolymer into a second elastic solid;

[0021] c. Peeling the second elastic solid from the silanized first elastic solid template, at which point a surface super-hydrophobic micro-nano structure is formed on the surface of the second elastic solid.

[0022] The template with the surface super-hydrophobic micro-nano structure can be made of natural materials, such as cicada wings, lotus leaves, etc., or can be prepared by artificial methods, such as surface etching of aluminum plates, surface etching of polymer plates, MEMS processing, surface enrichment of silica micro-nano particles, surface spraying of super-hydrophobic coatings, etc.

[0023] Furthermore, the method for preparing the polymer coating comprises the following steps:

[0024] S1. Mixing glycidyl methacrylate and other polymerizable monomers in water, adding tetramethylethylenediamine and an initiator, and performing a copolymerization reaction; after the reaction, the copolymer solution is dialyzed to remove unreacted monomers and initiators to obtain a polymer coating liquid;

[0025] S2. pre-treating the bottom surface to generate polar groups on the bottom surface, and then placing the pre-treated bottom surface in the polymer coating liquid for immersion treatment; then taking out the bottom surface, washing off the polymer coating liquid on the surface, and drying, so as to form a polymer coating on the bottom surface of the culture well;

[0026] Among them, the other polymerizable monomers include one or more of 2-(2-methoxyethoxy)ethyl methacrylate, methoxyethyl methacrylate, methyl dimethacrylate, hexafluorobutyl acrylate, and methyl methacrylate; and the polar groups include one or more of hydroxyl, amino, and carboxyl groups.

[0027] Furthermore, in step S1, the mixing time of glycidyl methacrylate and other polymerizable monomers in water is 5-30 minutes to ensure uniform mixing.

[0028] Furthermore, in step S1, the concentration of tetramethylethylenediamine added is 0.1%-3%, and the concentration of the initiator added is 0.1%-3%.

[0029] Furthermore, in step S1, the initiator may be a commonly used initiator in the art, preferably potassium persulfate. The copolymerization reaction time is preferably 30 minutes to 1 day.

[0030] Furthermore, in step S1, the dialysis is specifically: using a dialysis membrane to completely dialyze the copolymer solution for two days, changing the solution every 6-12 hours to remove monomers and small molecules that have not undergone polymerization reaction, and the solution in the dialysis bag is the polymer coating solution.

[0031] In step S2 of the present invention, by introducing groups such as hydroxyl, amino, carboxyl, etc. on the bottom surface of the culture well, the hydrophilic modification liquid can be bonded to the bottom surface of the culture well by covalent bonds, so that it is firmly bonded to the bottom surface of the culture well to form a polymer coating with micro-nano morphology. Preferably, the well plate is treated with plasma to introduce polar groups on the surface of the well plate. For example, oxygen plasma treatment is used to introduce hydroxyl groups.

[0032] Furthermore, in step S2, the soaking temperature is 30-70°C, and the soaking time is 30 minutes to 5 days; the drying temperature is 0-75°C, and the drying time is 30 minutes to 5 days.

[0033] In a preferred embodiment, a through hole is provided on the side or bottom of the tissue culture chamber, and the through hole connects the chamber to the atmosphere to maintain a continuous oxygen supply in the chamber.

[0034] In a preferred embodiment, a porous membrane is sandwiched between the lower substrate and the middle substrate, and the porous membrane is used to culture vascular endothelial cells and / or immune cells. Preferably, the porous membrane is fixed to the opening of the through hole on the lower substrate by pressing.

[0035] In the present invention, the bottom plate and the lower substrate are made of hard rigid materials, and the middle substrate is made of soft elastic materials.

[0036] The present invention also provides a multi-organ chip, comprising a plurality of the universal organ chip modules described above, wherein the organ chip modules are interconnected with the fluid interface on the top plate of another organ chip module through the longitudinal microchannels in the side walls of the lower substrate, forming a three-dimensional microchannel network to simulate the vascular network in the body.

[0037] In the present invention, various three-dimensional multi-organ chips can be formed by freely combining multiple organ chip modules, such as gradient structure multi-organ chips, spiral structure multi-organ chips, rectangular multi-organ chips, three-dimensional human organ chips, etc.

[0038] Furthermore, in the multi-organ chip, tissue culture chambers include but are not limited to brain culture chambers, eye culture chambers, nasal culture chambers, ear culture chambers, tongue culture chambers, tracheal culture chambers, heart culture chambers, lung culture chambers, liver-intestine culture chambers, kidney culture chambers, stomach culture chambers, pancreas culture chambers, spleen culture chambers, skin culture chambers, fat culture chambers, bone marrow culture chambers, muscle culture chambers, testicular culture chambers and tumor culture chambers.

[0039] The present invention also provides the application of the multi-organ chip in drug efficacy detection and toxicity detection.

[0040] Compared with the prior art, the present invention has the following beneficial effects:

[0041] 1. The present invention provides a universal organ chip module, which breaks through the limitation of the number of integrated organs in the traditional organ chip construction technology. By using the organ chip module of the present invention, a multi-organ chip with theoretically unlimited organs can be constructed.

[0042] 2. The universal organ chip module provided by the present invention realizes the modularization of multi-organ chips, thereby facilitating industrial large-scale production.

[0043] 3. The multi-organ chip in the present invention is composed of a free combination of organ chip modules, so it can be processed into any 3D shape, which provides convenience for the subsequent construction of supporting instruments. BRIEF DESCRIPTION OF THE DRAWINGS

[0044] Figure 1 This is an electron microscope photo of the surface of polydimethylsiloxane after it was molded twice through the lotus leaf;

[0045] Figure 2 The state of the droplet on the superhydrophobic PDMS surface;

[0046] Figure 3 This is a bright field photo of tumor spheres on the PDMS superhydrophobic surface;

[0047] Figure 4 The change of contact angle after repeated use for 12 times;

[0048] Figure 5 This is a surface electron microscope photo of a polydimethylsiloxane sheet containing silica microbubbles;

[0049] Figure 6 Bright field photograph of tumor spheres on the surface of polydimethylsiloxane sheet with silica microbubbles;

[0050] Figure 7 This is an electron microscope photo of the superhydrophobic PMMA surface;

[0051] Figure 8This is a bright field photo of tumor spheres on a superhydrophobic PMMA surface;

[0052] Fig. 9 This is an atomic force microscope image of the polymer coating surface;

[0053] Fig.10 The adsorption of fluorescent proteins on the modified PDMS surface (A) and the unmodified PDMS surface (B);

[0054] Fig.11 Tumor spheres formed on the surface of polydimethylsiloxane polymer coating;

[0055] Fig.12 Disintegration of renal microstructures on a normal surface;

[0056] Fig.13 The modular design of a single organ in Example 5;

[0057] Fig.14 The design diagram of the gradient four-organ chip in Example 6: (A) Schematic diagram of the tissue culture chamber; (B) porous membrane; (C) PDMS middle substrate with channels; (D) top plate;

[0058] Fig.15 This is a photo showing the actual gradient four-organ chip (without the porous membrane);

[0059] Fig.16 The toxicity evaluation results of the gradient four-organ chip (green represents live cells and red represents dead cells);

[0060] Fig.17 This is a schematic diagram (side view) of connecting adjacent organ modules in the spiral chip of Example 7;

[0061] Fig.18 It is a top view of the spiral chip structure;

[0062] Fig.19 This is a physical display of the spiral chip (without the porous membrane);

[0063] Fig. 20 The structure of the tissue culture chamber layer (A) and the PDMS channel layer (B) in the rectangular parallelepiped chip of Example 8;

[0064] Fig.21 This is a physical display of a rectangular chip (without the porous membrane);

[0065] Fig. 22 This is a schematic diagram of the tissue culture chamber and blood vessel structure of the three-dimensional human chip in Example 9;

[0066] Fig.23 A photo showing the physical object of the 3D human chip;

[0067] in, Fig.13 and Fig.17 Numbers in the figure: 1, bottom plate; 2, lower substrate; 3, porous membrane; 4, middle substrate; 5, top plate; 6, longitudinal microchannel; 7, microtissue; 8, pressing sheet; 9, horizontal microchannel; A, organ chip 1; B, organ chip 2;

[0068] Fig.14 Numbers in the figure: 1, liver tissue culture chamber; 2, kidney tissue culture chamber; 3, brain tissue culture chamber; 4, heart tissue culture chamber; a~a”', b~b”', c~c”', d~d”', e~e”', f~f”': mounting holes; S, K: fluid interfaces;

[0069] Fig.18 Numbers in the figure: a~e, organ module; S, sealing screw; t1~t4, screws; 1, 2~2', 3'~3", 4"~4"', 5"'~5"", 6"", fluid interface;

[0070] Fig. 20 Numbers in the figure: s, fixing bolt; t, longitudinal microchannel;

[0071] Fig.21 Numbers in the figure: 1. brain tissue culture chamber; 2. tracheal tissue culture chamber; 3. heart tissue culture chamber; 4, 5. lung tissue culture chamber; 6, 7. nerve and fat tissue culture chamber; 8. liver tissue culture chamber; 9. kidney tissue culture chamber; 10. pancreatic islet tissue culture chamber; 11. spleen tissue culture chamber; 12. bone marrow tissue culture chamber; 13. skin tissue culture chamber; 14. tumor tissue culture chamber; 15. cartilage tissue culture chamber; a~e, fluid interface. DETAILED DESCRIPTION

[0072] The present invention is further described below in conjunction with the accompanying drawings and specific embodiments so that those skilled in the art can better understand the present invention and implement it, but the embodiments are not intended to limit the present invention.

[0073] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those skilled in the art of the present invention. The terms used herein in the specification of the present invention are only for the purpose of describing specific embodiments and are not intended to limit the present invention. The term "and / or" used herein includes any and all combinations of one or more related listed items.

[0074] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods, and the materials, reagents, etc. used are all available from commercial sources unless otherwise specified.

[0075] Example 1: Using natural materials as templates to create super-hydrophobic surfaces

[0076] The lotus leaf was fixed on a glass plate, and liquid polydimethylsiloxane was poured on its surface. The liquid polydimethylsiloxane was polymerized overnight to form a solid. The solidified polydimethylsiloxane sheet was peeled off from the lotus leaf as a template for the next step. The surface of the polydimethylsiloxane template was then silanized, and liquid polydimethylsiloxane was poured on the template again. The temperature was raised for polymerization, and the newly solidified polydimethylsiloxane sheet was peeled off from the silanized polydimethylsiloxane sheet template. The electron microscope picture of its surface is shown in FIG. Figure 1 As shown, it has a super-hydrophobic micro-nanostructure with a contact angle of >120°( Figure 2 ), the super hydrophobic polydimethylsiloxane plate is completed.

[0077] Low Adhesion Characterization:

[0078] The surface was used for tumor cell spheroidization experiments. Human embryonic lung fibroblasts (MRC-5) and human lung adenocarcinoma cells (NCI-H1792) were mixed and cultured in RPMI 1640 + 10% FBS. The cells were placed in the wells with the super-hydrophobic surface. The ratio of fibroblasts to cancer cells was 1:1. The culture environment was 37°C and 5% CO2. After 3 days, the tumor cells did not adhere to the wall, but formed spheres ( Figure 3 ), demonstrating the low adhesion of the superhydrophobic surface.

[0079] Durability Characterization:

[0080] The test repeatability showed that the well was added with various cell culture media for 48 hours, then poured out, washed, dried, and the above experiment was repeated 12 times, and the contact angle was measured again, which was still greater than 120° ( Figure 4 ), indicating that the low adhesion surface can be reused at least 12 times and continuously used for more than 24 days.

[0081] Example 2: Directly fabricating a polydimethylsiloxane superhydrophobic surface

[0082] Immerse a piece of polydimethylsiloxane plate in tetraethyl orthosilicate at 50°C for 20 minutes. Then, take out the polydimethylsiloxane plate swollen with tetraethyl orthosilicate from the tetraethyl orthosilicate solution and immediately float it in an aqueous solution of 10% ethylenediamine, and silica microbubbles gradually form on the surface of the polydimethylsiloxane plate. After 10 hours, take out the polydimethylsiloxane plate with silica microbubbles and rinse it with deionized water 3 times. Finally, heat treat the polydimethylsiloxane plate with silica microbubbles in an oven for 1 hour. The electron microscope photograph of the prepared superhydrophobic surface is shown in the figure. Figure 5 shown.

[0083] Low Adhesion Characterization:

[0084] The surface was used for tumor cell spheroidization experiments. Human embryonic lung fibroblasts (MRC-5) and human lung adenocarcinoma cells (NCI-H1792) were mixed and cultured in RPMI 1640 + 10% FBS. The cells were placed in the wells with the super-hydrophobic surface. The ratio of fibroblasts to cancer cells was 1:1. The culture environment was 37°C and 5% CO2. After 3 days, the tumor cells did not adhere to the wall, but formed spheres ( Figure 6 ), demonstrating the low adhesion of the superhydrophobic surface.

[0085] Durability Characterization:

[0086] The test repeatability showed that cell culture medium (1640 culture medium + 10% fetal bovine serum) was added to the well for 48 hours, then poured out, washed, dried, and the above experiment was repeated 12 times. The final contact angle was measured to be 135.8°, which was still greater than 120°, indicating that the low adhesion surface can be reused at least 12 times, and the continuous use time is more than 24 days.

[0087] Example 3: Preparation of polymethyl methacrylate superhydrophobic surface

[0088] Weigh 0.25 g of hydrophobic nano-silica particles with a diameter of 20 nm, measure 50 ml of n-hexane and 2.5 ml of chloroform, mix them and perform ultrasonic treatment to make the nano-silica particles completely dispersed in the mixed solution, immerse the polymethyl methacrylate plate in the above solution for about 15 seconds, take it out and dry it, and the polymethyl methacrylate super-hydrophobic surface ( Figure 7 ).

[0089] Low Adhesion Characterization:

[0090] The surface was used for tumor cell spheroidization experiments. Human embryonic lung fibroblasts (MRC-5) and human lung adenocarcinoma cells (NCI-H1792) were mixed and cultured in RPMI 1640 + 10% FBS. The cells were placed in the wells with the super-hydrophobic surface. The ratio of fibroblasts to cancer cells was 1:1. The culture environment was 37°C and 5% CO2. After 3 days, the tumor cells did not adhere to the wall, but formed spheres ( Figure 8 ), demonstrating the low adhesion of the superhydrophobic surface.

[0091] Durability Characterization:

[0092] The test repeatability showed that the well plate was added with cell culture medium (1640 culture medium + 10% fetal bovine serum) for 48 hours, then poured out, washed, dried, and the above experiment was repeated 12 times. The final contact angle was measured to be 132.6°, which was still greater than 120°, indicating that the low adhesion surface can be reused at least 12 times, and the continuous use time is more than 24 days.

[0093] Example 4: Preparation of super hydrophilic polydimethylsiloxane surface

[0094] Evenly mix 2ml of methyl methacrylate, 0.04ml of poly(glycidyl methacrylate) and 38ml of water for 10 minutes, then add 0.4g of potassium persulfate and 2ml of water. After fully dissolved, add 0.04ml of TEMED and copolymerize at 25℃ for 15 minutes. After the solution color changes from transparent to milky white, stop the reaction and immediately transfer to a 1.5w dialysis bag for dialysis for two days. Change the solution every 12 hours to remove monomers and small molecules that have not undergone polymerization. The solution in the dialysis bag is the polymer coating solution.

[0095] First, hydroxyl groups were generated on the surface of polydimethylsiloxane by oxygen plasma, and then the surface was immersed in the polymer coating liquid for 1 hour, and then the polymer coating liquid was removed, washed three times with deionized water, and dried at 75° C. for 1 hour.

[0096] First, the surface was characterized by atomic force microscopy. Fig. 9 It can be clearly seen that after modification, there is a layer of polymer on the surface of polydimethylsiloxane, and the polymer forms a nano-groove structure on the surface.

[0097] Then, the protein adsorption prevention characterization was performed. The fluorescent labeled secondary antibody solution was injected onto the modified and unmodified surfaces respectively. It can be clearly seen that on the unmodified surface, the fluorescence intensity was very high, indicating that the protein adsorption was very serious, while on the modified surface, almost no fluorescence was detected, indicating that the coating effectively prevented the adsorption of proteins ( Fig.10 ).

[0098] Finally, anti-cell adhesion characterization was performed. Human embryonic lung fibroblasts (MRC-5) and human lung adenocarcinoma cells (NCI-H1792) were mixed and cultured in RPMI 1640 + 10% FBS. The culture ratio of fibroblasts to cancer cells was 1:1, and the culture environment was 37°C and 5% CO2. After 3 days, the cells did not adhere to the wall, but formed tumor spheres ( Fig.11 ), confirming the coating's ability to prevent cell adhesion. Five consecutive tumor sphere formation experiments were performed on the same surface, all of which were successful, demonstrating the durability of the low-adhesion surface.

[0099] When kidney tissue was placed on a non-low adhesion surface, it was found that the cells crawled out and the tissue quickly disintegrated ( Fig.12 ).

[0100] Example 5: Single organ module design

[0101] Fig.13 The designs of two organ modules are given. Fig.13In A, the porous membrane is directly sandwiched between the middle substrate and the substrate with through holes. Fig.13 In B, the porous membrane is fixed in the through hole by a pressing sheet. Endothelial cells and / or immune cells can be cultured on the porous membrane. Regardless of the design, the inner wall has vertical longitudinal microchannels. This organ module can be easily combined at will to form a multi-organ chip.

[0102] Example 6: Gradient Four-Organ Chip

[0103] This embodiment produces a gradient-type 4-organ chip that can contain four modules ( Fig.14 ). Fig.15 It is a real-life photo display. The chip is still a two-dimensional structure, so there are no vertical channels on the side walls of the chamber. Figure A is a PMMA tissue culture chamber. The four modules (1, 2, 3, 4) are respectively filled with liver, kidney, brain, and heart tissues. The bottom surfaces of the tissue culture chambers are all 4*4mm squares, with the highest point being 7mm and the lowest point being 4mm. Figure B is a porous membrane, which together covers the top surfaces of the four chambers. Figure C is a middle-layer substrate with a PDMS channel, with a channel diameter of 1mm, a height of 1mm, and a length of 26mm. Figure D is a top plate. In Figures A, B, C, and D, the 6 holes on the edge (a, b, c, d, e, and f series) are used to install fixing screws and nuts. In Figure D, S and K are fluid interfaces.

[0104] The method for obtaining the four types of micro-tissues, liver, kidney, brain, and heart, is: first, cut the organ into pieces with scissors, then mask, and finally sieve to retain tissue particles of 70-100 microns, which are the micro-tissues that simulate the organs in the chip. The culture medium used for each micro-tissue is shown in the table below.

[0105]

[0106]

[0107] The chip was used to measure the toxicity of doxorubicin, and it can be seen that doxorubicin has different toxicities to different organs ( Fig.16 ).

[0108] Example 7: Spiral Multi-Organ Chip

[0109] This example fabricates a three-dimensional spiral multi-organ chip ( Figure 17-19 ), showing that this method can make a three-dimensional multi-organ chip of any shape. The three-dimensional spiral multi-organ chip is composed of five organ modules stacked in a spiral. The horizontal channel in the upper organ module is connected to the horizontal microchannel in the adjacent organ module through the vertical microchannel in the side wall, so that the micro-tissues in different organ modules can communicate with each other through the microchannel. The connection between two adjacent organ modules is shown in FIG. Fig.17.

[0110] Fig.18 In the chip, a, b, c, d, and e are five organ modules from bottom to top, and the size of the tissue culture chamber is 4*4*4mm. S is the position of the individual sealing screws of each organ module. The t1 screw connects and fixes the a and b modules together, the t2 screw connects and fixes the b and c modules together, the t3 screw connects and fixes the c and d modules together, and the t4 screw connects and fixes the d and e modules together, finally forming a spiral multi-organ chip. In the a module, the cell culture fluid flows from 1 to 2 in the horizontal direction, then flows into the vertical channel in the side wall of the b module to 2', then flows horizontally to 3', then flows into the vertical channel in the side wall of the c module to 3", then flows horizontally to 4", then flows into the vertical channel in the side wall of the d module to 4"', then flows horizontally to 5"', then flows into the vertical channel in the side wall of the e module to 5", then flows horizontally to 6"", and then flows out from 6" and returns to 1, forming a circulation of the culture fluid to simulate blood circulation.

[0111] Fig.19 This is a photo showing the actual spiral chip.

[0112] Example 8: Cuboid Chip

[0113] Traditional multi-organ chips are relatively large, which is not conducive to the construction of portable instruments. This embodiment uses a modular multi-organ chip construction method to construct a rectangular organ chip that can accommodate 24 organs. The overall size of the chip is 34*34*29mm, which is the multi-organ chip with the highest organ density today, and is particularly convenient for the construction of portable organ chip instruments.

[0114] The entire cuboid chip is divided into three layers. Fig. 20 A shows a layer of a rectangular chip, which has 8 organ modules and the size of the tissue culture chamber is 4*4*4mm. Fig. 20 The horizontal microchannels of the middle substrate shown in B connect the eight tissue culture chambers on the same layer, and the horizontal channels of different middle substrates are interconnected through the longitudinal microchannels t on the side walls of the tissue culture chambers, so that the 24 organs can communicate with each other. s is the position of the fixing bolt.

[0115] Fig.21 This is a physical display picture of a rectangular chip.

[0116] Example 9: Three-dimensional human organ chip

[0117] The human body chip is the ultimate goal of the development of multi-organ chips. This embodiment uses the present invention to construct a modular three-dimensional human body chip that is similar to the real human body structure.

[0118] Fig. 22 The diagram shows the structure of the 3D human chip. Chamber 1 is used to store brain tissue, chamber 2 is used to store tracheal tissue, chamber 3 is used to store heart tissue, chambers 4 and 5 are used to store lung tissue, chambers 6 and 7 are used to store nerve and fat tissue, chamber 8 is used for liver tissue, chamber 9 is located at the back of chamber 8 and is used to store kidney tissue, chamber 10 is used to store pancreatic islet tissue, and chamber 11 is located at the back of chamber 10 and is used to store spleen tissue. Chamber 12 is used to store bone marrow tissue, chamber 13 is used to store skin tissue, chamber 14 is used to store tumor tissue, and chamber 15 is used to store cartilage tissue. The 3D distribution of these chambers, especially the key organ chambers, in the human chip is consistent with the distribution of real tissues in the 3D human chip.

[0119] Fig. 22 The thick solid lines in the figure represent microchannels, which simulate blood vessels. These microchannels pass through the surface of each chamber and are connected to the chamber. Thus, all tissues are connected together to form an organic whole. The cell culture fluid flows in from point a, flows out from points b, c, d, and e, and then returns to point a, forming a cell culture fluid circulation.

[0120] Fig.23 This is a physical display picture of the three-dimensional human body chip.

[0121] The above-described embodiments are only preferred embodiments for fully illustrating the present invention, and the protection scope of the present invention is not limited thereto. Equivalent substitutions or changes made by those skilled in the art based on the present invention are within the protection scope of the present invention. The protection scope of the present invention shall be subject to the claims.

Claims

1. A universal organ chip module, characterized in that: The module is composed of a bottom plate, a lower substrate, a middle substrate and a top plate; The lower substrate is stacked on the bottom plate, and a through hole is formed on the lower substrate. The through hole and the bottom plate form a tissue culture chamber. The tissue culture chamber is used to culture micro-tissues to simulate organs. The micro-tissues include primary mammalian tissues, cell spheres and organoids. The middle substrate is stacked on top of the lower substrate, a horizontal microchannel is etched on the lower side of the middle substrate in the horizontal direction, and the micro-tissues in the tissue culture chamber communicate through the fluid in the horizontal microchannel; The top plate is arranged on the middle substrate and is used for pressing the middle substrate and the lower substrate together; the top plate is provided with a fluid interface communicating with the horizontal microchannel; The side wall of the lower substrate is provided with a longitudinal microchannel, and the longitudinal microchannel is connected with the horizontal microchannel on the middle substrate; The bottom surface of the tissue culture chamber is modified with a polymer coating with a micro-nano morphology, and the contact angle between the polymer coating and the aqueous solution is less than 90°; The preparation method of the polymer coating comprises the following steps: S1. Glycidyl methacrylate and other polymerizable monomers are mixed in water, tetramethylethylenediamine and an initiator are added, and a copolymerization reaction is carried out; after the reaction is completed, the copolymer solution is dialyzed to remove unreacted monomers and initiators, and a polymer coating liquid is obtained; S2 pre-treating the bottom surface to produce polar groups on the bottom surface, and then placing the pre-treated bottom surface in the polymer coating liquid for immersion treatment; then remove the bottom surface, wash off the surface of the polymer coating liquid, after drying, that is, forming a polymer coating on the bottom surface; Among them, the other polymerizable monomers include one or more of 2-(2-methoxyethoxy)ethyl methacrylate, methoxyethyl methacrylate, methyl dimethacrylate, hexafluorobutyl acrylate, and methyl methacrylate; and the polar groups include one or more of hydroxyl, amino, and carboxyl groups.

2. A universal organ chip module according to claim 1, characterized in that: A through hole is provided on the side surface or the bottom surface of the tissue culture chamber.

3. A universal organ chip module according to claim 1, characterized in that: A layer of porous membrane is sandwiched between the lower substrate and the middle substrate, and the porous membrane is used for culturing vascular endothelial cells and / or immune cells.

4. A universal organ chip module according to claim 3, characterized in that: The porous membrane is fixed to the opening of the through hole on the lower substrate by pressing.

5. A universal organ chip module according to claim 1, characterized in that: The bottom plate and the lower substrate are made of hard rigid materials, and the middle substrate is made of soft elastic materials.

6. A multi-organ chip, comprising a plurality of universal organ chip modules according to any one of claims 1 to 5, characterized in that: The organ chip module is interconnected with the fluid interface on the top plate of another organ chip module through the longitudinal microchannel in the side wall of the lower substrate, forming a three-dimensional microchannel network to simulate the vascular network in the body.

7. The multi-organ chip according to claim 6, characterized in that: The tissue culture chambers in the universal organ chip module include brain culture chamber, eye culture chamber, nasal culture chamber, ear culture chamber, tongue culture chamber, tracheal culture chamber, heart culture chamber, lung culture chamber, liver-intestine culture chamber, kidney culture chamber, stomach culture chamber, pancreas culture chamber, spleen culture chamber, skin culture chamber, fat culture chamber, bone marrow culture chamber, muscle culture chamber, testis culture chamber and tumor culture chamber.

8. Use of the multi-organ chip according to claim 6 or 7 in drug efficacy detection and toxicity detection.

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