Micro-fluidic chip for in-vitro 3D hepatic organ co-culture

Abstract

The utility model discloses a micro-fluidic chip for in-vitro 3D hepatic organ co-culture, which belongs to the technical field of biomedical engineering and comprises a perfusion layer and a culture layer, and the perfusion layer is provided with a first culture solution inlet and a first culture solution outlet; the culture layer is provided with a circulating microfluidic perfusion culture system; the chip comprises an upper layer and a lower layer formed by bonding a perfusion layer and a culture layer; the circulating micro-fluidic perfusion culture system comprises two groups of micro-fluidic multi-chamber co-culture chips which are connected in series; the series micro-fluidic multi-chamber co-culture chip comprises a second culture solution inlet, a micro-fluid channel, a compound cell co-culture area and a second culture solution outlet, and the second culture solution inlet, the micro-fluid channel, the compound cell co-culture area and the second culture solution outlet are communicated with one another. The problem that in the prior art, while complex physiological environment simulation is maintained, the reliability and practicability of the model are improved is solved. The method has the advantages of high efficiency and accuracy.

Description

Technical Field

[0001] The utility model relates to the technical field of biomedical engineering, and in particular to a microfluidic chip for in vitro 3D liver organ co-culture. Background Art

[0002] In recent years, microfluidic chip technology has undergone significant development and has shown broad application potential in the fields of chemical analysis and biomedical research. In particular, in the field of liver tissue engineering, this technology has promoted the creation of highly simulated in vitro models of the liver in order to achieve rapid and efficient drug screening. Compared with traditional two-dimensional in vitro liver organ chips, the application of three-dimensional (3D) in vitro culture technology in liver chips has significantly enhanced cell-to-cell interactions, cell polarity, phenotypic expression and functionality, and more accurately simulated the natural physiological architecture and function of the liver. By reproducing the interactions and metabolic activities of the complex cell networks inside the liver, the 3D in vitro cell culture model not only deepens the understanding of the pathogenesis of the disease, but also improves the accuracy of predicting the in vivo effects of drugs.

[0003] The current research and development of 3D in vitro liver chip technology covers a variety of strategies, such as scaffold-free spheroid culture, scaffold-based cell culture, hollow fiber bioreactors, 3D printing technology, and organoid chips. Among them, most of the designed chips use hydrophilic polytetrafluoroethylene membranes to promote the co-culture of endothelial cells and hepatocytes, or use microarray chips to construct complex cell models, aiming to more faithfully simulate the in vivo environment. Although the use of scaffold materials is conducive to maintaining and promoting the specific phenotype of liver spheroids, their batch-to-batch variability has become a limiting factor, resulting in reduced experimental repeatability and comparability of results, thus posing challenges in the assessment of drug-induced liver injury. In addition, based on the scaffold model, it is necessary to carefully consider the potential impact of the scaffold material on drug transport kinetics, which is an indispensable part of ensuring the accuracy of drug screening and toxicity test results. In summary, 3D microfluidic liver chip technology provides an advanced platform for drug development and disease research by highly simulating the physiological characteristics of the liver, but optimization in standardization, material consistency, and drug diffusion kinetics remains a key direction for future research.

[0004] Therefore, there is an urgent need for a microfluidic chip for in vitro 3D liver organ co-culture as an efficient and accurate in vitro model, which will open up new avenues for drug screening, disease research, and exploration of physiological mechanisms. Utility Model Content

[0005] To this end, the utility model provides a microfluidic chip for in vitro 3D liver organ co-culture to solve the problem in the prior art of how to improve the reliability and practicality of the model while maintaining the simulation of a complex physiological environment.

[0006] In order to achieve the above purpose, the utility model provides the following technical solutions:

[0007] According to the first aspect of the utility model, a microfluidic chip for in vitro 3D liver organ co-culture is provided, comprising:

[0008] a perfusion layer, provided with a first culture solution inlet and a first culture solution outlet; and

[0009] The culture layer is provided with a circulating microfluidic perfusion culture system;

[0010] The chip is an upper and lower layer formed by bonding a perfusion layer and a culture layer.

[0011] Furthermore, the circulating microfluidic perfusion culture system includes two sets of serially connected microfluidic multi-chamber co-culture chips;

[0012] The serial microfluidic multi-chamber co-culture chip includes a second culture solution inlet, a microfluidic channel, a composite cell co-culture area, and a second culture solution outlet. The second culture solution inlet, the microfluidic channel, the composite cell co-culture area, and the second culture solution outlet are connected, and the second culture solution inlet is connected to the first culture solution inlet, and the second culture solution outlet is connected to the first culture solution outlet.

[0013] Furthermore, the composite cell co-culture area includes a HUVEC cell culture area and a 3D cell culture chamber, and the HUVEC cell culture area and the 3D cell culture chamber are connected through a microfluidic channel.

[0014] Furthermore, the microfluidic channel includes a first perfusion channel, a second perfusion channel and a third perfusion channel, the first perfusion channel is arranged between the second culture fluid inlet and the HUVEC cell culture area, the second perfusion channel is arranged between the HUVEC cell culture area and the 3D cell culture chamber, and the third perfusion channel is arranged between the 3D cell culture chamber and the second culture fluid outlet.

[0015] Furthermore, the second culture fluid inlet is vertically arranged at the end of the first perfusion channel and is connected to the first perfusion channel; the second culture fluid outlet is vertically arranged at the end of the third perfusion channel, and the third perfusion channel is connected to the second culture fluid outlet.

[0016] Furthermore, it also includes a detachable hose, which is connected between two groups of serially connected microfluidic multi-chamber co-culture chips.

[0017] Furthermore, the microfluidic chip has a length of 50 mm, a width of 25 mm, and a height of 8 mm; the perfusion layer has a height of 2 mm, and the culture layer has a height of 6 mm.

[0018] Furthermore, the HUVEC cell culture area has a length of 15 mm, a width of 4 mm, and a height of 5 mm; the 3D cell culture area has a diameter of 9 mm and a height of 5.5 mm.

[0019] Furthermore, the length of the first perfusion channel is 2.6 mm, the length of the second perfusion channel is 2.3 mm, the length of the third perfusion channel is 3.1 mm, and the width of the first perfusion channel, the second perfusion channel and the third perfusion channel are all 0.5 mm, and the height is all 1 mm.

[0020] The utility model has the following advantages:

[0021] 1. The 3D liver organ co-culture chip of the present application is selective. It can not only fully carry out 3D liver organoids, but also carry out multi-organ co-culture while simulating the first-pass effect of drugs in the liver, so as to more comprehensively explore the interaction of drugs in multiple organs of the human body.

[0022] 2. The microfluidic multi-cell co-culture chip of the present application has undergone structural optimization and controlled precise structural dimensions. At the same time, it uses high-permeability materials to facilitate observation and optimization of cell operations, improve the intuitiveness and efficiency of research, and improve the convenience of operation and visualization and positioning capabilities, making the experimental process more controllable, intuitive, and easy to manage.

[0023] 3. This application highly simulates the in vivo environment, improves the physiological relevance of drug screening and disease research, optimizes complex cell interactions, promotes more comprehensive metabolism, signaling research and mechanism understanding, precisely controls microfluidic technology, ensures the accuracy of cell culture, drug delivery, and reaction analysis, and adopts modular design to enhance experimental flexibility and adaptability to multiple scenarios, and promotes the construction of complex models. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] In order to more clearly illustrate the implementation of the utility model or the technical solution in the prior art, the following is a brief introduction to the drawings required for the implementation or the prior art description. Obviously, the drawings in the following description are only exemplary, and for ordinary technicians in this field, other implementation drawings can be derived from the provided drawings without creative work.

[0025] The structures, proportions, sizes, etc. illustrated in this specification are only used to match the contents disclosed in the specification so as to facilitate understanding and reading by persons familiar with the technology. They are not used to limit the conditions under which the present invention can be implemented, and therefore have no substantial technical significance. Any structural modification, change in proportion or adjustment of size shall still fall within the scope of the technical contents disclosed in the present invention without affecting the effects and purposes that can be achieved by the present invention.

[0026] Figure 1 This is a schematic diagram of the overall structure of a microfluidic chip for in vitro 3D liver organ co-culture provided by the utility model;

[0027] Figure 2 A schematic diagram of the perfusion layer structure provided by the utility model;

[0028] Figure 3 A schematic diagram of the structure of the circulating microfluidic perfusion culture system provided by the utility model;

[0029] In the figure: 1. perfusion layer; 11. first culture solution inlet; 12. first culture solution outlet; 2. culture layer; 21. circulating microfluidic perfusion culture system; 211. serial microfluidic multi-chamber co-culture chip; 2111. second culture solution inlet; 2112. microfluidic channel; 21121. first perfusion channel; 21122. second perfusion channel; 21123. third perfusion channel; 2113. composite cell co-culture area; 21131. HUVEC cell culture area; 21132. 3D cell culture chamber; 2114. second culture solution outlet; 212. detachable hose. DETAILED DESCRIPTION

[0030] The following is a specific embodiment of the present invention. People familiar with the technology can easily understand the other advantages and functions of the present invention from the contents disclosed in this specification. Obviously, the described embodiment is a part of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0031] The present application provides a microfluidic chip for in vitro 3D liver organ co-culture, comprising a perfusion layer 1 and a culture layer 2, such as Figure 1 As shown. The perfusion layer 1 is provided with a first culture fluid inlet 11 and a first culture fluid outlet 12; the culture layer 2 is provided with a circulating microfluidic perfusion culture system 21. The chip is an upper and lower layer in which the perfusion layer 1 and the culture layer 2 are seamlessly integrated through bonding technology, which ensures the tightness of the system and the stability of operation, while optimizing the microenvironment of cell culture.

[0032] In the perfusion layer 1, the first culture fluid inlet 11 is an input channel for the culture matrix, providing the cells with necessary nutrients and oxygen, while the first culture fluid outlet 12 is responsible for removing metabolic waste and maintaining the balance of the culture environment. The arrangement of the first culture fluid inlet 11 and the first culture fluid outlet 12 ensures efficient cell metabolism and simulates the basic functions of the physiological circulatory system in the body.

[0033] The culture layer 2 is equipped with a circulating microfluidic perfusion culture system 21, such as Figure 2; It not only contains microfluidic channels 2112 and supports three-dimensional co-culture of cells, but also accurately simulates the microenvironment of the liver, providing an ideal in vitro model for efficient drug screening. The circulating microfluidic perfusion culture system 21 can enhance the interaction between cells, and also simulates blood flow through microfluidic channels 2112 to ensure efficient exchange of nutrients and metabolites, as well as accurate delivery, delivery and effect evaluation of drugs or toxins.

[0034] The perfusion layer 1 and culture layer 2 of the microfluidic chip for in vitro 3D liver organ co-culture are bonded and designed. They are made of transparent, breathable and biocompatible polymer polydimethylsiloxane (PDMS). They are sterilized at high temperature and high pressure to ensure a sterile environment, and then washed and impregnated with culture medium for multiple times to ensure no contamination and prepare the best state for cell culture. In addition, the transparency of the microfluidic chip facilitates intuitive observation of cell dynamics, and the permeability facilitates gas exchange and maintains an appropriate oxygen level.

[0035] The circulating microfluidic perfusion culture system 21 includes two sets of serially connected microfluidic multi-chamber co-culture chips 211, which enhances its practicality and complex physiological simulation capabilities in cell culture, drug screening, disease model construction, etc., promotes close cooperation and interaction between cells, and optimizes the stability of the culture environment, making the experimental results closer to the actual situation in vivo.

[0036] like Figure 2 As shown, the serial microfluidic multi-chamber co-culture chip 211 includes a second culture solution inlet 2111, a microfluidic channel 2112, a composite cell co-culture area 2113 and a second culture solution outlet 2114. The second culture solution inlet 2111, the microfluidic channel 2112, the composite cell co-culture area 2113 and the second culture solution outlet 2114 are connected, and the second culture solution inlet 2111 is connected to the first culture solution inlet 11 for circulation, forming a closed circulation system, ensuring the continuous supply and renewal of the culture solution, while maintaining an effective balance between nutrients and metabolites. Fresh culture solution can enter from the first culture solution inlet 11, pass through the second culture solution inlet 2111, and finally return to the first culture solution outlet 12 via the second culture solution outlet 2114, forming a complete flow link, which not only ensures the uniform distribution of the culture solution, but also simulates the dynamics of the circulatory system in the body.

[0037] The composite cell co-culture area 2113 is the core of the system and is tightly connected by the microfluidic channel 2112. This section is designed for the coexistence of different types of cells, such as hepatocytes, endothelial cells, endothelial cells, etc. This design can better simulate the in vivo environment. The diversity of interactions and physiological functions between composite cells provides a highly simulated microenvironment for research. The composite cell co-culture area 2113 is directly connected to the second culture fluid inlet 2111 and outlet through the microfluidic channel 2112, which ensures the exchange of nutrients and metabolites, and is also convenient for the introduction of drugs or stimulants and the precise control of analytes. The second culture fluid outlet 2114 is connected to the first culture fluid outlet 12.

[0038] In summary, the circulating microfluidic perfusion culture system 21, through its serial microfluidic multi-chamber design, not only improves the physiological simulation of cell culture, but also optimizes the controllability and repeatability of the experiment, providing a powerful in vitro platform for drug screening and disease model research. The design details optimize the operation process, improve the efficiency of cell culture and the accuracy of the experiment, and also facilitate researchers to observe and analyze cell dynamics.

[0039] The composite cell co-culture area 2113 includes a HUVEC (i.e., human umbilical vein endothelial cell) cell culture area 21131 and a 3D cell culture chamber 21132, which are connected via a microfluidic channel 2112. The HUVEC cell culture area 21131, as a simulated vascular endothelial cell, is essential for maintaining blood and nutrient exchange and inflammatory response, while the 3D cell culture chamber 21132 accurately simulates the complex interactions and metabolic functions of liver parenchymal cells through its three-dimensional structure. The two are closely connected through a precisely designed microfluidic channel 2112, which not only ensures the two-way flow of nutrients and metabolites, but also promotes signal exchange and interaction between cells, simulating the real physiological dynamic environment in vivo.

[0040] The design of microfluidic channel 2112 not only takes into account fluid dynamics properties, such as flow rate and shear stress, to simulate the impact of blood flow on cells, but also precisely controls the distribution of liquid so that nutrients are evenly delivered to each cell, while recycling metabolic waste to maintain the purity and stability of the culture environment.

[0041] Through such a design, the microfluidic channel 2112 can provide precise application of drugs, growth factors or other stimuli, which is convenient for studying cell response monitoring and analysis. In addition, the design of the composite cell co-culture area 2113 also facilitates the observation and study of phenotypic changes of cells in long-term culture, as well as drug effects and responses of disease models, providing a highly controllable and repeatable experimental platform.

[0042] like Figure 2 and Figure 3 As shown, the microfluidic channel 2112 includes a first perfusion channel 21121, a second perfusion channel 21122 and a third perfusion channel 21123, which form a precise liquid circulation network, aiming to simulate the complex microenvironment in the liver and optimize the interaction and material exchange between cells. The first perfusion channel 21121 is arranged between the second culture fluid inlet 2111 and the HUVEC cell culture area 21131, and serves as the initial inlet for transporting nutrients and metabolites, ensuring that the nutrients carried by the fresh culture fluid can quickly and evenly reach the VEC cells, while providing a microenvironment similar to vascular endothelial cells for the VEC cells, promoting the expression of their physiological functions and the formation of metabolites.

[0043] The second perfusion channel 21122 is disposed between the HUVEC cell culture area 21131 and the 3D cell culture chamber 21132, forming a bridge for intercellular material transfer, which not only supports the material and signal exchange between VEC and 3D cells, such as the exchange of metabolites and cytokines, but also simulates the material and fluid exchange between different regions of the liver in vivo, further enhancing the overall physiological relevance. The third perfusion channel 21123 is disposed between the 3D cell culture chamber 21132 and the second culture fluid outlet 2114. As an outlet for metabolites, it effectively discharges waste and other metabolites generated by cell metabolism to maintain the cleanliness of the chamber. At the same time, this design also ensures the self-renewal and stability of the entire culture environment, providing continuity for long-term experiments.

[0044] Through such a microfluidic channel 2112 design, the directional flow of the culture medium is ensured, and precise regulation is achieved to simulate the physiological nutrient supply, waste removal process, and cell-to-cell interaction, providing a highly simulated in vitro platform for drug screening and disease models. The size and flow rate design of each channel are optimized to maximize cell growth, metabolism, and functional expression, minimize physical disturbances, and make the entire system a powerful tool for studying liver cell interactions, drug effects, and disease mechanisms.

[0045] The second culture fluid inlet 2111 is vertically arranged at the end of the first perfusion channel 21121 and is connected to the first perfusion channel 21121; this layout ensures that the culture fluid can flow from top to bottom, through the first perfusion channel 21121, and along the designed microfluidic channel 2112 to the HUVEC cell culture area 21131 uniformly and smoothly, simulating the natural flow of blood from blood vessels to the liver to transport nutrients and oxygen in the body, and providing cells with continuous fresh nutrient supply necessary for activity. The second culture fluid outlet 2114 is vertically arranged at the end of the third perfusion channel 21123, and the third perfusion channel 21123 is connected to the second culture fluid outlet 2114. This ensures that after the culture fluid is fully acted on and metabolized by the HUVEC cell culture area 21131 and the 3D cell culture chamber 21132, it carries the metabolites or drug reaction information, flows from bottom to top along the third perfusion channel 21123, and finally flows out through the second culture fluid outlet 2114. This design not only simulates the excretion process of metabolic waste in the body, but also facilitates the collection of samples for analysis, providing instant feedback for drug screening and toxicity assessment.

[0046] Through the above-mentioned precise layout, the entire perfusion channel system realizes a fully closed circulation of the culture fluid from the second culture fluid inlet 2111 to the HUVEC cell culture area 21131, then to the 3D cell culture chamber 21132, and finally to the second culture fluid outlet 2114, which not only ensures the steady state of the cell culture environment, but also facilitates the dynamic observation and control of the experiment, providing an efficient in vitro model for research.

[0047] Also included is a detachable hose 212, such as Figure 3 As shown. The detachable hose 212 is connected between two groups of serial microfluidic multi-chamber co-culture chips 211. The detachable hose 212 serves as a bridge connector, which tightly and flexibly connects the two groups of serial microfluidic multi-chamber co-culture chips 211. When in use, the detachable hose 212 connects the two serial microfluidic multi-chamber co-culture chips 211 through the first culture liquid inlet 11 and the first culture liquid outlet 12 of the perfusion layer 1. After the liquid flows out from the second culture liquid outlet 2114 of the first group of serial microfluidic multi-chamber co-culture chips 211, it flows into the second culture liquid inlet 2111 of the other group of serial microfluidic multi-chamber co-culture chips 211 through the detachable hose 212, and flows from the second culture liquid inlet 2111 to the second culture liquid outlet 2114 in the same flow area before flowing out; when the serial microfluidic multi-chamber co-culture chips 211 do not need to be connected, the detachable hose 212 can be removed.

[0048] This design not only maintains the continuity of the system in operation, making the exchange of liquids and nutrients, cell metabolites or drug test results between the two groups of chips unimpeded, but also enhances the overall operational flexibility of the system and the convenience of the experiment. The two groups of serial microfluidic multi-chamber co-culture chips 211 can be used in series or as two control groups and used separately.

[0049] Through the connection of the detachable hose 212, the user can easily realize the independent operation of the two sets of fluidic multi-chamber co-culture chips 211, such as staged drug treatment, observation of cell type differences, or specific environmental simulation, and can also ensure the rapid integration of the two sets of experimental data when necessary, conduct comparative analysis, verify drug efficacy, evaluate cell interactions, and even simulate disease models. The detachable hose 212 is made of non-toxic, good connection sealing, leak-proof, and high-temperature and high-pressure sterilization-resistant materials to ensure biocompatibility, ensure the sterile environment of the experiment, and ensure the accuracy and safety of the experiment.

[0050] Specifically, the microfluidic chip is 50 mm long, 25 mm wide and 8 mm high; the perfusion layer 1 is 2 mm high and is responsible for transporting and regulating the flow of culture fluid; the culture layer 2 is 6 mm high and provides sufficient growth space for cells. The combination of the two realizes the three-dimensional structure of the functional and fluid extracellular model.

[0051] Specifically, the HUVEC cell culture area 21131 is 15 mm long, 4 mm wide, and 5 mm high; this specification not only ensures that the HUVEC cells are fully spread, but also facilitates the simulation of the physiological functions of vascular endothelial cells. The 3D cell culture chamber 21132 is 9 mm in diameter and 5.5 mm in height; this three-dimensional spherical chamber not only provides a three-dimensional growth environment for cells and promotes natural interactions between cells, but also facilitates drug testing and the simulation of physiological reactions.

[0052] Specifically, the length of the first perfusion channel 21121 is 2.6 mm, the length of the second perfusion channel 21122 is 2.3 mm, and the length of the third perfusion channel 21123 is 3.1 mm. The width of the first perfusion channel 21121, the second perfusion channel 21122 and the third perfusion channel 21123 are all 0.5 mm, and the height is 1 mm. This ensures precise regulation of fluid dynamics, and effective control can be achieved from nutrient supply to waste removal to drug delivery. The channel size is optimized to reduce interference, minimize the impact on cells, and ensure the stability of the cell culture environment.

[0053] This design of microfluidic chip optimizes the cell growth environment through precise size and fluid channel settings, which not only simulates physiological conditions, but also provides an efficient and accurate in vitro platform for drug screening and disease model research. The precision of size and channel ensures the repeatability of the experiment and the reliability of the results.

[0054] Usage steps and operation process

[0055] 1. System pretreatment: cleaning and preparation of microfluidic chips, using culture medium to clean and soak multiple times to ensure no contamination and prepare the best state for cell culture;

[0056] 2. Cell inoculation: Inoculate the HUVEC cell culture area 21131 and the 3D cell culture chamber 21132, and introduce the cell suspension through the first culture medium inlet 11;

[0057] 3. Cycle start: introduce fresh culture fluid, flow through the first culture fluid inlet 11 and the second culture fluid inlet 2111 through the first perfusion channel 21121 to the HUVEC cell culture area 21131, then through the second perfusion channel 21122 to the 3D cell culture chamber 21132, and then flow out from the 3D cell culture chamber 21132, through the third perfusion channel 21123, the second culture fluid outlet 2114, and the first culture fluid outlet 12; if necessary, the liquid can be made to flow into the second culture fluid inlet 2111 of another set of serial microfluidic multi-chamber co-culture chips 211 through the detachable hose 212, and flow from the second culture fluid inlet 2111 to the second culture fluid outlet 2114 in the same flow area before flowing out.

[0058] 4. Observation and analysis: Using transparency, real-time monitoring of cell growth and morphological changes, collection of metabolites to analyze drug effects, and evaluation of cell function and drug response;

[0059] 5. Drug screening and modeling: Apply drugs through microfluidic channels 2112 to simulate in vivo drug effects, evaluate cell responses, and conduct efficient screening and disease mechanism research.

[0060] Although the utility model has been described in detail above with general descriptions and specific embodiments, it is obvious to those skilled in the art that some modifications or improvements can be made to the utility model. Therefore, these modifications or improvements made without departing from the spirit of the utility model are within the scope of protection claimed by the utility model.

Claims

1. A microfluidic chip for in vitro 3D liver organ co-culture, comprising a microfluidic chip, characterized in that: include: The perfusion layer (1) is provided with a first culture solution inlet (11) and a first culture solution outlet (12); and The culture layer (2) is provided with a circulating microfluidic perfusion culture system (21); The microfluidic chip is an upper and lower layer formed by bonding a perfusion layer (1) and a culture layer (2); The circulating microfluidic perfusion culture system (21) comprises two sets of serially connected microfluidic multi-chamber co-culture chips (211); The serial microfluidic multi-chamber co-culture chip (211) comprises a second culture solution inlet (2111), a microfluidic channel (2112), a composite cell co-culture area (2113) and a second culture solution outlet (2114); the second culture solution inlet (2111), the microfluidic channel (2112), the composite cell co-culture area (2113) and the second culture solution outlet (2114) are connected, and the second culture solution inlet (2111) is connected to the first culture solution inlet (11), and the second culture solution outlet (2114) is connected to the first culture solution outlet (12); The composite cell co-culture area (2113) includes a HUVEC cell culture area (21131) and a 3D cell culture chamber (21132), and the HUVEC cell culture area (21131) and the 3D cell culture chamber (21132) are connected via a microfluidic channel (2112).

2. The microfluidic chip for in vitro 3D liver organ co-culture according to claim 1, characterized in that: The microfluidic channel (2112) includes a first perfusion channel (21121), a second perfusion channel (21122) and a third perfusion channel (21123), wherein the first perfusion channel (21121) is arranged between the second culture fluid inlet (2111) and the HUVEC cell culture area (21131), the second perfusion channel (21122) is arranged between the HUVEC cell culture area (21131) and the 3D cell culture chamber (21132), and the third perfusion channel (21123) is arranged between the 3D cell culture chamber (21132) and the second culture fluid outlet (2114).

3. The microfluidic chip for in vitro 3D liver organ co-culture according to claim 2, characterized in that: The second culture fluid inlet (2111) is vertically arranged at the end of the first perfusion channel (21121) and is communicated with the first perfusion channel (21121); the second culture fluid outlet (2114) is vertically arranged at the end of the third perfusion channel (21123), and the third perfusion channel (21123) is communicated with the second culture fluid outlet (2114).

4. The in vitro 3D liver organ co-culture microfluidic chip according to claim 1, characterized in that: It also includes a detachable hose (212), which is connected between two groups of serially connected microfluidic multi-chamber co-culture chips (211).

5. The in vitro 3D liver organ co-culture microfluidic chip according to claim 1, characterized in that: The microfluidic chip has a length of 50 mm, a width of 25 mm and a height of 8 mm; the perfusion layer (1) is 2 mm high and the culture layer (2) is 6 mm high.

6. The in vitro 3D liver organ co-culture microfluidic chip according to claim 1, characterized in that: The HUVEC cell culture area (21131) has a length of 15 mm, a width of 4 mm, and a height of 5 mm; the 3D cell culture area has a diameter of 9 mm and a height of 5.5 mm.

7. The in vitro 3D liver organ co-culture microfluidic chip according to claim 3, characterized in that: The length of the first perfusion channel (21121) is 2.6 mm, the length of the second perfusion channel (21122) is 2.3 mm, the length of the third perfusion channel (21123) is 3.1 mm, and the widths of the first perfusion channel (21121), the second perfusion channel (21122) and the third perfusion channel (21123) are all 0.5 mm, and the heights are all 1 mm.

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