Organ chip for in vitro organoid-carc-t cell interaction study

Abstract

The utility model discloses a kind of organ chip for in vitro organoids and CAR-T cell interaction research, it is related to organ chip technical field, including bottom culture layer, cell culture layer and CAR-T cell storage layer, the upside of bottom culture layer is provided with cell culture layer, and the upside of cell culture layer is provided with cell storage layer, the inside of bottom culture layer is fixed with arrangement frame at equal distance, and the inside of arrangement frame is provided with culture micropore at equal distance.The organ chip for in vitro organoids and CAR-T cell interaction research, the construction of organoid research chip can be realized by the packaging of bottom culture layer, cell culture layer and CAR-T cell storage layer three-layer structure, only the culture micro-hole corresponding position of cell culture layer and bottom culture layer is cultured vascular endothelial cell on the basis of reticular structure, can be realized upper CAR-T cell penetration into bottom culture layer and in vitro organoids contact.

Description

Technical Field

[0001] This invention relates to the field of organ-on-a-chip technology, specifically to an organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro. Background Technology

[0002] Animal models have been used to test drug efficacy and safety, but it is a time-consuming and expensive method, and species-specific differences inevitably exist between animals and humans, meaning animal models cannot completely replace humans. In addition, ethical issues surrounding animal models have greatly limited their application in preclinical drug analysis in recent years. Organ-on-a-chip technology is an emerging scientific technology that integrates biological, chemical, and engineering laboratories to simulate and construct the microenvironment of tissues and organs in vitro, and has wide applications in disease simulation and new drug development. Developing tumor chips using organ-on-a-chip technology can provide new strategies for studying the occurrence, development, drug resistance, and development of novel tumor drugs.

[0003] Existing organ-on-a-chip systems are generally not convenient for setting up multiple groups to run simultaneously as needed, and cannot simultaneously conduct multi-effect target ratio experiments to reduce a large number of repetitive operations and space requirements, which in turn makes them prone to human error. Utility Model Content

[0004] The purpose of this invention is to provide an organ-on-a-chip for in vitro research on the interaction between organoids and CAR-T cells, in order to solve the problems mentioned in the background art.

[0005] To achieve the above objectives, this utility model provides the following technical solution: an organ-on-a-chip for in vitro organoid and CAR-T cell interaction research, comprising a bottom culture layer, a cell culture layer and a CAR-T cell storage layer, wherein a cell culture layer is disposed on the upper side of the bottom culture layer and a cell storage layer is disposed on the upper side of the cell culture layer, and an arrangement frame is fixed at equal intervals inside the bottom culture layer, and culture microwells are disposed at equal intervals inside the arrangement frame, and each row of culture microwells is interconnected by a connecting tube.

[0006] The leftmost culture microwell is connected to a liquid inlet tube, and a liquid inlet hole is provided on the outside of the liquid inlet tube.

[0007] The rightmost culture microwell is connected to a drain pipe, and a waste liquid hole is provided on the outside of the drain pipe.

[0008] The inlet tube, culture micropores, and outlet tube are interconnected, and both the inlet tube and outlet tube are microchannels.

[0009] The cell culture layer has a mesh structure, and the interior of the CAR-T cell storage layer is engraved with branched microchannels that mimic a capillary network.

[0010] The branched microchannels of the CAR-T cell storage layer contain a cell suspension of CAR-T cells, and medical-grade silicone tubes are connected to both sides of the outer side of the CAR-T cell storage layer.

[0011] This invention provides an organ-on-a-chip for in vitro research on the interaction between organoids and CAR-T cells, which has the following beneficial effects: This invention features a CAR-T cell storage layer. The construction of an organoid research chip is achieved through a three-layer encapsulation structure consisting of a bottom culture layer, a cell culture layer, and a CAR-T cell storage layer. The cell culture layer cultivates vascular endothelial cells on a mesh-like structure only at positions corresponding to the culture micropores of the bottom culture layer. This allows the upper CAR-T cells to penetrate into the bottom culture layer and contact the in vitro organoids, enabling specific targeting and killing of target tumor cells. The CAR-T cell storage layer contains branched microchannels etched inside, mimicking a capillary network, containing a suspension of CAR-T cells, allowing for fluid flow that mimics the flow of blood in the body.

[0012] This invention features a bottom culture layer, and multiple replicates of culture microwells can be set simultaneously to conduct multiple replicate experiments under the same conditions, reducing human error. Furthermore, multiple sets of arrangement frames can simultaneously culture various tumor organoids, enabling large-scale studies on the interaction between different tumor organoids and different CAR-T cells. Moreover, it can simultaneously conduct multi-effect-to-target ratio experiments, reducing a large number of repetitive operations and space requirements. Culture medium can be injected into several culture microwells through inlet holes and inlet tubes. Attached Figure Description

[0013] Figure 1 This is a schematic diagram of the exploded structure of an organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, according to this utility model. Figure 2 This is a schematic diagram of the overall structure of an organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, according to this utility model. Figure 3 This is a schematic diagram of the bottom culture layer structure of an organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, according to the present invention. Figure 4 This is a schematic diagram of the arrangement frame structure of an organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, according to the present invention.

[0014] In the diagram: 1. Bottom culture layer; 2. Arrangement frame; 3. Culture microwell; 4. Connecting tube; 5. Liquid inlet tube; 6. Liquid inlet hole; 7. Liquid outlet tube; 8. Waste liquid hole; 9. Cell culture layer; 10. CAR-T cell storage layer; 11. Medical-grade silicone tube. Detailed Implementation

[0015] The embodiments of this utility model will be described in further detail below with reference to the accompanying drawings and examples. The following examples are for illustrative purposes only and should not be construed as limiting the scope of this utility model.

[0016] like Figures 1-2 As shown, an organ-on-a-chip for in vitro organoid and CAR-T cell interaction research includes a bottom culture layer 1, a cell culture layer 9, and a CAR-T cell storage layer 10. The cell culture layer 9 is positioned above the bottom culture layer 1, and the cell storage layer 10 is positioned above the cell culture layer 9. The cell culture layer 9 has a mesh structure. The CAR-T cell storage layer 10 has branched microchannels etched inside, mimicking a capillary network. These branched microchannels contain a suspension of CAR-T cells. Medical-grade silicone tubes 11 are connected to both sides of the CAR-T cell storage layer 10. The organoid research chip can be constructed through the encapsulation of this three-layer structure: the bottom culture layer 1, the cell culture layer 9, and the CAR-T cell storage layer 10. Vascular endothelial cells are cultured on a mesh structure only at the positions corresponding to the culture micropores 3 of the bottom culture layer 1. This allows the upper CAR-T cells to penetrate into the bottom culture layer 1 and come into contact with organoids in vitro, killing tumor cells that can be specifically targeted. The CAR-T cell storage layer 10 has branched microchannels etched inside to simulate a capillary network, containing a cell suspension of CAR-T cells. The outer sides of the CAR-T cell storage layer 10 are connected to CAR-T suspension storage bags through medical-grade silicone tubes 11. Applying a pulsed air pressure of 0.08-0.12 MPa to the storage bag can push the suspension to flow along the tube at a speed that simulates aortic blood flow. The flow rate is linearly adjusted by the air pressure intensity. Conversely, switching to a low-pressure reflux chamber allows some of the suspension to flow back to simulate venous return, thereby making the liquid flow and mimicking the flow of blood in the body.

[0017] like Figure 1 , Figure 3 and Figure 4As shown, the bottom culture layer 1 has equidistant fixed arrangement frames 2, and the arrangement frames 2 have equidistant culture microwells 3. Each row of culture microwells 3 is interconnected by connecting tubes 4. The leftmost culture microwell 3 is connected to the left side of an inlet tube 5, and an inlet hole 6 is provided on the outside of the inlet tube 5. The rightmost culture microwell 3 is connected to the right side of an outlet tube 7, and a waste liquid hole 8 is provided on the outside of the outlet tube 7. The inlet tube 5, culture microwell 3, and outlet tube 7 are interconnected, and both the inlet tube 5 and outlet tube 7 are microchannels. Multiple replicates of culture microwells 3 can be set at the same time to conduct multiple replicate experiments under the same conditions, reducing human error. Multiple arrangement frames 2 can simultaneously culture multiple tumor organoids, conduct a large number of studies on the interaction between different tumor organoids and different CAR-T cells, and conduct multi-effect ratio experiments at the same time, reducing a large number of repetitive operations and space requirements. Culture medium can be injected into several culture microwells 3 through the inlet hole 6 and the inlet tube 5.

[0018] In summary, as Figures 1-4 As shown, this organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro can be constructed by first encapsulating a three-layer structure consisting of a bottom culture layer 1, a cell culture layer 9, and a CAR-T cell storage layer 10. The cell culture layer 9 cultured vascular endothelial cells on a mesh structure at the positions corresponding to the culture micropores 3 of the bottom culture layer 1, which allows the upper CAR-T cells to penetrate into the bottom culture layer 1 and come into contact with the in vitro organoids, killing tumor cells that can be specifically targeted. The CAR-T cell storage layer 10 has branched microchannels inside that simulate a capillary network, containing a cell suspension of CAR-T cells. The outer sides of the CAR-T cell storage layer 10 are connected to CAR-T suspension storage bags via medical-grade silicone tubes 11. Applying a pulsed air pressure of 0.08-0.12 MPa to the storage bag can push the suspension to flow along the tubing at a speed that simulates aortic blood flow. The flow rate is linearly adjusted by the air pressure intensity. Conversely, switching to a low-pressure return chamber allows some of the suspension to return to simulate venous return, thereby making the fluid flow and mimicking the flow of blood in the body. During use, microfluidic-linked 3D printing technology is used to extract cells from the target tissue derived from the patient. Then, microfluidic microdroplet encapsulation technology is used to prepare high-fidelity organoids with an immune cell microenvironment. Organoid microspheres are added to each microwell. Culture medium can be injected into several culture microwells 3 through the inlet hole 6 and the inlet tube 5. The injection is stopped immediately after the solution flows through the last microwell. After a suitable time, the medium is renewed by perfusion. Initially, vascular endothelial cells in cell culture layer 9 and organoids in the bottom culture layer 1 are cultured simultaneously. After the experimental material organoids and vascular endothelial cells are successfully cultured, cell suspension is infused into the CAR-T cell storage layer for subsequent experiments and observation. This completes the use of the organ-on-a-chip for in vitro research on the interaction between organoids and CAR-T cells.

[0019] The embodiments of this utility model are given for illustrative and descriptive purposes only, and are not intended to be exhaustive or to limit the utility model to the forms disclosed. Many modifications and variations will be apparent to those skilled in the art. The embodiments were chosen and described in order to better illustrate the principles and practical applications of this utility model, and to enable those skilled in the art to understand this utility model and design various embodiments with various modifications suitable for a particular purpose.

Claims

1. An organ-on-a-chip for in vitro study of organoid-CAR-T cell interaction, comprising a base culture layer (1), a cell culture layer (9), and a CAR-T cell storage layer (10), characterized in that, A cell culture layer (9) is provided on the upper side of the bottom culture layer (1), and a cell storage layer (10) is provided on the upper side of the cell culture layer (9). An arrangement frame (2) is fixed at equal intervals inside the bottom culture layer (1), and a culture micropore (3) is provided at equal intervals inside the arrangement frame (2). Each row of culture micropores (3) is interconnected through a connecting tube (4).

2. The organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, as described in claim 1, is characterized in that... The leftmost culture micropore (3) is connected to an inlet pipe (5), and an inlet hole (6) is provided on the outside of the inlet pipe (5).

3. An organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, as described in claim 2, is characterized in that... The right side of the culture microwell (3) is connected to a drain pipe (7), and a waste liquid hole (8) is provided on the outside of the drain pipe (7).

4. An organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, as described in claim 3, is characterized in that... The liquid inlet tube (5), culture micropore (3), and liquid outlet tube (7) are interconnected, and both the liquid inlet tube (5) and the liquid outlet tube (7) are microchannels.

5. An organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, as described in claim 1, characterized in that, The cell culture layer (9) has a mesh structure, and the CAR-T cell storage layer (10) has branched microchannels inside that simulate a capillary network.

6. An organ-on-a-chip for studying the interaction between organoids and CAR-T cells in vitro, as described in claim 1, is characterized in that... The branched microchannels of the CAR-T cell storage layer (10) contain a cell suspension of CAR-T cells, and medical-grade silicone tubes (11) are connected to both sides of the CAR-T cell storage layer (10).

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