Bioreactor for culturing nerve cells

Abstract

The invention discloses a bioreactor for nerve cell culture, and relates to the technical field of cell culture, a metal ball falls down to drive a culture dish to deflect, so that an azimuth magnetic field is generated, multi-stage energy storage is performed on the metal ball, the metal ball is bounced off after the energy is stored to a certain degree, and the bounced-off metal ball falls down disorderly to drive the culture dish to deflect disorderly. Therefore, the magnetic field is generated disorderly, irregular magnetic field stimulation is generated on nerve cells in the culture dish, and on one hand, the basic problems of cell adherence, cell block mass precipitation, uniform distribution of nutrient substances and oxygen, removal of metabolic waste and the like are solved; on the other hand, a stirring structure is not arranged, so that the problem of damage to cells caused by traditional stirring is avoided; and finally, the nerve cells are stimulated by combining a magnetic field disordered triggering mode with disordered deflection of the culture dish, so that the initiative of cell movement such as nerve cell axon growth, synaptic remodeling, cell migration and the like is stimulated.

Description

Technical Field

[0001] The invention relates to the technical field of cell culture, in particular to a bioreactor for culturing nerve cells. Background Art

[0002] Cell culture refers to a technical method that simulates the in vivo environment in vitro, removes cells from organisms, and makes them grow, reproduce and maintain their structure and function under sterile, appropriate temperature, pH and certain nutritional conditions. In this process, a bioreactor is required to provide cells with an optimized and controllable growth environment to achieve efficient cell culture and product production.

[0003] For example, a high-efficiency bioreactor for cell culture with publication number CN118599660A proposes that some fragile cells require low-shear stirring, while some high-density cultured cells require stronger stirring to ensure material exchange.

[0004] In fact, stirring of cell culture is necessary for the following main reasons:

[0005] 1. Agitation is very important in large-scale cell culture, such as in bioreactors used to produce vaccines or biopharmaceuticals; it helps maintain the cells in suspension, ensures even distribution of nutrients and oxygen, and removes metabolic waste;

[0006] 2. When culturing some suspension cells, stirring is necessary to help evenly distribute nutrients and oxygen in the culture medium while preventing cell sedimentation and aggregation. This is usually achieved by using a shaker or rotating culture device in the culture vessel;

[0007] 3. During the three-dimensional neural cell culture process, stirring can help maintain the stability of the three-dimensional structure and prevent cell clumps from settling.

[0008] A further question that needs to be raised is that when nerve cells are in the human body, as the neurons work, each nerve cell will undergo uninterrupted axon growth, synaptic remodeling and cell migration, etc. This is mainly because the human body is rich in a variety of growth factors, such as nerve growth factor, brain-derived neurotrophic factor, fibroblast growth factor, etc., as well as cell adhesion molecules and extracellular matrix proteins that are involved in regulating and promoting the axon growth, synaptic remodeling and cell migration of nerve cells. However, when nerve cells are cultured in vitro, due to the lack of stimulation from neurons, nerve cells lack autonomous migration, and therefore can only rely on simple stirring to force passive migration of cells. In fact, there is still an active and passive difference between in vitro culture and in vivo culture. In general, although the survival rate of cells cultured in vitro has been relatively high in recent years, compared with the human body, nerve cells cultured in vitro still have the problem of low cell motility in many aspects such as axon growth, synaptic remodeling and cell migration. This is also a difficult problem that needs to be overcome in the in vitro nerve cell culture in the current related fields.

[0009] In view of the above problems, it is urgent to carry out innovative designs based on the original bioreactors used for neural cell culture. Summary of the invention

[0010] The technical solution of the present invention aims at the technical problem that the existing technical solution is too single, and provides a solution that is significantly different from the existing technology. Specifically, the purpose of the present invention is to provide a bioreactor for culturing nerve cells to solve the above-mentioned background technology that when nerve cells are in the human body, each nerve cell will carry out uninterrupted axon growth, synaptic remodeling and cell migration as the neurons work. This is mainly due to the fact that the human body is rich in various growth factors, such as nerve growth factor, brain-derived neurotrophic factor, fibroblast growth factor, etc., as well as cell adhesion molecules and extracellular matrix proteins, which will participate in regulating and promoting the axon growth, synaptic remodeling and cell migration of nerve cells. However, when nerve cells are cultured in vitro, due to the promotion of neurons, nerve cells lack autonomous migration, so they can only rely on simple stirring to force passive migration of cells. In fact, there is still a difference between active and passive in vitro culture and in vivo culture. In general, although the survival rate of cells cultured in vitro has been relatively high in recent years, compared with the human body, nerve cells cultured in vitro still have the problem of low cell motility in many aspects such as axon growth, synaptic remodeling and cell migration.

[0011] To achieve the above object, the present invention provides the following technical solution: a bioreactor for culturing neural cells, comprising a bottom plate, wherein the inside of the bottom plate is provided with a gravity sensing plate with a through groove at the center, a culture dish with a spherical connector at the lower end, and a top cover, and an annular groove is provided at the edge of the bottom plate;

[0012] A disorderly switching electromagnetic induction component is installed between the annular groove and the culture dish;

[0013] The gravity sensing disk is fixed on the lower surface of the culture dish, and a disordered deflection component is arranged inside the gravity sensing disk.

[0014] Preferably, a spherical connecting nozzle located inside the central through groove of the gravity sensing disk is fixed to the upper surface inside the chassis, and the spherical connecting piece at the lower end of the culture dish is rotatably connected to the spherical connecting nozzle, and the spherical connecting nozzle is made of plastic material.

[0015] Preferably, the disordered switching electromagnetic induction component includes an electromagnet, a conductive ring, an induction coil, a rigid conductive wire and a circuit breaker, and a plurality of electromagnets whose lower ends are fixed to the inner wall of the annular groove are distributed at equal angles inside the annular groove, and a conductive ring is installed on the inner wall of the annular groove, an induction coil is wound around the outer wall of the electromagnet, and the lower end of the induction coil is fixedly connected to the lower surface of the conductive ring, the lower end of the rigid conductive wire is fixed to the upper surface of the conductive ring, and a circuit breaker is connected to the upper end of the rigid conductive wire and the upper end of the induction coil.

[0016] Preferably, the induction coil and the rigid conductive wire are electrically connected to the positive and negative poles of the conductive ring respectively, and a closed loop formed by the induction coil, the circuit breaker and the rigid conductive wire is connected in series with the conductive ring.

[0017] Preferably, a plurality of turns of induction coils not wound around the outer wall of the electromagnet are reserved above the electromagnet, and a section of the rigid conductive wire close to the conductive ring is spirally processed.

[0018] Preferably, a circuit breaker pressure rod is slidably installed inside the circuit breaker, and one end of a spring is welded to the lower end of the circuit breaker, and the other end of the spring is fixed to the inner wall of the circuit breaker, the conductive core of the induction coil and the hard conductive wire are symmetrically exposed inside the circuit breaker, and the end of the conductive core is on the same horizontal plane as the inner wall of the circuit breaker, the circuit breaker pressure rod consists of a vertical rod and a power terminal, and the circuit breaker and the vertical rod are both made of insulating material, and the power terminal is made of conductive material, and the outer wall of the culture dish is fixed with a pressure ring corresponding to the upper end of the circuit breaker pressure rod.

[0019] Preferably, the disordered deflection assembly includes an elastic telescopic rod, an arc-shaped baffle and a metal ball, and a circular cavity is opened in the gravity sensing disk, and elastic telescopic rods are installed at equal angles inside the cavity, the telescopic ends of the elastic telescopic rods are provided with arc-shaped baffles, and several arc-shaped baffles form a small circular chamber, and a freely movable metal ball is provided in the chamber.

[0020] Preferably, a flexible rubber sheet adapted to the dislocation movement of the arc-shaped baffles is provided between two adjacent arc-shaped baffles.

[0021] Compared with the prior art, the present invention has the following beneficial effects:

[0022] The present invention proposes a special "stirring" method, in which the falling metal ball drives the culture dish to deflect, so that the azimuth magnetic field is generated and the metal ball is subjected to multi-level energy storage. After the energy is stored to a certain level, the metal ball is bounced off, and the bounced metal ball falls disorderly, driving the culture dish to deflect disorderly, thereby generating disorderly magnetic field, and generating irregular magnetic field stimulation on the nerve cells inside the culture dish. On the one hand, it solves the basic problems such as cell adhesion, cell agglomeration precipitation, and ensuring the uniform distribution of nutrients and oxygen and the removal of metabolic waste; on the other hand, no stirring structure is set, thereby avoiding the problem of damage to cells caused by traditional stirring; finally, the disordered triggering method of the magnetic field is combined with the disordered deflection of the culture dish to stimulate nerve cells, so as to stimulate the initiative of cell movement such as axon growth, synaptic remodeling and cell migration of nerve cells (the magnetic field promotes the proliferation of nerve cells by affecting the signal transduction pathway in the cell, and the specific magnetic field strength and frequency can increase the proliferation rate of neural stem cells and neural progenitor cells, and the magnetic field helps the process of neural stem cells differentiating into neurons; the magnetic field can improve the growth of axons and dendrites, promote the growth of axons and dendrites of nerve cells, and help form a more complex neural network structure). BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 It is a schematic diagram of the three-dimensional structure of the present invention;

[0024] Figure 2 It is a schematic diagram of the explosion structure of the present invention;

[0025] Figure 3 It is a schematic diagram of the three-dimensional structure of the chassis of the present invention;

[0026] Figure 4 This is a schematic diagram of the series distribution structure of the electromagnet and the conductive ring of the present invention;

[0027] Figure 5 It is a schematic diagram of the electromagnetic induction structure of the present invention;

[0028] Figure 6 It is a structural schematic diagram of the disordered deflection assembly of the present invention;

[0029] Figure 7 This is the schematic diagram of the front view section of the initial state of the present invention;

[0030] Figure 8 For the present invention Figure 7 The enlarged structural diagram at A in the middle;

[0031] Fig. 9 This is the principle intention of the front view section of the metal ball after secondary acceleration of the present invention;

[0032] Fig.10 For the present invention Fig. 9 The enlarged structural diagram at B in the middle;

[0033] Fig.11 This is the principle intention of the front view section of the metal ball after three accelerations of the present invention;

[0034] Fig.12 For the present invention Fig.11 The enlarged structural diagram at C in the middle;

[0035] Fig.13 This is the principle intention of the front view section of the metal ball after it is released;

[0036] Fig.14 For the present invention Fig.13 Enlarged structural diagram at D in the middle.

[0037] In the figure: 1. chassis; 11. spherical connecting nozzle; 2. electromagnet; 21. conductive ring; 22. induction coil; 23. rigid conductive wire; 3. circuit breaker; 31. circuit breaker pressure rod; 32. spring; 4. culture dish; 41. pressure ring; 5. gravity sensing disk; 6. elastic telescopic rod; 7. arc baffle; 8. metal ball. DETAILED DESCRIPTION

[0038] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments 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.

[0039] See also Figure 1-14 The present invention provides a technical solution: a bioreactor for culturing neural cells, comprising a bottom plate 1, wherein the inside of the bottom plate 1 is provided with a gravity sensing plate 5 with a through groove at the center, a culture dish 4 with a spherical connector at the lower end, and a top cover, and an annular groove is provided at the edge of the bottom plate 1;

[0040] A disorderly switching electromagnetic induction component is installed between the annular groove and the culture dish 4;

[0041] The gravity sensing disk 5 is fixed to the lower surface of the culture dish 4 , and a disordered deflection component is arranged inside the gravity sensing disk 5 .

[0042] A spherical connection nozzle 11 located inside the central through groove of the gravity sensing disk 5 is fixed to the upper surface inside the chassis 1, and the spherical connection piece at the lower end of the culture dish 4 is rotatably connected to the spherical connection nozzle 11, and the spherical connection nozzle 11 is made of plastic material.

[0043] The disorderly switching electromagnetic induction component includes an electromagnet 2, a conductive ring 21, an induction coil 22, a rigid conductive wire 23 and a circuit breaker 3, and a plurality of electromagnets 2 whose lower ends are fixed to the inner wall of the annular groove are distributed at equal angles inside the annular groove, and a conductive ring 21 is installed on the inner wall of the annular groove, an induction coil 22 is wound around the outer wall of the electromagnet 2, and the lower end of the induction coil 22 is fixedly connected to the lower surface of the conductive ring 21, the lower end of the rigid conductive wire 23 is fixed to the upper surface of the conductive ring 21, and the upper end of the rigid conductive wire 23 and the upper end of the induction coil 22 are connected to the circuit breaker 3.

[0044] The induction coil 22 and the rigid conductive wire 23 are electrically connected to the positive and negative electrodes of the conductive ring 21 respectively, and the closed loop formed by the induction coil 22 , the circuit breaker 3 and the rigid conductive wire 23 is connected in series with the conductive ring 21 .

[0045] A plurality of turns of the induction coil 22 not wound around the outer wall of the electromagnet 2 are reserved above the electromagnet 2, and a section of the rigid conductive wire 23 close to the conductive ring 21 is spirally processed.

[0046] A circuit breaker pressure rod 31 is slidably installed inside the circuit breaker 3, and one end of a spring 32 is welded to the lower end of the circuit breaker pressure rod 31, and the other end of the spring 32 is fixed to the inner wall of the circuit breaker 3. The conductive cores of the induction coil 22 and the hard conductive wire 23 are symmetrically exposed inside the circuit breaker 3, and the ends of the conductive cores are on the same horizontal plane as the inner wall of the circuit breaker 3. The circuit breaker 3 and the vertical rod are both made of insulating material, and the power terminal is made of conductive material. A pressure ring 41 corresponding to the upper end of the circuit breaker pressure rod 31 is fixed to the outer wall of the culture dish 4.

[0047] The disordered deflection component includes an elastic telescopic rod 6, an arc-shaped baffle 7 and a metal ball 8, and a circular ring-shaped cavity is opened in the gravity sensing disk 5, and the elastic telescopic rod 6 is installed at equal angles inside the cavity, and the telescopic ends of the elastic telescopic rod 6 are all provided with arc-shaped baffles 7, and several arc-shaped baffles 7 form a small circular ring-shaped chamber, and a freely movable metal ball 8 is arranged in the chamber.

[0048] A flexible rubber sheet adapted to the dislocation movement of the arc-shaped baffles 7 is arranged between two adjacent arc-shaped baffles 7 .

[0049] Working principle: When using this bioreactor for neural cell culture, first, Figure 1 , Figure 2 and Figure 7 As shown, the raw materials, nerve cells and culture fluid required for nerve cell culture are placed inside the culture dish 4 and covered with a lid. Then, the spherical connector at the lower end of the culture dish 4 is inserted into the spherical connecting nozzle 11 by pressing so that the culture dish 4 and the chassis 1 form a stable small-angle rotation connection.

[0050] like Figure 7 and Figure 8 As shown, since the metal ball 8 can move freely inside the gravity sensing disk 5, after the installation is completed according to the above steps, the metal ball 8 will slide down along a certain direction until it contacts the arc baffle 7 at the corresponding direction. The sliding metal ball 8 will initially compress the corresponding elastic telescopic rod 6 through the arc baffle 7 under the acceleration of gravity.

[0051] Then, if Fig. 9 and Fig.10 As shown, the sliding of the metal ball 8 will drive the culture dish 4 to tilt toward the direction where the metal ball 8 is located and drive the culture liquid inside the culture dish 4 to flow at a small angle. At this time, the tilted culture dish 4 will drive the pressure ring 41 on the outer wall to contact and squeeze the circuit breaker pressure rod 31 at the corresponding direction, so that the power terminal compression spring 32 at the lower end of the circuit breaker pressure rod 31 falls and just connects the hard conductive wire 23 and the induction coil 22. Once the hard conductive wire 23 and the induction coil 22 are connected, a closed loop with stable power supply is formed between the hard conductive wire 23, the conductive ring 21 and the induction coil 22, and the electromagnet 2 generates an induced magnetic field. When the electromagnet 2 generates an induced magnetic field, it will generate additional attraction to the metal ball 8, so that the metal ball 8 further compresses the elastic telescopic rod 6 through the arc baffle 7 to store energy;

[0052] like Fig.11 and Fig.12 As shown, as the metal ball 8 further approaches the side of the culture dish 4, the weight of the tilted side of the culture dish 4 is further increased, so the pressure of the pressure ring 41 on the circuit breaker pressure rod 31 increases, and the resistance of the compressed spring 32 is greater than the elastic resistance of the hard conductive wire 23 and the induction coil 22. Therefore, when the pressure ring 41 further tilts with the culture dish 4, the circuit breaker pressure rod 31 and the circuit breaker 3 will remain relatively still and fall and compress the hard conductive wire 23 and the induction coil 22 synchronously, so that the spiral end of the hard conductive wire 23 is adaptively contracted, and at the same time, the coil of the induction coil 22 reserved above the electromagnet 2 is gradually sheathed on the outer wall of the electromagnet 2;

[0053] Finally, if Fig.13 and Fig.14As shown in the figure, when the coil reserved by the induction coil 22 above the electromagnet 2 is gradually sleeved on the outer wall of the electromagnet 2, the number of turns of the coil on the outer wall of the electromagnet 2 increases, and the current remains unchanged, that is, the induced magnetic field generated on the electromagnet 2 gradually increases. At this time, the attraction of the electromagnet 2 on the metal ball 8 is further increased, prompting the metal ball 8 to further compress the elastic telescopic rod 6 to store energy. Since the metal ball 8 further approaches the side of the culture dish 4, the weight of the inclined side of the culture dish 4 is further increased, so the pressure of the pressure ring 41 on the circuit breaker pressure rod 31 increases again. Since the hard conductive wire 23 and the induction coil 22 are gradually compressed, their resistance gradually becomes greater than the resistance of the compressed spring 32. At this time, the culture dish 4 will press the circuit breaker pressure rod 31 again through the pressure ring 41, so that the circuit breaker pressure rod 31 is as shown in the figure. Fig.14 As shown, the metal ball 8 is quickly ejected by overcoming the resistance of the spring 32 and breaking the closed loop between the induction coil 22 and the hard conductive wire 23. When the closed loop is broken, the magnetic field disappears instantly, and the elastic member inside the elastic telescopic rod 6 quickly releases energy to quickly eject the metal ball 8 through the arc baffle 7. Since the metal ball 8 does not eject along a specific trajectory in the annular cavity of the gravity sensing disk 5, the ejected metal ball 8 will move disorderly due to various factors such as the deflection of the culture dish 4 and the friction resistance of the annular cavity. After being ejected, the disorderly moving metal ball 8 will always fall at a certain position in the annular cavity, and will deflect the culture dish 4 again at the azimuth angle corresponding to the metal ball 8 according to the above steps, and this cycle will continue.

[0054] In simple terms, the falling metal ball 8 drives the culture dish 4 to deflect, thereby generating a magnetic field in that direction and performing multi-level energy storage on the metal ball 8. After the energy is stored to a certain level, the metal ball 8 is bounced off. The bounced metal ball 8 falls disorderly, driving the culture dish 4 to deflect disorderly, thereby generating disorderly magnetic fields and generating irregular magnetic field stimulation on the nerve cells inside the culture dish 4, so as to improve the growth of axons and dendrites, promote the growth of axons and dendrites of nerve cells, and promote the active migration of nerve cells. During this period, the culture dish 4 will deflect disorderly, and the deflection angle will be between 3°±0.5°. The disordered deflection frequency can be adjusted based on the cycle of the disordered switching electromagnetic induction component, so that the fluid inside the culture dish 4 will oscillate as a whole at a small angle, achieving a special stirring effect for the cell culture medium, and incidentally solving the basic problems of cell adhesion, cell clump precipitation, and ensuring the uniform distribution of nutrients and oxygen and the removal of metabolic waste, and effectively avoiding the cell damage problem caused by traditional stirring.

[0055] Although the present invention has been described in detail with reference to the aforementioned embodiments, it is still possible for those skilled in the art to modify the technical solutions described in the aforementioned embodiments, or to make equivalent substitutions for some of the technical features therein. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A bioreactor for culturing neural cells, comprising a chassis (1), characterized in that: The interior of the chassis (1) is provided with a gravity sensing plate (5) with a through groove at the center, a culture dish (4) with a spherical connector at the lower end, and a top cover in order from bottom to top, and an annular groove is provided at the edge of the chassis (1); A disorderly switching electromagnetic induction component is installed between the annular groove and the culture dish (4); The gravity sensing disk (5) is fixed to the lower surface of the culture dish (4), and a disordered deflection component is arranged inside the gravity sensing disk (5).

2. A bioreactor for culturing neural cells according to claim 1, characterized in that: A spherical connection nozzle (11) located inside the central through groove of the gravity sensing disk (5) is fixed on the upper surface inside the chassis (1), and a spherical connection piece at the lower end of the culture dish (4) is rotatably connected to the spherical connection nozzle (11), and the spherical connection nozzle (11) is made of plastic material.

3. A bioreactor for culturing neural cells according to claim 1, characterized in that: The disorderly switching electromagnetic induction component comprises an electromagnet (2), a conductive ring (21), an induction coil (22), a rigid conductive wire (23) and a circuit breaker (3), and a plurality of electromagnets (2) whose lower ends are fixed to the inner wall of the annular groove are distributed at equal angles inside the annular groove, and a conductive ring (21) is installed on the inner wall of the annular groove, an induction coil (22) is wound around the outer wall of the electromagnet (2), and the lower end of the induction coil (22) is fixedly connected to the lower surface of the conductive ring (21), the lower end of the rigid conductive wire (23) is fixed to the upper surface of the conductive ring (21), and the upper end of the rigid conductive wire (23) and the upper end of the induction coil (22) are connected to the circuit breaker (3).

4. A bioreactor for culturing neural cells according to claim 3, characterized in that: The induction coil (22) and the rigid conductive wire (23) are electrically connected to the positive and negative poles of the conductive ring (21) respectively, and a closed loop formed by the induction coil (22), the circuit breaker (3) and the rigid conductive wire (23) is connected in series with the conductive ring (21).

5. A bioreactor for culturing neural cells according to claim 3, characterized in that: A plurality of turns of an induction coil (22) not wound around the outer wall of the electromagnet (2) are reserved above the electromagnet (2), and a section of the hard conductive wire (23) close to the conductive ring (21) is spirally processed.

6. A bioreactor for culturing neural cells according to claim 3, characterized in that: A circuit breaker pressure rod (31) is slidably installed inside the circuit breaker (3), and one end of a spring (32) is welded to the lower end of the circuit breaker pressure rod (31), and the other end of the spring (32) is fixed to the inner wall of the circuit breaker (3). The conductive cores of the induction coil (22) and the hard conductive wire (23) are symmetrically exposed inside the circuit breaker (3), and the ends of the conductive cores are in the same horizontal plane as the inner wall of the circuit breaker (3). The circuit breaker pressure rod (31) consists of a vertical rod and a power terminal. The circuit breaker (3) and the vertical rod are both made of insulating material, and the power terminal is made of conductive material. A pressure ring (41) corresponding to the upper end of the circuit breaker pressure rod (31) is fixed to the outer wall of the culture dish (4).

7. A bioreactor for culturing neural cells according to claim 1, characterized in that: The disordered deflection component comprises an elastic telescopic rod (6), an arc-shaped baffle (7) and a metal ball (8), and a gravity sensing disk (5) is provided with an annular cavity, and the elastic telescopic rods (6) are installed at equal angles inside the cavity, and the telescopic ends of the elastic telescopic rods (6) are all provided with arc-shaped baffles (7), and a plurality of arc-shaped baffles (7) surround a small annular chamber, and a freely movable metal ball (8) is provided in the chamber.

8. A bioreactor for culturing neural cells according to claim 7, characterized in that: A flexible rubber sheet adapted to the dislocation movement of the arc-shaped baffles (7) is provided between two adjacent arc-shaped baffles (7).

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