Muscle tissue electricity-force co-stimulation system inspired by human muscle training and control method

Abstract

The invention provides a muscle tissue electricity-force co-stimulation system inspired by human muscle training and a control method. A uniform periodic electric field is generated through the annularly distributed multiple electrodes to stimulate muscle tissue contraction, meanwhile, the inherent elasticity of the spring structure provides load for engineering skeletal muscle tissue, dynamic mechanical stimulation is further provided, and by combining physical analysis software, dynamic mechanical stimulation is provided for the engineering skeletal muscle tissue. The centripetal contraction dynamics of the engineering skeletal muscle tissue can be monitored in real time by using the spring with the known elastic coefficient. The position of the tail end of the spring is dynamically adjusted through the micro servo electric cylinder to adjust the length of the engineering skeletal muscle tissue, so that a variable load is provided in the growth and development process of the muscle tissue, and skeleton extension in the growth process of human skeletal muscles is simulated. According to the invention, electrical stimulation and dynamic mechanical stimulation of the engineering skeletal muscle tissue can be realized at the same time, the centripetal contraction force of the engineering skeletal muscle tissue can be monitored in real time, and the mechanical load applied to the muscle tissue can be dynamically adjusted.

Description

Technical Field

[0001] The present invention belongs to the field of biomedical engineering, and specifically relates to a system inspired by the training model of human skeletal muscle tissue, which can electrically stimulate and dynamically mechanically stimulate engineered skeletal muscle tissue. The system is mainly used to promote myoblast fusion and differentiation and improve the maturity of skeletal muscle tissue. Background Art

[0002] Skeletal muscle is one of the most important motor units in the human body. There are more than 600 skeletal muscles in the human body, which are driven synergistically to enable the human body to perform complex and delicate movements. In addition, skeletal muscle plays an important role in maintaining body temperature and metabolic balance. With the rapid development of science and technology, researchers have explored the reconstruction of skeletal muscle tissue in vitro. This research has important practical significance in many fields. Such as skeletal muscle physiology, skeletal muscle disease modeling, drug development and testing, and biohybrid robots. However, due to the very complex multi-level structure of human skeletal muscle and the lack of relevant in vitro research methods, the current skeletal muscle tissue faces problems such as low in vitro differentiation rate.

[0003] In order to promote the growth and differentiation of muscle cells and improve the maturity of engineered skeletal muscle tissue, researchers have conducted a number of studies. The relevant results show that mechanical stretching stimulation can promote the directional arrangement of the cytoskeleton, protein synthesis and muscle cell hypertrophy in muscle tissue. On the other hand, electrical pulse stimulation is necessary for the normal growth and maturation of new fibers. Appropriate electrical stimulation helps the transformation of myoblasts into contractile myotubes, thereby improving the differentiation efficiency and driving force of skeletal muscle tissue in vitro.

[0004] Although the prior art has proposed some in vitro skeletal muscle tissue culture methods based on electrical or mechanical stimulation, they cannot effectively simulate the culture environment of human muscle tissue, so the degree of differentiation of engineered skeletal muscle tissue is still far lower than that of natural skeletal muscle tissue. In the human body, from embryo to adulthood, skeletal muscle tissue is always stimulated by a combination of neural electrical stimulation and dynamic mechanical stimulation, and its performance is improved or maintained. Therefore, the development of a new system that can co-stimulate in vitro skeletal muscle tissue will help promote further improvement in the performance of in vitro skeletal muscle tissue. Promote the further development of artificial skeletal muscle tissue research in the fields of skeletal muscle physiology, disease modeling, drug development and testing, and biohybrid robots. Summary of the invention

[0005] In view of the problem that the current stimulation methods are single and have poor effects, the purpose of the present invention is to provide an electrical-mechanical co-stimulation system for enhancing the performance of engineered skeletal muscle tissue. A periodic electric field is generated by annularly distributed multiple electrodes to cause muscle tissue contraction, and dynamic mechanical stimulation is provided by springs, thereby promoting myoblast differentiation and improving muscle tissue maturity and contractility.

[0006] The present invention relates to an electrical-mechanical co-stimulation system for enhancing the performance of engineered skeletal muscle tissue and a control method thereof, which can simultaneously perform electrical and dynamic mechanical co-stimulation and real-time centripetal contractile force measurement on engineered skeletal muscle tissue, can improve the efficiency of myoblast differentiation, and improve the maturity and contractile force of muscle tissue. Since the present invention can effectively improve the maturity of engineered skeletal muscle tissue, increase its contractile force, and measure the centripetal contractile force of muscle tissue, it has important application value in research fields such as skeletal muscle tissue engineering, drug screening and testing, contractile dynamics measurement, and biohybrid robots.

[0007] The present invention is realized by the following technical scheme: a muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training, comprising:

[0008] The electrodes are distributed in a ring shape within the edge of the culture dish and are connected to the multi-channel electrical stimulator to generate periodic electrical pulse stimulation acting on the muscle tissue through the periodic electrical pulse signal emitted by the multi-channel electrical stimulator;

[0009] A spring, the movable end of which is connected to the muscle tissue, and the fixed end of which is connected to the servo electric cylinder through a connector, for acting on the muscle tissue to generate dynamic mechanical stimulation;

[0010] The servo electric cylinder is connected to the host computer through a controller, and is used to realize the extension and retraction control of the servo electric cylinder according to the instructions of the host computer to drive the spring to move.

[0011] The electrode is a platinum sheet with a length of 15 mm, a width of 5 mm and a thickness of 0.1 mm. The electrodes are multiple and evenly distributed.

[0012] The spring is made of PDMS elastic material, has 3 turns, a spring pitch of 1.5 mm, a spring wire width of 0.8 mm, a spring wire thickness of 0.5 mm, and a spring elastic coefficient of 2.1 mN / mm.

[0013] One end of the muscle tissue is connected to the movable end of the spring, and the other end is connected to the micro-column fixed on the surface of the culture dish.

[0014] A control method of a muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training comprises the following steps:

[0015] Fix the muscle tissue to be tested on the micro-pillars on the surface of the culture dish and the active end of the spring;

[0016] During the growth cycle of muscle tissue, the muscle tissue drives the spring to move, and monitors its passive contraction force and centripetal contraction force. The servo electric cylinder adjusts the spring position in real time, thereby providing a variable load for the muscle tissue to ensure that the length of the muscle tissue remains unchanged.

[0017] The muscle tissue is cultured by electrical-mechanical co-stimulation; when the muscle tissue is electrically stimulated, the active end of the spring is driven to move periodically, and is subjected to periodic dynamic mechanical stimulation provided by the spring.

[0018] The passive contraction force and the centripetal contraction force are monitored, and the spring position is adjusted in real time by the servo electric cylinder, including the following steps:

[0019] In order to keep the length of the muscle tissue unchanged, the spring is lengthened by the servo electric cylinder to offset the passive contraction force that will shorten the length of the muscle tissue. The distance the spring is stretched is identified by visual images. At this time, the product of the distance the spring is stretched and the elastic coefficient is the passive contraction force.

[0020] The muscle tissue is subjected to electrical-mechanical co-stimulation culture as follows: the electrical stimulation frequency is 1 Hz, the electric field strength is 1.5 V / cm, the pulse width is 6 ms, stimulation is 7.5 minutes per hour, rest is 52.5 minutes, and the total stimulation is three days.

[0021] When the muscle tissue is electrically stimulated, the active end of the spring is driven to move periodically, and is subjected to periodic dynamic mechanical stimulation provided by the spring, including the following steps:

[0022] During electrical stimulation, the muscle tissue contracts to overcome the resistance of the spring, and the distance the spring is dynamically stretched is identified through visual images. At this time, the product of the distance the spring is stretched and the elastic coefficient is the centripetal contraction force.

[0023] The acquisition of muscle tissue comprises the following steps:

[0024] The matrix gel, collagen solution, NaOH solution, mouse C2C12 myoblasts and cell growth medium were fully mixed and injected into the muscle tissue culture mold;

[0025] On the fifth day of culture, when the muscle tissue showed spontaneous contraction, it was transferred to a culture dish.

[0026] The present invention produces the following beneficial effects and advantages:

[0027] 1. The electric-mechanical co-stimulation system for enhancing the performance of engineered skeletal muscle tissue described in the present invention stimulates muscle tissue contraction through a periodic electric field generated by a ring-shaped multi-electrode distribution, and uses the inherent elastic force of the spring to provide dynamic mechanical stimulation, and uses the spring to monitor the changes in the contractile force of the muscle tissue in real time, and uses a micro-servo electric cylinder to dynamically adjust the position of the active end of the spring, thereby providing a dynamically variable load for the muscle tissue. When the muscle tissue is electrically stimulated, it contracts and overcomes the spring resistance to drive the end of the spring to move. In this process, the muscle tissue is simultaneously electrically stimulated and dynamically mechanically stimulated, and its differentiation efficiency and driving performance are significantly improved.

[0028] 2. The present invention does not require complex mechanical systems and control methods. After electrical stimulation, the inherent elastic force of the spring can automatically achieve dynamic mechanical stimulation of muscle tissue. The invention can simplify the muscle tissue stimulation training process, and the spring structure can also be used for online real-time detection of muscle tissue contraction force without the need for additional contraction force characterization methods. The present invention is expected to provide new potential approaches for the study of skeletal muscle tissue in vitro models, drug testing, disease modeling, contraction dynamics measurement, and biohybrid robots based on skeletal muscle tissue. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] Figure 1 It is a schematic diagram of the system structure principle of the present invention.

[0030] Figure 2 It is an enlarged schematic diagram of the muscle tissue and spring part of the present invention.

[0031] Figure 3 The fluorescent staining images of myotubes of muscle tissue cultured by stimulation of the present invention are shown in Figure a). The blue part in Figure b) is the fluorescent image of cell nuclei, the green part in Figure c) is the fluorescent image of myosin heavy chain, and Figure c) is the merged image of Figure a) and Figure b).

[0032] Figure 4 The time-displacement curves of the engineered skeletal muscle tissue of the present invention driving springs with different elastic coefficients are shown in Figure a), wherein the time-displacement curve of the muscle tissue driving the spring with an elastic coefficient of 0.7 mN / mm, the time-displacement curve of the muscle tissue driving the spring with an elastic coefficient of 2.1 mN / mm, and the time-displacement curve of the muscle tissue driving the spring with an elastic coefficient of 3.9 mN / mm.

[0033] The meanings of the various marks in the accompanying drawings are: 1-micro servo cylinder; 2-connector; 3-ring-distributed multi-electrodes; 4-PDMS spring structure; 5-engineered skeletal muscle tissue; 6-standard culture dish with a diameter of 100 mm. DETAILED DESCRIPTION

[0034] The present invention is further described in detail below with reference to the accompanying drawings and examples.

[0035] The present invention is inspired by the human skeletal muscle training mode and provides an electrical-mechanical co-stimulation system and a control method for enhancing the performance of engineered skeletal muscle tissue. Through the multi-electrodes distributed in an annular pattern, a uniform periodic electric field is generated to stimulate the contraction of muscle tissue, while the inherent elasticity of the spring structure provides a load for the engineered skeletal muscle tissue, thereby providing dynamic mechanical stimulation, and the use of a spring with a known elastic coefficient can also monitor the centripetal contraction dynamics of the engineered skeletal muscle tissue in real time. The end position of the spring is dynamically adjusted by a micro-servo electric cylinder to adjust the length of the engineered skeletal muscle tissue, thereby providing a variable load during the growth and development of the muscle tissue, simulating the bone elongation during the growth of human skeletal muscle. The present invention can simultaneously realize electrical stimulation and dynamic mechanical stimulation of the engineered skeletal muscle tissue, and can monitor its centripetal contraction force in real time, and dynamically adjust the mechanical load applied to the muscle tissue. The present invention can significantly promote myoblast differentiation and muscle tissue maturation, and has important potential application value in the research fields of skeletal muscle tissue in vitro models, drug testing, disease modeling, contraction dynamics measurement, and biohybrid robots based on skeletal muscle tissue.

[0036] like Figure 1 , Figure 2 As shown, the present invention first provides a system for electrical-mechanical co-stimulation of engineered skeletal muscle tissue, comprising: a ring-distributed multi-electrode, a micro-servo electric cylinder, and a PDMS spring;

[0037] The annular distributed multi-electrodes are adhered to the edge of a standard culture dish by biocompatible glue and connected to a multi-channel electrical stimulator; the annular distributed multi-electrodes are 8 metal platinum sheets with a length of 15 mm, a width of 5 mm and a thickness of 0.1 mm;

[0038] The micro servo electric cylinder is fixed to the system base with bolts, the telescopic rod of the electric cylinder is connected to the connecting piece, and the micro servo electric cylinder is controlled by the host computer Matlab program through the USB controller; the connecting piece is a connecting rod, so that the axis of the movement direction of the electric cylinder is parallel to the axis of the telescopic direction of the spring; the spring is connected to the muscle tissue by anchoring and other connection methods.

[0039] One end of the spring structure is adhered to the matching mechanism using biocompatible glue and connected to the micro servo electric cylinder; the spring material is PDMS elastic material, the number of spring turns is 3 turns, the spring pitch is 1.5mm, the spring wire width is 0.8mm, the spring wire thickness is 0.5mm, and the spring elastic coefficient is 2.1mN / mm;

[0040] The present invention further provides a method for engineering skeletal muscle tissue culture and electrical-mechanical co-stimulation, comprising the following steps:

[0041] The matrix gel, collagen solution, NaOH solution, mouse C2C12 myoblasts and cell growth medium were fully mixed and injected into the muscle tissue culture mold;

[0042] In the first two days of muscle tissue formation, the culture medium is the growth medium. Under the action of cell traction, the matrix is ​​compacted and the volume of the muscle tissue decreases until it reaches a stable state. After two days of culture in the growth medium, the growth medium is replaced with the differentiation medium. On the third day of culture in the differentiation medium, the muscle tissue produces spontaneous contraction. On the fifth day, the muscle tissue is transferred to the electrical-mechanical co-stimulation system, one end of which is connected to the active end of the spring and the other end is connected to the micro-column on the surface of the culture dish.

[0043] The position of the spring's active end is adjusted by a micro servo electric cylinder to keep the muscle tissue length at its original length;

[0044] The electrical-mechanical co-stimulation specifically includes the following steps:

[0045] Through the software and electrical stimulation program, a multi-channel electrical stimulator was used to generate a periodic electric field through electrodes;

[0046] The electrical stimulation frequency was 1 Hz, the electric field strength was 1.5 V / cm, the pulse width was 6 ms, stimulation was for 7.5 minutes per hour, and rest was for 52.5 minutes, for a total of three days.

[0047] The muscle tissue contracts after receiving electrical stimulation, using the inherent elastic force of the spring to provide dynamic mechanical stimulation;

[0048] The centripetal contraction force of the muscle tissue is calculated by multiplying the displacement of the spring active end with the elastic coefficient, and the length of the muscle tissue is monitored in real time;

[0049] During the culture process, the muscle tissue grows in strength. At this time, the position of the spring is adjusted by a micro servo cylinder to always keep the length of the muscle tissue unchanged.

[0050] The electrical-mechanical co-stimulation system comprises a ring-shaped distributed multi-electrode for electrical stimulation of muscle tissue, a spring for dynamic mechanical stimulation of muscle tissue and measurement of concentric contraction force, and a micro servo electric cylinder for adjusting the position of the spring.

[0051] The annularly distributed multi-electrodes are controlled by a multi-channel electrical stimulator, and electric fields of different intensities, pulse widths and frequencies can be generated through the electrical stimulation program of the host computer software.

[0052] The PDMS spring has an elastic coefficient close to that of engineered skeletal muscle tissue, and its elastic coefficient can be adjusted to be consistent with muscle tissue by adjusting parameters such as the number of spring turns, spacing, line width and thickness.

[0053] The top of the electrical-mechanical co-stimulation system is transparent, allowing the visual monitoring system to collect information such as the displacement of the spring active end and the length of the muscle tissue in real time during the culture process.

[0054] The PDMS spring can reflect the contraction force of muscle tissue. Before electrical stimulation, the product of the stretch of the spring and the elastic coefficient can reflect the passive contraction force of the muscle tissue. During electrical stimulation, the product of the dynamic stretch of the spring and the elastic coefficient can reflect the centripetal contraction force of the muscle tissue.

[0055] The micro servo cylinder can be controlled by the host computer software through the USB controller to complete the telescopic movement. The stroke is 10mm, and the minimum single displacement is 5μm. It can accurately control the spring position while minimizing damage to muscle tissue.

[0056] The visual monitoring system measures the changes in the length of the muscle tissue during its growth, and a customized program is used to control the extension and retraction of the micro servo cylinder to adjust the length of the muscle tissue to its original length.

[0057] When not electrically stimulated, the muscle will have a passive contraction force due to the traction of the cells in the muscle. If not restrained, the muscle will atrophy spontaneously, showing a shortened length or even failure. Therefore, the spring can provide constraints, but when the spring and muscle are assembled at their original length, the spring will inevitably be stretched and the muscle will shorten. However, since the spring is further stretched, the elastic force it generates is sufficient to counteract the passive force, causing the muscle to not fail, but it will still affect the function. Therefore, it is very necessary to use a micro-electric cylinder to adjust the position of the spring to keep the muscle at its original length. The adjustment process is to stretch the spring so that the muscle returns to its original length. The stretched part of the spring × the elastic coefficient is the passive contraction force of the muscle.

[0058] When the electrode generates an electric field and the muscle is electrically stimulated, it produces regular centripetal contraction, driving the spring to move. In this process, the distance the spring is stretched multiplied by the elastic coefficient is the centripetal contraction force of the muscle.

[0059] Embodiment 1

[0060] The present invention uses mouse C2C12 myoblast-derived engineered skeletal muscle tissue as an example for demonstration. Before manufacturing skeletal muscle tissue, it is necessary to expand the experimental cells to a certain number (5×10 6 C2C12 cells were revived in a standard culture dish with a diameter of 100 mm, 6 mL of growth medium (89% volume of high-glucose DMEM medium + 10% volume of premium fetal bovine serum + 1% penicillin-streptomycin mixture) was added, and cultured in an incubator at 37° C. and 5% CO 2 .

[0061] When the cell density reaches about 70%, subculture is performed, and the cells in one culture dish are dispersed into three culture dishes for culture.

[0062] After passaging, when the cell density reached about 90%, the cells were digested with 0.25% trypsin solution and centrifuged at 1000 rpm for 5 minutes.

[0063] Use 330 μL growth medium to resuspend the cells, 240 μL matrix gel, 240 μL collagen type I (concentration of 4 mg / mL), 0.1% NaOH solution, blow the mixture evenly, inject it into the modular strip skeletal muscle tissue culture mold, and then transfer it to a cell culture incubator with a temperature of 37 degrees Celsius and 5% carbon dioxide for one hour. After the mixture solidifies, add 8 ml of cell growth medium and put it back into the cell culture incubator for culture.

[0064] After 48 hours of culture, the growth medium was replaced with a cell differentiation medium (97% volume of high-glucose DMEM medium + 2% volume of special horse serum + 1% penicillin-streptomycin mixture), and the cells were continued to be cultured in a cell culture incubator.

[0065] After 96 h of culture in differentiation medium, the muscle tissue was transferred to the electrical-mechanical co-stimulation system, one end of which was connected to the active end of the spring and the other end was assembled to the fixed micro-column of the culture dish.

[0066] The spring material is PDMS. By setting the number of spring turns to 3, the spacing to 1.5 mm, the line width to 0.8 mm, and the thickness to 0.5 mm, an elastic coefficient close to that of muscle tissue can be obtained to simulate the antagonistic effect of the biceps and triceps.

[0067] The micro servo electric cylinder is controlled by the upper computer software to adjust the position of the spring active end to adjust the length of the muscle tissue to the original length.

[0068] Start the electrical stimulator and set the electrical stimulation program through the supporting software of the host computer. The electrical pulse parameters are electric field intensity 1.5V / cm, frequency 1Hz, and pulse width 6ms.

[0069] Each stimulation lasted for 7.5 minutes, followed by a rest period of 52.5 minutes, for a total of 48 hours. During this period, the culture medium was replaced every 12 hours to prevent muscle tissue damage caused by electrolysis.

[0070] During the electrical-mechanical co-stimulation culture stage, the spring displacement was monitored in real time by a visual monitoring system to measure the muscle tissue length and contraction force.

[0071] According to the changes in muscle tissue length, the spring position is adjusted in real time through a micro servo electric cylinder to ensure that the muscle tissue length always remains at its original length.

[0072] Take out an in vitro skeletal muscle tissue that has been trained with electrical-mechanical co-stimulation, observe the differentiation of myotubes, perform immunofluorescence staining, and use a fluorescence microscope to image it. Finally, analyze the fluorescence image ( Figure 3 ).

[0073] The muscle tissue after electrical-mechanical co-stimulation has a good differentiation effect, and its internal sarcomere structure can be observed through fluorescent staining images, which is one of the important signs of skeletal muscle maturation.

[0074] Finally, the time-displacement curves of the muscle tissue cultured by electrical-mechanical co-stimulation driving springs with different elastic coefficients were measured to evaluate the concentric contraction dynamics of the muscle tissue under different loads.

[0075] like Figure 4 As shown in the figure, as the elastic coefficient of the spring increases, the displacement driven by the muscle tissue to move the end of the spring decreases, the peak tension time (the time from the beginning of contraction to the peak force of contraction) is prolonged, and the contraction speed (the slope of the contraction curve) is reduced. This result is consistent with the biological law of skeletal muscle contraction under different loads.

Claims

1. A muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training, including: The electrodes are distributed in a ring shape within the edge of the culture dish and are connected to the multi-channel electrical stimulator to generate periodic electrical pulse stimulation acting on the muscle tissue through the periodic electrical pulse signal emitted by the multi-channel electrical stimulator; A spring, the movable end of which is connected to the muscle tissue, and the fixed end of which is connected to the servo electric cylinder through a connector, for acting on the muscle tissue to generate dynamic mechanical stimulation; The servo electric cylinder is connected to the host computer through a controller, and is used to realize the extension and retraction control of the servo electric cylinder according to the instructions of the host computer to drive the spring to move.

2. The muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training according to claim 1, characterized in that: The electrode is a platinum sheet with a length of 15 mm, a width of 5 mm and a thickness of 0.1 mm. The electrodes are multiple and evenly distributed.

3. The muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training according to claim 1, characterized in that: The spring is made of PDMS elastic material, has 3 turns, a spring pitch of 1.5 mm, a spring wire width of 0.8 mm, a spring wire thickness of 0.5 mm, and a spring elastic coefficient of 2.1 mN / mm.

4. The muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training according to claim 1, characterized in that: One end of the muscle tissue is connected to the movable end of the spring, and the other end is connected to the micro-column fixed on the surface of the culture dish.

5. The control method of the muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training according to any one of claims 1-4, characterized in that: The steps include: Fix the muscle tissue to be tested on the micro-pillars on the surface of the culture dish and the active end of the spring; During the growth cycle of muscle tissue, the muscle tissue drives the spring to move, and monitors its passive contraction force and centripetal contraction force. The servo electric cylinder adjusts the spring position in real time, thereby providing a variable load for the muscle tissue to ensure that the length of the muscle tissue remains unchanged. The muscle tissue is cultured by electrical-mechanical co-stimulation; when the muscle tissue is electrically stimulated, the active end of the spring is driven to move periodically, and is subjected to periodic dynamic mechanical stimulation provided by the spring.

6. The control method of the muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training according to claim 5, characterized in that: The passive contraction force and the centripetal contraction force are monitored, and the spring position is adjusted in real time by the servo electric cylinder, including the following steps: In order to keep the length of the muscle tissue unchanged, the spring is lengthened by the servo electric cylinder to offset the passive contraction force that will shorten the length of the muscle tissue. The distance the spring is stretched is identified by visual images. At this time, the product of the distance the spring is stretched and the elastic coefficient is the passive contraction force.

7. The control method of the muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training according to claim 5, characterized in that: The muscle tissue is subjected to electrical-mechanical co-stimulation culture as follows: the electrical stimulation frequency is 1 Hz, the electric field strength is 1.5 V / cm, the pulse width is 6 ms, stimulation is 7.5 minutes per hour, rest is 52.5 minutes, and the total stimulation is three days.

8. The control method of the muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training according to claim 5, characterized in that: When the muscle tissue is electrically stimulated, the active end of the spring is driven to move periodically, and is subjected to periodic dynamic mechanical stimulation provided by the spring, including the following steps: During electrical stimulation, the muscle tissue contracts to overcome the resistance of the spring, and the distance the spring is dynamically stretched is identified through visual images. At this time, the product of the distance the spring is stretched and the elastic coefficient is the centripetal contraction force.

9. The control method of the muscle tissue electrical-mechanical co-stimulation system inspired by human muscle training according to claim 5, characterized in that: The acquisition of muscle tissue comprises the following steps: The matrix gel, collagen solution, NaOH solution, mouse C2C12 myoblasts and cell growth medium were fully mixed and injected into the muscle tissue culture mold; On the fifth day of culture, when the muscle tissue showed spontaneous contraction, it was transferred to a culture dish.

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