Single cell dynamic mechanical loading system and force regulation and control method based on magnetic beads and bidirectional static magnetic field

Abstract

The invention belongs to the technical field of cell mechanics, and particularly relates to a single cell dynamic mechanical loading system based on magnetic beads and a bidirectional static magnetic field and a force regulation and control method. The invention provides a single cell dynamic mechanical loading system based on magnetic beads and a bidirectional static magnetic field and a force regulation and control method. The system is composed of a magnetic field system module, a dynamic regulation and control module and a cell stress feedback module. Through the system, single-cell-level pulling / pressing two-way mechanical stimulation is achieved for the first time, magnetic bead concentration-magnetic field direction cooperative adjustment is achieved, and limitation of a traditional multi-cell loading mechanical mode is broken through; the system is used for carrying out mechanical regulation and control on single cells, can be used for single cell mechanical response research (such as stem cell differentiation and tumor cell migration), drug screening (detecting the influence of drugs on cell mechanical characteristics) and construction of a dynamic mechanical microenvironment in tissue engineering, and has a good application prospect.

Description

Technical Field

[0001] This invention belongs to the field of cell mechanics technology, specifically relating to a single-cell dynamic mechanical loading system and force control method based on magnetic beads and a bidirectional static magnetic field. Background Technology

[0002] Cells exist within a complex mechanical microenvironment in vivo, encountering forces such as vascular shear and tissue compressive forces. These mechanical signals have a direct and profound impact on cell differentiation, migration, and disease progression. Therefore, in-depth research into the mechanical responses of single cells is of paramount scientific and clinical value for understanding cellular behavior mechanisms, revealing the patterns of disease development, and developing new therapeutic strategies. However, existing single-cell mechanical loading techniques still have many limitations, making it difficult to meet the needs of precise research into single-cell mechanical responses.

[0003] Currently, common single-cell mechanical loading techniques mainly include the following: First, the Flexcell cell stretching device, which applies tensile force to cells through mechanical means. However, this device is usually designed for cell populations, making it difficult to precisely control individual cells and achieve dynamic mechanical loading. Second, the method of directly pressing cells with a glass plate. While this can apply pressure to cells, the operation is relatively crude, with low force application precision, and it cannot achieve bidirectional tensile / compressive force loading. Furthermore, it is difficult to avoid interference between multiple cells. In addition, existing technologies suffer from the following problems: insufficient force application precision; existing magnetic bead-magnetic field systems are mostly designed for cell populations, with a large magnetic field gradient range, making it impossible to focus on individual cells; poor targeting; the binding of magnetic beads to cells relies on non-specific adsorption, leading to multi-cell interference; a single mechanical mode; dynamic magnetic fields (such as alternating magnetic fields) are prone to thermal effects; and a unidirectional magnetic field is difficult to achieve bidirectional tensile / compressive force loading, making it difficult to simulate complex mechanical environments. These shortcomings severely limit the precise study of single-cell mechanical responses and hinder the development of the field of cell mechanics.

[0004] Given the shortcomings of existing technologies, it is particularly necessary to develop a precise, dynamic, and safe mechanical loading system for single cells. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a single-cell dynamic mechanical loading system and force control method based on magnetic beads and a bidirectional static magnetic field.

[0006] This invention provides a single-cell dynamic mechanical loading system, which includes... The magnetic field system module is configured to adjust the permanent magnet array according to the mechanical control mode, wherein the magnetic field strength of the permanent magnet array at a single cell is 0.5-2T; The dynamic control module is configured to adjust the magnitude of the force applied to a single cell by controlling the magnetic bead concentration gradient, and to adjust the direction of the force on a single cell by controlling the direction of the magnetic field.

[0007] The cell force feedback module is configured to reflect the actual force on a single cell in real time.

[0008] Preferably, in the magnetic field system module, the magnetic field strength of the permanent magnet array at a single cell is 1T.

[0009] Preferably, in the magnetic field system module, when the mechanical control mode is the tension mode, a permanent magnet array is placed above; when the mechanical control mode is the pressure mode, a permanent magnet array is placed below.

[0010] Preferably, in the dynamic control module, the magnetic bead concentration is 0-200 μg / mL.

[0011] Preferably, the magnetic beads are magnetic beads co-modified with RGD and polyglycerol.

[0012] Preferably, the RGD and polyglycerol co-modified magnetic beads are prepared according to the following steps; Step 1: Glycidyl ether is prepared by reacting glycidyl ether with ethyl vinyl ether; Step 2: Ethyl glycidyl ether and allyl glycidyl ether undergo an anionic ring-opening copolymerization to generate PG- b -PAGE block copolymer; Step 3, via photo-initiated thiol-ene click reaction, PG- b -PAGE block copolymer coupled with cysteine ​​hydrochloride; Step 4: The coupled product undergoes a condensation reaction with 3,4-dihydroxyhydrocinnamic acid to prepare catechol-functionalized polyglycerol PG-Cat. Step 5: PG-Cat reacts with activated RGD via a cycloaddition reaction to obtain RGD-PG-Cat; Step 6: RGD-PG-Cat reacts with amino-modified magnetic beads to obtain the final product.

[0013] Preferably, in the cell force feedback module, the measurement of the actual force on a single cell includes measurement according to Coulomb's law of magnetic force. In the magnetic field system module, the permanent magnet is selected from neodymium iron boron permanent magnets. In the dynamic control module, the diameter of the magnetic bead is 1-2 μm, and the saturation magnetization of the magnetic bead is 3-6 emu / g.

[0014] This invention applies a force control method for a single-cell dynamic mechanical loading system as described in any of the preceding claims, comprising: Step 1: Cells are co-incubated with magnetic beads, and the concentration gradient of the magnetic beads is controlled by a dynamic regulation module; Step 2, set the mechanical control mode; Step 3: Adjust the permanent magnet array in the magnetic field system module; Step 4: Observe the force situation of a single cell through the cell force feedback module.

[0015] Preferably, in step 1, the magnetic bead concentration is controlled by a gradient from low to high using a dynamic control module; And / or, in step 4, the magnetic bead concentration of the dynamic control module is adjusted according to the force situation of the single cell.

[0016] The present invention provides an apparatus integrating a single-cell dynamic mechanical loading system as described in any of the preceding claims.

[0017] This invention provides a single-cell dynamic mechanical loading system and force control method based on magnetic beads and a bidirectional static magnetic field by screening magnetic field parameters, magnetic bead modification methods, and dosages. The system consists of a magnetic field system module, a dynamic control module, and a cell force feedback module. Through this system, this invention achieves, for the first time, bidirectional tensile / compressive mechanical stimulation at the single-cell level, realizing the coordinated regulation of magnetic bead concentration and magnetic field strength, breaking through the limitations of traditional single mechanical modes. The force control of single cells using this system can be applied to single-cell mechanical response research (such as stem cell differentiation and tumor cell migration), drug screening (detecting the effect of drugs on cell mechanical properties), and the construction of dynamic mechanical microenvironments in tissue engineering, showing promising application prospects.

[0018] Obviously, based on the above description of the present invention, and according to common technical knowledge and conventional methods in the field, various other modifications, substitutions or alterations can be made without departing from the basic technical concept of the present invention.

[0019] The following detailed embodiments further illustrate the above-described content of the present invention. However, this should not be construed as limiting the scope of the present invention to the following examples. All technologies implemented based on the above-described content of the present invention fall within the scope of the present invention. Attached Figure Description

[0020] Figure 1 Image a shows the SEM results of magnetic beads of different concentrations binding with bone marrow-derived macrophages (BMDMs). Figure 1 b shows the SEM results of magnetic beads at different concentrations combined with bone marrow mesenchymal stem cells (BMSCs). Figure 1 Figure c shows the quantitative analysis results of the binding of magnetic beads of different concentrations to bone marrow-derived macrophages (BMDMs). Figure 1 d is a statistical diagram of the forces on a single cell after magnetic beads of different concentrations bind to bone marrow-derived macrophages (BMDMs); Figure 2 The image shows the fluorescence staining results of different magnetic field intensities on the spread of bone marrow-derived macrophages (BMDMs). Figure 3 a shows the spread of bone marrow-derived macrophages (BMDMs) after magnetic fields were placed above and below them, with different concentrations of magnetic beads binding to them. Figure 3 b is a statistical result of cell spreading after placing a magnet on top of the cells; Figure 3 c is a statistical result of cell spreading after placing a magnet under the cells; Figure 4 a shows the spread of bone marrow-derived mesenchymal stem cells (BMSCs) after magnetic fields were placed above and below them, with different concentrations of magnetic beads combined with them. Figure 4 b is a statistical result of cell spreading after placing a magnet on top of the cells; Figure 4 c is a statistical result of cell spreading after placing a magnet under the cell. In the figure, NF means no magnetic field, which means no force; TF means a magnetic field is placed above the cell, which means tension; and CF means a magnetic field is placed below the cell, which means pressure.

[0021] Figure 5 A schematic diagram of osteoclast generation after 50 μg / mL magnetic beads were bound to bone marrow-derived macrophages (BMDMs), and magnetic fields were placed above and below them for 24 hours. After osteoclast differentiation was induced for 4-5 days, the diagram shows the osteoclast generation. Figure 5 b is a statistical result of osteoclastogenesis after placing magnetic fields above and below the cells, respectively; Figure 5 c represents the binding of 50 μg / mL magnetic beads to bone marrow-derived mesenchymal stem cells (BMSCs). After being placed above and below the cells with magnetic fields for 24 hours, the cells were cultured in osteogenic differentiation induction medium for 7 days. Schematic diagram of alkaline phosphatase staining (ALP). Figure 5 d is a statistical graph showing the osteogenic differentiation of cells after placing magnetic fields above and below the cells, respectively. In the graph, NF indicates no magnetic field, which means no force; TF indicates a magnetic field above the cell, which means tension; and CF indicates a magnetic field below the cell, which means pressure. Detailed Implementation

[0022] Unless otherwise specified, all reagents and materials used in the following examples and experimental cases are commercially available.

[0023] Example 1: A Single-Cell Dynamic Mechanical Loading System Based on Magnetic Beads and a Two-Way Static Magnetic Field and Its Control Method I. Single-cell dynamic mechanical loading system based on magnetic beads and bidirectional static magnetic field 1. Preparation of RGD-modified micron magnetic beads (RGD-PG-MBs) (1) Preparation of micron magnetic beads Ferric salts were dissolved in a polyol solvent, and sodium acetate and a polymeric dispersant were added to form a reaction solution. The reaction solution was subjected to a solvothermal reaction at 200 °C for 12 h. After the reaction, the mixture was magnetically separated, washed, and dried to obtain Fe3O4 superparamagnetic microspheres with a particle size of 1.7 μm. Then, the Fe3O4 microspheres were dispersed in an ethanol / water system, and tetraethoxysilane was added dropwise under alkaline conditions adjusted by ammonia, causing hydrolysis and condensation to form a SiO2 shell on the Fe3O4 surface. The reaction temperature was 35 °C for 8 h. After magnetic separation, washing, and drying, Fe3O4@SiO2 particles were obtained. Finally, the Fe3O4@SiO2 particles were dispersed in anhydrous alcohol or aromatic hydrocarbon solvent, and 3-aminopropyltriethoxysilane was added for silanization. The reaction conditions were room temperature for 8 h or reflux at 80 °C for 8 h. After the reaction, the mixture was magnetically separated, washed, and dried to obtain micron-sized magnetic beads with -NH2 groups on the surface.

[0024] (2) Synthesis of ethyl glycidyl ether monomer (EEGE) 40.0 g of 2,3-propanediol was dissolved in 200 mL of vinyl ether and stirred in an ice-water bath. 1 g of p-toluenesulfonic acid was added, and the mixture was stirred for 3 hours while maintaining the temperature below 40 °C. The mixture was washed with a saturated sodium bicarbonate aqueous solution, the phases were separated, and the organic phase was dried. The product was obtained by filtration, vacuum concentration to remove the vinyl ether, and final distillation.

[0025] (3) Poly(glycidyl)-block-poly(acryloyl glycidyl ether) (PG- b Synthesis of -PAGE Tetrabutylammonium azide (285 mg) was dried under vacuum at 80 °C and dissolved in dry toluene (140 mL). Ethyl ethylene oxide (EEGE, 19.3 mL, 137 mmol) and triisobutylaluminum (1.8 mL, 2 mmol) were added, and polymerization was initiated at 0 °C. After 3 hours, alkenyl ethylene oxide (AGE, 0.9 g) and additional triisobutylaluminum (1.8 mL, 2 mmol) were added, and the mixture was stirred for 12 hours at room temperature. The mixture was quenched in water (1 mL), dried, concentrated under vacuum, dissolved in diethyl ether, centrifuged, and dialyzed in dichloromethane to obtain the polymer (90% yield). After removing the protecting group, hydroxyl-containing polyglycerol blocks were obtained. These blocks were dissolved in THF (100 mL), and 37% concentrated hydrochloric acid (5-10 mL) was added. The mixture was stirred for 10 hours, washed with THF, and dialyzed against methanol to obtain PG- b -PAGE.

[0026] (4) Synthesis of catechol-functionalized coating polymer (PG-Cat) The thiol-alkene reaction was carried out at room temperature. PG- b -PAGE and cysteine ​​hydrochloride (4 molar equivalents) were dissolved in methanol, and Irgacure 651 (2% equivalent) was added. The mixture was irradiated with 365 nm UV light for 12 hours. PG- was purified by methanol dialysis. b -PAmGE. (This refers to PG-) b - PAmGE and EDCI (3 molar equivalents) were dissolved in MES buffer / methanol (1:1 v / v), DHHA (3 molar equivalents) were added, and the mixture was stirred at room temperature for 12 hours. Finally, PG-Cat was purified by methanol dialysis.

[0027] (5) Synthesis of RGD-modified magnetic beads (RGD-PG-MBs) CycloRGDfK (50 mg) and BCN (bicyclo[6.1.0]nonyne, 30.6 mg) were dissolved in anhydrous DMF, and a catalytic amount of triethylamine (Et3N) was added, followed by stirring for 3 hours. After solvent removal under vacuum, RGD activated with BCN was used for further functionalization. PG-Cat (200 mg) and BCN-RGD (14.79 mg) were dissolved in methanol, stirred for 4 hours, and finally purified by methanol dialysis. Finally, RGD-PG-MBs were synthesized by reacting the amino groups on magnetic beads with the catechol groups of RGD-PG-Cat.

[0028] This embodiment successfully synthesized magnetic beads with RGD (cyclic RGDfK). The polyglycerol (PG) segment effectively prevents the magnetic beads from being phagocytosed by cells; RGD binds to integrins on the cell surface, fixing MBs to the cell, thereby achieving efficient transmission and transduction of mechanical force. Therefore, RGD-PG-MBs not only adhere firmly to cells but are also not easily internalized.

[0029] 2. Mixing magnetic beads and cells RGD-PG-MBs were mixed evenly with cells at concentration gradients of 25, 50, 100, and 200 μg / mL.

[0030] 3. Two-way static magnetic field loading device The bidirectional static magnetic field loading device is equipped with a bidirectional static magnetic field loading system, which consists of a magnetic field system module, a dynamic control module, and a cell force feedback module.

[0031] (1) A magnetic field system module is configured to adjust the upper and lower symmetrical neodymium iron boron permanent magnet arrays according to the mechanical control mode, wherein the magnetic field strength of the permanent magnet array in the upper and lower central regions is 1T; When the mechanical control mode is the tension mode, a neodymium iron boron permanent magnet array is placed on the upper side; When the mechanical control mode is pressure mode, a neodymium iron boron permanent magnet array is placed on the lower side; (2) The dynamic control module is configured to change the number of magnetic beads bound to the surface of a single cell and adjust the basal force value by controlling the magnetic bead concentration gradient (within the range of 0-200 μg / mL); the magnetic bead concentration gradient is set to 25, 50, 100 and 200 μg / mL respectively.

[0032] (3) The cell force feedback module is configured to measure the actual force on a single cell in real time; The actual force on a single cell is measured according to Coulomb's law of magnetism, using the following formula: II. Force Control Methods (1) Place the cell on the worktable in the central region of the upper and lower symmetrical NdFeB permanent magnet array of the bidirectional static magnetic field loading device.

[0033] (2) The dynamic control module controls the magnetic bead concentration gradient from low to high, changing the number of magnetic beads bound to the surface of a single cell in order to regulate the basic force value; (3) Set the mechanical control mode to: tension mode and pressure mode; (4) Observe the force of a single cell through the cell force feedback module; dynamically adjust the concentration of magnetic beads by observing the force of a single cell in real time.

[0034] The technical solution of the present invention will be further explained through experiments below.

[0035] Experimental Example 1: Preparation of RGD-modified magnetic beads I. Experimental Methods Following the method of Example 1, firstly, glycidyl ether was reacted with ethyl vinyl ether under ice bath conditions using toluenesulfonic acid as a catalyst to obtain ethyl glycidyl ether (EEGE). Next, anionic ring-opening copolymerization of EEGE and allyl glycidyl ether (AGE) was carried out in anhydrous toluene using tetrabutylammonium azide as an initiator and triisobutylaluminum as an activator to obtain PG- b -PAGE block copolymer. Then, PG-PAGE block copolymers are produced via UV-initiated thiol-ene click reaction. b - PAGE was coupled with cysteine ​​hydrochloride to introduce amino groups, and then condensed with 3,4-dihydroxyhydrocinnamic acid under EDCI-mediated reaction to obtain catechol-functionalized polyglycerol (PG-Cat). Finally, BCN-RGD was combined with PG-Cat via a cycloaddition reaction, and the catechol groups of PG-Cat were covalently bonded to the amino groups on the surface of superparamagnetic iron oxide nanoparticles (MBs) to prepare RGD-modified magnetic beads (RGD-PG-MBs).

[0036] II. Experimental Results Magnetic beads with RGD (cyclic RGDfK) were successfully synthesized. The polyglycerol (PG) segment effectively prevents the magnetic beads from being phagocytosed by cells. Through dynamic binding with integrins on the cell surface, RGD anchors MBs to the cell, thereby achieving efficient transmission and transduction of mechanical forces. Therefore, RGD-PG-MBs not only adhere firmly to cells but are also not easily internalized.

[0037] Experiment Example 2: Screening of Magnetic Bead Concentration I. Experimental Methods RGD-modified magnetic beads (prepared in Example 1) at concentrations of 25 μg / mL, 50 μg / mL, 100 μg / mL, and 200 μg / mL were co-incubated with bone marrow-derived macrophages (BMDM) or bone marrow mesenchymal stem cells (BMSC) at 37 °C for 24 h. The binding of cells to magnetic beads was observed by scanning electron microscopy (SEM). After the magnetic beads bound to the cells, they were placed in the magnetic field of Example 1, and the theoretical force acting on a single cell was calculated using the equation (magnetic Coulomb's law).

[0038] II. Experimental Results SEM results are as follows Figure 1 As shown, RGD-PG-MBs specifically bind to the surface of BMDM or BMSC cells; different concentrations of magnetic nanobeads result in different numbers distributed on the cell surface. Quantitative analysis showed that the binding of RGD-PG-MBs to cells was dose-dependent. At concentrations of 25 μg / mL, 50 μg / mL, 100 μg / mL, and 200 μg / mL, the average number of magnetic beads bound per cell was 24.8 ± 1.62, 53.9 ± 4.36, 117.3 ± 4.08, and 118.1 ± 5.90 (n = 10), respectively. Figure 1 (C) indicates that when the concentration is 100 μg / mL, the number of magnetic beads effectively bound by a single cell has reached saturation. Even if the cell concentration is increased, the number of magnetic beads bound will not change. Therefore, the effective concentrations are 25 μg / mL, 50 μg / mL, and 100 μg / mL, and 100 μg / mL is the maximum binding concentration.

[0039] The results of quantitative analysis of forces on single cells are as follows Figure 1As shown in Figure d, the forces applied to single cells at magnetic bead concentrations of 25 μg / mL, 50 μg / mL, and 100 μg / mL were 88.92 ± 11.26 pN, 181.08 ± 20.15 pN, 422.28 ± 14.70 pN, and 425.16 ± 21.23 pN, respectively. The results indicate that the force applied to single cells can be controlled by adjusting the magnetic bead concentration, and within the concentration range of 0-100 μg / mL, a higher magnetic bead concentration results in a greater force applied to the single cells.

[0040] Experiment Example 3: Effects of different magnetic field strengths on cells I. Experimental Methods 50 μg / mL of RGD-PG-MBs (prepared in Example 1) were co-incubated with BMDM cells, and then placed in magnetic field environments with magnetic field strengths of 0.5T, 1T, and 2T, respectively. Cell morphology was observed and recorded using a fluorescence microscope.

[0041] II. Experimental Results The results are as follows Figure 2 As shown, different magnetic field strengths have a significant impact on cell spreading. The cell spreading area is largest when the magnetic field strength is 1T, but the spreading area decreases when the magnetic field strength is continuously increased to 2T. Therefore, the optimal magnetic field strength for the bidirectional static magnetic field loading system of this invention is 1T.

[0042] Experiment 4 was used to regulate cell spreading. I. Experimental Methods RGD-PG-MBs (prepared in Example 1) at concentrations of 25 μg / mL, 50 μg / mL, and 100 μg / mL were co-incubated with bone marrow-derived macrophages (BMDM) or bone marrow mesenchymal stem cells (BMSC), respectively, and then placed in the magnetic field of Example 1. Cell spreading was observed and recorded using a fluorescence microscope after F-actin staining. Cell morphology under magnetic bead concentration gradients was analyzed by controlling the direction of the applied magnetic field.

[0043] II. Experimental Results The results are as follows Figure 3 , 4 As shown, under the influence of a magnetic field, the cell spreading area increased significantly, and the increase was even more significant when the concentration of RGD-PG-MBs was 50 μg / mL, indicating that the cells were subjected to different degrees of force. This further demonstrates the successful construction of the single-cell force-applying system of the present invention.

[0044] Experiment 5: Regulating Cell Differentiation I. Experimental Methods RGD-PG-MBs (prepared in Example 1) at a concentration of 50 μg / mL were co-incubated with bone marrow-derived macrophages (BMDMs) or bone marrow mesenchymal stem cells (BMSCs), and then placed in the magnetic field of Example 1 for 24 h. BMDMs were cultured for 4-5 days with 100 ng / mL receptor activator of nuclear factor κB (RANKL) and 40 ng / mL macrophage colony-stimulating factor (M-CSF), and osteoclastogenesis was detected by tartrate-resistant acid phosphatase staining (Trap staining). BMSCs were induced in osteogenic differentiation medium for 7 days, and osteogenic formation was detected afterward.

[0045] II. Experimental Results The results are as follows Figure 5 As shown, under the influence of a magnetic field, both tensile and compressive forces inhibit osteoclast differentiation and promote osteogenic differentiation, indicating that the single-cell force application system of this invention is successful and can further regulate cell differentiation.

[0046] As can be seen from the above embodiments and experimental examples, this invention provides a single-cell dynamic mechanical loading system and force control method based on magnetic beads and a bidirectional static magnetic field by screening magnetic field parameters, magnetic bead modification methods, and dosages. This system consists of a magnetic field system module, a dynamic control module, and a cell force feedback module. Through this system, this invention achieves, for the first time, bidirectional tensile / compressive mechanical stimulation at the single-cell level, realizing the coordinated regulation of magnetic bead concentration and magnetic field strength, breaking through the limitations of traditional single mechanical modes. Using this system to control the force of single cells, it can be used for single-cell mechanical response research (such as stem cell differentiation and tumor cell migration), drug screening (detecting the effect of drugs on cell mechanical properties), and constructing dynamic mechanical microenvironments in tissue engineering, showing promising application prospects.

Claims

1. A single-cell dynamic mechanical loading system, characterized in that: It includes The magnetic field system module is configured to adjust the permanent magnet array according to the mechanical control mode, wherein the magnetic field strength of the permanent magnet array at a single cell is 0.5-2T; The dynamic control module is configured to adjust the magnitude of the force applied to a single cell by controlling the magnetic bead concentration gradient, and to adjust the direction of the force on a single cell by controlling the direction of the magnetic field. The cell force feedback module is configured to reflect the actual force on a single cell in real time.

2. The single-cell dynamic mechanical loading system according to claim 1, characterized in that: In the magnetic field system module, the magnetic field strength of the permanent magnet array at a single cell is 1T.

3. The single-cell dynamic mechanical loading system according to claim 1, characterized in that: In the magnetic field system module, when the mechanical control mode is the tension mode, a permanent magnet array is placed above; when the mechanical control mode is the pressure mode, a permanent magnet array is placed below.

4. The single-cell dynamic mechanical loading system according to claim 1, characterized in that: In the dynamic control module, the magnetic bead concentration is 0-200 μg / mL.

5. The single-cell dynamic mechanical loading system according to claim 1, characterized in that: The magnetic beads are RGD and polyglycerol co-modified magnetic beads.

6. The single-cell dynamic mechanical loading system according to claim 5, characterized in that: The magnetic beads co-modified with RGD and polyglycerol were prepared according to the following steps; Step 1: Glycidyl ether is prepared by reacting glycidyl ether with ethyl vinyl ether; Step 2: Ethyl glycidyl ether and allyl glycidyl ether undergo an anionic ring-opening copolymerization to generate PG- b -PAGE block copolymer; Step 3, via photo-initiated thiol-ene click reaction, PG- b -PAGE block copolymer coupled with cysteine ​​hydrochloride; Step 4: The coupled product undergoes a condensation reaction with 3,4-dihydroxyhydrocinnamic acid to prepare catechol-functionalized polyglycerol PG-Cat. Step 5: PG-Cat reacts with activated RGD via a cycloaddition reaction to obtain RGD-PG-Cat; Step 6: RGD-PG-Cat reacts with amino-modified magnetic beads to obtain the final product.

7. The single-cell dynamic mechanical loading system according to claim 1, characterized in that: In the cell force feedback module, the measurement of the actual force on a single cell includes the measurement according to Coulomb's law of magnetic force. In the magnetic field system module, the permanent magnet is selected from neodymium iron boron permanent magnets. In the dynamic control module, the diameter of the magnetic bead is 1-2 μm, and the saturation magnetization of the magnetic bead is 3-6 emu / g.

8. A force control method using the single-cell dynamic mechanical loading system according to any one of claims 1-7, characterized in that, It includes: Step 1: Cells are co-incubated with magnetic beads, and the concentration gradient of the magnetic beads is controlled by a dynamic regulation module; Step 2, set the mechanical control mode; Step 3: Adjust the permanent magnet array in the magnetic field system module; Step 4: Observe the force situation of a single cell through the cell force feedback module.

9. The force control method according to claim 8, characterized in that, In step 1, the magnetic bead concentration is controlled by a gradient from low to high using a dynamic control module; And / or, in step 4, the magnetic bead concentration of the dynamic control module is adjusted according to the force situation of the single cell.

10. An apparatus integrating the single-cell dynamic mechanical loading system according to any one of claims 1-7.

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