Heterogeneous composite structure microcarrier and controllable preparation method thereof

Abstract

The invention discloses a heterogeneous composite structure microcarrier and a controllable preparation method thereof. The preparation method comprises the following steps: dissolving collagen and a coating drug in dilute acetic acid to obtain a solution A; dissolving PLCL and a pore-foaming agent in an organic solvent to obtain a solution B; mixing the solution A and the solution B, and mechanically homogenizing to obtain a mixed emulsion as a dispersed phase O; dissolving a surfactant in deionized water to respectively serve as a continuous phase W and a collection liquid Wo; adjusting the pH values of the continuous phase W and the collected liquid Wo to 7.5-9.0; respectively injecting the continuous phase W and the dispersed phase O into a two-phase micro-channel, shearing the dispersed phase O into micro-emulsion drops with uniform size, receiving the micro-emulsion drops in a collecting solution Wo, standing or horizontally rotating and curing, washing and freeze-drying to obtain the heterogeneous composite structure microcarrier; the microcarrier prepared by the invention has concentrated particle size distribution, good initial cell adhesion efficiency and biodegradability, can provide adhesion sites and mechanical support for cells at the same time, and meets carrier functions required by in-vitro culture and in-vivo implantation of cells.

Description

Technical Field

[0001] The invention relates to the technical field of biomedical material preparation, and in particular to a heterogeneous composite structure microcarrier and a controllable preparation method thereof. Background Art

[0002] Microcarriers usually refer to microspheres with a diameter of 90-400 microns and a density slightly greater than that of water, which can support cell attachment and growth and can achieve high cell density in a relatively low volume. Microcarrier culture, as a three-dimensional culture method suitable for adherent cells, combines the advantages of planar culture and suspension culture, can provide higher cell proliferation efficiency and maintain good cell morphology. According to the structural classification of microcarriers, they can be mainly divided into solid microcarriers and porous microcarriers. Among them, the porous microcarriers have rich and interconnected pore structures inside and outside, which give them a larger specific surface area and space capacity, which is convenient for cell growth and migration; at the same time, the interconnected porous structure is also conducive to protein adsorption, elimination of metabolic waste, transport of nutrients and transfer of biological factors. However, there are still many problems in the scientific research and clinical application of microcarriers, such as the monodispersity of microcarrier particle size, precise regulation of microcarrier structure, effective modification of microcarriers and application expansion.

[0003] Collagen, as a natural biomacromolecule, is one of the main components of the human extracellular matrix (ECM). It can not only provide adhesion sites for cells, but also simulate the in vivo environment for cells and promote cell adhesion. However, microcarriers prepared with collagen as raw material show problems of insufficient mechanical strength and rapid degradation in subsequent in vitro and in vivo applications. Synthetic polymers with good biocompatibility are another widely used microcarrier raw material. This type of material has excellent mechanical properties and a long degradation cycle, but has low cell affinity, which affects cell adhesion to a certain extent. How to combine the advantages of the two materials, while meeting the requirements of providing cell adhesion sites and rigid support, and having a suitable degradation cycle and non-cytotoxic functionalized composite microcarriers, is still an urgent problem to be solved.

[0004] Traditional preparation methods for microcarriers mainly include emulsification / solvent evaporation method, spray drying method, supercritical CO2 method, etc. Different preparation methods lead to differences in the performance of the obtained microcarriers, including morphology, internal structure, cell adhesion rate, degradation cycle, etc., and these performance differences determine whether the obtained microcarriers can effectively carry out cell adhesion, proliferation and differentiation, tissue repair and other further biomedical effects. Therefore, the selection of microcarrier preparation methods and the optimization of operating parameters are crucial. It should be emphasized that the more uniform the diameter of the microcarriers and the more controllable the preparation process, the more accurate the evaluation of the cell response to them, which is conducive to optimizing the best biological effect. In addition, how to take into account the formation and control of the coating at the same time in the process of microcarrier preparation, and avoid the problems of microcarrier adhesion agglomeration and coating shedding caused by the traditional immersion modification method, is also a challenge to be solved in the field of microcarriers.

[0005] Therefore, in order to solve the above problems, the present invention provides a heterogeneous composite structure microcarrier and a controllable preparation method thereof, which can be used for cell expansion and tissue regeneration. The present invention preferably meets the production requirements and has biocompatible high molecular polymer components and biomacromolecules, and has an easily removable porogen, and through reasonable component ratio and structural regulation, the composite microcarrier can promote cell adhesion / proliferation, thereby achieving cell expansion and having anti-inflammatory function. Summary of the invention

[0006] The purpose of the present invention is to provide a heterogeneous composite structure microcarrier and a controllable preparation method thereof, which utilizes the principles of water-in-oil-in-water emulsion, solvent volatilization, phase separation and surface polymerization, and through the combination of two-phase microchannels used in the preparation process, the control of the orifice diameter and the fluid flow rate, the composite microcarrier has a single dispersion and is conducive to cell adhesion and a stable and controllable structure.

[0007] To achieve the above object, the present invention provides the following technical solutions:

[0008] A heterogeneous composite structure microcarrier, which consists of a spherical body composed of synthetic high molecular polymer and biological macromolecules, a coating drug coated on the surface of the spherical body and a connected pore structure; the synthetic high molecular polymer is an oil-soluble high molecular polymer, the biological macromolecule is a water-soluble biological macromolecule, and the coating drug is polydopamine.

[0009] Preferably, calculated based on the total mass percentage (dry weight) of the heterogeneous composite structure microcarrier, the oil-soluble high molecular polymer is poly (L-lactic acid-caprolactone copolymer) (PLCL), and its content is 50% to 70%, the water-soluble biomacromolecule is collagen, and the collagen is type I fish collagen, and its content is 20% to 48%, and the content of polydopamine is 2% to 10%.

[0010] Preferably, the monomer molar ratio of LA to CL in the poly (L-lactic acid-caprolactone) copolymer (PLCL) is 90-10:10-90.

[0011] Preferably, the heterogeneous composite structure microcarrier has a particle size of 120 to 350 microns, a pore size of 3 to 50 microns, and a porosity of 10% to 80%.

[0012] The present application also claims a method for preparing a heterogeneous composite structure microcarrier, which is a controllable preparation method comprising the following steps:

[0013] (1) Dissolving collagen and coating drugs in dilute acetic acid to obtain a hydrophilic solution A;

[0014] (2) dissolving the poly (L-lactic acid-caprolactone) copolymer and the porogen in a volatile organic solvent to obtain a lipophilic solution B;

[0015] (3) mixing the solution A prepared in step (1) and the solution B prepared in step (2) in a certain proportion and then mechanically homogenizing them to obtain a mixed emulsion as dispersed phase O;

[0016] (4) dissolving the surfactant in deionized water to obtain a surfactant aqueous solution which serves as the continuous phase W and the collection liquid Wo of the microemulsion droplets;

[0017] (5) adjusting the pH of the continuous phase W and the collected liquid Wo to 7.5-9.0;

[0018] (6) The continuous phase W and the dispersed phase O are injected into the two-phase microchannel respectively, and the dispersed phase O is sheared into microemulsion droplets of uniform size;

[0019] (7) The microemulsion droplets obtained in step (6) are received in a collecting liquid Wo, and allowed to stand or rotate horizontally to solidify, followed by washing and freeze-drying to obtain heterogeneous composite structure microcarriers.

[0020] Preferably, the porogen is selected from one of isooctane, camphene, n-tridecane and toluene.

[0021] Preferably, the particle size of the heterogeneous composite structure microcarrier is regulated by the flow rate of the dispersed phase O and the continuous phase W or the size of the two-phase microchannel or the pipeline combination; the flow rate of the dispersed phase O is controlled to be 1mL / h~20mL / h, and the flow rate of the continuous phase W is controlled to be 3mL / h~60mL / h; the inner channel diameter of the two-phase microchannel is 50 microns to 500 microns, and the outer channel diameter is 300 microns to 800 microns; the pipeline combination is selected from one of Y-type, T-type, cross-type, co-flow type, and flow focusing type.

[0022] Preferably, the pore structure of the heterogeneous composite structure microcarrier is regulated by the concentration of the porogen, the curing time and the curing method; the structural morphology of the heterogeneous composite structure microcarrier is regulated by the volume ratio of solution A to solution B and the concentration of collagen in solution A; the curing time is 24 to 72 hours, and when the curing method is horizontal rotation curing, the rotation speed is 20 to 80 rpm.

[0023] Preferably, the volatile organic solvent is selected from one of dichloromethane, chloroform and tetrahydrofuran; the coating drug is dopamine hydrochloride, and the concentration of dopamine hydrochloride is 1-20 mg / ml; the concentration of collagen in the solution A is 0.3wt%-5wt%; the concentration of poly-L-lactic acid-caprolactone copolymer (PLCL) in the solution B is 1.5wt%-5wt%, and the concentration of the porogen is 1wt%-5wt%; solution A and solution B are mixed in a volume ratio of 1-5:4-5 and then mechanically homogenized.

[0024] Preferably, in step (4), the surfactant is selected from one of sodium polyacrylate (ASAP), polyvinyl alcohol (PVA), and polyvinyl pyrrolidone (PVP); and the concentration of the surfactant aqueous solution is 0.5wt% to 4wt%.

[0025] Due to the application of the above technical solution, the present invention has the following beneficial effects compared with the prior art:

[0026] 1. The present invention utilizes the shear force and surface tension between microfluids to prepare microemulsion droplets with uniform particle size and consistent composition;

[0027] 2. The present invention adopts a static solidification or horizontal rotation solidification method to fully volatilize the organic solvent in the oil phase, and the composite microemulsion droplets are gradually phase-separated into a Janus structure and shrunk into a composite microcarrier; a deionized water washing method is adopted to remove the surfactant and porogen remaining on the surface of the composite microcarrier; and a freeze-drying method is adopted to pre-freeze the composite microcarrier with deionized water in the solution to form a solid state, and freeze-drying is performed to remove water, and the dried composite microcarrier can be collected, and the obtained heterogeneous composite structure microcarrier has uniform size;

[0028] 3. The present invention preferably selects synthetic polymer components, biomacromolecules and coating drugs with good performance that meet production requirements and have biocompatibility and biodegradability, and through reasonable component ratio and structural regulation of porogens, the microcarriers have unique heterogeneous morphology and excellent biological properties, as well as adjustable degradation cycles. Compared with conventional microcarriers, the present invention is based on the principles of water-in-oil-in-water emulsion, solvent volatilization, phase separation and surface polymerization, through the combination of two-phase microchannels used in the preparation process, the control of the orifice diameter and fluid flow rate, and the regulation of the polymer ratio, so that the composite microcarrier has a single dispersion and a stable and controllable structure that is conducive to cell adhesion;

[0029] 4. The heterogeneous composite structure microcarrier prepared by the present invention has a concentrated particle size distribution, good initial cell adhesion efficiency and biodegradability. By changing the mass ratio of high molecular polymers and biomacromolecules and the oil-water phase ratio, the morphology and structure of the microcarrier and the degradation cycle can be precisely controlled. It can also provide adhesion sites and mechanical support for cells, meeting the carrier function required for cell culture in vitro and implantation in vivo. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] In order to more clearly illustrate the specific implementation methods of the present invention or the technical solutions in the prior art, the drawings required for use in the specific implementation methods or the description of the prior art will be briefly introduced below. Obviously, some of the drawings described below are some implementation methods of the present invention. For ordinary technicians in this field, other drawings can be made based on these drawings without paying any creative work.

[0031] Figure 1 is a schematic diagram of a two-phase microchannel of Example 1 of the present invention;

[0032] Figure 2 is an optical microscope image and size distribution diagram of the heterogeneous composite structure microcarrier microemulsion droplets with a lower polydopamine content according to Example 1 of the present invention;

[0033] Figure 3 is an optical microscope image and size distribution diagram of the heterogeneous composite structure microcarrier with a lower polydopamine content according to Example 1 of the present invention;

[0034] Figure 4 is a scanning electron microscope image of a heterogeneous composite structure microcarrier with a lower polydopamine content according to Example 1 of the present invention;

[0035] Figure 5 is a scanning electron microscope image of a heterogeneous composite structure microcarrier with a higher polydopamine content according to Example 2 of the present invention;

[0036] Figure 6 It is a scanning electron microscope image of the heterogeneous composite structure microcarrier of comparative example 1 of the present invention. DETAILED DESCRIPTION

[0037] In order to have a clearer understanding of the technical features, purposes and effects of the present invention, a specific implementation scheme is now described in detail.

[0038] The present invention is further described below in conjunction with the examples, but the present invention is not limited to the following examples. The implementation conditions used in the examples can be further adjusted according to the different requirements of specific use, and the implementation conditions not indicated are conventional conditions in the industry. The technical features involved in each embodiment of the present invention can be combined with each other as long as they do not conflict with each other.

[0039] Example 1

[0040] See attached Figure 1 -Attached Figure 4 This embodiment provides a heterogeneous composite structure microcarrier and a controllable preparation method thereof. This embodiment is for preparing a heterogeneous composite structure microcarrier with a low polydopamine content, comprising the following steps:

[0041] (1) Dissolve 0.075 g collagen and 10 mg dopamine hydrochloride in 5 mL dilute acetic acid (1% v / v) to match the claim, including the units, to obtain a hydrophilic solution A;

[0042] (2) Dissolve 1.8 g PLCL and 0.15 ml n-tridecane in dichloromethane to obtain a lipophilic solution B;

[0043] (3) 2 mL of solution A prepared in step (1) and 5 mL of solution B prepared in step (2) were mechanically homogenized at 6000 rpm for 2 min to obtain a mixed emulsion as dispersed phase O;

[0044] (4) dissolving polyvinyl alcohol in deionized water at 75° C. to prepare a 2 wt % polyvinyl alcohol solution which is used as the continuous phase W and the collection liquid Wo of the microemulsion droplets;

[0045] (5) Using NaHCO3, adjust the pH of the continuous phase W and the collected liquid Wo to 8.5;

[0046] (6) Injecting the dispersed phase O into the two-phase microchannel (such as Figure 1 ) with an inner channel having a diameter of 380 μm and an injection rate of 10 mL / h; the continuous phase W is injected into the outer channel of the two-phase microchannel, with a diameter of 780 μm and an injection rate of 30 mL / h; the dispersed phase O is sheared into microemulsion droplets of uniform size by the shearing action of the fluid;

[0047] (7) The microemulsion droplets obtained in step (6) are collected in a collecting liquid Wo, and the diameter of the heterogeneous composite structure microcarrier microemulsion droplets with a low polydopamine content before solidification is observed by optical microscopy. The droplets are about 534 μm in diameter, and the size is highly monodisperse (e.g. Figure 2 );

[0048] The microemulsion droplets were allowed to solidify for 48 h and washed five times with deionized water. The cured heterogeneous composite microcarriers with a lower polydopamine content were observed under an optical microscope. The diameter of the microcarriers shrank to about 259 μm, and a high degree of monodispersity was maintained (e.g. Figure 3 );

[0049] The heterogeneous composite microcarriers with low polydopamine content were freeze-dried for 24 hours and observed under a scanning electron microscope. They showed a half-half heterogeneous structure, with the PLCL side being a porous spherical structure and the collagen side being an uneven, loose, and porous structure (e.g. Figure 4 ).

[0050] Example 2

[0051] See attached Figure 5 This embodiment provides a heterogeneous composite structure microcarrier and a controllable preparation method thereof. This embodiment is for preparing a heterogeneous composite structure microcarrier with a high polydopamine content, comprising the following steps:

[0052] (1) Dissolve 0.075 g collagen and 50 mg dopamine hydrochloride in 5 mL dilute acetic acid (1% v / v) to obtain a hydrophilic solution A;

[0053] (2) Dissolve 1.8 g PLCL and 0.15 ml n-tridecane in dichloromethane to obtain a lipophilic solution B;

[0054] (3) 2 mL of solution A prepared in step (1) and 5 mL of solution B prepared in step (2) were mechanically homogenized at 6000 rpm for 2 min to obtain a mixed emulsion as dispersed phase O;

[0055] (4) dissolving polyvinyl alcohol in deionized water at 75° C. to prepare a 2 wt % polyvinyl alcohol solution which is used as the continuous phase W and the collection liquid Wo of the microemulsion droplets;

[0056] (5) Using NaHCO3, adjust the pH of the continuous phase W and the collected liquid Wo to 8.5;

[0057] (6) The dispersed phase O is injected into the inner channel of the two-phase microchannel, the diameter of the inner channel is set to 380 μm, and the injection rate is 10 mL / h; the continuous phase W is injected into the outer channel of the two-phase microchannel, the diameter of the outer channel is set to 780 μm, and the injection rate is 30 mL / h; the dispersed phase O is sheared into microemulsion droplets of uniform size by the shearing action of the fluid;

[0058] (7) receiving the microemulsion droplets obtained in step (6) in a collection liquid Wo, allowing them to stand and solidify for 48 hours, and washing them with deionized water five times;

[0059] After freeze drying for 24 hours, a heterogeneous composite microcarrier with a high polydopamine content was obtained. Scanning electron microscopy showed a half-half heterogeneous structure. The PLCL side was a smooth spherical structure wrapped with a layer of collagen membrane to make the inner pores invisible. The collagen side was an uneven porous structure. After magnification, the polydopamine coating on the surface (such as Figure 5 ); This is because the increase in dopamine hydrochloride concentration changes the hydrophilicity of the system, thereby changing the rate of phase separation.

[0060] Comparative Example 1

[0061] See attached Figure 6 This comparative example provides a heterogeneous composite structure microcarrier and a controllable preparation method thereof. This comparative example is for preparing a heterogeneous composite structure microcarrier without polydopamine, comprising the following steps:

[0062] (1) Dissolve 0.075 g of collagen in 5 mL of dilute acetic acid (1% v / v) to obtain a hydrophilic solution A;

[0063] (2) Dissolve 1.8 g PLCL and 0.15 ml n-tridecane in dichloromethane to obtain a lipophilic solution B;

[0064] (3) 2 mL of solution A prepared in step (1) and 5 mL of solution B prepared in step (2) were mechanically homogenized at 6000 rpm for 2 min to obtain a mixed emulsion as dispersed phase O;

[0065] (4) dissolving polyvinyl alcohol in deionized water at 75° C. to prepare a 2 wt % polyvinyl alcohol solution which is used as the continuous phase W and the collection liquid Wo of the microemulsion droplets;

[0066] (5) The dispersed phase O is injected into the inner channel of the two-phase microchannel, the diameter of the inner channel is set to 300 μm, and the injection rate is 10 mL / h; the continuous phase W is injected into the outer channel of the two-phase microchannel, the diameter of the outer channel is set to 780 μm, and the injection rate is 30 mL / h; the dispersed phase O is sheared into microemulsion droplets of uniform size by the shearing action of the fluid;

[0067] (6) The microemulsion droplets obtained in step (5) were collected in the collection liquid Wo, allowed to stand for 48 hours, washed with deionized water five times, and freeze-dried for 24 hours to obtain PLCL / collagen composite microcarriers. Scanning electron microscopy showed that the microcarriers exhibited a heterogeneous porous structure, with a compact structure on one side of the collagen (e.g. Figure 6 ).

[0068] In summary, the present invention utilizes the shear force and surface tension between microfluids to prepare microemulsion droplets with uniform particle size and consistent composition; the present invention adopts a static solidification or horizontal rotation solidification method to fully volatilize the organic solvent in the oil phase, and the composite microemulsion droplets are gradually phase-separated into a Janus structure and shrunk into a composite microcarrier; a deionized water rinsing method is adopted to remove the surfactant and porogen remaining on the surface of the composite microcarrier; and a freeze-drying method is adopted to pre-freeze the composite microcarrier with deionized water in a solution to form a solid state, freeze-dry to remove moisture, and then collect the dried composite microcarrier, and the prepared heterogeneous composite structure microcarrier has uniform size; the present invention preferably selects artificial synthetic high molecular polymer components, biological macromolecules and coating drugs with good performance that meet production requirements and have biocompatibility and biodegradability, and through a reasonable component ratio and porogen The microcarrier has a unique heterogeneous morphology and excellent biological properties, as well as an adjustable degradation cycle. Compared with conventional microcarriers, the present invention is based on the principles of water-in-oil-in-water emulsion, solvent volatilization, phase separation and surface polymerization. Through the combination of two-phase microchannels used in the preparation process, the control of the orifice diameter and the fluid flow rate, and the regulation of the polymer ratio, the composite microcarrier has a single dispersion and a stable and controllable structure that is conducive to cell adhesion; the heterogeneous composite structure microcarrier prepared by the present invention has a concentrated particle size distribution, good initial cell adhesion efficiency and biodegradability. By changing the mass ratio of high molecular polymers and biomacromolecules and the oil-water phase ratio, the morphology and structure of the microcarrier and the degradation cycle can be precisely regulated, and adhesion sites and mechanical support can be provided for cells at the same time, meeting the carrier function required for in vitro cell culture and in vivo implantation.

[0069] The above-mentioned embodiments only express several implementation methods of the present invention, and the descriptions thereof are relatively specific and detailed, but they cannot be understood as limiting the scope of the invention patent. It should be pointed out that, for ordinary technicians in this field, several variations and improvements can be made without departing from the concept of the present invention, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention shall be subject to the attached claims.

Claims

1. A heterogeneous composite structure microcarrier, characterized in that: The heterogeneous composite structure microcarrier is composed of a spherical body composed of synthetic high molecular polymers and biological macromolecules, a coating drug coated on the surface of the spherical body and a connected pore structure; the synthetic high molecular polymer is an oil-soluble high molecular polymer, the biological macromolecule is a water-soluble biological macromolecule, and the coating drug is polydopamine.

2. A heterogeneous composite structure microcarrier according to claim 1, characterized in that: Calculated based on the total mass percentage of the heterogeneous composite structure microcarrier, the oil-soluble high molecular polymer is poly (L-lactic acid-caprolactone copolymer), and its content is 50% to 70%. The water-soluble biomacromolecule is collagen, and the collagen is type I fish collagen, and its content is 20% to 48%. The content of polydopamine is 2% to 10%.

3. A heterogeneous composite structure microcarrier according to claim 2, characterized in that: The molar ratio of LA to CL in the poly (L-lactic acid-caprolactone) copolymer is 90-10:10-90.

4. A heterogeneous composite structure microcarrier according to claim 1, characterized in that: The particle size of the heterogeneous composite structure microcarrier is 120 to 350 microns, the pore size is 3 to 50 microns, and the porosity is 10% to 80%.

5. The controllable preparation method of heterogeneous composite structure microcarrier according to any one of claims 1 to 4, characterized in that: The steps include: (1) Dissolving collagen and coating drugs in dilute acetic acid to obtain a hydrophilic solution A; (2) dissolving the poly (L-lactic acid-caprolactone) copolymer and the porogen in a volatile organic solvent to obtain a lipophilic solution B; (3) mixing the solution A prepared in step (1) and the solution B prepared in step (2) in a certain proportion and then mechanically homogenizing them to obtain a mixed emulsion as dispersed phase O; (4) dissolving the surfactant in deionized water to obtain a surfactant aqueous solution which serves as the continuous phase W and the collection liquid Wo of the microemulsion droplets; (5) adjusting the pH of the continuous phase W and the collected liquid Wo to 7.5-9.0; (6) The continuous phase W and the dispersed phase O are injected into the two-phase microchannel respectively, and the dispersed phase O is sheared into microemulsion droplets of uniform size; (7) The microemulsion droplets obtained in step (6) are received in a collecting liquid Wo, and allowed to stand or rotate horizontally to solidify, followed by washing and freeze-drying to obtain heterogeneous composite structure microcarriers.

6. The controllable preparation method of heterogeneous composite structure microcarrier according to claim 5, characterized in that: The porogen is selected from one of isooctane, camphene, n-tridecane and toluene.

7. The controllable preparation method of heterogeneous composite structure microcarrier according to claim 5, characterized in that: The particle size of the heterogeneous composite structure microcarrier is regulated by the flow rate of the dispersed phase O and the continuous phase W or the size of the two-phase microchannel or the pipeline combination; the flow rate of the dispersed phase O is controlled to be 1mL / h~20mL / h, and the flow rate of the continuous phase W is controlled to be 3mL / h~60mL / h; the inner channel diameter of the two-phase microchannel is 50 microns~500 microns, and the outer channel diameter is 300 microns~800 microns; the pipeline combination is selected from one of Y-type, T-type, cross-type, co-flow type, and flow focusing type.

8. The controllable preparation method of heterogeneous composite structure microcarrier according to claim 5, characterized in that: The pore structure of the heterogeneous composite structure microcarrier is regulated by the concentration of the porogen, the curing time and the curing method; the structural morphology of the heterogeneous composite structure microcarrier is regulated by the volume ratio of solution A to solution B and the concentration of collagen in solution A; the curing time is 24 to 72 hours, and when the curing method is horizontal rotation curing, the rotation speed is 20 to 80 rpm.

9. The controllable preparation method of heterogeneous composite structure microcarrier according to claim 5, characterized in that: The volatile organic solvent is selected from one of dichloromethane, chloroform and tetrahydrofuran; the coating drug is dopamine hydrochloride, and the concentration of the dopamine hydrochloride is 1-20 mg / ml; the concentration of collagen in the solution A is 0.3wt%-5wt%; the concentration of the poly-L-lactic acid-caprolactone copolymer in the solution B is 1.5wt%-5wt%, and the concentration of the porogen is 1wt%-5wt%; the solution A and the solution B are mixed in a volume ratio of 1-5:4-5 and then mechanically homogenized.

10. The controllable preparation method of heterogeneous composite structure microcarrier according to claim 5, characterized in that: In step (4), the surfactant is selected from one of sodium polyacrylate, polyvinyl alcohol and polyvinyl pyrrolidone; and the concentration of the surfactant aqueous solution is 0.5wt% to 4wt%.

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