CN114854042A

CN114854042A - Suspension medium for suspension 3D biological printing and preparation method and application

Info

Publication number: CN114854042A
Application number: CN202210490735.1A
Authority: CN
Inventors: 鲁路; 刘璐; 邱震; 周长忍
Original assignee: Jinan University
Current assignee: Jinan University
Priority date: 2022-05-07
Filing date: 2022-05-07
Publication date: 2022-08-05

Abstract

The invention belongs to the technical field of biological materials, and discloses a suspension medium for suspension 3D biological printing, and a preparation method and application thereof. The suspension medium comprises the following components in percentage by mass: 0-10% of gellan gum, 0-10% of thiolated gellan gum and the balance of solvent; and the contents of gellan gum and thiolated gellan gum are not zero at the same time. The suspension medium has good thixotropy and self-repairing capability, so that a spray head of the 3D biological printer can move freely in the suspension medium, and the integrity of a 3D biological printing structure can be kept; the problem that natural polymer materials such as collagen and fibrin can not realize self-supporting printing in the traditional printing is solved, the printing precision can reach 200 micrometers, the fidelity reaches 99%, the printing method is expected to be used for printing tissue and organ structures loaded with cells and active substances in vitro, and the printing method has a good industrialization prospect.

Description

Suspension medium for suspension 3D biological printing and preparation method and application

Technical Field

The invention belongs to the technical field of biological materials, and particularly relates to a suspension medium for suspension 3D biological printing, and a preparation method and application thereof.

Background

3D biological printing is used as a tissue engineering technology, and on the basis of 3D printing, living cells and biological materials are used as raw materials, the positioning of the biological materials, the living cells, functional components and the like in each layer is accurately controlled, and a 3D scaffold which can be used for transplantation is formed by printing layer by layer.

In 3D bioprinting, the main technologies currently used for biomaterial accumulation and molding are inkjet printing, micro-extrusion printing, and photo-assisted printing. The heating nozzle of the temperature-controlled ink-jet printer can enable the nozzle to generate pulse air pressure to print liquid drops, and the acoustic printer generates pulse pressing through piezoelectric type or ultrasonic type; a micro-extrusion printer uses a pneumatic or mechanical (piston or screw) drive system to extrude a continuous bead or strand containing material and cells; photo-assisted printing utilizes light focused on an energy absorbing substrate to provide a driving force for printing of biological materials. However, these conventional 3D bio-printing methods have certain drawbacks. For example, printing inks in inkjet printing must form solid 3D structures with structural organization and function. In extrusion 3D bioprinting, auxiliary support structures are often required to fabricate stents with complex 3D structures, such as the gastric tract, the tubule trachea, and the tumor vascular network, and it is therefore difficult to prepare vascularized structures with spatially defined features.

Suspension 3D printing is to use suspension medium material to provide support, extrude liquid phase material from a syringe into a support bath, and form a scaffold with a complex 3D structure. The working principle of the suspended 3D printing technique requires that the ink material must gel rapidly into a filament without spreading. The technology improves the forming stability of 3D biological printing ink with weak mechanical strength in the printing process without additionally providing a printing support structure. The currently commonly used suspension medium material is mainly gelatin microparticles, the size and uniformity of the microparticles directly influence the printing precision, and the preparation process of the support bath is complex. Therefore, the development of suspension media materials with good printing properties is one of the key issues that contribute to the development and implementation of this technology.

Gellan gum is an anionic extracellular bacterial polysaccharide, the repeating units are formed by connecting 1, 3-beta-D-glucose, 1, 4-beta-D-glucuronic acid, 1, 4-beta-D-glucose and 1, 4-alpha-L-rhamnose through glycosidic bonds. The natural gellan gum shows obvious rheological behavior of Bingham fluid, has yield stress, rapidly reduces viscosity under the action of certain stress, has good thixotropy, and has good biocompatibility, thereby laying a foundation for the application of the gellan gum in 3D printing.

Gellan gum can form a gel by: at high temperature, the gellan gum is in a random coil conformation; when the temperature is lowered, a thermally reversible double helix structural transition occurs, which is a prerequisite for the formation of a gel. Structures are then formed that consist of antiparallel double helices that self-assemble into directed bundles, called junction regions. The unwound regions of polysaccharide chains connect the junction regions in the form of helically extending chains, thereby allowing the formation of a three-dimensional network of gel. By utilizing the thermal reversibility of the gellan gum, after the 3D structure is printed, the gellan gum gel bath is melted by heating, and the printed structure is taken out. No gellan based suspension medium has been reported.

Disclosure of Invention

To overcome the above-mentioned drawbacks and deficiencies of the prior art, it is a primary object of the present invention to provide a suspension medium for suspending 3D bio-printing.

Another object of the present invention is to provide a process for the preparation of the suspension medium described above.

It is a further object of the present invention to provide the use of a suspension medium as described above.

The purpose of the invention is realized by the following scheme:

a suspension medium for suspending 3D bioprinting, comprising, but not limited to, the following components in mass percent: 0-10% of gellan gum, 0-10% of thiolated gellan gum and the balance of solvent; and the contents of the gellan gum and the thiolated gellan gum are not zero at the same time.

Furthermore, the thiolated gellan gum is obtained by introducing a sulfhydryl group into the gellan gum.

Further, the suspension medium for suspending 3D bioprinting of the present invention includes, but is not limited to, the following components in mass percent: 0-5% of gellan gum, 0-5% of thiolated gellan gum and the balance of solvent; and the contents of the gellan gum and the thiolated gellan gum are not zero at the same time.

Furthermore, the suspension medium of the invention can also contain the following components in percentage by mass: 0-2% of pH sensitivity regulator, 0-10% of temperature sensitive agent, 0-2% of thickening agent and 0-2% of reinforcing agent.

In the present invention, the pH-sensitive modifier may be a pH modifier conventionally used in the art, such as at least one of sodium alginate, chitosan and derivatives thereof, and the like.

In the present invention, the temperature sensitive agent is a temperature sensitive agent conventionally used in the art, and may include, but is not limited to, at least one of gelatin, collagen, agar, and derivatives of the above compounds.

In the present invention, the thickener is a water-soluble polymer thickener conventionally used in the art, and may include, but is not limited to, at least one of conventional natural gums (e.g., hyaluronic acid, agar, alginate, gelatin, etc.), celluloses (e.g., carboxyethyl cellulose, carboxymethyl cellulose, etc.), polyacrylics (e.g., sodium polyacrylate, etc.), polyurethanes (e.g., water-soluble polyurethanes, etc.), polyoxyethylene thickeners, and modified derivatives thereof.

In the present invention, the enhancer is an enhancer conventionally used in the art, and may include, but is not limited to, at least one of silk fibroin, chitin, derivatives thereof, and the like.

In the present invention, the solvent may include, but is not limited to, at least one of water, PBS, cell culture medium, and the like.

The suspension medium is a system based on gellan gum and/or sulfhydrylation gellan gum, shows obvious rheological behavior of Bingham fluid, has yield stress, rapidly reduces viscosity under the action of certain stress, has good thixotropy, and shows rapid self-repairing performance after the stress is eliminated; meanwhile, disulfide bonds can be formed among the gellan gums due to the introduction of the sulfydryl groups, the gellan gums belong to dynamic chemical bonds, the mechanical property, the thixotropy and the self-healing property of the gellan gums are improved, the gelation speed of the gellan gums can be improved, the gellan gums have better mechanical property compared with the existing gelatin system, and the gellan gum is more suitable for suspended 3D bioprinting.

The invention also provides a preparation method of the suspension medium, which is prepared by mixing the components in proportion, heating for dissolving, then preserving heat and gelating.

The temperature for heating and dissolving can be 60-90 ℃.

The temperature of the heat preservation can be 20-30 ℃, and is preferably 25 ℃.

The heat preservation time can be 10min-2 h.

The invention also provides application of the suspension medium in suspension printing. In the suspension medium, an extrusion type 3D biological printer is adopted, biological ink is extruded in the extrusion type 3D biological printer for 3D printing, a 3D model with a specific structure is constructed, the printing structure is further crosslinked and solidified, and the printing structure is obtained after the suspension medium is removed.

The crosslinking curing refers to that the biological ink can be further crosslinked and cured by adjusting the pH, the temperature, the ionic strength, the photocuring and the like of the system at the same time of printing or after the printing is finished.

The bio-ink can be natural polymer bio-material with weak mechanical strength, such as collagen, chitosan, alginic acid, gelatin and derivatives thereof.

The suspension medium has good thixotropy and self-repairing capability, so that a spray head of the 3D biological printer can move freely in the suspension medium, and the integrity of a 3D biological printing structure can be kept; after printing is completed, the suspension medium can be removed by utilizing the thermal reversibility of the suspension medium or by performing alkali treatment, and the printed structure can be taken out.

The suspension medium disclosed by the invention effectively improves the mechanical property of a system by constructing the hydrogel with the double-network structure, simultaneously keeps the thixotropy and the gel property of the gellan gum/thiolated gellan gum, can realize 3D biological printing of self-supporting materials, directly constructs micron-sized fine structures such as spiral structures, columnar structures and other structures containing suspended parts or hollow structures, solves the problem that natural high polymer materials with weak mechanical strength such as collagen and fibrin in the traditional printing cannot realize self-supporting printing, and is expected to be used for in-vitro printing of tissue and organ structures loaded with cells and active substances.

Compared with the existing microsphere suspension medium, the suspension medium based on gellan gum and/or thiolated gellan gum simplifies the process flow of suspension medium preparation, and has printing precision reaching 200 micron level, fidelity reaching 99 percent and good industrialization prospect.

Drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings needed to be used in the embodiments will be briefly described below, it should be understood that the following drawings only illustrate some embodiments of the present invention and therefore should not be considered as limiting the scope, and for those skilled in the art, other related drawings can be obtained according to the drawings without inventive efforts.

FIG. 1 is a graph of gellan gum viscosity versus shear rate.

FIG. 2 is a graph of gellan gum viscosity versus time.

FIG. 3 is a graph of shear modulus of gellan gum versus time.

FIG. 4 is a schematic view of a suspended 3D bioprinting apparatus; wherein, 1 is a printing ink tube, 2 is a printing needle, 3 is a suspension medium, and 4 is a printed model.

FIG. 5 is a diagram of a model printed in example 1.

FIG. 6 is a graph of compressive stress-strain curves for gellan gum and thiolated gellan gum.

FIG. 7 is a graph showing the relationship between Young's modulus and compressive strength of gellan gum and thiolated gellan gum.

FIG. 8 is a temperature scan of gellan gum and thiolated gellan gum.

FIG. 9 is a diagram of a model printed in example 5.

FIG. 10 is a diagram of a model printed in example 7.

Detailed Description

The present invention will be described in further detail with reference to examples, but the embodiments of the present invention are not limited thereto. The materials referred to in the following examples are commercially available without specific reference. The method is a conventional method unless otherwise specified.

The thiolated gellan gum used in the following examples was prepared by introducing a thiol group into gellan gum, either as is commercially available or by conventional methods in the art, or by the following method, as shown in formula one below:

heating and dissolving gellan gum in water to obtain a solution with the concentration of 1.5 wt%, wherein the temperature is 80 ℃; then gelatinizing at 25 ℃; the obtained gel block was immersed in a mixed solution (0.5 mmol of 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride, 0.5mmol of N-hydroxysuccinimide and 0.5mmol of L-cysteine dissolved in 20mL of water) at 4 ℃ for 12 hours. The entire sulfhydrylation reaction of gellan gum is carried out in the gel state. Followed by purification by dialysis, freeze-drying and further use.

Example 1

A suspension medium for suspending 3D bio-printing comprises the following components in percentage by mass: 2.8 percent of gellan gum and the balance of water.

The preparation method comprises the following steps: dissolving gellan gum in water of 80 ℃ according to mass percent, stirring uniformly, fully dissolving, and cooling for 45min at 25 ℃ until the gellan gum becomes a gel state, thereby obtaining the suspension medium. The resulting media had very good thixotropic properties and the specific rheological data are shown in figures 1-3.

As can be seen from FIG. 1, the viscosity of gellan gum decreases with increasing shear rate, reflecting the rheological behavior of gellan gum with a pronounced Bingham fluid. To better understand the rheological properties of the suspension medium during printing and to compare the recovery and support properties of different concentrations of gellan gum, a static shear recovery test and an oscillation time sweep test were performed. The shear rate during printing can be calculated from the diameter and speed of movement of the needle. Shear rate of about 30s during printing ^-1 -35s ^-1 In the experiment, the shear rate was switched in 3 steps from low to high to low, and the results are shown in fig. 2 and 3. The result shows that the gellan gum has good viscosity repairing performance, the viscosity recovery degree is better along with the reduction of the concentration, the performance of the gellan gum as a suspension medium is better, but the strength and the supporting capability are weaker under the consideration of the lower concentration, and therefore, the influence of the concentration on the performance is larger. Figure 3 shows that the gel-sol transition of gellan gum is almost instantaneous and the storage modulus increases significantly with increasing gellan gum concentration.

The application comprises the following steps:

(1) according to the mass fraction, 20% GelMA is dissolved in deionized water at 60 ℃, evenly stirred, fully dissolved and stored at 25 ℃ to obtain the hydrogel ink required by printing.

(2) The suspension medium of the invention is used as a printing suspension medium for 3D biological printing, and the printing schematic diagram is shown in figure 4. With 20 wt% GelMA waterPlacing the solution as printing ink in a printing ink tube, printing a target structure model in the suspension medium with a printing needle diameter of 200 μm, and irradiating the material in an ultraviolet reactor for 5min after printing, wherein the ultraviolet wavelength is 312nm and the radiation intensity is 50J/cm ² The suspension medium was placed in hot water at 60 ℃ together with the model, and the suspension medium was removed to obtain a model as shown in FIG. 5.

(3) And (3) carrying out body type microscope shooting on the printed model, and determining the fidelity according to the ratio of the actual height/diameter of the printed model to the theoretical height/diameter, wherein the result shows that the fidelity is 89%.

Example 2

A suspension medium for suspending 3D bio-printing comprises the following components in percentage by mass: 2% of sulfhydrylation gellan gum and the balance of water.

The preparation method comprises the following steps: according to the mass percentage, the thiolated gellan gum is dissolved in water at 80 ℃, stirred evenly and cooled for 25min at 25 ℃ after being fully dissolved until the thiolated gellan gum becomes a gel state, and then the suspension medium is obtained.

The gellan gum of example 1 and the thiolated gellan gum of this example were subjected to mechanical property tests, and the specific mechanical property data are shown in fig. 6 to 8, which shows that the thiolated gellan gum has better mechanical properties than gellan gum.

From fig. 6-8, it can be seen that the young's modulus of the thiolated gellan gum is about three times that of the gellan gum, and the compressive strength is very close. The fracture strain of the thiolated gellan gum is lower than that of the gellan gum, which is mainly due to the increase of the crosslinking density in the double-network hydrogel, so that the toughness of the thiolated gellan gum can be effectively improved by adding the gellan gum. Temperature scanning tests show that the thiolated gellan gum and the gellan gum have similar sol-gel transition behaviors, the transition temperature is about 37 ℃, and the gellan gum and the thiolated gellan gum sol can be converted into gel at the temperature below 37 ℃.

The application comprises the following steps:

(1) according to the mass percentage, 5 percent of chitosan is dissolved in 2 percent of acetic acid and is uniformly mixed to obtain the hydrogel ink required by printing.

(2) The suspension medium is used as a printing suspension medium to carry out 3D biological printing, the hydrogel ink is placed in an ink tube of a 3D biological printer, the diameter of a printing needle is 200 microns, a target structure model is printed in the suspension medium, the suspension medium and the printing model are placed into a 2% NaOH solution after printing, the suspension medium is removed, and the printing structure is solidified.

(3) And (4) carrying out fidelity measurement on the obtained curing printing model, wherein the result shows that the fidelity can reach 95%.

Example 3

A suspension medium for suspending 3D bio-printing comprises the following components in percentage by mass: 1% of gellan gum, 2% of thiolated gellan gum and the balance of water.

The preparation method comprises the following steps: dissolving 1% gellan gum and 2% sulfhydrylation gellan gum in water at 80 ℃ by mass fraction, stirring uniformly, fully dissolving, and cooling at 25 ℃ for 30min until the gel state is achieved, thus obtaining the suspension medium.

The application comprises the following steps:

(1) according to the mass percentage, 20 percent of GelMA is dissolved in deionized water at 60 ℃, evenly stirred and stored at 25 ℃ after being fully dissolved, so that the hydrogel ink required by printing is obtained.

(2) 3D biological printing is carried out by taking the suspension medium as a printing suspension medium, the hydrogel ink is placed in an ink tube of a 3D biological printer, the diameter of a printing needle is 200 micrometers, a target structure model is printed in the suspension medium, after printing, the suspension medium and the printing model are placed in an ultraviolet reactor to be irradiated for 5min, the ultraviolet wavelength is 312nm, and the radiation intensity is 50J/cm ² And putting the suspension medium and the model into hot water of 80 ℃ together, and removing the suspension medium to obtain the printed model.

(3) And the fidelity of the obtained curing printing model is measured, and the result shows that the fidelity can reach 97%.

Example 4

The application comprises the following steps:

(1) diluting 35mg/mL collagen and 0.24Mol/L acetic acid at a volume ratio of 2:1, mixing well, and centrifuging at 3000g for 5 minutes to remove air bubbles to obtain the acidified collagen bio-ink.

(2) The suspension medium is used as a printing suspension medium to carry out 3D biological printing, the hydrogel ink is placed in an ink tube of a 3D biological printer, the diameter of a printing needle is 200 microns, a target structure model is printed in the suspension medium, after printing, the suspension medium and the printing model are placed in an ultraviolet reactor to be irradiated for 5min, the ultraviolet wavelength is 312nm, the radiation intensity is 50J/cm2, the suspension medium and the model are placed in hot water of 80 ℃, and the suspension medium is removed to obtain the printed model.

Example 5

A suspension medium for suspending 3D bio-printing comprises the following components in percentage by mass: gellan gum 2%, thiolated gellan gum 1.5%, pH sensitive regulator 2% (sodium alginate), and water in balance.

The preparation method comprises the following steps: dissolving 2% of gellan gum and 1.5% of thiolated gellan gum in water at 80 ℃ by mass, stirring uniformly, adding 2% of sodium alginate into the mixture to dissolve fully, and cooling at 25 ℃ for 30min until the mixture becomes a gel state, thereby obtaining the suspension medium.

The application comprises the following steps:

(2) 3D biological printing is carried out by taking the suspension medium of the invention as a printing suspension medium, the hydrogel ink is placed in an ink tube of a 3D biological printer, the diameter of a printing needle is 200 microns, a target structure model is printed in the suspension medium, after printing, the suspension medium and the printing model are placed into a 2% NaOH solution, the suspension medium is removed, and the printing structure is solidified, so that the obtained model is shown in figure 9.

(3) And the fidelity of the obtained curing printing model is measured, and the result shows that the fidelity can reach 99%.

Example 6

A suspension medium for suspending 3D bio-printing comprises the following components in percentage by mass: gellan gum 2%, thiolated gellan gum 1%, temperature sensitive agent 2% (gelatin), and water in balance.

The preparation method comprises the following steps: dissolving 2% gellan gum and 1% sulfhydrylation gellan gum in water at 80 ℃ by mass, stirring uniformly, adding 2% gelatin into the mixture to dissolve fully, and cooling at 25 ℃ for 30min until the gelatin state is formed, thus obtaining the suspension medium.

The application comprises the following steps:

(1) dissolving 20% of photo-crosslinkable methacrylate collagen (ColMA) in water by mass percent, and uniformly mixing to obtain the hydrogel ink required by printing.

(2) 3D biological printing is carried out by taking the suspension medium as a printing suspension medium, the hydrogel ink is placed in an ink tube of a 3D biological printer, the diameter of a printing needle is 200 microns, a target structure model is printed in the suspension medium, the suspension medium and the printing model are placed in an ultraviolet reactor to be irradiated for 5min after printing, the ultraviolet wavelength is 312nm, and the radiation intensity is 50J/cm ² And putting the suspension medium and the model into hot water of 60 ℃ together, and removing the suspension medium to obtain the printed model.

(3) And (4) carrying out fidelity measurement on the obtained curing printing model, wherein the result shows that the fidelity can reach 98%.

Example 7

A suspension medium for suspending 3D bio-printing comprises the following components in percentage by mass: gellan gum 2%, thiolated gellan gum 1%, reinforcing agent 2% (silk fibroin), and water in balance.

The preparation method comprises the following steps: dissolving 2% gellan gum and 1% sulfhydrylation gellan gum in water at 80 ℃ by mass, stirring uniformly, adding 2% silk fibroin into the solution to dissolve fully, and cooling at 25 ℃ for 30min until the solution becomes a gel state, thus obtaining the suspension medium.

The application comprises the following steps:

(2) 3D biological printing is carried out by taking the suspension medium as a printing suspension medium, the hydrogel ink is placed in an ink tube of a 3D biological printer, the diameter of a printing needle is 200 microns, a target structure model is printed in the suspension medium, the suspension medium and the printing model are placed in an ultraviolet reactor to be irradiated for 5min after printing, the ultraviolet wavelength is 312nm, and the radiation intensity is 50J/cm ² The suspension medium was placed in hot water at 80 ℃ along with the mold, and the suspension medium was removed to obtain a printed mold as shown in FIG. 10.

Comparative example 1

Printing was performed with gelatin microparticles as the suspension medium and compared to the fidelity of the printed model of gellan and/or thiolated gellan suspension media.

A suspension medium for suspending 3D bio-printing comprises the following components in percentage by mass: gelatin 2%, 0.25%

F-127, 0.1 percent of Arabic gum and the balance of ethanol and hydrochloric acid.

The preparation method comprises the following steps: according to the mass percentage, 2.0 percent of gelatin and 0.25 percent of

F-127 and 0.1% gum arabic were dissolved in a beaker containing 50% ethanol solution at 45 deg.C and added byThe pH was adjusted to 6.25 by addition of 1Mol/L hydrochloric acid. The beaker was placed under a stirrer, sealed with plastic wrap to minimize evaporation, and cooled to room temperature while stirring overnight. The resulting slurries were then poured separately into conical tubes and centrifuged for 5 minutes to compact the gelatin microparticles. The supernatant was then removed and the gelatin microparticles were resuspended in phosphate buffered saline at pH 7.4 to remove ethanol and

f-127. The gelatin slurry was then washed three times with phosphate buffered saline at pH 7.4 for 2 minutes. Before printing, the slurry was evacuated for 15 minutes in a vacuum chamber and then centrifuged at 2000g for 5 minutes. The supernatant was removed to give gelatin microparticles as the suspension medium.

The application comprises the following steps:

(2) And (2) taking the gelatin particle suspension medium as a printing suspension medium, placing the acidified collagen in an ink tube of a 3D bioprinter, printing a target structure model with the diameter of a printing needle being 200 microns, placing the suspension medium and the printing model in an ultraviolet reactor to irradiate for 5min after printing, wherein the ultraviolet wavelength is 312nm, and the radiation intensity is 50J/cm2, and stripping the suspension medium by a physical means to obtain the printing model.

(3) The resulting cured print model was subjected to fidelity measurement, which showed a fidelity of 78%. The two-component system of the present invention has higher fidelity compared to the lower fidelity of gelatin microparticles.

The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.

Claims

1. A suspension medium for suspending 3D bioprinting, characterized by comprising but not limited to the following components in mass percent: 0-10% of gellan gum, 0-10% of thiolated gellan gum and the balance of solvent; and the contents of the gellan gum and the thiolated gellan gum are not zero at the same time.

2. The suspension medium for suspended 3D bioprinting of claim 1, further comprising the following components in mass percent: 0-2% of pH sensitivity regulator, 0-10% of temperature sensitive agent, 0-2% of thickening agent and 0-2% of reinforcing agent.

3. The suspension medium for suspended 3D bioprinting of claim 2, wherein: the pH sensitivity regulator comprises at least one of sodium alginate, chitosan and derivatives thereof.

4. The suspension medium for suspended 3D bioprinting of claim 2, wherein: the temperature sensitive agent comprises at least one of gelatin, collagen, agar and derivatives of the above compounds.

5. The suspension medium for suspended 3D bioprinting of claim 2, wherein: the thickening agent comprises at least one of natural gum, cellulose, polyacrylic acid, polyurethane, polyoxyethylene thickening agent and modified derivatives thereof.

6. The suspension medium for suspended 3D bioprinting of claim 2, wherein: the reinforcing agent comprises at least one of silk fibroin, chitin and derivatives thereof.

7. The suspension medium for suspended 3D bioprinting of claim 2, wherein: the solvent comprises at least one of water, PBS and cell culture medium.

8. A method for preparing a suspension medium for suspended 3D bioprinting according to any one of claims 1 to 7, wherein the suspension medium is obtained by mixing the components in proportion, heating to dissolve, then holding the temperature, and allowing to gel.

9. Use of the suspension medium for suspended 3D bioprinting of any of claims 1-7 in suspended printing.

10. Use according to claim 9, characterized in that it is in particular: in the suspension medium for suspension 3D bioprinting according to any one of claims 1 to 7, an extrusion type 3D bioprinter is used, wherein the extrusion type 3D bioprinter extrudes bio-ink to perform 3D printing to construct a 3D model of a specific structure, the printed structure is further cross-linked and cured, and after the suspension medium is removed, the printed structure is obtained.