KR101947154B1 - Fabrication method and device for three-dimensional electrospun scaffold - Google Patents

Abstract

A method and an apparatus for manufacturing a three-dimensional electrospinning scaffold are provided. A three-dimensional electrospinning scaffold manufacturing apparatus comprises a chamber for containing a medium containing cells, at least two driving sources for applying at least two kinds of physical forces to the medium to cause at least two kinds of mixed flow, A syringe positioned above the chamber and containing the scaffold material and supplied with a discharge pressure from the syringe pump, a high voltage power source electrically connected to the syringe, and a ground electrode for grounding the medium.

Description

TECHNICAL FIELD [0001] The present invention relates to a three-dimensional electrospinning scaffold,

The present invention relates to a three-dimensional electrospinning scaffold, and more particularly, to a method and apparatus for manufacturing a three-dimensional electrospinning scaffold in which uniform distribution and intrusion of cells are improved.

Electrospinning is a simple and widely used method of fabricating micrometer or nanometer scale fibers using electric fields. Especially, microfibers obtained by electrospinning have a size and distribution similar to those of extracellular matrix, so they can be used as a scaffold, which is an artificial support for controlling tissue functions, tissue growth, cell growth and tissue function in tissue engineering.

Most microfibrous scaffolds require a lot of time to fabricate and have limited pore size because they form a three-dimensional structure by laminating the fibers. Since the pore structure of the scaffold greatly affects the inoculation, attachment and cultivation of cells, various studies are being conducted to overcome this problem.

The inventors of the present application proposed a method of manufacturing a scaffold by eccentrically rotating a medium containing cells in an earlier study (non-patent reference of the following prior art document) and applying an electric field. According to this method, the procedure from preparation to cell culture can be shortened, and scaffold contamination can be reduced because no additional process is required.

&Quot; Enhanced cellular distribution and infiltration in a wet electrospun three-dimensional fibrous scaffold using eccentric rotation-based hydrodynamic conditions ", Sensors and Actuators B: Chemical Volume 226, April 2016, Pages 357-363 The inventor of the present application intends to propose a method for easily creating a scaffold of various characteristics suitable for a desired intention as a follow-up study. Scaffolds that can replace organs of the human body are different in characteristics from each other, and continuous improvement of performance is required.

The present invention provides a method and apparatus for manufacturing a three-dimensional electrospinning scaffold capable of producing an improved scaffold having various mechanical properties, porosity, biodegradability, and cell-friendly surface characteristics suitable for the required characteristics.

A method of manufacturing a three dimensional electrospinning scaffold according to an embodiment of the present invention includes a preparation step, a flow generation step, and an electrospinning step. The preparation step consists of placing the medium containing the cells in a chamber and placing the syringe with the scaffold material on top of the medium. The flow generating step consists in applying at least two kinds of physical forces to the medium to cause a non-uniform compound flow in which at least two kinds of flows are mixed in which the driving source is different. The electrospinning step consists of generating a voltage difference between the medium and the syringe and activating the syringe pump to emit the scaffold material in the form of fibers toward the surface of the medium where complex flow occurs from the syringe. In the electrospinning step, the scaffold material is entangled three-dimensionally to form a fibrous structure, and cells penetrate into the fibrous structure to form a scaffold.

In the preparation stage, a vortex generator and an ultrasonic device may be installed in the chamber. In the flow generating step, ultrasonic waves can be applied at the same time that the flow is generated by the vortex generator in the medium. The ultrasonic waves applied to the medium may be ultrasonic waves for treatment having a frequency of 1 MHz to 3 MHz and an intensity of 0.1 W / cm 2 to 2.0 W / cm 2.

On the other hand, a vortex generator and a bubble generator may be installed in the chamber during the preparation step. The flow caused by the vortex generator and the flow caused by the bubble diffusion can occur at the same time in the flow generating step and the oxygen saturation of the medium can be increased.

On the other hand, a vortex generator and a magnet rotating device may be installed in the chamber during the preparation step. The magnet rotating device may include a bar magnet contained in the medium and a magnetic field applying section located outside the chamber. In the flow generating step, a flow caused by the vortex generator in the medium and a vortex flow caused by the rotation of the bar magnet can occur at the same time.

After the electrospinning step, the medium can be replaced with another kind of medium containing different kinds of cells, and the flow generation step and the electrospinning step can be repeated. In the second electrospinning step, the fibrous structure can be divided into a central portion where the first cells are located and an outer portion where the second cells are located.

In the preparation step, growth aid granules may be added to the medium.

The preparation in the preparation step may consist of a plurality of syringes containing different scaffold materials. In the electrospinning step, a plurality of syringes can be simultaneously applied with a high voltage, and simultaneously, a discharge pressure can be supplied from a syringe pump to simultaneously radiate the scaffold material toward the medium, and two or more kinds of scaffold materials are entangled, .

On the other hand, in the electrospinning step, a plurality of syringes are sequentially applied with a high voltage, and at the same time, the discharge pressure is sequentially supplied from the syringe pump, and the scaffold material can be sequentially radiated toward the medium. The material can form a fibrous structure while forming its own layer.

The apparatus for manufacturing a three-dimensional electrospinning scaffold according to an embodiment of the present invention includes a chamber for containing a medium containing cells, and a non-uniform compounded mixture in which at least two types of flows are mixed by applying at least two kinds of physical forces to the medium. A high voltage power source electrically connected to the syringe, a ground electrode for grounding the medium, at least two drive sources for generating a flow, a syringe positioned above the chamber and containing a scaffold material and supplied with a discharge pressure from the syringe pump, . The at least two driving sources include a vortex generator, and further include at least one of an ultrasonic device, a bubble generator, and a magnet rotating device.

The ultrasonic apparatus may include an ultrasonic driving unit and an ultrasonic transducer connected to the ultrasonic driving unit and in contact with the medium. The ultrasonic transducer may be provided with a treatment ultrasonic wave having a frequency of 1 MHz to 3 MHz and a strength of 0.1 W / can do.

The magnet rotating device may include a bar magnet contained in the medium and a magnetic field applying portion located outside the chamber, and may cause a swirl flow in the medium by rotating the bar magnet.

The syringes may comprise a plurality of syringes containing different scaffold materials. The plurality of syringes may simultaneously spin the scaffold material toward the medium or sequentially spin the scaffold material for a predetermined time difference.

According to the embodiments, the cells can evenly penetrate into the scaffold, and even distribution of cells to the entire scaffold is possible, maximizing the functionality of the scaffold. In addition, scaffold fabrication and cell seeding can be performed at the same time as conventional scaffold fabrication and cell seeding processes, simplifying manufacturing and facilitating mass production.

1 and 2 are block diagrams of a scaffold manufacturing apparatus according to a first embodiment of the present invention.
FIG. 3 is a block diagram showing a movement path of a rotating shaft included in the vortex generator in the scaffold manufacturing apparatus shown in FIG. 1. FIG.
Fig. 4 is a block diagram showing the state before and after voltage application of a syringe in the scaffold manufacturing apparatus shown in Fig. 1. Fig.
5 is a flowchart showing a method of manufacturing a scaffold of the first embodiment using the apparatus for producing scaffolds shown in Fig.
6 is a configuration diagram of a scaffold manufacturing apparatus according to a second embodiment of the present invention.
7 is a partial enlarged view of Fig.
Fig. 8 is a diagram showing the comparison of the cell proliferation in the case of the comparative example in which the oxygen saturation is not raised and in the case of the second embodiment in which the oxygen saturation degree is raised.
FIG. 9 and FIG. 10 are block diagrams of a scaffold manufacturing apparatus according to a third embodiment of the present invention.
FIG. 11 is a block diagram for explaining a scaffold manufacturing method according to a fifth embodiment of the present invention.
12 is a schematic configuration diagram of a scaffold fabricated by the method of the fifth embodiment.
FIG. 13 is a structural view for explaining a scaffold manufacturing method according to a sixth embodiment of the present invention.
14 and 15 are block diagrams of a scaffold manufacturing apparatus according to a seventh embodiment of the present invention.
16A to 16D are block diagrams showing a method for manufacturing a scaffold according to an eighth embodiment using the apparatus for manufacturing a scaffold of the seventh embodiment.
17 is a schematic view showing a scaffold manufactured by the method of the eighth embodiment.

Hereinafter, exemplary embodiments of the present invention will be described in detail with reference to the accompanying drawings, which will be readily apparent to those skilled in the art to which the present invention pertains. The present invention may be embodied in many different forms and is not limited to the embodiments described herein.

FIG. 1 and FIG. 2 are block diagrams of an apparatus for fabricating a three-dimensional electrospinning scaffold according to a first embodiment of the present invention (hereinafter referred to as a "scaffold manufacturing apparatus").

1 and 2, the scaffold manufacturing apparatus 100 of the first embodiment includes a chamber 10 for accommodating a medium 12 containing cells 11, a vortex generator (not shown) installed in the chamber 10, A high voltage power supply 51 for generating a voltage difference necessary for electrospinning in the medium 12 and the syringe 40; And the ground electrode 52. [

The medium 12 is selected as a cell culture medium to be infiltrated into the scaffold, and the medium 12 is selected. Fetal bovine serum in the medium 12 may be added only during incubation of the cells 11 and not added to avoid foaming due to flow during scaffold fabrication. May include a rotating shaft 21 coupled to the lower side of the chamber 10 and a driving unit 22 for driving the rotating shaft 21. When the rotary shaft 21 is rotated by the operation of the driving unit 22, the chamber 10 coupled to the rotary shaft 21 rotates together to cause a flow in the medium 12 accommodated in the chamber 10. At this time, the rotary shaft 21 of the vortex generator 20 can move along a predetermined path.

FIG. 3 is a block diagram showing a movement path of a rotating shaft included in the vortex generator in the scaffold manufacturing apparatus shown in FIG. 1. FIG. Referring to FIG. 3, the rotating shaft of the vortex generator can move along a predetermined circular path at the same time as rotating. In this case, the vortex generators cause a strong eccentric rotation in the chamber, resulting in non-uniform flow in the medium.

Referring again to FIGS. 1 and 2, scattering of the cells 11 and a large number of water droplets occurs on the surface of the medium 12 where uneven flow occurs. Uneven flow of the medium 12 promotes the formation of the fibrous structure 13 intricately entangled three-dimensionally in the fiber spinning process described below, resulting in intricate mixing between the cell 11 and the fibrous structure 13.

The ultrasound device 30, along with the vortex generator 20, serves as an additional source of flow that causes a flow to the medium 12. That is, the vortex generator 20 and the ultrasonic device 30 apply two types of physical force to the medium 12 contained in the chamber 10 to cause a non-uniform composite flow in which the two kinds of flows of the driving source are mixed.

The ultrasonic device 30 may include an ultrasonic driving unit 31 and an ultrasonic transducer 32 connected to the ultrasonic driving unit 31 and in contact with the medium 12. The ultrasonic transducer 32 applies ultrasonic waves to the medium 12 where uneven flow occurs by the vortex generator 20.

The ultrasound transmits mechanical energy to the cells 11 contained in the medium 12 as an energy form having a frequency outside the hearing range of the human being. Ultrasound can be classified into diagnostic, cleaning, and therapeutic depending on frequency and intensity. Therapeutic ultrasonic waves have a frequency of about 1 to 3 MHz and an intensity of 0.1 to 2.0 W / cm 2, and have a physical effect on cells and tissues by thermal and non-thermal mechanisms.

The ultrasonic device 30 can transmit ultrasound to the medium 12 at a frequency of approximately 1 to 3 MHz and an intensity of 0.1 to 2.0 W / cm 2 corresponding to the therapeutic ultrasonic wave. In this case, the ultrasonic device 30 can cause the ultrasonic wave to flow to the medium 12 and cause a physical effect on the cells 11 contained in the medium 12.

Especially, the low intensity ultrasound energy field and the electromagnetic field of special wavelength are known to promote cell growth. For example, osteogenic cells that are sensitive to physical stimuli can promote bone formation in low current and low intensity ultrasound energy fields and electromagnetic fields of special wavelengths.

The syringe 40 receives a scaffold material (polymer solution) therein and is connected to a syringe pump 41 for applying pressure to the scaffold material to push out the scaffold material. The high voltage power supply 51 applies a voltage of several kV to the syringe 40 and the ground electrode 52 can be placed on the bottom plate of the chamber 10 and locked to the medium 12. [ A voltage difference necessary for electrospinning is generated between the medium 12 and the syringe 40 by the high voltage power source 51 and the ground electrode 52. [

4 is a configuration diagram showing states before and after voltage application of a syringe. Referring to FIG. 4, when the polymer solution is discharged to the tip of the syringe 40, the polymer solution forms a round shape at the bottom of the tip due to the surface tension. At this time, when a high voltage is applied to the syringe 40, a high positive charge begins to accumulate in the polymer solution.

When the positive charge builds up above the limit, the repulsive force between the charges in the polymer solution gradually increases, and when the increased repulsive force exceeds the surface tension, the polymer solution from the tip of the syringe 40 forms an approximate Taylor cone shape And is radiated.

Referring again to FIG. 2, the radiated polymer solution is in the form of fibers as the solvent dries, and the spun fibers fall on the surface of the medium 12 where uneven flow is occurring and are mixed with the cells 11.

The fibers radiated from the syringe 40 become a fibrous structure 13 that is three-dimensionally intricately entangled by the nonuniform composite flow of the medium 12 and the fibrous structure 13 The cells 11 of the medium 12 uniformly permeate.

Assuming that the scaffold material is spun on a medium that is free of flow or that is being flowed by a single simple drive source, voids may not be uniformly generated inside the spindle material when the spunbond material forms a fiber structure have. In this case, cells of the medium can not penetrate evenly into the fibrous structure, cells are not evenly distributed throughout the scaffold, and the functionality as a scaffold is lowered.

However, in the first embodiment, two kinds of physical forces are applied to the medium 12 to cause a non-uniform composite flow in which two different kinds of flows are mixed in the driving source. As a result, when the radiated scaffold material falls to the medium 12, the scaffold material forms a three-dimensionally complex fibrous structure 13 by the composite flow, and the voids are uniformly formed in the fiber structure 13, The cells 11 of the culture medium 12 can be evenly distributed.

That is, the scaffold material falling into the medium 12 is entangled in three dimensions by the complex flow and has a substantially uniform pore size and a volume, and the volume of the fibrous structure 13 grows in the medium 12, The cells 11 can evenly penetrate into the pores of the fibrous structure 13. [

5 is a flowchart showing a method of manufacturing a scaffold of the first embodiment using the apparatus for producing scaffolds shown in Fig.

Referring to FIG. 5, a first step (S10) is a preparation step of placing a medium containing cells in a chamber and disposing a syringe containing the scaffold material in the upper part of the chamber. The second step S20 is a flow generating step in which at least two kinds of physical forces are applied to the medium contained in the chamber to cause a non-uniform compound flow in which the drive source is mixed with at least two kinds of flows.

The third step (S30) is to generate a voltage difference for electrospinning between the medium and the syringe and to operate the syringe pump to emit the scaffold material in the form of fibers from the syringe towards the surface of the medium where the compound flow occurs It is an electrospinning step.

In the second step S20, the two types of physical forces can be made by a combination of two mechanical forces by a vortex generator and an ultrasonic device. In the third step S30, the scaffold material radiated in the form of fibers is uniformly penetrated into the fibrous structure while forming a fibrous structure entangled intricately in three dimensions by the nonuniform composite flow of the medium, Constitute a scaffold.

The method of manufacturing the scaffold of the first embodiment can uniformly penetrate the cells into the scaffold and uniform distribution of the cells to the entire scaffold, maximizing the functionality of the scaffold, and applicable to various fields. In addition, scaffold fabrication and cell seeding can be performed at the same time as conventional scaffold fabrication and cell seeding processes, simplifying manufacturing and facilitating mass production.

FIG. 6 is a configuration diagram of a scaffold manufacturing apparatus according to a second embodiment of the present invention, and FIG. 7 is a partially enlarged view of FIG.

6 and 7, the scaffold manufacturing apparatus 200 of the second embodiment is configured by replacing the ultrasonic wave device 30 of the first embodiment with a bubble generator 60. [ The bubble generator 60 may include an air pump 61 and a bubble injection nozzle 62 connected to the air pump 61 and in contact with the medium 12. The bubble injecting nozzle 62 discharges the air of a predetermined pressure supplied from the air pump 61 into the medium 12 in the form of bubbles.

A large amount of bubbles 14 discharged into the medium 12 causes another kind of flow to the medium 12 along with the flow by the vortex generator 20. That is, in the medium 12, in addition to the non-uniform flow caused by the vortex generator 20, an internal flow due to the diffusion of the bubbles 14 occurs, resulting in a non-uniform composite flow in which two kinds of flows are mixed.

The method of manufacturing a scaffold according to the second embodiment is similar to the method of the first embodiment shown in Fig. 5 except that in the second step S20, two kinds of physical forces are applied to the vortex generator 20 and the bubble generator 60 ) Is a combination of two mechanical forces.

The large amount of bubbles 14 ejected from the bubble injection nozzle 62 helps the scaffold material that has fallen into the medium 12 to accumulate in three dimensions with the flow by the vortex generator 20 more quickly. A large amount of bubbles 14 are interposed between the fibrous structures 13 to form a space and provide an additional space inside the fibrous structure 13 in which the medium 12 and the cells 11 can enter more do.

The scaffolds produced by the method of the second embodiment include the voids created by electrospinning, the voids created by the uneven flow of the media 12 by the vortex generator 20, and the large bubble 14 diffusion The pores generated by the flow are mixed.

For example, the scaffold fabricated by the method of the second embodiment is mixed with various voids of several to several hundred micrometers (占 퐉) in size. The various sizes of voids formed within the scaffold facilitate cell migration and attachment within the scaffold and aid cell growth.

Furthermore, the air injected into the medium 12 increases the oxygen saturation of the medium 12. The increase in oxygen saturation of the medium 12 aids in the proliferation of the cells 11 and enhances the viability of the cells 11. [

Fig. 8 is a diagram showing the comparison of the cell proliferation in the case of the comparative example in which the oxygen saturation is not raised and in the case of the second embodiment in which the oxygen saturation degree is raised. 8, the case of the second embodiment in which the oxygen saturation degree of the culture medium is increased is more effective for cell proliferation than the comparative example which does not.

FIG. 9 and FIG. 10 are block diagrams of a scaffold manufacturing apparatus according to a third embodiment of the present invention.

9 and 10, the scaffold manufacturing apparatus 300 of the third embodiment is configured by replacing the ultrasonic wave device 30 of the first embodiment with a magnet rotating device 70. FIG. The magnet rotating device 70 may include a bar magnet 71 immersed in a medium and a magnetic field applying part 72 located outside the chamber 10. [ The magnetic field applying unit 72 may be embedded in the driving unit 22 of the vortex generator 20.

The rod magnet 71 is inserted into the chamber 10 after disinfection and can be locked in the medium 12 and causes the medium 12 to vortex while rotating inside the medium 12 by the magnetic field applying unit 72. The rotation of the bar magnet 71 causes another kind of flow in the medium 12 along with the flow by the vortex generator 20.

In other words, in the culture medium 12, uneven flow by the vortex generator 20 and vortex flow due to the rotation of the bar magnet 71 occur at the same time, resulting in a non-uniform composite flow in which two kinds of flows are mixed.

The method of manufacturing a scaffold according to the third embodiment is similar to the method of the first embodiment shown in Fig. 5 except that in the second step S20, two kinds of physical forces are applied to the vortex generator 20 and the magnet rotating apparatus 70) is a combination of two mechanical forces.

The swirling flow of the medium 12 due to the rotation of the bar magnets 71 facilitates rapid aggregation of the scaffold material that has fallen into the medium 12 in three dimensions along with the flow by the vortex generators 20, ) To improve the internal penetration of the cell (11).

The scaffold manufacturing apparatus (not shown) of the fourth embodiment may include at least two of the above-described ultrasonic apparatus 30, the bubble generator 60, and the magnet rotating apparatus 70 in addition to the vortex generator 20 . In the method of manufacturing a scaffold according to the fourth embodiment, at most four kinds of physical forces can be simultaneously applied to the medium 12 to generate a non-uniform composite flow in which a maximum of four kinds of flows having different driving sources are mixed.

FIG. 11 is a structural view for explaining a scaffold manufacturing method according to a fifth embodiment of the present invention, and FIG. 12 is a schematic structural view of a scaffold manufactured by the method of the fifth embodiment.

11 and 12, the scaffold manufacturing method of the fifth embodiment is similar to that of the above-described embodiment except that the medium 12a contained in the chamber 10 is replaced with another kind of medium 12b, Which is the same as any one of the methods of the first to fourth embodiments. 11 shows the first embodiment in which the vortex generator 20 and the ultrasonic device 30 are provided in the chamber 10 in a basic configuration.

The first medium 12a containing the first cells 11a is placed in the chamber 10 and at least two kinds of physical forces are applied to the first medium 12a so that at least two kinds of flows, Resulting in non-uniform composite flow. A voltage difference is generated between the first medium 12a and the syringe 40 and the syringe pump 41 is operated to emit the scaffold material toward the first medium 12a.

Then, a first scaffold made up of the fibrous structures 13 intricately entangled three-dimensionally in the first medium 12a and the first cells 11a penetrating the inside of the fibrous structure 13 is produced.

The first medium 12a is then removed from the chamber 10 and the second medium 12b containing the second cells 11b is placed in the chamber 10. At least two types of physical forces are applied to the second medium 12b to cause a non-uniform composite flow in which at least two kinds of flows different in driving source are mixed. A voltage difference is generated between the second medium 12b and the syringe 40 and the syringe pump 41 is operated to emit the scaffold material toward the second medium 12b.

Then, the fibers are intricately entangled outside the first scaffold to form the fibrous structure 13, and the second cells 11b penetrate into the fibrous structure 13 to constitute the second scaffold. The completed scaffold is divided into a central portion (first scaffold portion) where the first cells 11a are located and an outer frame portion (second scaffold portion) where the second cells 11b are located .

Biological tissue is composed of various cells. By using the method of the fifth embodiment, various kinds of cells can be infiltrated into one scaffold, and a desired kind of cells can be positioned at desired positions, so that a scaffold having excellent biocompatibility can be produced.

FIG. 13 is a structural view for explaining a scaffold manufacturing method according to a sixth embodiment of the present invention.

13, the method of manufacturing a scaffold of the sixth embodiment is the same as the method of any of the first to fifth embodiments described above except that the growth aid grains 15 are added to the medium 12 And the same process. In Fig. 13, the configuration of the first embodiment is shown as a basic configuration.

The growth aid particles 15 can be added to the medium 12 in a frozen state and are moved by the non-uniform composite flow of the medium 12 to help the three-dimensional fibrous structure 13 to be generated more quickly. The growth aid particles 15 may be composed of, for example, an Epidermal Growth Factor (EGF), in which case the growth aid particles 15 may dissolve and promote the growth of the cells 11 have.

14 and 15 are block diagrams of a scaffold manufacturing apparatus according to a seventh embodiment of the present invention.

14 and 15, the scaffold manufacturing apparatus 700 of the seventh embodiment includes a plurality of syringes 40 containing different scaffold materials (polymer solution) And the apparatus of the fourth embodiment. In Figs. 14 and 15, the configuration of the first embodiment is shown as a basic configuration.

A plurality of syringes 40 are connected to the syringe pump 41 and are electrically connected to the high voltage power source 51. Although four types of syringes 40 are shown in Figs. 14 and 15, the number of syringes 40 is not limited to the illustrated example.

In the scaffold manufacturing method according to the seventh embodiment, a voltage difference is generated between the medium 12 and the plurality of syringes 40, and the syringe pump 41 is operated to move the plurality of scaffolds The substance is emitted simultaneously. Two or more scaffold materials are entangled in three dimensions to form a fibrous structure 13, and the cells 11 penetrate into the fibrous structure 13 to form a scaffold.

The fibrous structure 13 of the scaffold fabricated by the method of the seventh embodiment may include various polymer components, and excellent characteristics can be realized by complementing the pros and cons of each material.

16A to 16D are block diagrams showing a method for manufacturing a scaffold according to an eighth embodiment using the apparatus for manufacturing a scaffold of the seventh embodiment.

Referring to Figs. 16A to 16D, in the method of manufacturing a scaffold according to the eighth embodiment, a plurality of scaffold materials are sequentially emitted toward the medium 12. The plurality of syringes 40 may include a first syringe 401, a second syringe 402, a third syringe 403, and a fourth syringe 404.

The high voltage power source 51 applies a high voltage in order of the first syringe 401, the second syringe 402, the third syringe 403 and the fourth syringe 404 in a predetermined time difference can do. The syringe pump 41 is connected to the first syringe 401, the second syringe 402, the third syringe 403, and the fourth syringe 402 in synchronization with the high voltage power supply 51, And the discharge pressure can be provided in the order of the paper 404.

Then, the four polymer materials are sequentially entangled to form a four-layered fiber structure 13, and cells penetrate into the fiber structure to form a scaffold.

17 is a schematic view showing a scaffold manufactured by the method of the eighth embodiment. Referring to FIG. 17, the scaffold manufactured by the method of the eighth embodiment has a multi-layer structure having the same number of stacked layers as the number of sliver.

Specifically, the scaffold includes a first layer 13a positioned at the center, a second layer 13b surrounding the first layer 13a, a third layer 13c surrounding the second layer 13b, And a fourth layer 13d surrounding the third layer 13c. Each of the first layer 13a, the second layer 13b, the third layer 13c and the fourth layer 13d includes a first syringe 401, a second syringe 402, The second syringe 403, and the fourth syringe 404, respectively.

The first to fourth layers 13a, 13b, 13c, and 13d may be made of a polymer material having a different degradation, and a multi-layer scaffold having a different resolution may be manufactured.

While the present invention has been described in connection with what is presently considered to be practical exemplary embodiments, it is to be understood that the invention is not limited to the disclosed embodiments, but, on the contrary, Of course.

100, 200, 300, 700: Scaffold manufacturing apparatus
10: chamber 11: cell
12: Medium 13: Fiber structure
20: vortex generator 30: ultrasonic device
40: Syringe 41: Syringe pump
51: high voltage power source 52: ground electrode
60: Bubble generator 70: Magnetic rotating device

Claims (14)

Preparing a medium containing the cells in a chamber and placing a syringe containing the scaffold material on top of the medium;
A flow generating step of applying at least two types of physical forces to the medium to cause a non-uniform compound flow in which at least two kinds of flows different from each other are mixed; And
An electrospinning step of generating a voltage difference between the medium and the syringe and activating the syringe pump to spin the scaffold material into a fiber form toward the surface of the medium from which the compound flow occurs,
/ RTI >
Wherein in the electrospinning step, the scaffold material is entangled in three dimensions to form a fibrous structure, and the cells penetrate into the fibrous structure to form a scaffold. The method according to claim 1,
In the preparation step, the vortex generator and the ultrasonic device are installed in the chamber,
Wherein in the flow generating step, flow is generated in the medium by the vortex generator and ultrasonic waves are applied to the medium. 3. The method of claim 2,
Wherein the ultrasound applied to the culture medium has a frequency of 1 MHz to 3 MHz and an intensity of 0.1 W / cm 2 to 2.0 W / cm 2. The method according to claim 1,
In the preparing step, a vortex generator and a bubble generator are installed in the chamber,
Wherein said flow generating step simultaneously causes flow in said medium by said vortex generator and flow by bubble diffusion to increase oxygen saturation of said medium. The method according to claim 1,
In the preparing step, a vortex generator and a magnet rotating device are installed in the chamber,
Wherein the magnet rotating device includes a bar magnet contained in the medium and a magnetic field applying part located outside the chamber,
Wherein in the flow generating step, a flow by the vortex generator and a vortex flow by the rotation of the bar magnet occur simultaneously in the culture medium. 6. The method according to any one of claims 1 to 5,
After the electrospinning step, the medium is replaced with another kind of medium containing different kinds of cells, the flow generation step and the electrospinning step are repeated,
In the second electrospinning step, the fiber structure is divided into a central portion where first cells are located and an outer portion where second cells are located. 6. The method according to any one of claims 1 to 5,
Wherein in said preparation step growth aid particles are added to said medium. 6. The method according to any one of claims 1 to 5,
Wherein in the preparing step, the syringe is comprised of a plurality of syringes containing different scaffold materials. 9. The method of claim 8,
In the electrospinning step, the plurality of syringes are simultaneously applied with a high voltage, simultaneously receive a discharge pressure from the syringe pump and simultaneously radiate the scaffold material toward the medium, and two or more types of scaffold materials are entangled, A method of manufacturing a three dimensional electrospinning scaffold for forming a structure. 9. The method of claim 8,
In the electrospinning step, the plurality of syringes are sequentially applied with a high voltage, and sequentially receive the discharge pressure from the syringe pump, sequentially radiating the scaffold material toward the medium, And forming the fibrous structure while forming layers of each of the three-dimensional electrospinning scaffolds. A chamber for containing the medium containing the cells;
At least two drive sources applying at least two types of physical forces to the medium to cause a non-uniform composite flow of at least two streams mixed;
A syringe positioned above the chamber and containing a scaffold material and supplied with a discharge pressure from a syringe pump;
A high voltage power source electrically connected to the syringe; And
And a ground electrode for grounding the medium,
Wherein the at least two driving sources include an eddy current generator, and further comprising at least one of an ultrasonic device, a bubble generator, and a magnet rotating device. 12. The method of claim 11,
The ultrasonic apparatus includes an ultrasonic driving unit and an ultrasonic transducer connected to the ultrasonic driving unit and contacting the medium. The ultrasonic transducer has a frequency of 1 MHz to 3 MHz and a strength of 0.1 W / cm2 to 2.0 W / Dimensional electrostatic radiation scaffold. 12. The method of claim 11,
Wherein the magnet rotating device includes a bar magnet contained in the medium and a magnetic field applying part located on the outer side of the chamber and rotating the bar magnet to cause a swirl flow in the medium. 14. The method according to any one of claims 11 to 13,
Wherein the syringe is comprised of a plurality of syringes containing different scaffold materials,
Wherein the plurality of syringes simultaneously spin the scaffold material toward the medium or sequentially spin the scaffold material for a predetermined period of time.

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