WO2023101034A1

WO2023101034A1 - 3d printed structure for implantation in human body and method for preparing same

Info

Publication number: WO2023101034A1
Application number: PCT/KR2021/017815
Authority: WO
Inventors: 김준규; 이기원; 김형구; 이환철
Original assignee: 주식회사 엘앤씨바이오
Priority date: 2021-11-30
Filing date: 2021-11-30
Publication date: 2023-06-08
Also published as: KR20230080621A

Abstract

The present invention relates to a 3D printed structure for implantation in a human body and a method for preparing same. According to the present invention, a structure is prepared using both biodegradable polymer ink and biopolymer ink, and thus the structure can be prepared into a 3D printed structure having various physical properties by adjusting the thickness of a nozzle and a spacing between lines formed of the ink. In addition, a structure for implantation is prepared by adjusting the size of a spacing between lines formed of the biodegradable polymer ink and by filling a space formed by the spacing with the biopolymer ink without any empty space, whereby the structure can be manufactured into a structure having various physical properties without a change in the size of the entire structure. The structure prepared in the method described above can have physical properties, such as mechanical strength, elasticity, biological properties, and the like, suitable for the physical environment of a body tissue into which the structure is to be implanted.

Description

3D printed structure for human body implantation and its manufacturing method

The present invention relates to a 3D printed structure for implantation in the human body and a method for manufacturing the same, and to a 3D printed structure for implantation in the human body and a method for manufacturing the same, which have mechanical strength and elasticity suitable for the transplanted tissue and biological characteristics suitable for the surrounding environment of the tissue. it's about

The 3D printing structure and method for manufacturing the same of the present invention is a technology for manufacturing a 3D printing structure having various physical properties by adjusting the spacing and nozzle thickness of a plurality of inks. The printing method is used, and biopolymer ink, which is a mixture of human and animal tissue-derived extracellular matrix and biocompatible polymer, is sprayed after selecting a nozzle suitable for the size of the gap created with the biodegradable polymer ink, and then a transplant structure suitable for the tissue or organ. will manufacture

According to the 3D printing structure and its manufacturing method of the present invention, through the above process, 1) physical properties such as mechanical strength and elasticity of the 3D printing structure can be changed, and 2) a line in which the biopolymer ink is made of biodegradable polymer ink After setting it to pass over the gap, replacing the nozzle with a nozzle suitable for the size of the gap so that the biopolymer ink can fill the gap and spraying it, 3) Filling the structure with biodegradable polymer ink and biopolymer ink, The content of the biopolymer ink can be increased without changing the size.

3D printing technology is drawing attention as a technology that can bring innovation to the future manufacturing industry as a technology that builds up materials layer by layer as if printing according to a 3D blueprint to create a product in a desired 3D shape. In particular, it has been recognized for its potential for use in terms of its ability to produce personalized manufacturing, and in the fields of tissue engineering and regenerative medicine, it is used for implant production, scaffold production, and cell printing for artificial organ development.

In 3D printing, which is used in tissue engineering and regenerative medicine, structures are manufactured using various materials such as ceramics, metals, and polyethylene (PE). Depending on the characteristics of the material, the manufactured structure has different physical properties such as mechanical strength, and as a result, the fields in which it can be used are different. In particular, in the field of tissue engineering, polylactic acid (PLA) and polyglycolic acid (PGA), which are synthetic polymers with biodegradability, are used as polymers used for human tissue and organ production through the development of 3D bioprinting technology. and its copolymer (PLGA), polycarprolactone (PCL), etc. are mainly used.

Among the synthetic polymers used, PCL is a material with elasticity and rigidity, and is used as a material for medical devices to be implanted into the human body. However, constructs made only from PCL perform shape-maintaining functions such as physical properties and support, but have disadvantages in that the function of cell attachment, proliferation, and differentiation in tissues after transplantation is poor. In order to compensate for these disadvantages, cases in which biopolymers such as human and animal tissue-derived extracellular matrices and hydrogels are used as inks and used together with PCL are increasing. Attempts to manufacture a 3D printing structure that improves biofunctionality while having physical properties, support, and shape retention suitable for a living environment by using the two materials together as ink have been actively attempted.

Tissues and organs in the human body have various physical properties, and the physical environment in the body is different depending on the tissue and organ, so a method of transplanting a structure suitable for the environment is required. In order to manufacture with 3D printing, it is also necessary to use a plurality of inks and change the spraying method of the two inks at the same time.

Registered Patent No. 10-1067827 suggests a method of making a lattice shape by alternately stacking two layers using a biodegradable synthetic polymer and a hydrogel. However, since this method is a printing method that leaves voids without filling the gaps while maintaining the line gap size of the biodegradable synthetic polymer ink that functions to maintain the shape, there are limitations in changing the physical properties of the structure. In addition, there is a disadvantage in that the amount of hydrogel ink for application to various tissues cannot be changed by spraying the hydrogel using a single fixed nozzle.

Therefore, in the present invention, a 3D printing structure for human body implantation is manufactured using both biodegradable polymer ink and biopolymer ink, but 3D printing structure having various physical properties by adjusting the spacing and nozzle thickness of lines made of the two inks, and We would like to present the manufacturing method.

Through the 3D printing jetting method using multiple inks, it is possible to manufacture a structure with various physical properties by adjusting the size of the line gap of the biodegradable polymer ink, and select and replace a nozzle suitable for the size of the line gap formed with the biopolymer ink to spray Accordingly, a structure capable of increasing the content of biopolymer ink without changing the size of the structure and a manufacturing method thereof are proposed.

[Prior art literature]

[Patent Literature]

Korean Patent Registration No. 10-1067827

[Non-Patent Literature]

Griffin M et al. J Vis Exp. 118, 54872 (2016)

An object of the present invention is to manufacture a structure by using a plurality of inks, that is, biodegradable polymer ink and biopolymer ink together, and to provide a 3D printing structure having various physical properties by adjusting the thickness of the nozzle and the spacing between lines made of the inks, and It is to provide a manufacturing method.

Another object of the present invention is to provide a printing method that can be filled with biopolymer ink by selecting and replacing nozzles with a thickness suitable for the line gap size generated by the biodegradable polymer ink during the 3D printing process and spraying the biopolymer ink. It is an object of the present invention to provide a structure capable of increasing the content of biopolymer ink without changing the size of the structure and a method for manufacturing the same.

The 3D printed structure for transplantation into the human body and the method of manufacturing the same according to the present invention can have biological properties suitable for the surrounding environment, such as mechanical strength and elasticity suitable for the tissue and organ to be transplanted, and tissue. According to the present invention, the size of the line spacing of biodegradable polymer ink is adjusted, and the characteristics suitable for transplanted tissues and organs are obtained by changing the nozzle thickness of biopolymer ink in which human and animal tissue-derived extracellular matrices and biocompatible polymers are mixed. will be granted

The 3D printing structure for human body implantation of the present invention as described above includes two or more biodegradable polymer lines comprising a biodegradable polymer; and a biopolymer line formed of biopolymer ink in a space between the biodegradable polymer lines.

As the biodegradable polymer, polylactic acid (PLA), polyglycolic acid (PGA) and copolymers thereof (PLGA), and polycaprolactone (PCL) may be preferably used.

The biopolymer may include a human or animal-derived extracellular matrix and a hydrogel.

The 3D printing structure for human body implantation includes, for example, two or more unit layers formed of the biodegradable polymer line formed in one direction and the biopolymer line formed between the biodegradable polymer lines, and the unit layers adjacent to each other The biodegradable polymer line and the biopolymer line may be formed while crossing each other. Through this, it is possible to maintain the overall mechanical balance while forming a structure for larger tissue production.

The 3D printed structure for transplantation into the human body may be used as an implant, a support, or an artificial organ.

The width of the biodegradable polymer line is preferably 100 to 1000 μm, and the width of the biopolymer line is preferably 100 to 4000 μm. Here, the ratio of the width of the biopolymer line to the width of the biodegradable polymer line may be 1:1 to 1:4.

Widths of the biodegradable polymer line and the biopolymer line may be adjusted according to required properties.

In addition, the manufacturing method of the 3D printing structure for human body implantation of the present invention, forming two or more biodegradable polymer lines with a distance from each other with a biodegradable polymer (S1); and forming a biopolymer line with biopolymer ink in a space formed between the biodegradable polymer lines (S2).

The biopolymer may include a human or animal tissue-derived extracellular matrix and a hydrogel.

In the manufacturing method, the step S1 and the step S2 are performed one or more times on the unit layer formed through the steps S1 and S2 so that two or more unit layers are stacked, but the unit layers adjacent to each other are the biodegradable polymer line. And the biopolymer lines may be formed while crossing each other.

The 3D printed structure manufactured through the manufacturing method may be used as an implant, a support, or an artificial organ.

In the step S2, the biopolymer line may be formed by spraying the biopolymer ink while adjusting the thickness of the nozzle according to the interval between the biodegradable polymer lines.

In the present invention, a structure having various physical properties can be manufactured without changing the size of the entire structure by adjusting the size of the line spacing of the biodegradable polymer ink and filling the space formed by the spacing with the biopolymer ink to prepare a structure for transplantation. there is. The structure prepared in this way may have physical properties such as mechanical strength, elasticity, and biological properties suitable for the physical environment of the body tissue to be implanted.

Unlike conventional 3D printing methods that utilize multiple inks and leave line gaps, cell adhesion by tissue-derived extracellular matrix to be implanted by increasing the content of biocompatible ink in the structure by using a method of filling all gaps with biopolymer ink However, the field of application can be expanded by promoting proliferation and differentiation. Unlike attempts to induce cell inflow by intentionally leaving gaps in existing methods, the present invention utilizes a method of changing the size of line gaps and filling them with biopolymer ink, so that cells in the tissue after transplantation do not change the overall size of the structure. It is possible to increase the content of the biopolymer ink providing an environment capable of attaching, proliferating and differentiating.

1 is a side cross-sectional view of a unit layer of a 3D printed structure for human body implantation according to an embodiment of the present invention.

2 is a plan view of a unit layer of a 3D printed structure for human body implantation according to an embodiment of the present invention.

3 is a photograph of a structure manufactured with single ink (biodegradable polymer ink) and multiple inks (biodegradable polymer ink + biopolymer ink) according to a 3D printing method according to an embodiment of the present invention (from the left, biodegradable polymer ink). This is an example of a structure prepared by changing the ratio of line width to line spacing of ink to 1:1, 1:2, 1:3, and 1:4).

4 is a diagram showing a comparison of infill and line spacing size ratios in 3D printing and showing size changes in structure spacing units according to line spacing size ratios.

5 and 6 are pictures showing a method of stacking biodegradable polymer ink and biocompatible polymer ink during the manufacturing process of a 3D printed structure for human body implantation according to an embodiment of the present invention.

7 is a diagram showing the nozzle thickness of the biopolymer ink selected according to the size of the line spacing generated by the biodegradable polymer ink.

8 is a view for explaining a manufacturing process of the 3D printed structure of Example 1, and is a diagram illustrating a process of manufacturing first and second unit layers.

9 is a graph showing a change in ink content used for manufacturing each line according to a ratio of a biodegradable polymer line and a biopolymer line of a 3D printed structure for human body implantation according to an embodiment of the present invention.

10 and 11 are graphs showing the tensile strength and Young's modulus of the structure according to the change in the line spacing size ratio of the structure manufactured according to Example 1, respectively.

The present invention will be described in more detail with reference to the following embodiments with reference to the drawings. However, the following examples are only intended to specifically illustrate the present invention, and the scope of the present invention is not limited by the following examples.

According to the 3D printing structure and its manufacturing method of the present invention, a structure is manufactured using a plurality of inks, that is, biodegradable polymer ink and biopolymer ink together, but the structure is to be implanted by adjusting the spacing and nozzle thickness of the plurality of inks. It can have physical properties such as mechanical strength, elasticity, and bio-characteristics suitable for the physical environment of internal tissues.

In addition, in the present invention, a structure having various physical properties can be manufactured without changing the size of the entire structure by manufacturing a structure for implantation by filling the space formed by the line spacing of the biodegradable polymer ink without empty space.

1 and 2 are a cross-sectional side view and a plan view of a unit layer of a 3D printed structure for human body implantation according to an embodiment of the present invention.

As shown in Figures 1 and 2, the 3D printing structure for human body implantation according to an embodiment of the present invention is a biodegradable polymer line (gray) formed of a biodegradable polymer and a biodegradable polymer line in the space between the lines. It includes biopolymer lines (yellow) formed by filling polymer ink.

Here, the biodegradable polymer forming the biodegradable polymer line is, for example, polylactic acid (PLA), polyglycolic acid (PGA) and its copolymer (PLGA), polycaprolactone (Polycarprolactone, PCL) etc. can be used. However, it is not limited thereto.

In addition, the biopolymer ink may be prepared using a human or animal tissue-derived extracellular matrix and a hydrogel, and the extracellular matrix may be prepared using various human or animal tissues according to the required characteristics of the structure to be manufactured. there is.

The biodegradable polymer line may be formed in one direction as shown in FIGS. 1 and 2, and the biopolymer line may be formed in one direction in the space formed by the gap between the biodegradable polymer lines, but is not necessarily limited thereto. It can be formed in various ways according to the required characteristics of.

Preferably, the width of the biodegradable polymer line is 100 to 1000 μm, the biopolymer line may be formed to be 100 to 4000 μm, and the ratio of the biodegradable polymer line to the biopolymer line is in the range of 1:1 to 1:4. It may be, but is not limited thereto.

Although the present embodiment shows an example in which the biodegradable polymer line and the biopolymer line are formed in one layer, that is, a unit layer, it is not limited thereto and may be manufactured in multiple layers according to the tissue to be manufactured.

In the case of manufacturing a structure with multilayers in which unit layers are stacked, the unit layers adjacent to each other may be formed while the biodegradable polymer line and the biopolymer line cross each other, but are not limited thereto. In addition, the meaning of mutually intersecting does not necessarily mean that they are formed in a vertical direction.

3 is a photograph of a structure manufactured with single ink (biodegradable polymer ink) and multiple inks (biodegradable polymer ink + biopolymer ink) according to a 3D printing method according to an embodiment of the present invention (from the left, biodegradable polymer ink). This is an example of a structure manufactured by changing the ratio of line width to line spacing of ink to 1:1, 1:2, 1:3, and 1:4), Figure 4 shows the ratio of filling and line spacing in 3D printing It is a figure showing the size change of the structure spacing unit according to the line spacing size ratio.

As shown in FIGS. 3 and 4, the biopolymer ink is filled in the space between the biodegradable polymer lines formed with the biodegradable polymer ink, and as the interval between the biodegradable polymer lines increases, the empty space increases, and the biopolymer ink The volume of the biopolymer line to be formed increases. As such, the characteristics of the structure can be controlled by adjusting the volume of the biodegradable polymer line and the biopolymer line.

5 shows a process of forming a unit layer of a structure by forming biopolymer lines (yellow) with biopolymer ink in the space between the biodegradable polymer lines (gray).

6 is a biodegradable polymer line (gray) formed by preparing a first layer with a biodegradable polymer and then a second layer with a biodegradable polymer thereon, and biopolymer ink in the space formed between the biodegradable polymer lines. It is a figure showing an example of forming a biopolymer line (yellow) with . The reason why the biodegradable polymer line is formed by stacking the biodegradable polymer in two layers is because, due to the nature of the biodegradable polymer, if the biodegradable polymer line cannot be formed at the required height, the biodegradable polymer line must be formed in multiple layers of two or more layers. Because.

As shown in FIG. 7 , the thickness of the nozzle spraying the biopolymer ink can be adjusted according to the distance between the biodegradable polymer lines, that is, the width of the biopolymer line, so that the biopolymer ink can be completely filled without empty spaces.

Hereinafter, a process of preparing a structure by forming a biodegradable polymer line and then forming a biopolymer line according to an embodiment of the present invention, and evaluating the mechanical properties of the prepared structure will be described.

실시예 1. 구조체 제조방법Example 1. Structure manufacturing method

1.1 3D 프린팅 기술을 이용한 생분해성 고분자 잉크 분사1.1 Spraying biodegradable polymer ink using 3D printing technology

PCL (Evonik, Essen, Germany) was placed in a 3D printed custom syringe with thermal conductivity and mounted on Head 1, and the temperature was set to 90° C. to 120° C. through the system. The dissolved PCL was sprayed through a precision nozzle with a pressure of 100 to 700 kPa and an inner diameter of 100 to 500 μm. The spacing of the lines generated by the spraying PCL was adjusted to 1000, 1500, 2000, and 2500 μm through 3D printer settings and g code. The process of determining the size of the line spacing generated according to the adjusted line spacing and the method of calculating the filling according to the distance ratio are shown in FIG. 4 . After completing the spraying of the first layer to have a height of 100 to 300 μm according to the ratio of the set distance, the second layer was completed so that the height of the line was 200 to 600 μm by spraying in the same direction and manner.

1.2 생체고분자 잉크의 제조1.2 Preparation of biopolymer ink

(1) Manufacture of micronized human tissue

After trimming the human-derived tissue, fat was removed using isopropyl alcohol (IPA) and hexane. After that, the cells were removed using sodium hydroxide (NaOH) and washed with alcohol and distilled water. Thereafter, water was removed by freeze-drying. Freeze-dried decellularized human-derived tissue was pulverized using a freeze grinder. After the grinding, tissue powder having a diameter of 500 μm or less was obtained by passing through a sieve having a diameter of 100 to 500 μm.

(2) Preparation of cross-linked hyaluronic acid (HA)-carboxymethylcellulose (CMC) carrier

Hyaluronic acid (HA) and carboxymethyl cellulose (CMC) were mixed at a predetermined ratio, 1,4-butanediial diglycidyl ether (BDDE) as a cross-linking agent was added, NaOH was added, mixed, and cross-linking was performed. After cross-linking, HA-CMC was washed using a dialysis membrane. The washed HA-CMC was freeze-dried to remove moisture. The freeze-dried HA-CMC was pulverized, mixed with sterile physiological saline in proportion, and then prepared as HA-CMC gel.

(3) Manufacture of biopolymer ink

Biopolymer ink was prepared by mixing micronized human tissue and HA-CMC gel in proportion. Thereafter, the biopolymer ink was put into a syringe and centrifuged to ensure that it was constantly discharged during 3D printing.

1.3 생체고분자 잉크의 분사1.3 Jetting of biopolymer ink

After putting the biopolymer ink prepared by the method described in 1.2 above into a syringe and selecting nozzles of 22, 18, 16, and 14G sizes according to the line spacing size ratio set according to the line spacing formed by the biodegradable polymer ink, attached to the syringe. The line spacing size ratio and nozzle selection of the biopolymer ink according to the ratio are shown in Table 1 below. 3D printing settings and g code were written so that the biopolymer ink can be sprayed while passing through the gap created by the biodegradable polymer ink. After mounting the syringe on Head 2, biopolymer ink was sprayed according to the setting while applying a pressure of 100 to 200 kPa. A first unit layer was formed by filling a two-layer height gap made of biodegradable polymer ink with biopolymer ink, and a second unit layer was prepared by crossing the first unit layer thereon in the same manner. 8 is a diagram schematically illustrating a process of manufacturing up to a second unit layer in this embodiment. Thereafter, the structure of this embodiment was manufactured by stacking up to the eighth unit layer.

실시예 2. 구조체 형태 및 물성 차이 확인Example 2. Confirmation of structure shape and difference in physical properties

2.1 구조체 형태 확인2.1 Check the shape of the structure

The 3D printing method in Example 1 uses a single biodegradable polymer ink and a biodegradable polymer ink + multiple biopolymer inks, and has a size of width X length X height (20 X 20 X 3 mm) and a line spacing size ratio (1 :1, 1:2, 1:3, 1:4) were prepared for different structures. The shape of the fabricated structure is shown in FIG. 3 . In addition, the contents of the biodegradable polymer ink and biopolymer ink included in the fabricated structure were calculated and shown in a graph in FIG. 9, and the contents of the biodegradable polymer ink and biopolymer ink in FIG. 9 are shown in Table 2 below.

[Correction under Rule 91 20.12.2021]

As shown in FIG. 9 and Table 2, the content of each ink used varies according to the ratio between the width of the biodegradable polymer line and the width of the biopolymer line, and accordingly, the physical properties of the structure may be changed. In addition, it can be seen that the content of the biopolymer can be increased while increasing the distance between the biodegradable polymer lines without changing the size of the 3D printed structure.

2.2 구조체 물성 차이 확인2.2 Confirmation of differences in structural properties

In order to confirm the mechanical strength of the structure prepared in Example 1, a tensile strength test was performed using a measuring device (EZ Test, Shimadzu Corporation, Kyoto, Japan) that performs tension at a rate of 8 mm/min. The specimen used in the test (manufactured according to ASTM D 638 No. 4 standard, manufactured in dog bone form) was manufactured according to the line spacing size ratio by 3D printing. Table 3 shows the results of decreasing tensile strength and Young's modulus (modulus of elasticity) as the line spacing size ratio increases.

10 and 11 are graphs showing the tensile strength and Young's modulus of the structure according to the change in the line spacing size ratio of the structure manufactured according to Example 1, respectively. 10 and 11, it can be confirmed that as the line spacing size ratio increases, the rigidity of the structure decreases, but the ductility at which deformation occurs until failure increases.

Claims

Two or more biodegradable polymer lines comprising a biodegradable polymer; and

A 3D printing structure for human body implantation comprising a biopolymer line formed of biopolymer ink in a space between the biodegradable polymer lines.
According to claim 1,

The biodegradable polymer is a human body, characterized in that at least one selected from polylactic acid (PLA), polyglycolic acid (PGA) and its copolymer (PLGA), polycarprolactone (PCL) 3D printed structures for implantation.
According to claim 1,

The biopolymer is a 3D printing structure for human transplantation, characterized in that consisting of a human or animal tissue-derived extracellular matrix and a hydrogel.
According to claim 1,

At least two unit layers formed of the biodegradable polymer line formed in one direction and the biopolymer line formed in one direction,

The adjacent unit layers are 3D printed structures for human implantation, characterized in that formed while the biodegradable polymer line and the biopolymer line cross each other.
According to claim 1,

A 3D printed structure for implantation in the human body, characterized in that it is used as an implant, support or artificial organ.
According to claim 1,

3D printing structure for human body implantation, characterized in that the width of the biodegradable polymer line is 100 to 1000 μm.
According to claim 1,

3D printing structure for human body implantation, characterized in that the width of the biopolymer line is 100 to 4000 μm.
According to claim 1,

3D printing structure for human body implantation, characterized in that the width of the biodegradable polymer line and the biopolymer line can be adjusted according to the required characteristics.
Forming two or more biodegradable polymer lines with a biodegradable polymer line apart from each other (S1); and

A method of manufacturing a 3D printing structure for implantation into a human body, comprising: forming a biopolymer line with biopolymer ink in a space formed between the biodegradable polymer lines (S2).
According to claim 9,

The biodegradable polymer is a human body, characterized in that at least one selected from polylactic acid (PLA), polyglycolic acid (PGA) and its copolymer (PLGA), polycarprolactone (PCL) A method of manufacturing a 3D printed structure for implantation.
According to claim 9,

The biopolymer is a method of manufacturing a 3D printing structure for human implantation, characterized in that comprising a human or animal-derived extracellular matrix and a hydrogel.
According to claim 9,

Steps S1 and S2 are performed one or more times on the unit layer formed through steps S1 and S2 so that two or more unit layers are laminated,

The adjacent unit layers are a method of manufacturing a 3D printing structure for human body implantation, characterized in that the biodegradable polymer line and the biopolymer line are formed while crossing each other.
According to claim 9,

A method of manufacturing a 3D printed structure for human implantation, characterized in that it is used as an implant, support or artificial organ.
The method of manufacturing a 3D printing structure for human body implantation, characterized in that the width of the biodegradable polymer line is 100 to 1000μm.
According to claim 1,

The method of manufacturing a 3D printing structure for human body implantation, characterized in that the width of the biopolymer line is 100 to 4000μm.
According to claim 1,

In the step S2, the biopolymer ink is sprayed while adjusting the thickness of the nozzle according to the interval between the biodegradable polymer lines to form the biopolymer line.