Manufacturing Method and Device for Three-Dimensional Engineered Tissue

Abstract

A method for fabricating a cellular structure is disclosed. The method includes feeding cell aggregates onto a plurality of thread or needle shaped members in each of a plurality of supports. Each member forms net or mesh shaped spaces in the respective support. The method also includes layering the supports with the cell aggregates fed therein, culturing the cell aggregates to fuse, and removing the plurality of thread or needle shaped members from the fused cellular structure.

1 . A support for fabricating a three-dimensional cellular structure, the support comprising:
 a first frame placed in an x-direction, a first frame placed in a y-direction, a second frame placed in the x-direction, and a second frame placed in the y-direction,   a plurality of string members placed in the x-direction, wherein each string member within the plurality of string members placed in the x-direction is connected to the first frame and/or the second frame placed in the y-direction, and wherein a gap distance (Px) between each string member within the plurality of string members placed in the x-direction is between about 0.01 mm and about 3 mm;   a plurality of string members placed in the y-direction, wherein each string member within the plurality of string members placed in the y-direction is connected to the first frame and/or the second frame placed in the x-direction, and wherein a gap distance (Py) between each string member within the plurality of string members placed in the y-direction is between about 0.01 mm and about 3 mm;   wherein the plurality of string members placed in the x-direction and the plurality of string members placed in the y-direction are perpendicular to each other in a same plane and form a plurality of rectangular net shaped or matrix shaped spaces in the same plane inside each support of the one or more support;   wherein the plurality of net shaped or matrix shaped spaces defines a plurality of physical boundaries capable of holding cell aggregates; and   wherein the plurality of string members placed in the x-direction and the plurality of string members placed in the y-direction are separable from the frames.   
     
     
         2 . The support of  claim 1 , wherein the plurality of string members placed in the x-direction and the plurality of string members placed in the y-direction are made of a material that is non-adhesive with respect to the cell aggregates. 
     
     
         3 . The support of  claim 1 , wherein the plurality of string members placed in the x-direction and the plurality of string members placed in the y-direction are coated with a material that is non-adhesive with respect to the cell aggregates. 
     
     
         4 . The support of  claim 1 , further comprising an alignment mechanism for aligning each support when a plurality of supports is layered. 
     
     
         5 . The support of  claim 1 , wherein the gap distance Px is uniform. 
     
     
         6 . The support of  claim 1 , wherein the gap distance Py is uniform. 
     
     
         7 . The support of  claim 1 , wherein the gap distance Px is equal to the gap distance Py. 
     
     
         8 . The support of  claim 1 , wherein the gap distance Px is greater than the gap distance Py. 
     
     
         9 . The support of  claim 1 , wherein the gap distance Py is greater than the gap distance Px. 
     
     
         10 . The support of  claim 1 , wherein a ratio between the gap distance Px and the gap distance Py is between 0.1 and 1.0. 
     
     
         11 . The support of  claim 1 , wherein a ratio between the gap distance Py and the gap distance Px is between 0.1 and 1.0. 
     
     
         12 . The support of  claim 1 , wherein the plurality of net shaped or matrix shaped spaces includes a plurality of net shaped or matrix shaped spaces smaller than the size of the cell aggregates (spheroids) and a plurality of net shaped or matrix shaped spaces larger than the size of the cell aggregates (spheroids). 
     
     
         13 . An apparatus for fabricating the three-dimensional cellular structure, comprising a plurality of supports of  claim 1 , wherein the plurality of supports comprise one or more upper support and one or more lower support. 
     
     
         14 . The apparatus of  claim 13 , wherein the apparatus is configured so that the cell aggregates (spheroids) held in the plurality of net shaped or matrix shaped spaces in an upper support of the plurality of supports are in contact with the cell aggregates (spheroids) held in the plurality of net shaped or matrix shaped spaces in a lower support of the plurality of supports. 
     
     
         15 . The apparatus of  claim 13 , further comprising a guide member for defining the cell aggregates (spheroids) held in the plurality of net shaped or matrix shaped spaces. 
     
     
         16 . The apparatus of  claim 13 , further comprising an alignment mechanism for aligning each support when the plurality of supports is layered. 
     
     
         17 . The apparatus of  claim 13 , wherein a height of the plurality of layered supports corresponds to a height of the three-dimensional cellular structure. 
     
     
         18 . The apparatus of  claim 13 , wherein the net shaped or the matrix shaped spaces in the upper support of the plurality of supports are larger than the net shaped or the matrix shaped spaces in the lower support of the plurality of supports. 
     
     
         19 . The apparatus of  claim 13 , wherein the gap distance Px between each string member within the plurality of string members placed in the x-direction and the gap distance Py between each string member within the plurality of string members placed in the y-direction in the upper support of the plurality of supports are larger than the gap distance Px between each string member within the plurality of string members placed in the x-direction and the gap distance Py between each string member within the plurality of string members placed in the y-direction in the lower support of the plurality of supports. 
     
     
         20 . The apparatus of  claim 13 , wherein the gap distance Px between each string member within the plurality of string members placed in the x-direction and the gap distance Py between each string member within the plurality of string members placed in the y-direction in the upper support of the plurality of supports are larger than the size of the cell aggregates (spheroids) held in these spaces, and the gap distance Px between each string member within the plurality of string members placed in the x-direction and the gap distance Py between each string member within the plurality of string members placed in the y-direction in the lower support of the plurality of supports are smaller than the size of the cell aggregates (spheroids) held in these spaces. 
     
     
         21 . The apparatus of  claim 13 , wherein the gap distance Px between each string member within the plurality of string members placed in the x-direction and the gap distance Py between each string member within the plurality of string members placed in the y-direction in the upper support of the plurality of supports includes a gap distance Px and a gap distance Py suitable for holding the cell aggregates (spheroids) and a gap distance Px and a gap distance Py suitable for dropping the cell aggregates (spheroids). 
     
     
         22 . The apparatus of  claim 13 , wherein one support of the plurality of supports comprises the plurality of net shaped or the matrix shaped spaces formed by the plurality of string members placed in the x-direction and another support of the plurality of supports comprises the plurality of net shaped or matrix shaped spaces formed by the plurality of string members placed in the y-direction. 
     
     
         23 . The apparatus of  claim 13 , wherein one support of the plurality of supports comprises the plurality of net shaped or the matrix shaped spaces formed by the plurality of string members placed in the x-direction that provide a physical boundary for the three-dimensional cellular structure in the x-direction and another support of the plurality of supports comprises the plurality of net shaped or matrix shaped spaces formed by the plurality of string members placed in the y-direction that provide a physical boundary for the three-dimensional cellular structure in the y-direction. 
     
     
         24 . A method for making the support of  claim 1 , the method comprising:
 forming a mask layer on a thin plate member; and   forming the plurality of string members in the x-direction and the plurality of string members in the y-direction by etching the thin plate member exposed by the mask layer.   
     
     
         25 . A method for making the support of  claim 1 , the method comprising forming the plurality of string members in the x-direction and the plurality of string members in the y-direction by bonding wires. 
     
     
         26 . A method for making the support of  claim 1 , the method comprising forming the plurality of string members in the x-direction and the plurality of string members in the y-direction by using a 3D printer. 
     
     
         27 . A method for fabricating a three-dimensional cellular structure, the method comprising:
 (a) feeding the cell aggregates (spheroids) onto one or more support of  claim 1 , thus obtaining a plurality of supports;   (b) layering the plurality of supports obtained in step (a);   (c) culturing the cell aggregates (spheroids) to fuse; and   (d) removing the plurality of string members placed in the x-direction and the plurality of string members placed in the y-direction inside the plurality of frames of each support within the plurality of supports from the fused three-dimensional cellular structure;   
       thereby fabricating the three-dimensional cellular structure. 
     
     
         28 . The method of  claim 27 , wherein the feeding comprises loading the cell aggregates (spheroids) into a dispenser and scanning the dispenser to dispense the cell aggregates (spheroids) onto the plurality of net shaped or matrix shaped spaces. 
     
     
         29 . The method of  claim 27 , wherein the feeding comprises placing a guide member onto the one or more support of the plurality of supports, the guide member defining the shape of the three-dimensional cellular structure, wherein the cell aggregates (spheroids) are fed within the guide member. 
     
     
         30 . The method of  claim 27 , wherein the feeding comprises scooping up a liquid containing the cell aggregates (spheroids) retained in a container by using the one or more support of the plurality of supports. 
     
     
         31 . The method of  claim 27 , further comprising immersing the one or more support of the plurality of supports with the fed cell aggregates (spheroids) into container holding a liquid containing nutrients and applying vibrations or swings to the one or more support of the plurality of supports or the container. 
     
     
         32 . The method of  claim 27 , wherein the removing comprises drawing the plurality of string members placed in the x-direction from the cell aggregates (spheroids) in the x-direction and drawing the plurality of string members placed in the y-direction from the cell aggregates (spheroids) in the y-direction. 
     
     
         33 . The method of  claim 27 , wherein a first plurality of net shaped or matrix shaped spaces that are smaller than the size of the cell aggregates (spheroids) and a second plurality of net shaped or matrix shaped spaces that are larger than the size of the cell aggregates (spheroids) are formed inside the plurality of frames of each support of the plurality of supports, wherein the first plurality and the second plurality define the plurality of physical boundaries for the cell aggregates (spheroids) to be fused. 
     
     
         34 . A method for fabricating a three-dimensional cellular structure, the method comprising:
 (a) layering one or more support of  claim 1 , thus obtaining a plurality of layered supports;   (b) feeding the cell aggregates (spheroids) onto the plurality of layered supports obtained in step (a);   (c) culturing the cell aggregates (spheroids) to fuse; and   (d) removing the plurality of string members placed in the x-direction and the plurality of string members placed in the y-direction inside the plurality of frames of each support with the plurality of supports from the fused three-dimensional cellular structure;   thereby fabricating the three-dimensional cellular structure.   
     
     
         35 . The method of  claim 34 , wherein the feeding comprises loading the cell aggregates (spheroids) into a dispenser and scanning the dispenser to dispense the cell aggregates (spheroids) onto the plurality of net shaped or matrix shaped spaces. 
     
     
         36 . The method of  claim 34 , wherein the feeding comprises scooping up a liquid containing the cell aggregates (spheroids) retained in a container by using the plurality of layered supports. 
     
     
         37 . The method of  claim 34 , further comprising immersing the plurality of layered supports with the fed cell aggregates (spheroids) into container holding a liquid containing nutrients and applying vibrations or swings to the one or more support of the plurality of layered supports or the container. 
     
     
         38 . The method of  claim 34 , wherein removing comprises drawing the plurality of string members placed in the x-direction from the cell aggregates (spheroids) in the x-direction and drawing the plurality of string members placed in the y-direction from the cell aggregates (spheroids) in the y-direction. 
     
     
         39 . The support of  claim 1 , wherein the support is stackable.