Method of printing a tissue construct with embedded vasculature

Patent metadata

Field	Value
Publication number	US-2016287756-A1
Application number	US-201415146613-A
Country	US
Kind code	A1
Filing date	Nov 4, 2014
Priority date	Nov 5, 2013
Publication date	Oct 6, 2016
Grant date	—

Abstract

A printed tissue construct comprises one or more tissue patterns, where each tissue pattern comprises a plurality of viable cells of one or more predetermined cell types. A network of vascular channels interpenetrates the one or more tissue patterns. An extracellular matrix composition at least partially surrounds the one or more tissue patterns and the network of vascular channels. A method of printing a tissue construct with embedded vasculature comprises depositing one or more cell-laden filaments, each comprising a plurality of viable cells, on a substrate to form one or more tissue patterns. Each of the one or more tissue patterns comprises one or more predetermined cell types. One or more sacrificial filaments, each comprising a fugitive ink, are deposited on the substrate to form a vascular pattern interpenetrating the one or more tissue patterns. The vascular pattern and the one or more tissue patterns are at least partially surrounded with an extracellular matrix composition. The fugitive ink is then removed to create vascular channels in the extracellular matrix composition, thereby forming an interpenetrating vascular network in a tissue construct.

First claim

1 . A method of printing a tissue construct with embedded vasculature, the method comprising:
depositing one or more cell-laden filaments each comprising a plurality of viable cells on a substrate to form one or more tissue patterns, each of the tissue patterns comprising one or more predetermined cell types; depositing one or more sacrificial filaments on the substrate to form a vascular pattern interpenetrating the one or more tissue patterns, each of the sacrificial filaments comprising a fugitive ink; at least partially surrounding the one or more tissue patterns and the vascular pattern with an extracellular matrix composition, removing the fugitive ink to create vascular channels in the extracellular matrix composition, thereby forming an interpenetrating vascular network in a tissue construct.

2 . The method of claim 1 , wherein each of the one or more cell-laden filaments further comprise an extracellular matrix material comprising one or more of gelatin, fibrin, and gelatin methacrylate.

3 . (canceled)

4 . The method of claim 1 ,
wherein at least one of the one or more cell-laden filaments further comprises one or more functional chemical substances selected from the group consisting of: drugs, small molecules, toxins, proteins, and hormones.

5 .- 9 . (canceled)

10 . The method of claim 1 , wherein each of the one or more cell-laden filaments and each of the one or more sacrificial filaments are extruded from a nozzle at room temperature before being deposited on the substrate.

11 . (canceled)

12 . The method of claim 1 , wherein removing the fugitive ink comprises cooling the one or more sacrificial filaments.

13 . The method of claim 1 , further comprising at least one of the following steps:
depositing additional viable cells with the one or more sacrificial filaments, wherein at least a portion of the additional viable cells remains in the vascular channels after removal of the fugitive ink, and wherein the additional viable cells comprise endothelial cells; after removing the fugitive ink, injecting a suspension of endothelial cells into the vascular channels; or depositing one or more structural filaments layer by layer on the substrate to form a mold prior to at least partially surrounding the one or more tissue patterns and the vascular pattern with the extracellular matrix composition.

14 .- 16 . (canceled)

17 . The method of claim 13 , wherein the mold comprises flow channels in fluid communication with the vascular channels for perfusion thereof after removal of the fugitive ink.

18 . The method of claim 13 , further comprising:
depositing one or more additional sacrificial filaments on the substrate to form a functional channel pattern, each of the additional sacrificial filaments comprising a second fugitive ink; at least partially surrounding the functional channel pattern with the extracellular matrix composition, and removing the second fugitive ink to create one or more functional channels in the extracellular matrix composition, thereby forming a functional channel network in the tissue construct, wherein, optionally, additional viable cells are deposited with the one or more additional sacrificial filaments, and wherein at least a portion of the additional viable cells remain in the one or more functional channels after removal of the second fugitive ink.

19 . (canceled)

20 . The method of claim 18 , further comprising, after removing the second fugitive ink, injecting a suspension of additional viable cells into the one or more functional channels, wherein the additional viable cells comprise epithelial cells.

21 . (canceled)

22 . The method of claim 18 , wherein the at least partial surrounding of the one or more tissue patterns and the vascular pattern with the extracellular matrix composition occurs during deposition of the one or more cell-laden filaments and the one or more sacrificial filaments, the one or more tissue patterns and the vascular pattern thereby being formed and embedded simultaneously in the extracellular matrix composition.

23 .- 24 . (canceled)

25 . A printed tissue construct with embedded vasculature, the printed tissue construct comprising:
one or more tissue patterns, each tissue pattern comprising a plurality of viable cells of one or more predetermined cell types; a network of vascular channels interpenetrating the one or more tissue patterns; and an extracellular matrix composition at least partially surrounding the one or more tissue patterns and the network of vascular channels.

26 - 27 . (canceled)

28 . The printed tissue construct of claim 25 , wherein the extracellular matrix composition comprises a synthetic or naturally derived biomaterial.

29 .- 31 . (canceled)

32 . The printed tissue construct of claim 25 , wherein the extracellular matrix composition fully surrounds the network of vascular channels.

33 . The printed tissue construct of claim 25 , wherein the viable cells are distributed uniformly throughout each of the one or more tissue patterns.

34 .- 35 . (canceled)

36 . The printed tissue construct of claim 25 , wherein each of the one or more tissue patterns is defined by an arrangement of one or more cell-laden filaments comprising the viable cells, wherein the one or more cell-laden filaments optionally comprise at least one of
an extracellular matrix material, wherein the extracellular matrix material comprises one or more of gelatin, fibrin, and gelatin methacrylate, or one or more functional chemical substances selected from the group consisting of: drugs, small molecules, toxins, proteins, and hormones.

37 .- 43 . (canceled)

44 . The printed tissue construct of claim 25 , wherein one or more of the vascular channels comprise an endothelial layer thereon, the endothelial layer comprising at least about 70% confluency.

45 . (canceled)

46 . The printed tissue construct of claim 44 , wherein one or more of the vascular channels further comprise a stromal layer or a smooth muscle cell layer on the endothelial layer.

47 . The printed tissue construct of claim 25 , further comprising one or more functional channels in the extracellular matrix composition, wherein the one or more functional channels comprise an epithelial layer thereon.

48 . The printed tissue construct of claim 47 , wherein the one or more functional channels comprise an epithelial layer thereon, and, optionally, a stromal layer on the epithelial layer.

49 . (canceled)

50 . The printed tissue construct of claim 25 , further comprising an interface structure at least partially surrounding the extracellular matrix composition and comprising flow channels in fluid communication with the network of vascular channels.

51 . A method of printing an epithelial tissue construct, the method comprising:
depositing one or more sacrificial filaments on a substrate to form a functional channel pattern, each of the sacrificial filaments comprising a fugitive ink and a plurality of epithelial cells; at least partially surrounding the functional channel pattern with an extracellular matrix composition, removing the fugitive ink to create one or more functional channels in the extracellular matrix composition, at least a portion of the epithelial cells re

Assignees

• Harvard College

Inventors

• Lewis Jennifer A
• Kolesky David B
• Skylar-Scott Mark A
• Homan Kimberly A
• Truby Ryan L
• Gladman Amelia Sydney

Classifications

• A61L27/507 Primary

  for artificial blood vessels (apparatus for applying cells on a blood vessel prosthesis A61F2/062) · CPC title

• A61F2/105

  Skin implants, e.g. artificial skin · CPC title

• C12M33/00

  Means for introduction, transport, positioning, extraction, harvesting, peeling or sampling of biological material in or from the apparatus (chemical or physical laboratory apparatus in general B01L, devices for taking cell samples A61B10/0045, withdrawing or distributing predetermined quantities of fluid B01L99/00) · CPC title

• A61F2/06

  Blood vessels · CPC title

• A61L27/222

  Gelatin · CPC title