Method for creating perfusable microvessel systems

• US 9,446,169 B2
• Filed: 08/17/2011
• Issued: 09/20/2016
• Est. Priority Date: 03/24/2006
• Status: Active Grant

• Current Assignee: Nortis, Inc.
• Original Assignee: Nortis, Inc.
• Inventors: Neumann, Thomas
• Primary Examiner(s): HURST, JONATHAN M
• Application Number:
  US13/212,107
• Publication Number:
  US 20110300616A1
• Time in Patent Office: 1,861 Days
• Field of Search: 435/297.4, 435/400, 435/284.1, 435/283.1, 435/283.4
• US Class Current: 1/1
• CPC Class Codes: A61L 27/3808 Endothelial cells
• CPC Class Codes: C12M 21/08 for producing artificial tissue or for ex-vivo cultivation of tissue prostheses A61F2/00, grafts A61L27/00
• CPC Class Codes: C12M 23/06 Tubular C12M23/08, C12M23/16 take precedence
• CPC Class Codes: C12M 23/20 Material Coatings immunocoatings C12M25/00
• CPC Class Codes: C12M 25/14 Scaffolds; Matrices in general C12N5/0068
• CPC Class Codes: C12M 29/10 Perfusion
• CPC Class Codes: C12N 2533/50 Proteins
• CPC Class Codes: C12N 5/0068 General culture methods using substrates for specific animal cell type C12N5/06
• CPC Class Codes: C12N 5/0691 Vascular smooth muscle cells; 3D culture thereof, e.g. models of blood vessels

Abstract

A method for the creation of endothelial parent vessels from human vascular endothelial cells in vitro in a culture perfusion device (CPD) including a collagen chamber, inlet ports, a capillary tube, and an outlet port. A collagen solution is injected into the collagen chamber through a syringe needle until the chamber is filled with collagen. The CPD is perfused by filling the inlet ports and sequentially priming the inlet ports, and the outlet ports. A perfusable channel is created in the collagen chamber and a concentrated suspension of endothelial cells is injected into the inlet ports. The endothelial cells are injected into the at least one perfusable channel and incubated to attach to the walls of the perfusable channel. The cells are distributed within the CPD; and perfused to form a parent vessel having homogeneous monolayers of cells.

Claims

1. A device for creating perfusable vessels in vitro comprising:

a device body including a silicone layer sandwiched between a first layer and a second layer;

a plurality of fluid ports embedded in the silicone layer;

a plurality of port connecting channels embedded in the silicone layer;

at least one cell reservoir port embedded in the silicone layer, the cell reservoir port being connected by a first port connecting channel to an inlet port and connected by a second port connecting channel to an outlet port;

a matrix chamber containing collagen embedded in the silicone layer, the matrix chamber having one or more perfusion channels formed in the collagen;

a plurality of non-permeable capillary tubes each coupled at one end to one or more of the plurality of fluid ports and having a second end;

wherein only the second end of each non-permeable capillary tube reaches into the matrix chamber; and

wherein the first end of each non-permeable capillary tube is located to be pulled back into the at least one cell reservoir port.

2. The device of claim 1 further comprising a perfusion system connected to a pair of said inlet and outlet ports which are individually connected to opposing ends of at least one of the plurality of port connecting channels.

3. The device of claim 2 where the perfusion system comprises at least one syringe pump.

4. The device of claim 2 where the perfusion system comprises at least one peristaltic pump.

5. The device of claim 2 the perfusion system comprises a gravity flow system.

6. The device of claim 1 wherein the plurality of non-permeable capillary tubes comprise polyimide-coated fused-silica capillaries.