Three-dimensional tissue equivalent using macromass culture
The present invention provides a three-dimensional tissue equivalent for in-vivo and in-vitro uses. The three dimensional tissue equivalent of the present invention is a non-contractile cellular sheet cultured over a porous scaffold using a macromass culturing technique, for example where the cellular sheet is entirely on one side of a porous sponge. In one embodiment, the present invention provides a dermal wound dressing that comprises a high density cellular sheet of dermal fibroblast cells.
1. A process for preparing a three dimensional non-contractile tissue equivalent comprising a macromass cellular sheet of dermal fibroblast cells adhered to a porous scaffold or matrix, wherein the method comprises:
a) absorbing a liquid blocking agent into pores of the porous scaffold or matrix,
b) solidifying the blocking agent;
c) culturing dermal fibroblast cells onto only one side of the porous scaffold or matrix to form a multilayered high density macromass cellular sheet, wherein the porous scaffold or matrix comprises pores that do not contact the macromass cellular sheet and are devoid of the cells;
d) desolidifying and removing the blocking agent; and
e) forming a three dimensional non-contractile tissue equivalent comprising the macromass cellular sheet adhered to the porous scaffold or matrix.
2. The method of claim 1 , wherein the dermal fibroblast cells comprise neonatal human dermal fibroblast cells.
3. The method of claim 1 , wherein the blocking agent comprises material chosen from gelatin, alginate, pectin, agar and agarose.
4. The method of claim 1 , wherein the porous scaffold or matrix comprises material chosen from chitosan, collagen, polyglycolic acid, and polylactic acid.
5. The method of claim 1 , wherein the dermal fibroblast cells do not leak or pass through the pores of the porous scaffold or matrix during the culturing step.
6. The method of claim 1 , wherein at least 20×10 6 dermal fibroblast cells are seeded onto the porous scaffold or matrix during the culturing step, and wherein the macromass cellular sheet has a cell density of 1×10 6 cells per cm 2 to 12×10 6 cells per cm 2 of the porous scaffold or matrix.
7. The method of claim 1 , wherein the cells in the macromass cellular sheet express VEGF and/or IL-8 in an amount greater than would be expressed by the same cells in a monolayer having the same diameter as the macromass cellular sheet.