Bioengineered human corneal stromal tissue
Provided herein is a method of making an aligned ECM scaffold useful in refractive correction of the eye and repair of the cornea. Methods of use of the scaffold as well as a scaffold construct are provided.
1. A method of preparing an aligned extracellular matrix (ECM) scaffold comprising culturing functional keratocytes on a template scaffold comprising one or more layers comprising aligned fibers of a biocompatible, biodegradable polymeric composition for a length of time sufficient for the cells to produce a product ECM scaffold having oriented fibers, wherein the biocompatible, biodegradable polymeric composition comprises:
a polyurethane, a polyester, a polyether, a polyacrylamide, and/or a polycarbonate;
a polyurethane, a polyester, a polyether, a polyacrylamide, and/or a polycarbonate-containing block copolymer, or
a copolymer formed from one or more acrylic monomers, acrylamide monomers, succinimide monomers, glycolide monomers, caprolactone monomers, dioxanone monomers, lactide monomers, and/or carbonate monomers, and
wherein the template scaffold degrades so that the product ECM scaffold does not contain the biocompatible, biodegradable polymeric composition of the template scaffold.
2. The method of claim 1 , in which the biocompatible, biodegradable polymeric composition comprises a poly(ester urethane) urea elastomer.
3. The method of claim 1 , in which the biocompatible, biodegradable polymeric composition comprises a polymer composition having a Lower Critical Solution Temperature of 35° C. or less.
4. The method of claim 3 , in which the biocompatible, biodegradable polymeric composition comprises poly(N isopropyl acrylamide).
5. The method of claim 1 , in which the template scaffold comprises a plurality of the one or more layers and wherein, the aligned fibers of a first layer of the plurality of the one or more layers are arranged at a different angle with respect to the aligned fibers of a second layer of the plurality of the one or more layers that is adjacent to the first layer.
6. The method of claim 5 , in which the aligned fibers of a first layer of the plurality of layers are arranged at a 20° to 90° angle with respect to the aligned fibers of a second layer of the plurality of the one or more layers that is adjacent to the first layer.
7. The method of claim 1 , further comprising preparing the template scaffold by electrospinning the biocompatible, biodegradable polymeric composition to prepare the aligned fibers of the one or more layers.
8. The method of claim 1 , further comprising preparing the template scaffold by seeding one or more of the one or more layers with functional keratocytes or functional keratocyte precursors that are differentiated on the template scaffold to produce the functional keratocytes.
9. The method of claim 8 , in which the one or more layers comprises a plurality of layers and further comprising stacking the one or more layers so that at least one layer is arranged so that its fibers are oriented at a different angle with respect to fibers of an adjacent layer.
10. The method of claim 8 , in which the functional keratocytes or functional keratocyte precursors are seeded by electrospraying.
11. The method of claim 8 , in which the functional keratocytes or functional keratocyte precursors are seeded by depositing cells onto the one or more layers in culture.
12. The method of claim 8 , in which the functional keratocyte precursors are corneal stroma stem cells.
13. The method of claim 8 , in which the functional keratocyte precursors are adipose-derived stem cells.
14. The method of claim 1 , further comprising decellularizing the product ECM scaffold.
15. The method of claim 1 , wherein the biocompatible, biodegradable polymeric composition comprises a polyurethane, a polyester, or a polymer composition having a Lower Critical Solution Temperature of 35° C. or less.