Modalities for the treatment of degenerative diseases of the retina
This invention relates to methods for improved cell-based therapies for retinal degeneration and for differentiating human embryonic stem cells and human embryo-derived into retinal pigment epithelium (RPE) cells and other retinal progenitor cells.
1. A method of producing a composition comprising isolated human RPE cells suitable for human therapy, comprising:
(a) culturing human pluripotent cells to form clusters of cells;
(b) culturing the clusters of cells of step (a) under adherent conditions that do not maintain the undifferentiated state of the pluripotent cells for a sufficient time for the appearance of pigmented cells comprising brown pigment dispersed in their cytoplasm; and
(c) isolating and culturing the pigmented cells to form a monolayer comprising cells having a cobblestone, polygonal, epithelial-like appearance and brown pigment dispersed in their cytoplasm, and expressing RPE65 and bestrophin, thereby obtaining human RPE cells.
2. The method of claim 1 wherein said clusters of cells of step (a) are trypsinized and cultured on adherent plates.
3. The method of claim 1 , wherein the monolayer of step (c) is dissociated to form a suspension of RPE cells suitable for subretinal transplantation in a human patient.
4. The method of claim 1 , wherein the monolayer of step (c) is disposed on a matrix or substrate to form a monolayer of RPE cells suitable for subretinal transplantation in a human patient.