Abstract
Purpose: :
To determine if bone marrow–derived stem cells (BMSC) have the capacity in vitro and in vivo to express retinal pigment epithelial (RPE)–like markers.
Methods: :
In vitro, mouse Sca–1+ GFP+ cells of bone marrow origin were used in coculture with adult mouse RPE cells. The coculture in a 1:1 ratio was performed with and without cell–cell–contact for up to 3 weeks. Mouse fibroblasts served as a control. Immunocytochemical analysis was performed using monoclonal antibodies (mAbs) against specific RPE markers – cytokeratin, RPE65, MITF – as well as non–RPE markers – opsin (photoreceptors) and glial fibrillary acidic protein (GFAP; glia). In vivo, sodium iodate (NaIO3) was used to damage the RPE. For this study, C57BL/6 mice were injected i.v. with 35 mg/kg NaIO3 followed by the subretinal (s.r.) injection of 3x104 Sca–1+ GFP+ BMSC on day 3. The mice were sacrificed on days 7, 14, 21, and 28 after transplantation. Whole eye flat mounts (FM) were prepared and examined for GFP+ cells under a laser scanning microscope (argon laser). Immunocytochemical analysis was performed on FM, as well as on paraffin cross sections, using mAbs against the specific RPE marker RPE65.
Results: :
In vitro, BMSC changed from round to flattened, polygonal cells and expressed cytokeratin, RPE65 and MITF when cocultured in direct contact with RPE cells. In vivo, with 35 mg/ml of NaIO3 patchy RPE damage was observed starting on day 14. At the same time point the transplanted GFP+ Sca–1+ BMSC in the subretinal space at the sites of RPE loss expressed RPE65.
Conclusions: :
BMSC are capable of expressing RPE–like markers in coculture with adult RPE cells, as well as in the subretinal space in a murine model of RPE cell loss. However, it is not yet known whether these cells are capable of replacing the damaged RPE functionally.
Keywords: retinal pigment epithelium • retinal degenerations: cell biology • differentiation