Abstract
Purpose: :
To identify the components of the innate immune response in subretinal transplants by using a retinal pigment epithelium (RPE) graft model. To date all cell-based therapies (RPE, Retinal Progenitors and Stem Cells) have been accompanied by significant donor cell loss. The mechanism for this loss is currently unknown. Subretinal transplants have developed into one of the most promising potential treatments for currently incurable retinal disorders such as age related macular degeneration and retinitis pigmentosa. We postulate that the innate immune system plays a major role in graft failure and is not allospecific.
Methods: :
Subretinal transplants of 105 cells from a C57BL/10.RIII.H-2r mouse derived SV40 T +ve RPE cell line were performed in healthy C57BL/10 (syngeneic) mice and harvested at post operative day (POD) 1, 3, 7 and 28 days, sham treatments were also applied (n=4 animals/timepoint). Eyes were fixed and snap frozen for immunocytochemistry. Frozen sections were processed and immunostained for macrophage markers CD11b & F4/80 and neutrophil marker Gr-1. Glial fibrillary acidic protein (GFAP) immunostaining was used to identify the graft site. Donor cells were visualized using a SV40 T antibody.
Results: :
The subretinal bolus at POD 1&3 comprised largely of SV40T +ve donor cells. CD11b, F480 and Gr1 cells increased as a proportion of the bolus from POD 1 to predominate over SV40 T cells by POD 7. Co-localization of SV40 T +ve cells was only seen at POD 7 predominately with F4/80, to a lesser degree with CD11b. By POD 28 the subretinal bolus was no longer in evidence and only occasional grafted cells could be detected. From POD 3 to 28, graft sites in shams and transplants could be detected by GFAP up-regulation.
Conclusions: :
Subretinal syngeneic RPE grafts are subjected, as are allograft cells, to rapid attack by cells of the innate immune system, which infiltrate the graft by POD 1. This culminates in marked graft cell loss and colocalization of surviving graft cells with macrophages by POD 7. We have identified a mechanism of rapid graft cell loss in syngeneic as well as allogeneic subretinal transplant approaches. These will have to be overcome for long term graft survival in future cell-based therapies including stem cells.
Keywords: transplantation • retina • immune tolerance/privilege