IMMUNOHISTOCHEMICAL AND HISTOPATHOLOGICAL CHARACTERISATION OF ALLOGENIC GFP+ PIG RETINAL PROGENITOR CELLS AFTER TRANSPLANTATION INTO THE SUB-RETINAL SPACE

Abstract

Purpose: The development of photoreceptor replacement therapy for retinal degenerative disorders requires transplantation experiments to evaluate the potency of the donor cells.Although xenotransplants of human cells are possible, they are complicated by the host immune response, which often causes rejection before examination of the graft can be performed. Moreover, preclinical studies of a cell line analogous to human cells is an accepted path for demonstration of safety and efficacy.We therefore have isolated a porcine retinal progenitor cell (RPC) line that is equivalent to an established human RPC line. Here we investigate the outcome of allogeneic pig RPC transplantation into the subretinal space of control and immunosuppressed pigs.

Methods: RPCs were derived from the neural retina of GFP transgenic pigs, characterized and transplanted to the subretinal space of hybrid pigs.The recipients were divided into two groups: six pigs received only sub-retinal transplantation, and six pigs received sub-retinal injections of RPCs followed by local immunosuppression with rapamycin, delivered by intravitreal injections following transplantation. Four weeks after transplantation the eyes were removed and analyzed by ICC and histopathology.

Results: We were able to demonstrate the presence of RPC by immune staining for GFP in the inner segment and in the outer nuclear layer (ONL) in all the retinal samples subjected to staining. A few retinas also displayed small bipolar-like cells and cell processes in the ONL.In the 2 samples investigated RPCs did not express Ki-67; host tissue demonstrated a low level of proliferation with very small numbers of Ki-67 cells scattered throughout the retinal layers. GFAP, recoverin and rhodopsin were shown to co-localise with GFP labelled RPCs.Histopathological analyses did not show any evident macrophages or inflammatory cells.

Conclusions: No abnormal pathology was identified in any of the cells grafted to the retina.There was no evidence of immune response, hypercellularity or autofluorescent macrophages, regardless of the immunosuppression treatment.Taken together, the good level of RPC survival, position of surviving transplanted RP cells, lack of graft proliferation and absence of immune response at 4 weeks post implantation suggests allogeneic RPC cell transplantation in pigs warrants further investigation.

Keywords: 696 retinal degenerations: hereditary • 412 age-related macular degeneration • 741 transplantation