Subretinal survival of retinal progenitors (RPs) derived from human embryonic stem cells (hESCs) in different animal models

Abstract

Purpose : To examine the survival of RPs derived from hESCs following subretinal (SR) transplantation in immunodeficient rodent models.

Methods : hESCs (HAD102 line) engineered to express GFP were differentiated in-vitro towards a retinal fate using 2 protocols: To obtain RPs, hESCs were grown with bFGF, nicotinamide and CHIR while Retinal Pigmented Epithelium (RPE) cells were derived using our previously published protocol (Idelson et al., Cell Stem Cells, 2009).
Between 75-150K cells were transplanted as a suspension to the SR space of 3 animal models: Royal College of Surgeons (RCS) rats (n=66 eyes), Nude-RCS rats (n=18 eyes) and NSG mice (n=53 eyes). RCS rats were treated with Cyclosporine in the drinking water to reduce rejection. Nude-RCS rats are athymic and lack T cells, while NSG mice lack mature T, B and natural killer cells.
Success of engraftment was determined at 4 weeks (4w) post-transplant by in-vivo visualization of GFP expression using a Micron III Retinal Microscope. Further survival, graft rejection or tumor formation was examined at set time points using the Micron III and in select cases by Spectralis OCT imaging in vivo as well as by retinal histology.

Results : In RCS rats receiving hESC-derived RPs (n=42), cell rejection (often manifest as development of retinal detachment) was observed in 25% of eyes at 6w, rising to 70% rejection at 8w. In order to try and reduce cells rejection, RPs were then transplanted in Nude-RCS eyes (n=18). However, in this model rejection was also noted at 6w in 62% of eyes.
Interestingly, hESC-derived RPE cells survived in RCS rats for 8w or more with no signs of acute rejection (n=24).
Survival of RPs in NSG mice (n=27) was much improved, with 100% survival at 6w, 90% at 10w and 80% survival at 16w. However, in 2 out of 4 eyes further followed for 22w, marked proliferation of the cells occurred with tumor development. In similarity to transplants in RCS rats, no signs of acute rejection were noted when hESC-derived RPE cells were transplanted in NSG mice (n=26).

Conclusions : Immune status of recipient animals influences transplant survival. NSG, a highly immunodeficient mouse, provides a better model for studying long term survival, differentiation and possible integration of RPs. hESC-derived RPs invoke much higher levels of rejection than RPE cells derived from the same hESC line even when transplanted in immune-deficient animals.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.