Transplanted human photoreceptors transfer cytoplasmic material but not to the recipient mouse retina

Abstract

Background The discovery of material transfer between transplanted and host mouse photoreceptors has expanded the possibilities for utilizing transplanted photoreceptors as potential vehicles for delivering therapeutic cargo. However, previous research has not directly explored the capacity for human photoreceptors to engage in material transfer, as human photoreceptor transplantation has primarily been investigated in rodent models of late-stage retinal disease, which lack host photoreceptors. Methods In this study, we transplanted human stem-cell derived photoreceptors purified from human retinal organoids at different ontological ages (weeks 10, 14, or 20) into mouse models with intact photoreceptors and assessed transfer of human proteins and organelles to mouse photoreceptors. Results Unexpectedly, regardless of donor age or mouse recipient background, human photoreceptors did not transfer material in the mouse retina, though a rare subset of donor cells (<5%) integrated into the mouse photoreceptor cell layer. To investigate the possibility that a species barrier impeded transfer, we used a flow cytometric assay to examine material transfer in vitro. Interestingly, dissociated human photoreceptors transferred fluorescent protein with each other in vitro, yet no transfer was detected in co-cultures of human and mouse photoreceptors, suggesting that material transfer is species specific. Conclusions While xenograft models are not a tractable system to study material transfer of human photoreceptors, these findings demonstrate that human retinal organoid-derived photoreceptors are competent donors for material transfer and thus may be useful to treat retinal degenerative disease.

1. Introduction

Photoreceptor degeneration is irreversible due to the limited regenerative capacity of the mammalian central nervous system. Cell therapy to the retina has been an ongoing pursuit since the 1980s with the hope that the new healthy donor cells deposited in the subretinal space would engraft and form new synaptic connections with the host retina to restore vision [1, 2]. The amount of GFP-labelled donor cells found inside the mouse outer nuclear layer (ONL), termed cellular integration, was used to evaluate transplant success. In 2006, seminal work established that retinal dissociates from postnatal day 4–6 mice were the optimal donor cell age for transplantation [3, 4], leading to the adoption of this donor cell age for many subsequent murine to murine photoreceptor transplantations [5–10].

Many groups have since reported that transplantation of post-mitotic donor photoreceptors resulted in visual recovery, and these functional improvements were thought to be correlated to the number of donor cells migrating and integrating into the mouse ONL [6–8]. However, later studies demonstrated that transplanted mouse photoreceptors rarely integrate with the recipient retina and, instead, transfer cytoplasmic contents, such as the fluorescent GFP tag used to label the donor cells, to the host retina, in a process termed material transfer [11–15]. Further investigations into this phenomenon have demonstrated that transfer is bidirectional (donor to host and host to donor), requires direct contact, and is mediated through the formation of nanotube-like protrusions between photoreceptor cells [11–17].

Though cell integration may not the main mechanism for visual recovery, cell transplantation to the retina still holds promise. Material transfer has the potential to be leveraged as a therapy to deliver organelles or proteins involved in visual transduction from healthy donor cells to rescue diseased photoreceptors. Mouse photoreceptors have been shown to transfer RNA, cytoplasmic proteins, and organelles; however, for this to be a feasible strategy, it is important to determine if human photoreceptors can engage in material transfer. The majority of reported studies on human photoreceptor transplants have used mouse models of end-stage retinal disease.