Transplanted hESC-Derived Retina Organoid Sheets Differentiate, Integrate, and Improve Visual Function in Retinal Degenerate Rats

Abstract

PurposeTo investigate whether sheets of retina organoids derived from human embryonic stem cells (hESCs) can differentiate, integrate, and improve visual function in an immunodeficient rat model of severe retinal degeneration (RD).Methods3D hESC-derived retina organoids were analyzed by quantitative PCR and immunofluorescence. Sheets dissected from retina organoids (30–65 days of differentiation) were transplanted into the subretinal space of immunodeficient rho S334ter-3 rats. Visual function was tested by optokinetic testing and electrophysiologic recording in the superior colliculus. Transplants were analyzed at 54 to 300 days postsurgery by immunohistochemistry for donor and retinal markers.ResultsRetina organoids contained multiple retinal cell types, including progenitor populations capable of developing new cones and rods. After transplantation into an immunodeficient rat model of severe RD, the transplanted sheets differentiated, integrated, and produced functional photoreceptors and other retinal cells, according to the longer human developmental timetable. Maturation of the transplanted retinal cells created visual improvements that were measured by optokinetic testing and electrophysiologic recording in the superior colliculus. Immunohistochemistry analysis indicated that the donor cells were synaptically active. Extensive transplant projections could be seen within the host RD retina. Optical coherence tomography imaging monitored long-term transplant growth and survival up to 10 months postsurgery.ConclusionsThese data demonstrate that the transplantation of sheets dissected from hESC-derived retina organoids is a potential therapeutic method for restoring vision in advanced stages of RD.

Reproducibility and staging of 3D human retinal organoids across multiple pluripotent stem cell lines

“…The capacity to generate bona fide fetal-stage neural retinal cell types and tissues from human embryonic and induced pluripotent stem cells (hESCs and hiPSCs; collectively, hPSCs) has spurred their use in disease modeling (Tucker et al, 2011(Tucker et al, , 2013Jin et al, 2012;Phillips et al, 2014;Yoshida et al, 2015;Arno et al, 2016;Parfitt et al, 2016;Megaw et al, 2017;Schwarz et al, 2017;Sharma et al, 2017;Shimada et al, 2017;Deng et al, 2018) and photoreceptor (PR) replacement efforts (Lamba et al, 2009(Lamba et al, , 2010Hambright et al, 2012;Barnea-Cramer et al, 2016;Shirai et al, 2016;Chao et al, 2017;Gonzalez-Cordero et al, 2017;Mandai et al, 2017;Zhu et al, 2017Zhu et al, , 2018Iraha et al, 2018;Lakowski et al, 2018;McLelland et al, 2018;Gagliardi et al, 2018). Most of these studies employ hPSC differentiation methods that propagate retinal progeny as isolated 3D optic vesicle-like structures (OVs), also known as retinal organoids, in suspension culture (Meyer et al, 2009(Meyer et al, , 2011Nakano et al, 2012;Phillips et al, 2012Phillips et al, , 2018bSridhar et al, 2016;Reichman et al, 2014;Zhong et al, 2014;Kuwahara et al, 2015;Mellough et al, 2015;Singh et al, 2015;Lowe et al, 2016;…”

Photoreceptor cell replacement in macular degeneration and retinitis pigmentosa: A pluripotent stem cell-based approach

“…The transplanted sheets differentiated into functional photoreceptors, leading to visual improvements as evaluated by the optokinetic testing and the electrophysiologic recording in the superior colliculus at 2 to 4 months after transplantation. Interestingly, up to 7 months after transplantation, when no active host photoreceptors were supposed to remain, 9 of 13 rats showed responses to flashes of light in the superior colliculus, with no responses detected in the controls (McLelland et al, 2018).…”

“…Human ESC and iPSC-derived retinal sheets have also been demonstrated to survive and mature following transplantation in a degenerated retinal environment in different animal models (Iraha et al, 2018;McLelland et al, 2018;Shirai et al, 2016;Tu et al, 2019). In a context of xenografts, the development of immunodeficient retinal dystrophic rodents as recipient models has allowed to monitor the long-term transplant growth and survival.…”

“…The majority of the grafts observed at D240-270 developed ONL in rosette-like structures, where the development of inner segment/outer segment structures can be observed, and the regular presence of human INL cells in the graft prevented the direct contact of human photoreceptors with host bipolar cells (Shirai et al, 2016). Very recently, similar immunodeficient retinal dystrophic rats have been used to evaluate the potential of human ESC-derived retinal sheets to restore the visual function up to 10 months post-surgery (McLelland et al, 2018). Based on the differentiation protocol adapted from Zhong and colleagues (Zhong et al, 2014), retinal sheets were prepared from retinal organoids between D30 and 65 of differentiation, when no mature photoreceptors should be present.…”

“…Nevertheless, it is still unclear how to regulate this mechanism. The use of immunecompromised photoreceptor degenerative models in the case of xenografts is becoming more common (Iraha et al, 2018;McLelland et al, 2018;Shirai et al, 2016). This strategy seems more reliable when long-term immunosuppression is needed compared to immunosuppressive drugs and should avoid the severe side effects caused by long-term immunosuppressive…”