Genetically engineered stem cell-derived retinal grafts for improved retinal reconstruction after transplantation

Abstract

Summary ESC/iPSC-retinal sheet transplantation, which supplies photoreceptors as well as other retinal cells, has been shown to be able to restore visual function in mice with end-stage retinal degeneration. Here, by introducing a novel type of genetically engineered mouse ESC/iPSC-retinal sheet with reduced numbers of secondary retinal neurons but intact photoreceptor cell layer structure, we reinforced the evidence that ESC/iPSC-retinal sheet transplantation can establish synaptic connections with the host, restore light responsiveness, and reduce aberrant retinal ganglion cell spiking in mice. Furthermore, we show that genetically engineered grafts can substantially improve the outcome of the treatment by improving neural integration. We speculate that this leads to reduced spontaneous activity in the host which in turn contributes to a better visual recovery.

A Genetic modification that reduces ON-bipolar cells in hESC-derived retinas enhances functional integration after transplantation

“…There was no effect of host sex ( Figure 6 H sex). Similar to our previous finding in mice ( Matsuyama et al., 2021 ), there was a small but distinct effect of stimulus strength , with spontaneous activities increased after brighter stimulation ( Figure 6 H stimulus). We also found that spontaneous activity clearly increased upon L-AP4 blockade ( Figure 6 H L-AP4 treatment).…”

“…Figure 6 G shows the distribution of averaged spontaneous firing rate (spontaneous spiking before light stimulation). The spontaneous activity has a lognormal distribution, as we have reported previously ( Matsuyama et al., 2021 ). We estimated the effect of different parameters (group, sex, L-AP4 treatment, stimulus strength, and sample bias) on the spontaneous activity using hierarchical Bayesian inference.…”

“…Previous studies showed that Bhlhb4 gene deletion specifically reduced rod bipolar cells ( Bramblett et al., 2004 ), whereas Isl1 conditional KO mice presented a considerable reduction of ON-bipolar cells, cholinergic amacrine cells, and RGC ( Elshatory et al., 2007 ). Consistently, using mouse ESC/iPSC-retinas, we observed a drastic reduction of rod bipolar cells in both KO graft lines and of subsets of cone bipolar cells in Isl1 KO ( Isl1 −/− ) retina grafts, both with enhanced host-graft synaptic formation, signal-to-noise ratio in RGC response, and improved light responsive behavior ( Matsuyama et al., 2021 ). In the current study, in view of clinical application, we deleted the ISL1 −/− gene to confirm the reproducibility of these features using hESC-retinas.…”

“…To better characterize and compare the light responsiveness in transplanted retinas with robust RGC spontaneous firing, we classified the detected host RGCs, in consideration of their sensitivity to L-AP4 blockade and recovery after L-AP4 washout, into unresponsive , onXoff , on , and adapted on types ( Figure 6 F; see STAR Methods for further details) ( Matsuyama et al., 2021 ). Most of the responding cells in transplanted retinas were on or adapted on types with similar on to adapted on ratios in both types of transplanted retinas.…”

“…Based on these observations, in order to enhance host-graft contact by reducing graft bipolar cells while retaining the cellular and structural benefits of the retinal sheet, we recently reported the knockout (KO) effect of Bhlhb4 and Islet-1 (Isl1) genes that are related to the fate-determination and maturation of bipolar cells, in mouse ESC/iPSC-retinal sheet ( Matsuyama et al., 2021 ). Previous studies showed that Bhlhb4 gene deletion specifically reduced rod bipolar cells ( Bramblett et al., 2004 ), whereas Isl1 conditional KO mice presented a considerable reduction of ON-bipolar cells, cholinergic amacrine cells, and RGC ( Elshatory et al., 2007 ).…”