Daylight Vision Repair by Cell Transplantation

Abstract

Human daylight vision depends on cone photoreceptors and their degeneration results in visual impairment and blindness as observed in several eye diseases including age-related macular degeneration, cone-rod dystrophies, or late stage retinitis pigmentosa, with no cure available. Preclinical cell replacement approaches in mouse retina have been focusing on rod dystrophies, due to the availability of sufficient donor material from the rod-dominated mouse retina, leaving the development of treatment options for cone degenerations not well studied. Thus, an abundant and traceable source for donor cone-like photoreceptors was generated by crossing neural retina leucine zipper-deficient (Nrl 2/2 ) mice with an ubiquitous green fluorescent protein (GFP) reporter line resulting in double transgenic tg(Nrl 2/2 ; aGFP) mice. In Nrl 2/2 retinas, all rods are converted into cone-like photoreceptors that express CD73 allowing their enrichment by CD73-based magnetic activated cell sorting prior transplantation into the subretinal space of adult wild-type, cone-only (Nrl 2/2 ), or cone photoreceptor function loss 1 (Cpfl1) mice. Donor cells correctly integrated into host retinas, acquired mature photoreceptor morphology, expressed cone-specific markers, and survived for up to 6 months, with significantly increased integration rates in the cone-only Nrl 2/2 retina. Individual retinal ganglion cell recordings demonstrated the restoration of photopic responses in cone degeneration mice following transplantation suggesting, for the first time, the feasibility of daylight vision repair by cell replacement in the adult mammalian retina. STEM CELLS 2015;33:79-90

Rebuilding the Missing Part—A Review on Photoreceptor Transplantation

“…Despite heterogeneous expression of GFP throughout more than one cell population (cones and rod bipolars), GFP-positive cones are solely labeled until P14, allowing their enrichment using FACS. Transplantation of E17.5 Ccdc136 GFP/+ sorted cell fractions resulted in integrated photoreceptors in C57BL/6j wild-type hosts but these cells lacked the expression of cone-specific photo-transduction markers (Smiley et al, 2016) in contrast to Santos-Ferreira et al (2015) who showed cone markers (cone arrestin, s-opsin) in reporter labeled cells following cone-like photoreceptor transplantation into CPFL1 hosts. Interestingly, in both studies the reporter labeled cells within the host ONL frequently exhibited rod-like morphologies, that might be explained by the exchange of cell material between donor and host photoreceptors (see below).…”

“…Functional analysis was performed by micro-electrode arrays (MEA), that allow the simultaneous read out of hundreds of retinal ganglion cells (RGCs). These measurements showed spike train responses in RGCs under daylight conditions (Santos-Ferreira et al, 2015). Importantly, sham or rod photoreceptor injected CPFL1 hosts were unable to detect high-intensity light stimuli.…”

Statistics for real-time deformability cytometry: Clustering, dimensionality reduction, and significance testing

“…After dissection of the eyes, the retina was isolated and transferred to a papain solution (Worthington, Lakewood, USA) and incubated for 35 min at 37 °C 38 . Retinas dissociated to a single cell suspension were centrifuged for 5 min at 300 rcf, re-suspended in Fluorescence-activated cell sorting buffer (2 mM EDTA and 1% bovine serum albumin in PBS) and passed through a 40  μ m nylon cell strainer (BD Biosciences, Heidelberg, Germany) before FAC-sorting.…”

Challenges in Retinal Circuit Regeneration

“…7,[25][26][27][28] Photoreceptor precursors at specific ontogenic stages (P4-P6) integrated into recipient adult retinas. 7,27) We theorized that if ES cells could differentiate into photoreceptors that correspond to the P4-P6 stages, those cells might integrate into the host retina and restore visual function.…”