QuantifiableIn VivoImaging Biomarkers of Retinal Regeneration by Photoreceptor Cell Transplantation

Abstract

32Purpose: Short-term improvements in retinal anatomy are known to occur in preclinical models 33 of photoreceptor transplantation. However, correlative changes over the long term are poorly 34 understood. We aimed to develop a quantifiable imaging biomarker grading scheme, using non-35 invasive multimodal confocal scanning laser ophthalmoscopy (cSLO) imaging, to enable serial 36 evaluation of photoreceptor transplantation over the long term. 37Methods: Yellow-green fluorescent microspheres were transplanted into the vitreous cavity 38 and/or subretinal space of C57/BL6J mice. Photoreceptor cell suspensions or sheets from 39 rhodopsin-green fluorescent protein mice were transplanted subretinally, into either NOD.CB17-40 Prkdc scid /J or C3H/HeJ-Pde6b rd1 mice. Multimodal cSLO imaging was performed serially for up 41 to three months after transplantation. Imaging biomarkers were scored, and a grade was defined 42 for each eye by integrating the scores. Image grades were correlated with immunohistochemistry 43 (IHC) data. 44 Results: Multimodal imaging enabled the extraction of quantitative imaging biomarkers 45 including graft size, GFP intensity, graft length, on-target graft placement, intra-graft lamination, 46 hemorrhage, retinal atrophy, and peri-retinal proliferation. Migration of transplanted material 47 increasing the efficiency of preclinical retinal cell transplantation studies in rodents and other 53 animal models. 54 Key words: degenerative retinal diseases, age-related macular degeneration, stem cell therapy, 55 xenotransplantation, photoreceptor cell, retinal organoid, confocal scanning laser 56 ophthalmoscopy 57 Photoreceptor transplantation is being developed as a therapeutic modality to restore vision in 59 people affected by retinal degenerative diseases, including retinitis pigmentosa (RP) and age-60 related macular degeneration (AMD) 1-6 . Short term improvements in outer retinal anatomy after 61 photoreceptor cell transplantation have been observed, mainly by histological staining in 62 preclinical models of retinal degeneration 4,7 . However, histology is a relatively inefficient 63 method to track graft and recipient anatomy longitudinally over the long term. Histological 64 assays are labor-intensive, require large initial cohorts of recipients, and face the challenge of 65 recipient attrition over time. Non-invasive imaging could facilitate the longitudinal evaluation of 66 retinal anatomy in relatively smaller cohorts of recipient animals over time, without the need to 67 sacrifice animals at every assessment time point. 68 Recent advances in imaging techniques have enabled detailed imaging studies in mouse models 69 of retinal disease and regeneration 8-10 . Confocal scanning laser ophthalmoscopy (cSLO) can be 70 used to capture images in multiple imaging modes, including short-wavelength fluorescence 71 (SWF) excitation (488 nm) to detect photoreceptor cells labeled with green fluorescent protein 72 (GFP) in mouse recipients 1 . Multicolor reflectance (MR) imaging combines blue (488 nm)...

The Prospects for Retinal Organoids in Treatment of Retinal Diseases

“…They were able to observe migration of the transplanted tissue as well. 38 While these 2 techniques demonstrated high resolution and dynamic imaging, it relied on genetically engineered reporter cell lines (CRX +/tdTomato and Rho +/GFP ) to emit fluorescent light, which is not applicable for future clinical use in human subjects.…”