Previous studies evaluating neural stem cells transplanted into the mature retina have demonstrated limited levels of graft-host integration and photoreceptor differentiation. The purpose of this investigation is to enhance photoreceptor cell differentiation and integration of retinal progenitor cells (RPC) following subretinal transplantation into retinal degenerate rats by optimization of isolation, expansion, and transplantation procedures.RPCs were isolated from human placental alkaline phosphatase (hPAP)-positive embryonic day 17 (E17) rat retina and expanded in serum-free defined media. RPCs at passage 2 underwent in vitro induction with all trans retinoic acid or were transplanted into the subretinal space of post-natal day (P) 17 S334ter-3 and S334ter-5 transgenic rats. Animals were examined post-operatively by ophthalmoscopy and optical coherence tomography (OCT) at weeks 1 and 4. Differentiation profiles of RPCs, both in vitro and in vivo were analysed microscopically by immunohistochemistry for various retinal cell specific markers. Our results demonstrated that the majority of passage 2 RPCs differentiated into retina-specific neurons expressing rhodopsin after in vitro induction. Following subretinal transplantation, grafted cells formed a multi-layer cellular sheet in the subretinal space in both S334ter-3 and S334ter-5 rats. Prominent retina-specific neuronal differentiation was observed in both rat lines as evidenced by recoverin or rhodopsin staining in 80% of grafted cells. Less than 5% of the grafted cells expressed glial fibrillary acidic protein. Synapsin-1 (label for nerve terminals) positive neural processes were present at the graft-host interface. Expression profiles of the grafted RPCs were similar to those of RPCs induced to differentiate in vitro using all-trans retinoic acid.In contrast to our previous study, grafted RPCs can demonstrate extensive rhodopsin expression, organize into layers, and show some features of apparent integration with the host retina following subretinal transplantation in slow and fast retinal degenerate rats. The similarity of the in vitro and in vivo RPC differentiation profiles suggests that intrinsic signals may have a significant contribution to RPC cell fate determination. q
Purpose. To correlate the functional outcomes with histologic findings following transplantation of fetal retinal sheets in rd mice, and to investigate the mechanisms of visual function restoration.Methods. Twenty-one postnatal day 31 -38 rd/rd (C3H/HeJ) mice were transplanted in one eye with retinal sheets (1·0 £ 0·4 mm) obtained from embryonic day (E) 17 enhanced-green-fluorescent protein (eGFP) mice. Five mice underwent sham surgery without insertion of tissue. Four to five weeks after transplantation, visual responses to a light flash were recorded across the superior colliculus (SC) in seven eyes of seven transplanted mice that had clear corneas and lenses, and in all five sham surgery mice. Following the SC recording, the eyes were enucleated and processed for immunohistochemistry and examined using confocal microscopy.Results. In three out of the seven eyes (43%), positive responses were recorded in the SC in an area topographically corresponding to the placement of the transplant in the host retina. No responses were recorded in the untreated eyes of 5-week-old and 9-week-old rd/rd mice, and in the 9-week-old sham surgery mice. In contrast, visual responses were recorded over the entire SC in normal eyes. The response onset latencies of the 3 transplanted mice with responses were similar to those of normal control mice. The organization of the graft did not appear to correlate as expected with the electrophysiology results, as eyes with well-organized, laminated grafts showed no response whereas the three light-responsive eyes had rosetted or disorganized grafts. All three light-responsive eyes demonstrated much higher levels of recoverin immunoreactivity in the host retina overlying the graft compared with untreated age-matched rd/rd mice.Conclusion. Restoration of the SC visual response does not appear to depend on a well-organized transplant in the rd mouse. Increased recoverin-staining in the host retina in light-responsive animals suggested that host cone rescue was the likely mechanism of vision restoration in this transplant model. q
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.