Investigating the mechanisms of molecular exchange in between retinal neurons

Background: Retinal degeneration due to the loss of photoreceptors (PRs) is the leading cause of untreatable blindness. Repair by transplantation of healthy PRs is a promising therapeutic tool. Previous studies have shown that transplantation of PR precursors can rescue visual function in some model...

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Bibliographic Details
Published in:Journal of extracellular vesicles 2018-01, Vol.7, p.227-227
Main Authors: Kalargyrou, Aikaterini, Ali, Robin, Pearson, Rachael
Format: Article
Language:English
Subjects:
Actin
Animal models
Blindness
Cell adhesion
Cell culture
Horizontal cells
Immunofluorescence
Neural stem cells
Photoreceptors
Recombination
Retina
Retinal degeneration
Transplantation
Ultracentrifugation
Visual perception
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Summary:Background: Retinal degeneration due to the loss of photoreceptors (PRs) is the leading cause of untreatable blindness. Repair by transplantation of healthy PRs is a promising therapeutic tool. Previous studies have shown that transplantation of PR precursors can rescue visual function in some models of retinae dystrophy. Previously, this was thought to arise from donor PRs integrating within the host retina. However, we have recently shown that, where some host PRs remain, many reporter-labelled cells previously interpreted as integrated donor cells, were actually host PRs that acquired the label through molecular exchange or material transfer, between donor and host cells. This exchange is robust and permits acquisition by the host cell of many proteins expressed only by the donor. Since extracellular vesicles (EVs) are increasingly recognized as key players of molecular communication, we hypothesized that material transfer is mediated by the exchange of molecular information packaged in EVs. Methods: Rod PRs were isolated from postnatal day (P)4 wildtype mouse retinae using MACS and cultured for 14-21 days. EVs were isolated from culture medium using differential ultracentrifugation. Large, medium and small EVs retrieved by 2K, 10K and 100K spins were analysed with DLS, TEM, Western blot, dot-blot and RTqPCR. MVB analysis in whole eyes was performed using TEM. TNTs were analysed with confocal imaging. Functional exchange was assessed using with a Cre-loxP recombination read-out system. Results: Cultured PRs release a variety of EVs in a developmentally dependent manner. Small EVs (sEVs) bear proteins typical of PRs and of endocytic origin. When separated in a transwell co-culture system, Cre+ photoreceptors can mediate recombination of underlying reporter retinal cells through a mechanism that does not require sustained cell-cell contact. In culture, primary PRs extend filamentous actin+ protrusions within the first 24 h. These changes over time, and immunofluorescence analysis reveals the presence of vesicular like forms within them. Summary/Conclusion: Primary PRs release sEVs with morphological and molecular profiles typical of neuronal EVs in a developmentally dependent manner. These sEVs appear capable of mediating horizontal signalling with other retinal cells in vitro.
ISSN:2001-3078