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Andrew Osborne, Alessia Tassoni, Julie Sanderson, Keith R Martin; Neuroprotection of human retinal ganglion cells by human mesenchymal stem cells and platelet derived growth factor. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4401.
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© ARVO (1962-2015); The Authors (2016-present)
It is unknown if human mesenchymal stem cells (hMSCs) can protect human retinal ganglion cells (RGCs) after injury. We tested if hMSCs, or platelet derived growth factor (PDGF) as produced by hMSC, could delay RGC death in human retinal explants. Potential neuroprotective downstream signalling pathways were also investigated.
84 human retinal explants from 17 post mortem eyes were cultured for up to 7 days in Neurobasal-A medium. Explants were either co-cultured with 5,000 hMSCs on the RGC layer, maintained in medium containing 50-150ng/ml recombinant PDGF-AA and/or –AB, or co-cultured with inhibitors of molecular signalling pathways. Immunohistochemistry was used to quantify survival of RGC layer cells, to visualise location/activation of PDGF receptors and to investigate downstream signalling. Signalling pathway activation was also assayed by Western blot of retinal lysates 1 and 3 days post treatment.
Transplantation of hMSCs onto human retinal explants increased RGC survival 2.41±0.29-fold (n=5; p<0.05) vs controls. PDGF-AB increased RGC survival 1.82±0.19-fold (n=5; p<0.05). No such protection was observed with PDGF-AA. MSC or PDGF-AB treatment reduced apoptosis (25.5±1.71% and 46.6±3.69% of total RGCs) vs controls (57.0±2.89%) 7 days ex vivo (n=5; p<0.002 and 0.05). Activation of PDGFa and b receptors was detected in the retina after PDGF or MSC treatments with PDGFb expression restricted to the RGC layer. Downstream signalling of survival pathways PI3K, AKT, STAT3 and ERK was upregulated 1 and 3 days after MSC or PDGF-AB treatment vs controls (PI3K 2.22±0.42 and 2.65±0.54, AKT 3.82±1.26 and 2.59±0.78, STAT3 1.77±0.28 and 1.77±0.28, ERK 1.26±0.05 and 1.23±0.04-fold change at day 3; n=3; p<0.05) which correlated with reduced retinal pro-apoptotic BAX expression (0.81±0.05 and 0.75± 0.05-fold change; n=3; p<0.05). Immunohistochemistry showed signalling was occurring in the RGC layer. Inhibition of each pathway reduced retinal BAX expression with PDGF mediated protection abolished in the presence of anti-PDGF (n=3; p<0.05). Protection by MSCs was however still possible in the presence of anti-PDGF.
hMSCs and PDGF protect injured RGCs in human retinal explants. Both MSCs and PDGF triggered activation of similar survival pathways which may offer therapeutic targets to prevent degenerative visual loss.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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