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Thomas V. Johnson, Victoria A. Levasseur, Nicholas W. Dekorver, Radu I. Cojocaru, Stanislav I. Tomarev; Retinal Ganglion Cell Neuroprotection By Mesenchymal Stem Cell-derived Factors. Invest. Ophthalmol. Vis. Sci. 2012;53(14):4682.
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© ARVO (1962-2015); The Authors (2016-present)
We have shown that bone marrow-derived mesenchymal stem cells (MSCs) confer neuroprotection to retinal ganglion cells (RGCs) in vivo and in organotypic retinal culture. This work assesses the neuroprotective properties of factors secreted by MSCs.
Adult rat retinal explant cultures were maintained for 7 days ex vivo using an established protocol. Explants were co-cultured with MSCs or with indicated factors diluted in culture media. RGC survival was assessed using immunohistochemistry to quantify Islet-1+ cell survival in the RGC layer. MSC gene expression was compared before and after retinal explant co-culture using Human Genome U133 Plus 2.0 arrays.
Transplantation of human MSCs onto retinal explants resulted in 67±11% greater RGC survival (p<0.001) compared to controls; treatment of explants with MSC conditioned medium increased RGC survival by only 25±8% (p<0.05). Sephacryl gel fractionation of MSC conditioned media yielded one fraction with improved neuroprotective capacity, which increased RGC survival by 48±11% (p<0.05). To avert confounding effects of unidentified secreted factors, explants were treated with a cocktail of 13 purified proteins, previously identified as being enriched for in the MSC secretome (Dekorver et al., ARVO 2011, #2603), which increased RGC survival by 29±6% (p<0.05). Subgrouping proteins according to signaling pathway revealed that factors acting through gp130 (IL-6, IL-11, and LIF) reduced RGC survival by 15±2% (p<0.01) whereas other subgroups significantly protected RGCs; the strongest effect was seen for PDGF family proteins, which increased RGC survival by 62±9% (p<0.001). To identify changes in MSC phenotype resulting from interactions with retinal tissue, MSC gene expression was assayed using gene microarrays, which revealed that 1,351 genes were significantly (p<0.05) up- or down-regulated more than 2-fold in MSCs following co-culture with retinal explants. Ingenuity Pathway Analysis of differentially regulated genes revealed overrepresentation of genes involved in growth factor signaling, inflammation, endocytosis and axon guidance that might potentially contribute to neuroprotection by MSCs.
MSCs secrete a variety of bioactive factors, some of which are neuroprotective to RGCs in organotypic retinal explant culture. Ongoing works aims to identify the most important factors mediating this neuroprotection and to determine how changes in gene expression induced by exposure to retinal tissue promote the neuroprotective effects of MSCs.
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