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Christophe Roubeix, David Godefroy, Ivana IVKOVIC, Fradot Valerie, Serge A Picaud, Stephane Melik Parsadaniantz, Françoise Baudouin, Christophe Baudouin; The secretome of Mesenchymal stem cells protects both primary trabecular meshwork cells and retina ganglion cells. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):2411.
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© ARVO (1962-2015); The Authors (2016-present)
To investigate the effect of Mesenchymal Stem Cells (MSCs) secretome (MSC-CM) on primary human trabecular meshwork cells and primary retinal ganglion cells
Primary human trabecular meshwork (hTM) cells (ScienceCell) from passage 1 to 8 were cultured in 6-well plates till 90% confluence before exposure to MSC-CM, TGF-β2 (20µg/ml) or both for 3 or 72 hours. We investigated the hTM-myofibroblast phenotype transition induced by TGF-β2+/-MSC-CM, and α-SMA, col3 and col4 mRNA level expression after 72h. Next, hTM exposure to TGF-β2+/-MSC-CM for 3h, we looked for the Akt- and myosin (MLC) phosporylation using Western-Blot assays. Purified primary RGCs was obtained from rat retina using immunopanning methods. Cells were seeded in 96-well plates at 12 000cells/well and maintained for 6 days in neurobasal+glutamine (NBAg), MSC-CM or NBAg+B27 medium. Cell viability was evaluated by calcein green fluorescence using live-dead test kit and a cell counting software.
MSC-CM displayed favorable effects on viability, contractibility and phenotype conservation of hTM cells especially by impeding the deleterious effect of TGF-β2. MSC-CM clearly triggered the phosphorylation of the antiapoptotic Akt pathway resulting in an increased survival as proved in an in vitro BAK cytotoxic model. MSC-CM significantly inhibited the TGF-β2-dependent MLC phosphorylation on hTM and α-SMA, col3 and col4 mRNA increase. Furthermore, MSC-CM promoted cell viability and growth in a purified primary culture of rat RGC in vitro.
Through these in vitro approaches, we highlighted the beneficial effects of MSC-CM on primary cells directly involved in the physiopathology features of glaucoma. These results support the promising concept of MSCs-based therapy to treat glaucoma disease.
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