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In vitro Differentiation of Adult Adipose Mesenchymal Stem Cells into Retinal Progenitor Cells


Introduction: It has been previously shown that adult mes- enchymal stem cells (MSCs) differentiate into neural progen- itor cells (NPCs) and that the differentiation process was completed in 24–48 h. In this previous study, MSCs from a bone marrow or fat source were co-incubated with homolo- gous autoaggressive cells (ECs) against nerve tissue, and these NPCs were successfully used in human regenerative therapeutic approaches.

The present study was conducted to investigate whether a similar differentiation method could be used to obtain autologous retinal progenitor cells (RPCs). Methods: Human Th1 cells against retinal tissue were obtained by challenging human blood mononuclear cells with an eye lysate of bovine origin; negative selection was performed using a specific immunomagnetic bead cocktail. Fat MSCs were obtained from a human donor through me- chanical and enzymatic dissociation of a surgical sample. The ECs and MSCs were co-cultured in a serum-free medium without the addition of cytokines for 0, 24, 48 and 72 h. The plastic adherent cells were morphologically examined using inverted-phase microscopy and characterized by immuno- fluorescent staining using antibodies against Pax 6, TUBB3,

GFAP, Bestrophin 2, RPE 65, OPN1 SW, and rhodopsin anti- gens. Results: The early signs of MSC differentiation into RPCs were observed at 24 h of co-culture, and the early dif- ferentiated retinal linage cells appeared at 72 h (neurons, rods, Müller cells, retinal ganglion cells and retinal pigment- ed epithelial cells). These changes increased during further culture. Conclusion: The results reported here support the development of a method to obtain a large number of au- tologous adult RPCs, which could be used to treat different retinopathies.

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Figure 1: Phase contrast morphology of the cultured cells. a Non-stimulated MSC. b After 24 h of co-culture it is possible to observe cells with the shape of RPCs. c, d After 72 h of co-culture cells with the shape of retinal ganglion cells (RGC), Müller cells (MC), retinal pigmented epithelium cells (RPE) and rods can be observed.
Figure 2: Confocal microscopy analysis of the cultured cells. a GFAP-positive cells. b Bestrophin 2-positive cells. c Merge image of a and b. d Tubulin beta III-positive cells. e OPN1 SW-positive cells.

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