Reprogramming of MSCs to express neural markers using exosome treatments

Neural stem cells (NSCs), capable of self-renew and differentiate into neural cells, hold promise for use in studies and treatments for neurological diseases. However, current approaches to obtain NSCs from a live brain are risky and invasive, since NSCs reside in the subventricular zone and the in the hippocampus dentate gyrus. Alternatively, mesenchymal stem cells (MSCs) could be a more available cell source due to their abundance in tissues and easier to access. However, MSCs are committed to producing mesenchymal tissue and are not capable of spontaneously differentiating into neural cells. Thus, the process of reprogramming of MSCs into neural cells to use in clinical and scientific settings has significantly impacted the advancement of regenerative medicine.

Previously, University of Central Florida researchers reported trans-differentiation of MSCs to neural cells through the induced pluripotent stem (iPS) cells state, which was produced by overexpression of the embryonic stem cell gene NANOG. In the current study, they demonstrate that treatment with exosomes derived from NSCs makes MSCs capable of expressing neural cell markers bypassing the generation of iPS cells. An epigenetic modifier, decitabine (5-aza-2′-deoxycytidine), enhanced the process. This novel Xeno and transgene-free trans-differentiation technology eliminates the issues associated with iPS cells, such as tumorigenesis. Thus, it may accelerate the development of neurodegenerative therapies and in vitro neurological disorder models for personalized medicine.

Generation of cells expressing neural markers under Xeno-free conditions

Panel A shows the schematic procedure to produce iNSCs from MSCs. Method 1; treatment with NSCs derived exosomes alone. Method 2; treatment with NSCs derived exosomes in the presence of 10μM decitabine. Method 3; treatment with exosomes derived from both NSCs and iPS cells. Method 4; treatment with exosomes derived from both NSCs and iPS cells in the presence of 10μM decitabine. Panel B shows phase-contrast microscopic images for 3 different stages, production (15 DIV), Isolation (21 DIV), and differentiation in various methods 1–4. XIII. No treatment: without any treatment. XIV. Treatment alone: decitabine treated MSCs. XV. Treatment + iPS exosome: MSCs treated with iPS cells derived exosome with decitabine treatment are shown as controls. Panel C shows the number of primary neurospheres formed at 21 DIV. The decitabine treated cells developed significantly higher numbers of spheroids as compared to those without the treatment. Without NSCs derived exosome treatment, any spheroid formation was not observed. Panel D shows the differences in the nucleic size of the cells at 21 DIV. The iNSC produced in all four methods showed significantly reduced nuclei size as compared to the naive MSCs. Panel E shows immunocytochemistry of iNSCs and MSCs (control) after differentiation for NSC marker, Musashi (MSI1, green), and MSC marker (CD73, Red). The nuclei were counterstained with DAPI (Blue). The scale bar is 50μm. Panel F shows semi-quantitative fluorescence intensity measurement by Image J. *P<0.05, **P<0.01, ***P<0.005, ****P<0.0001.