| Summary: | Human adipose-derived mesenchymal stem cells (hASCs) may be used in some nervous system pathologies, although obtaining an adequate degree of neuronal differentiation is an important barrier to their applicability. This requires a deep understanding of the expression and epigenetic changes of the most important genes involved in their differentiation. We used hASCs from human lipoaspirates to induce neuronal-like cells through three protocols (Neu1, 2, and 3), determined the degree of neuronal differentiation using specific biomarkers in culture cells and neurospheres, and analyzed epigenetic changes of genes involved in this differentiation. Furthermore, we selected the <i>Hoxa-5</i> gene to determine its potential to improve neuronal differentiation. Our results showed that an excellent hASC neuronal differentiation process using Neu1 which efficiently modulated NES, CHAT, SNAP25, or SCN9A neuronal marker expression. In addition, epigenetic studies showed relevant changes in <i>Hoxa-5</i>, <i>GRM4</i>, <i>FGFR1</i>, <i>RTEL1</i>, <i>METRN</i>, and <i>PAX9</i> genes. Functional studies of the <i>Hoxa-5</i> gene using CRISPR/dCas9 and lentiviral systems showed that its overexpression induced hASCs neuronal differentiation that was accelerated with the exposure to Neu1. These results suggest that <i>Hoxa-5</i> is an essential gene in hASCs neuronal differentiation and therefore, a potential candidate for the development of cell therapy strategies in neurological disorders.
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