Abstract
Standardization of methods for obtaining iPS cells from the human somatic cells and then their successful differentiation are important in the context of their possible application in personalized cell therapy and the development of toxicological and pharmacological tests. In the present study, the influence of the small molecules representing epigenetic modulators (histone deacetylase inhibitor Trichostatin A and DNA methyltransferase inhibitor RG-108) on the process of reverting neural progenitors from HUCB-NSC (Human Umbilical Cord Blood Neural Stem Cell) line to the pluripotent state was tested. The experiments were conducted in low oxygen tension, in three different experimental layouts: (1) in the presence of reprogramming/recombinant polyarginine-tailed proteins; (2) with recombinant proteins and small molecules; (3) only in the presence of small molecules. We wanted to find out, whether it will be possible to induce pluripotent state of neural stem cells only by epigenetic modulators. Our results revealed that the inhibitors of DNA methylation and histone deacetylation used along with 5 percent oxygen tension can only transiently induce or elevate some pluripotency genes in neural progenitors with different pattern, but were not sufficient for stable reprogramming. The iPS cells from neural progenitor cells of HUCBNSC were obtained only when TSA, RG-108 and reprogramming proteins have been applied simultaneously. These cells were tested for the expression of the selected pluripotency genes and in functional assays to prove their pluripotency stage. The obtained data show that the small molecules in conjunction with reprogramming factors are the potent tools in cell reprogramming.