5-Azacytidine Makes Human Preadipocytes Able to Differentiate into Mesoderm-Derived Cell Lineages

In the present study we have evaluated whether (i) 5-azacytidine (AZA), a well-known demethylating agent, could be able to modify the phenotype of human preadipocytes and (ii) the modified cells could possess multilineage differentiation potential. Human preadipocytes at the 3rd passage were treated for 48 or 96 h with 10 mu M AZA and then expanded up to passage 5. Stem cell markers, such as OCT-4, Nanog, and Sox2, were upregulated after 96 h of treatment with the demethylating treatment. Further, decreases in the expression of genes, such as adipose differentiation-related protein, characterizing the preadipocytes were noted. Our data showed that AZA-treated preadipocytes differentiated into cell lineages derived from mesoderm. Indeed, after incubation with inductive media for 3 weeks, osteblast-, chondrocyte-, and myoblast-like cells were detected in the cultures. Interestingly, both upregulation of stem cell markers and differentiation potential were maintained by the treated cultures expanded until the 5th passage. Taken together, our results suggest that AZA, without the use of transduction methods, convert preadipocytes to a less differentiated state that can be induced, under suitable stimuli, to the formation of mesoderm-derived cell lineages.