Production of human CNS neurons from embryonal carcinoma cells using a cell aggregation method

When treated with retinoic acid (RA), a human embryonal carcinoma (EC) cell line, NTera2 cl.D/1 (NT2), differentiates into several morphologically dis tinct cell types, which include terminally differentiated postmitotic centr al nervous system (CNS) neurons. Accumulating evidence has demonstrated the significant potential of NT2 cells in studies related to cancer therapy an d neurodegenerative diseases However, preparation of enriched NT2 neurons o ften requires a lengthy period (co. five,weeks) and depends largely on tedi ous techniques similar to those used for primary neuronal cultures. Here, w e report a rapid protocol for the preparation of these human CNS neurons. U sing the method of cell aggregation, enriched NT2 neurons can he obtained i n approximately two weeks. We also demonstrated that cell aggregation reduc ed the time normally required for the induction of neuronal differentiation as revealed by the early expression of neuronal markers. The period of RA treatment could also be reduced if NT2 cells were maintained cls aggregates for a sufficient period of time. Taken together our findings demonstrated that cell aggregation promoted RA-induced neuronal differentiation of NT2 c ells and provided a rapid protocol for the efficient production of NT2 neur ons. The ability to produce large quantities of human CNS neurons should fa cilitate future rise of these neurons for basic research and applications i n cell therapy.