Developmental regulation of neurogenesis in the pluripotent human embryonal carcinoma cell line NTERA-2

Embryonal carcinoma (EC) cells provide a caricature of pluripotent embryoni c stem (ES) cells and may be used as surrogates for investigating the mecha nisms that regulate cell differentiation during embryonic development. NTER A-2 is a human EC cell line that differentiates in response to retinoic aci d yielding cells that include terminally differentiated neurons. The expres sion of genes known to be involved in the formation of the vertebrate nervo us system was examined during retinoic acid-induced NTERA-2 differentiation . Differentiation of these human EC cells into neurons could be divided int o three sequential phases. During phase 1, in the first week of differentia tion, hath1 mRNA showed a small transient increase that correlated with the rapid accumulation of nestin message, a marker of neuroprogenitors. Transc ripts of nestin were quickly downregulated during phase 2 as expression of neuroD1, characteristic of neuroprogenitors exiting the cell cycle, was ind uced. A neural cell surface antigen, detected by the monoclonal antibody A2 B5, was expressed by cells exiting the cell cycle, correlating with the exp ression of neuroD1 as the cells became post-mitotic. Markers of mature neur al cells (e.g. synaptophysin and neuron-specific enolase) were subsequently increased during phase 3 and were maintained. This regulated pattern of ge ne expression and commitment to the neural lineage indicates that different iation of NTERA-2 neurons in vitro follows a similar pathway to that observ ed by neural ectodermal precursors during vertebrate neurogenesis in vivo.