Differentiation of embryonic stem cells to a neural fate: a route to re-building the nervous system?

The many and varied proposed applications of cell replacement therapies in the treatment of human disease states, particularly those arising from cell loss or dysfunction, have been discussed widely in both the scientific and popular press. Although an attractive concept, cell therapies require the development of a readily available source of donor cells suitable for transplantation. Embryonic stem (ES) cells, with proven ability to differentiate to all cell populations of the embryo and adult in vitro, provide a potential source of therapeutic cells. The differentiation capability of mouse ES cells in vitro has been studied extensively over the last 20 years and the formation of neural precursors and neural cell lineages from mouse ES cells is well established. Cell populations highly enriched/homogenous in neural precursors have been achieved using a variety of chemical or biological inducing agents coupled with selective growth conditions. Preliminary reports suggest that similar neural enrichment is seen when these methodologies are applied to primate and human ES cells. ES cell-derived neural precursors have been analyzed in vitro and in vivo and found to be functionally normal and, after introduction into rodent models of human neurodegenerative diseases, capable of effecting measurable disease recovery. We review progress in the formation of neural precursors from mouse ES cells, particularly the recent reports of directed differentiation of ES in response to biological inductive factors, and assess the transfer of these approaches to human ES cells.

Hypoxia-inducible factor-1alpha mRNA contains an internal ribosome entry site that allows efficient translation during normoxia and hypoxia.

HIF-1alpha is the regulated subunit of the HIF-1 transcription factor, which induces transcription of a number of genes involved in the cellular response to hypoxia. The HIF-1alpha protein is rapidly degraded in cells supplied with adequate oxygen but is stabilized in hypoxic cells. Using polysome profile analysis, we found that translation of HIF-1alpha mRNA in NIH3T3 cells is spared the general reduction in translation rate that occurs during hypoxia. To assess whether the 5'UTR of the HIF-1alpha mRNA contains an internal ribosome entry site (IRES), we constructed a dicistronic reporter with the HIF-1alpha 5'UTR inserted between two reporter coding regions. We found that the HIF-1alpha 5'UTR promoted translation of the downstream reporter, indicating the presence of an IRES. The IRES had activity comparable to that of the well-characterized c-myc IRES. IRES activity was not affected by hypoxic conditions that caused a reduction in cap-dependent translation, and IRES activity was less affected by serum-starvation than was cap-dependent translation. These data indicate that the presence of an IRES in the HIF-1alpha 5'UTR allows translation to be maintained under conditions that are inhibitory to cap-dependent translation.