Study of transforming growth factor alpha for the maintenance of human embryonic stem cells

Human embryonic stem cells (hESCs) have great potential for regenerative medicine as they have selfregenerative and pluripotent properties. Feeder cells or their conditioned medium are required for the maintenance of hESC in the undifferentiated state. Feeder cells have been postulated to produce growth factors and extracellular molecules for maintaining hESC in culture. The present study has aimed at identifying these molecules. The gene expression of supportive feeder cells, namely human foreskin fibroblast (hFF-1) and non-supportive human lung fibroblast (WI-38) was analyzed by microarray and 445 genes were found to be differentially expressed. Gene ontology analysis showed that 20.9% and 15.5% of the products of these genes belonged to the extracellular region and regulation of transcription activity, respectively. After validation of selected differentially expressed genes in both human and mouse feeder cells, transforming growth factor a (TGFa) was chosen for functional study. The results demonstrated that knockdown or protein neutralization of TGFa in hFF-1 led to increased expression of early differentiation markers and lower attachment rates of hESC. More importantly, TGFa maintained pluripotent gene expression levels, attachment rates and pluripotency by the in vitro differentiation of H9 under non-supportive conditions. TGFa treatment activated the p44/42MAPK pathway but not the PI3K/Akt pathway. In addition, TGFa treatment increased the expression of pluripotent markers, NANOG and SSEA-3 but had no effects on the proliferation of hESCs. This study of the functional role of TGFa provides insights for the development of clinical grade hESCs for therapeutic applications.