Human embryonic stem cells (hESCs) have the potential to differentiate into various cell types, and the three germ layers in vivo and in vitro. They are therefore useful in transplantation and tissue engineering. Here, we describe the expression patterns of selected steroid receptor mRNAs - estrogen receptor-alpha (ER-alpha), ER-beta, glucocorticoid receptor (GR), and progesterone receptor (PR) - in undifferentiated hESCs and embryoid bodies (EBs) cultured for 2, 4, and 6 d, as assessed by real-time PCR, in order to define the possible influence of steroid hormones on the differentiation of hESCs. These receptor mRNAs were expressed in undifferentiated hESCs and EBs. The expression of PR mRNA only decreased during the differentiation of EBs but not of hESCs. Immunohistochemical analysis gave strong staining of ER-alpha, ER-beta, and GR proteins in the nuclei of hESCs and EBs, whereas PR was not detected. We also examined the potential of these steroid hormones to direct the differentiation of hESCs in vitro. The expression of 11 cell-specific markers representing 3 germ layers and 5 tissue types was used to assess the differentiation of hESCs. We found that certain endodermal marker genes were either only expressed in the estrogen-treated group or their expression was stimulated in that group, suggesting that steroid hormones can control the differentiation of hESCs into various cell types.