Sex hormones and growth factors have been implicated in the pathogenesis of uterine leiomyomas. The uterus is also an abundant source of the glucocorticoid receptor but its role and function have been largely ignored. Human samples of uterine leiomyomas and matched myometrium retain expression of the glucocorticoid receptor (GR) suggesting a potential role for GR in leiomyoma function. However, hormone responsive gene expression varies between normal myometrium and leiomyoma cells. We now employ genome-wide microarray studies comparing glucocorticoid and estrogen-treated human uterine leiomyoma cells to those treated with both steroids to identify the potential role of glucocorticoids in uterine leiomyoma cells. Treatment with the synthetic glucocorticoid dexamethasone (Dex) regulated 3,128 probes. Estrogen (E(2)) treatment identified 2,094 probes, and in the presence of both hormones, 4,626 probes were regulated. Of the 552 probes identified, the majority of genes co-regulated by Dex, E(2), and Dex + E(2) exhibited co-downregulation. Interestingly, a small group of 17 genes displayed antagonistic regulation by Dex and E(2), where all genes in this group, Dex reversed the E(2) effect with. Ingenuity Pathway Analysis of the data identified cell growth, development, and differentiation as significant glucocorticoid regulated pathways. Flow cytometry confirmed that glucocorticoids regulated cell proliferation and significantly reduced the percentage of S-phase cells either in the presence or absence of estrogen in leiomyomas but not smooth muscle cells. Translation of our results suggest that glucocorticoids may play a significant role in regulating uterine leiomyoma gene expression and cell growth, and thus may have implications for therapeutic development of uterine leiomyoma treatment.