Primary cultures of human uterine smooth muscle cells have been widely used as a model system to evaluate agents that may play a role in the regulation of both normal and abnormal proliferative responses. We have used this in vitro system to determine if human uterine smooth muscle cells are responsive to treatment with a potent natural derivative of vitamin A, all-trans retinoic acid (ATRA). These studies were also designed to determine if there is a difference in retinoid responsiveness between normal smooth muscle and adjacent leiomyoma (a benign tumor of uterine smooth muscle). When cells were cultured in the presence of ATRA, a dose-dependent inhibition in proliferation was observed. This inhibition in proliferation was accompanied by an alteration in smooth muscle cell morphology. Both the inhibition in proliferation and the altered morphology were reversible when ATRA treatment was discontinued. Responsiveness to retinoids is determined, in part, by the expression of ligand-specific receptors belonging to the steroid/thyroid superfamily (RARs and RXRs); we have therefore identified the pattern of retinoid receptor transcript expression in human uterine smooth muscle cells. The data indicate that human uterine smooth muscle cells express retinoic acid receptors RAR alpha, beta, and gamma, and retinoid X receptors RXR alpha and beta. No difference in retinoid responsiveness or in the pattern of retinoid receptor expression was observed between normal smooth muscle and adjacent leiomyoma. This is the first observation of an antiproliferative effect of ATRA in uterine smooth muscle cells and the first report of retinoid receptor expression patterns in this cell type. Since retinoids are common pharmacologic tools in the treatment of a wide variety of hyperproliferative disorders, these observations may have both therapeutic and toxicologic implications.