The presence of growth substance(s) for smooth muscle cells and fibroblasts was documented in extracts of human leiomyoma tissues (Koutsilieris et al, Am. J. Obst. Gynecol. 163: 1665-1670, 1990). In this study leiomyoma tissues (700 grs) obtained at surgery were extracted and purified using CM-sepharose, heparin-sepharose and reverse-phase high performance liquid chromatography (r-HPLC). The mouse NIH/3T3 cell fibroblasts (ATCC-CRL 1658) and the rat smooth muscle cells A10 (ATCC-CRL 1476) were used as indicator bioassay systems assessing the proliferative effect (tritiated-thymidine incorporation and cell number) of leiomyoma extracts. The profile of peptides purified by cm-sepharose, heparin-sepharose and by r-HPLC was analyzed by standard SDS gel electrophoresis. An overall 50,000 fold purification was achieved of a preferentially acting material on rat aorta smooth muscle cells (A10 cells). This material possessed mitogenic activity equivalent to epidermal growth factor (EGF) on A10 smooth muscle cells, in vitro. We believe that the final chemical definition of such leiomyoma-derived growth substance with preferential action on smooth muscle cells may elucidate the paracrine and/or autocrine interactions among myometrial cells and thus could provide new clues for the pathogenesis of uterine leiomyomas.