Female hormone-dependent cancers and other diseases pose a serious health threat for women, and low-risk medicines against such cancers have not yet been discovered. The present study examines the effects of the traditional Chinese herbal mixture, Tokishakuyakusan (TS) and 17β-estradiol on the uterus of parous ovariectomized rats. Uterine atrophy that causes a reduction in uterine tissue and the uterine cavity area, was induced by ovariectomy, and slightly recovered by the daily oral administration of TS for two weeks (1000 mg/kg body weight). TS restored the decreased plasma estradiol concentration due to ovariectomy. However the yeast two-hybrid assay showed that TS did not bind estrogen receptors α and β and immunohistochemical staining revealed that 17β-estradiol stimulated the protein expression of estrogen receptor α, progesterone receptor, c-fos and c-jun in the uterus, whereas TS did not. These results suggest that TS might be useful for treating menopausal syndromes among women, as well as for patients when hormone replacement therapy (HRT) with estrogen is contraindicated.