Using messenger RNA (mRNA) in situ hybridization, we investigated estrogen receptor-beta (ERbeta) mRNA levels in normal mammary, benign breast tumor (BBT), breast cancer (BC), and metastatic lymph node tissues to verify the role of ERbeta in BC development and progression. ERbeta expression was significantly decreased in BC and metastatic lymph node tissues compared with normal mammary and BBT tissues (p < 0.01). The intensity and extent of ERbeta mRNA signals were also significantly lower in BC and metastatic lymph node tissues than in the normal mammary and BBT tissues (p < 0.01). An inverse relationship was found between ERbeta mRNA level and both histologic grade (p = 0.091) and progesterone receptor expression (p = 0.052) with marginal significance, but no significant association was noted between ERbeta expression in cancer tissues and the other clinico-pathologic data. The 3-year distant relapse-free survival probability was found to be independent of ERbeta expression. Collectively, ERbeta mRNA decreases in the process of BC development, but seems to be associated with poor differentiation.