The syncytiotrophoblast is the major component of the human placenta, involved in feto-maternal exchanges and secretion of pregnancy-specific hormones. Multinucleated syncytiotro- phoblast arises from fusion of mononuclear cytotrophoblast cells. In trisomy 21-affected placentas, we recently have shown that there is a defect in syncytiotrophoblast formation and a decrease in the production of pregnancy-specific hor- mones. Due to the role of oxygen free radicals in trophoblast cell differentiation, we investigated the role of the key anti- oxidant enzyme, copper/zinc superoxide dismutase, encoded by chromosome 21 in in vitro trophoblast differentiation. We first observed that overexpression of superoxide dismutase in normal cytotrophoblasts impaired syncytiotrophoblast for- mation. This was associated with a significant decrease in mRNA transcript levels and secretion of hCG and other hor- monal markers of syncytiotrophoblast. We confirmed abnor- mal cell fusion by overexpression of green fluorescence pro- tein-tagged superoxide dismutase in cytotrophoblasts. In addition, a significant decrease in syncytin transcript levels was observed in superoxide dismutase-transfected cells. We then examined superoxide dismutase expression and activity in isolated trophoblast cells from trisomy 21-affected placen- tas. Superoxide dismutase mRNA expression (P < 0.05), pro- tein levels (P < 0.01), and activity (P < 0.05) were significantly higher in trophoblast cells isolated from trisomy 21-affected placentas than in those from normal placentas. These results suggest that superoxide dismutase overexpression may di- rectly impair trophoblast cell differentiation and fusion, and superoxide dismutase overexpression in Down's syndrome may be responsible at least in part for the failure of syncy- tiotrophoblast formation observed in trisomy 21-affected placentas. (Endocrinology 142: 3638 -3648, 2001)