Trophoblast cells do not normally express major histocompatibility complex (MHC) class II antigens during placental development in either mice or rats. We have previously observed that in vive treatment of pregnant mice with interferon-gamma (IFN gamma) induces immunohistochemically detectable class II cell surface expression in many maternal cell types, but not on placental cells or other cells of extra-embryonic origin. Both IFN gamma- and 5-azacytidine-induced placental class II expression have been reported in mice by other scientists, however, which made it important to further clarify this issue. The present study was performed to analyze whether treatment of pregnant mice with recombinant IFN gamma or the drug 5-azacytidine in vive can induce detectable MHC class II Ab mRNA expression. A strain of transgenic mice carrying a cytomegalovirus-regulated MHC class II Abs transgene, which was strongly expressed in the placenta, was used as a positive control in all in situ hybridizations and ribonuclease protection analyses. All mice were analyzed on gestation Days 12.5 and 17.5. Treatment of pregnant mice with IFN gamma did not induce detectable class II expression in the placental cells, whereas the maternal decidua showed expression both at the mRNA and protein level. Similarly, treatment with 5-azacytidine did not induce class II expression in the placenta, while a slight increase in mRNA expression was detected in the maternal decidual and uterine tissues. These results strengthen the opinion that MMC class II mRNA cannot normally be induced in murine placental cells after IFN gamma or 5-azacytidine treatments.