Characterization of pure human first-trimester cytotrophoblast cells in long-term culture: Growth pattern, markers, and hormone production

Pure long-term cytotrophoblast cultures were established from human first-trimester placentas by growing chorionic villus explants without enzymatic digestion. Cytoplasmic human chorionic gonadotropin was detectable in all (100%) cells in culture when labeled with a polyclonal anti-human chorionic gonadotropin antibody and in 71% to 83% of cells labeled with a monoclonal anti-α-human chorionic gonadotropin antibody. Most of the cells expressed cytokeratin and surface Trop-1 and Trop-2 antigens (89% to 95%), but none expressed cytoplasmic vimentin or surface 63D3 antigens. Study of the ultrastructure of the cells demonstrated epithelial morphologic features. The average doubling time of the-trophoblast was 48 to 96 hours. Some of the lines have been continuously propagated for 8 months. They produced variable amounts of human chorionic gonadotropin (50 to 710 mlU/ml per 105 cells per 24 hours). The basal level of progesterone secreted by trophoblast (444.4 ± 32.4 pg/ml per 105 cells per 24 hours) doubled in the presence of pure human chorionic gonadotropin (100 ng/ml). They produced small amounts of 17β-estradiol (<20 pg/ml per 105 cells per 24 hours); human chorionic gonadotropin had no effect on the estradiol production. Trophoblast-derived human chorionic gonadotropin acted as a growth factor because trophoblast proliferation (measured by uptake of thymidine labeled with tritium) was reduced by 60% in the presence of an anti-human chorionic gonadotropin antibody. Availability of pure, functionally competent human cytotrophoblast in long-term cultures is relevant for further studies in reproduction biology. Pure long-term cytotrophoblast cultures were established from human first-trimester placentas by growing chorionic villus explants without enzymatic digestion. Cytoplasmic human chorionic gonadotropin was detectable in all (100%) cells in culture when labeled with a polyclonal anti-human chorionic gonadotropin antibody and in 71% to 83% of cells labeled with a monoclonal anti-α-human chorionic gonadotropin antibody. Most of the cells expressed cytokeratin and surface Trop-1 and Trop-2 antigens (89% to 95%), but none expressed cytoplasmic vimentin or surface 63D3 antigens. Study of the ultrastructure of the cells demonstrated epithelial morphologic features. The average doubling time of the-trophoblast was 48 to 96 hours. Some of the lines have been continuously propagated for 8 months. They produced variable amounts of human chorionic gonadotropin (50 to 710 mlU/ml per 105 cells per 24 hours). The basal level of progesterone secreted by trophoblast (444.4 ± 32.4 pg/ml per 105 cells per 24 hours) doubled in the presence of pure human chorionic gonadotropin (100 ng/ml). They produced small amounts of 17β-estradiol (<20 pg/ml per 105 cells per 24 hours); human chorionic gonadotropin had no effect on the estradiol production. Trophoblast-derived human chorionic gonadotropin acted as a growth factor because trophoblast proliferation (measured by uptake of thymidine labeled with tritium) was reduced by 60% in the presence of an anti-human chorionic gonadotropin antibody. Availability of pure, functionally competent human cytotrophoblast in long-term cultures is relevant for further studies in reproduction biology.