Morphological and biochemical changes occurring in rabbit endometrial epithelial cells when placed in culture were investigated. Cells were examined by scanning- and transmission electron microscopy and freeze-fracture. Morphologically, cultured cells are shorter and broader than the columnar epithelial cells in vivo, but retain their polarity as indicated by the presence of apical microvilli and a well-developed junctional belt. To study changes in biochemical function, proteins synthesized by cells in primary culture were analyzed by two-dimensional gel electrophoresis. Proteins were labeled during a 24-h incubation with35S-methionine and gels examined by fluorography. The pattern of proteins changed after cells had been in culture for 48 h. On day 3 new proteins were synthesized and several protein species labeled during days 1 or 2 of culture, including uteroglobin, no longer appeared. On days 3–8 of culture the protein patterns were similar. Addition of progesterone, estradiol, prolactin, or combinations of these hormones to the culture medium for 24–144 h failed to elicit consistent changes in the pattern of labeled proteins established after 3 days of culture. Minor differences in protein patterns among unrelated cultures appear to have been derived from the original cells of the culture. These results indicate that after 48 h in primary culture, cells grown in vitro resemble endometrial epithelial cells morphologically, but no longer reflect functionally the character of epithelial cells in the ute
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