Establishment of endometrial glandular epithelial cell subculture in a serum-free, hormonally defined medium, on a basement membrane matrix.
Epithelial glands were isolated from guinea-pig endometrium. In order to reduce the requirement for a serum supplement and the contamination by non epithelial cells in primary culture, various coatings of the culture dishes were tested using serum-free Ham's F12 containing defined chemicals including 17 beta-estradiol. While epithelial glands seeded on culture dishes coated with Matrigel, a basement membrane matrix-failed to spread, they formed on poly-D-lysine plus serum-coated dishes, a subconfluent monolayer (5-7 days) enriched in cytokeratin-immunostained cells (78%). Cells from subconfluent primary cultures, obtained on poly-D-lysine plus serum-coated dishes in serum-free hormonally defined medium, were passaged on Matrigel-coated dishes in serum-free hormonally defined medium. These subcultures contained, at confluence (4-5 days), a high percentage (greater than 95%) of cytokeratin-immunostained cells. These monolayers consisted of well-differentiated cells which exhibited ultrastructural features characteristic of endometrial epithelial cells. Moreover, these confluent cells contained 50% immunostained nuclei for progesterone receptors. Progesterone receptor amounts decreased in confluent subcultures treated with progesterone and became undetectable after long-term treatment, suggesting responsiveness of these cells to progesterone. This culture system provides a well-defined model for the study of protein synthesis and secretion by endometrial glandular epithelial cells under hormonal control.
Mahfoudi
A
A
INSERM U 198, Besançon, France.
Nicollier
M
M
Propper
A Y
AY
Coumes-Marquet
S
S
Adessi
G L
GL
D000818:Animals; D002478:Cells, Cultured / Q000187:drug effects / Q000648:ultrastructure; D003094:Collagen / Q000494:pharmacology; D016895:Culture Media, Serum-Free; D046508:Culture Techniques / Q000379:methods*; D004338:Drug Combinations; D004717:Endometrium / Q000166:cytology* / Q000648:ultrastructure; D004958:Estradiol / Q000494:pharmacology; D005260:Female; D005455:Fluorescent Antibody Technique; D006168:Guinea Pigs; D007381:Intermediate Filament Proteins / Q000032:analysis; D007382:Intermediate Filaments / Q000737:chemistry; D007797:Laminin / Q000494:pharmacology; D008854:Microscopy, Electron; D011374:Progesterone / Q000494:pharmacology; D011509:Proteoglycans / Q000494:pharmacology; D011980:Receptors, Progesterone / Q000032:analysis; D014746:Vimentin / Q000032:analysis
D016895:Culture Media, Serum-Free; D004338:Drug Combinations; D007381:Intermediate Filament Proteins; D007797:Laminin; D011509:Proteoglycans; D011980:Receptors, Progesterone; D014746:Vimentin; C059485:matrigel; D011374:Progesterone; D004958:Estradiol; D003094:Collagen