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Abstract. The present study describes a culture environment in which luminal epithelial cells isolated from immature rat uteri and cultured on a matrix-coated permeable surface, with separate apical and basal secretory compartments, proliferate to confluence. Subsequently the cells undergo a process of differentiation accompanied by progressive development of functional polarity. Ultrastructural and immunocytochemical evidence verifies the ability of these primary cultures to regain polar organization, separate membrane domains, and form functional tight junctions as demonstrated by the development of transepithelial resistance. The appearance of uvomorulin is restricted to the lateral cell surface. Coordinated indices of functional polarity that develop progressively in postconfluent cultures include the preferential uptake of [35S]methionine from the basal surface and a rise in uterine epithelial cell secretory activity characterized by a progressive preference for apical secretion. The time dependent development of polarity was characterized by differences in the protein profiles of the apical and basolateral secretory compartments. The maintenance of hormone responsiveness by the cultured cells was validated by the secretion of two proteins identified as secretory markers of estrogen response in the intact uterus. The technique of culturing the cells on a matrix-coated permeable surface with separate secretory compartments produces a uterine epithelial cell that morphologically and functionally resembles its in situ equivalent. The culture method and analytical approach used in this present study may be applied to primary cultures of a variety of natural epithelia, which have hitherto proven resistant to more conventional culture methodologies.

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