Abstract
Establishment of apical-basal polarity is crucial for epithelial sheets that form a compartment in the body, which function to maintain the environment in the compartment. Effects of impaired polarization are easily observed in three-dimensional (3-D) culture systems rather than in two-dimensional (2-D) culture systems. Although the mechanisms for establishing the polarity are not completely understood, signals from the extracellular matrix (ECM) are considered to be essential for determining the basal side and eventually generating polarity in the epithelial cells. To elucidate the common features and differences in polarity establishment among various epithelial cells, we analyzed the formation of epithelial apical-basal polarity using three cell lines of different origin: MDCK II cells (dog renal tubules), EpH4 cells (mouse mammary gland), and R2/7 cells (human colon) expressing wild-type α-catenin (R2/7 α-Cate cells). These cells showed clear apical-basal polarity in 2-D cultures. In 3-D cultures, however, each cell line displayed different responses to the same ECM. In MDCK II cells, spheroids with a single lumen formed in both Matrigel and collagen gel. In R2/7 α-Cate cells, spheroids showed similar apical-basal polarity as that seen in MDCK II cells, but had multiple lumens. In EpH4 cells, the spheroids displayed an apical-basal polarity that was opposite to that seen in the other two cell types in both ECM gels, at least during the culture period. On the other hand, the three cell lines showed the same apical-basal polarity both in 2-D cultures and in 3-D cultures using the hanging drop method. The three lines also had similar cellular responses to ECM secreted by the cells themselves. Therefore, appropriate culture conditions should be carefully determined in advance when using various epithelial cells to analyze cell polarity or 3-D morphogenesis.
Highlights
Epithelial sheets in multicellular organisms form physiological barriers separating the internal environment from the external environment [1]
Apical-basal polarities of epithelial cells in 2-D culture To determine the common features and variations in the mechanism of apical-basal polarity establishment seen in many epithelial cells, focusing especially on the role of extracellular matrix (ECM), we chose three types of epithelial cell lines: MDCK II cells, EpH4 cells, and
Scrib [25], a member of the scribble complex consisting of Scrib, lethal (2) giant larvae homologue (LGL) and discs-large homologue (DLG) localized clearly to basolateral membranes but not to apical membranes in all three cell lines (Fig. 2, red in the merged panels). atypical protein kinase C (aPKC), a member of the Par polarity complex consisting of Par3 and Par6, is known to play an essential role in cell polarity [25], and one aPKC member, PKC zeta, was found to localize preferentially to apical membranes in all cases (Fig. 2, green in the merged panels)
Summary
Epithelial sheets in multicellular organisms form physiological barriers separating the internal environment from the external environment [1]. The epithelial plasma membrane is divided into two regions, an apical membrane facing the lumen or external environment and a basolateral membrane contacting adjacent cells and the underlying extracellular matrix (ECM). These two membrane regions have distinct functions and molecular constituents. At the border of these two regions, in the vicinity of the most apical position along the basolateral membrane, are apical junctions composed of tight and adherens junctions (Fig. 1A). Cell structures such as cilia or microvilli show biased localization. ZO-1 is a scaffoliding protein localized to tight junctions in polarized epithelial cells [1]
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