Galectin-3 expression and effects on cyst enlargement and tubulogenesis in kidney epithelial MDCK cells cultured in three-dimensional matrices in vitro.
Galectin-3 is a member of a closely related family of beta-galactoside-binding soluble proteins found in many vertebrate epithelial and myeloid cell types. The developmentally regulated presence of galectin-3 in tissues, for example kidney, and an affinity for many cell-surface and matrix glycoproteins indicate its importance in extracellular biological processes. Since a polarised expression and secretion of galectin-3 was observed in monolayer-cultured MDCK cells, an understanding of the secretion and distribution of this lectin in a three-dimensional in vitro model would help to uncover its role(s) in the interplay between cell-surface adhesion molecules and extracellular matrix components occurring during cell aggregation and polarisation in tissue formation. In this study, the cellular distribution and secretion of galectin-3 were examined in MDCK cells cultured within a gel matrix. MDCK cells were cultured within type I collagen or Matrigel to obtain multicellular cysts, and tubule formation was induced in collagen gels with hepatocyte growth factor. Immunofluorescent staining of these structures using antibodies against galectin-3 and other cell-surface domain markers was carried out either in situ or on cryosections and was visualised by confocal and conventional epifluorescence microscopy. Our results show that MDCK cells suspended in hydrated collagen gels or Matrigel exhibit differential and polarised galectin-3 expression on the baso-lateral surface domains of cells lining the cysts. The lectin is colocalised with laminin on the basal surface. In tubule-forming cysts, galectin-3 is excluded from the initial spikes and the progressing tips of the tubules although its basolateral expression on the cyst body remains. Galectin-3 added exogenously to cultures, as well as antibodies against laminin subunits and integrin beta 1 subunit, exerted an inhibitory effect on cyst enlargement of MDCK cells in 3-D Matrigel while galectin-3-specific antibodies could promote this process. The results suggest that galectin-3 exerts its effect on MDCK cells in a three-dimensional environment through modulation of both cell-cell and cell-substratum adhesions, and the interplay between these adhesions is important in the growth of multicellular aggregates and extensions occurring during normal kidney tubulogenesis.
Bao
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National Institute for Medical Research, Mill Hill, London, UK.
Hughes
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RC
D000818:Animals; D000906:Antibodies; D000943:Antigens, Differentiation / Q000096:biosynthesis* / Q000502:physiology*; D002460:Cell Line; D003094:Collagen; D046508:Culture Techniques / Q000379:methods; D004285:Dogs; D004338:Drug Combinations; D004847:Epithelial Cells; D004848:Epithelium / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D037502:Galectin 3; D005782:Gels; D015870:Gene Expression / Q000187:drug effects; D017228:Hepatocyte Growth Factor / Q000494:pharmacology; D007668:Kidney; D007700:Kinetics; D007797:Laminin / Q000032:analysis; D008562:Membrane Glycoproteins / Q000096:biosynthesis; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008858:Microscopy, Phase-Contrast; D011509:Proteoglycans; D013997:Time Factors
D000906:Antibodies; D000943:Antigens, Differentiation; D004338:Drug Combinations; D037502:Galectin 3; D005782:Gels; D007797:Laminin; D008562:Membrane Glycoproteins; D011509:Proteoglycans; C059485:matrigel; D017228:Hepatocyte Growth Factor; D003094:Collagen