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• W2036690815 endingPage "67" @default.
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• W2036690815 abstract "In search of guiding principles involved in the branching of epithelial tubes in the developing kidney, we analyzed branching of the ureteric bud (UB) in whole kidney culture as well as in isolated UB culture independent of mesenchyme but in the presence of mesenchymally derived soluble factors. Microinjection of the UB lumen (both in the isolated UB and in the whole kidney) with fluorescently labeled dextran sulfate demonstrated that branching occurred via smooth tubular epithelial outpouches with a lumen continuous with that of the original structure. Epithelial cells within these outpouches cells were wedge-shaped with actin, myosin-2 and ezrin localized to the luminal side, raising the possibility of a “purse-string” mechanism. Electron microscopy and decoration of heparan sulfates with biotinylated FGF2 revealed that the basolateral surface of the cells remained intact, without the type of cytoplasmic extensions (invadopodia) that are seen in three-dimensional MDCK, mIMCD, and UB cell culture models of branching tubulogenesis. Several growth factor receptors (i.e., FGFR1, FGFR2, c-Ret) and metalloproteases (i.e., MT1-MMP) were localized toward branching UB tips. A large survey of markers revealed the ER chaperone BiP to be highly expressed at UB tips, which, by electron microscopy, are enriched in rough endoplasmic reticulum and Golgi, supporting high activity in the synthesis of transmembrane and secretory proteins at UB tips. After early diffuse proliferation, proliferating and mitotic cells were mostly found within the branching ampullae, whereas apoptotic cells were mostly found in stalks. Gene array experiments, together with protein expression analysis by immunoblotting, revealed a differential spatiotemporal distribution of several proteins associated with epithelial maturation and polarization, including intercellular junctional proteins (e.g., ZO-1, claudin-3, E-cadherin) and the subapical cytoskeletal/microvillar protein ezrin. In addition, Ksp-cadherin was found at UB ampullary cells next to developing outpouches, suggesting a role in epithelial–mesenchymal interactions. These data from the isolated UB culture system support a model where UB branching occurs through outpouching possibly mediated by wedge-shaped cells created through an apical cytoskeletal purse-string mechanism. Additional potential mechanisms include (1) differential localization of growth factor receptors and metalloproteases at tips relative to stalks; (2) creation of a secretory epithelium, in part manifested by increased expression of the ER chaperone BiP, at tips relative to stalks; (3) after initial diffuse proliferation, coexistence of a balance of proliferation vs. apoptosis favoring tip growth with a very different balance in elongating stalks; and (4) differential maturation of the tight and adherens junctions as the structures develop. Because, without mesenchyme, both lateral and bifid branching occurs (including the ureter), the mesenchyme probably restricts lateral branching and provides guidance cues in vivo for directional branching and elongation as well as functioning to modulate tubular caliber and induce differentiation. Selective cadherin, claudin, and microvillar protein expression as the UB matures likely enables the formation of a tight, polarized differentiated epithelium. Although, in vivo, metanephric mesenchyme development occurs simultaneously with UB branching, these studies shed light on how (mesenchymally derived) soluble factors alone regulate spatial and temporal expression of morphogenetic molecules and processes (proliferation, apoptosis, etc.) postulated to be essential to the UB branching program as it forms an arborized structure with a continuous lumen." @default.
• W2036690815 created "2016-06-24" @default.
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• W2036690815 date "2004-11-01" @default.
• W2036690815 modified "2023-09-26" @default.
• W2036690815 title "Spatiotemporal regulation of morphogenetic molecules during in vitro branching of the isolated ureteric bud: toward a model of branching through budding in the developing kidney" @default.
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