The expression of specific proteins in cultured renal collecting duct cells

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Abstract
Summary It is still uncertain whether cell cultures attain the functional maturity of corresponding in vivo cells. The degree of differentiation of cultured collecting-duct (CD) epithelium cells was therefore examined using immunohistochemical procedures. Three monoclonal antibodies (mabs CD 1, CD 2, and CD 3) were raised against proteins (PCD) isolated from the renal papilla. At Western-blot analysis, each of these antibodies reacted with a specific protein that was distinguishable according to its molecular weight [PCD1, 190 kilodaltons (kDa); PCD2, 210 kDa; PCD3, 50 kDa]. Using immunofluorescence, these proteins were found to be localized exclusively in the renal CD system. Other renal structures, such as the proximal or distal tubular portions, the glomeruli and the interstitial network, were not reactive. The mabs, CD 2 and CD 3, labeled both the cortical and medullary CD in a uniform way, whereas mab CD 1 produced heterogeneous immunolabeling along the length of the cortical, medullary, and papillary CD. As revealed by immunohistochemistry, the mabs revealed differences with respect to the expression of the specific renal proteins in cultured CD cells. In polar-differentiated epithelium cultured for 5 days on a specific renal support, mab CD 1 was unreactive, whereas mabs CD 2 and CD 3 were positive. This demonstrated the biochemical immaturity of this cultured epithelium with respect to CD 1 reactivity. In morphologically dedifferentiated CD monolayer cells grown on the bottom of a culture dish, only a weak reaction for mab CD 3 was observed. The loss of epithelial polarization in CD monolayer cells obviously coincides with the absence of the renal proteins PCD1 and PCD2. Thus, our mabs proved to be valuable tools for the investigation of the differentiation and dedifferentiation of renal CD cells in culture.