A fluorescent metal-sensor study provides evidence for iron transport by transcytosis in the intestinal epithelial cells.
JOURNAL OF CLINICAL BIOCHEMISTRY AND NUTRITION(2017)
摘要
Iron transport across the intestinal epithelium is facilitated by the divalent metal transporter 1 (DMT1) on the brush border membrane (BBM). The fluorescent metal sensor calcein, which is hydrophilic, membrane-impermeable and quenched by chelation with iron, was used to test our hypothesis that intestinal iron absorption is through the endocytic processes and is involved in a pathway where BBM-derived vesicles fuse with basolateral membrane (BLM)-derived vesicles. To monitor the flux of iron via transcytosis, Caco-2 cells were employed as a polarized cell layer in Transwell chambers. When calcein was added to the basal chamber along with apo-transferrin (apo-Tf), calcein rapidly underwent endocytosis and co-localized with apo-Tf. Calcein was quenched by adding an iron-ascorbate complex and then restored by adding 2,2'-dipyridyl into the apical chamber. These results were confirmed by live-cell imaging. When hemin from the apical surface and calcein from the basal chamber were added to the Caco-2 cells, internalization of DMT1 and quenching of calcein were not observed until 2 h later. These results indicated that absorbed hemin required processing before hemin-derived iron was available to BLM-derived vesicles. These studies suggest that iron is transported in Caco-2 cells by transcytosis with apical-derived vesicles that are fused to BLM-derived vesicles.
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关键词
iron absorption,calcein,iron transport,Caco-2,hemin
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