Human erythropoietin (Epo) is a 30 kDa glycoprotein hormone produced in the kidney (and to a lesser extent the liver) in response to hypoxic stimuli, which promotes the proliferation and differentiation of erythroid precursors. Erythroid development and synthesis of haemoglobin creates a demand for iron that is met by efficient recycling of body iron and also enhanced intestinal absorption. The mechanisms regulating intestinal iron absorption in response to increased erythropoietic requirements are unknown but one potential pathway could involve a direct action of Epo on intestinal enterocytes. Therefore, in the current study we have investigated the effects of Epo on the absorption of non-haem iron using Caco-2 cells as a model of human intestinal epithelial cells.
Caco-2 cells were grown for 21 days on Transwell inserts at which time they exhibited a fully differentiated small intestinal enterocyte-like phenotype. Human recombinant Epo (50 mU ml-1) was added to the basolateral medium for the final 24 h of the culture period and cells were subsequently used to measure either pH-dependent 55Fe2+ uptake or the expression of the intestinal iron transporter DMT1 by Western blotting.
Iron uptake across the apical membrane of Caco-2 cells was significantly increased following 24 h incubation with Epo (control, 366.3 ± 25.3 pmol cm-2 h-1, n = 3; Epo, 462.4 ± 21.5 pmol cm-2 h-1, n = 3; mean ± S.E.M.; P = 0.04, Student’s unpaired t test). DMT1 protein density in the apical membrane of Caco-2 cells was 42.4 a.u. in control cells and 85.6 a.u. following Epo treatment. These data suggest that Epo could act as an erythroid regulator of intestinal iron uptake by controlling the expression of the intestinal iron transporter DMT1. The cellular mechanisms involved in this response remain to be elucidated, but may involve an intracellular signalling cascade downstream from the erythropoietin receptor, which was detected in Caco-2 cells as a 69 kDa protein by Western blotting. These possibilities are currently under investigation.