Appropriately regulated intestinal iron absorption is crucial for the maintenance of body iron homeostasis as there is no regulated excretory mechanism for iron in humans. Despite the importance of this process, the way in which the body regulates iron absorption is poorly understood. The studies described in this thesis investigate the function and regulation of recently described molecules involved in the absorption process. This was achieved by utilizing various animal models in which iron absorption was altered.
Chapter one contains a review of the literature on body iron homeostasis with particular emphasis on intestinal iron absorption. It begins with a description of general cellular iron metabolism before proceeding with an in depth analysis of the current literature concerning the molecules involved in iron absorption. This is followed by description of liver iron metabolism and the iron overload disorder
haemochromatosis. Finally there is a brief summary of the research presented in this thesis.
Chapter two describes the general materials and methods used in the studies described in this thesis. Protocols used only in specific studies are described in the relevant chapters.
Chapter three describes the expression and regulation of the recently described membrane-bound ferroxidase hephaestin which is thought to play a role in the basolateral transfer of iron from the intestinal enterocyte into the body. The rat hephaestion gene was cloned and found to encode a protein 96% identical to mouse hephaestin. Expression analysis revealed that hephaestin is present at high levels throughout the small intestine and colon, and has a supranuclear localisation in these tissues. Variations in iron status had a small but non-significant inverse correlation with hephaestin expression in the duodenum. The high sequence conservation between rat and mouse
hephaestin is consistent with this protein playing a central role in intestinal iron absorption although its precise function remains to be determined.
Chapter four examines iron absorption and gene expression in rats following a large oral dose of iron. Such treatment has been previously shown to decrease the absorption of a subsequent smaller dose of iron in a phenomenon known as mucosal block. Intestinal iron absorption decreased following such treatment and this correlated with a rapid reduction in the expression of the brush border iron transporters DMT1 and Dcytb. No such change was seen in the expression of hephaestin or the basolateral iron transport molecule Ireg1. These data indicate that brush border, but not basolateral, iron transport components are regulated by local enterocyte iron levels and support the hypothesis that systemic stimuli exert their primary effect on basolateral transport.
Chapter five investigates
changes in gene expression and iron absorption in rats switched from a control to an iron deficient diet. Such changes lead to an increase in iron absorption within six days and were accompanied by an increase in the duodenal expression of Dcytb, DMT1 and Ireg1. These changes precisely correlated with decreases in hepatic hepcidin expression and transferrin saturation. These results demonstrate a close relationship between the expression of hepcidin, duodenal iron transporters and iron absorption. Both hepcidin expression and iron absorption can be regulated before iron stores and erythropoiesis are affected, and transferrin saturation may signal such changes.
Chapter six examines the cause of the extremely high iron absorption that occurs during the neonatal period. Analysis of sections of the gastrointestinal tract in neonatal animals showed an increase in both absorption and the expression of Dcytb, DMT1 and Ireg1 in the distal small intestine and colon
when compared to adults. Absorption in the duodenum remained unchanged throughout the study. These data suggest that the distal gastrointestinal tract plays a role in the high absorption seen in neonatal animals.
Chapter seven provides a discussion of the results presented in this thesis. In addition, a model for the regulation of iron absorption is proposed. Central to this hypothesis are the suggestions that the liver is of prime importance in the regulation of iron absorption and that the signal produced by the liver to alter absorption is detected in the mature villus enterocytes rather than by those of the crypts. This model is able to explain the regulation of iron absorption under normal conditions and also the disrupted iron metabolism that occurs in many pathological states.