Duodenal ascorbate and ferric reductase in human iron deficiency

BACKGROUND: The first step in iron absorption requires the reduction of ferric iron to ferrous iron, a change that is catalyzed by duodenal ferric reductase. Iron deficiency is associated with high iron absorption, high ferric reductase activity, and high duodenal ascorbate concentrations in experimental animals, but it is not known whether a relation between reductase and ascorbate is evident in humans. OBJECTIVE: The objective of the study was to assess the relation between ferric reductase activity in human duodenal biopsy specimens and ascorbate concentrations in iron-replete and iron-deficient subjects. DESIGN: Patients and control subjects were overnight-fasted adults presenting sequentially for upper gastrointestinal endoscopic investigation. Ferric reductase activity in duodenal biopsy specimens was assayed by using nitroblue tetrazolium. Ascorbate was assayed in duodenal biopsy specimens and plasma. RESULTS: Iron-deficient patients had significantly higher reductase activity (n = 6-9; P < 0.05) and duodenal (n = 20; P < 0.001) and plasma (n = 6; P < 0.001) ascorbate concentrations than did control subjects. Incubation of biopsy specimens with dehydroascorbate (to boost cellular ascorbate) increased reductase activity in the tissues that initially had normal activity (n = 9; P < 0.01) but inhibited reductase activity in the tissues that already had high reductase activity (n = 13; P < 0.001). CONCLUSIONS: Iron deficiency in humans is associated with increased duodenal ascorbate concentrations. This finding suggests that increased reductase activity is partly due to an increase in this substrate for duodenal cytochrome b reductase 1.     

Duodenal cytochrome B expression stimulates iron uptake by human intestinal epithelial cells

Duodenal cytochrome B (Dcytb) is localized principally in the apical membrane of the enterocyte. It is thought to act as a ferric reductase that furnishes Fe(II), the specific and selective iron species transported by divalent metal transporter 1 (DMT1) in the duodenal enterocytes. Expression of both genes is strongly iron regulated and is thought to be required for transcellular iron trafficking in concert in response to physiological requirements. We tested this hypothesis by expressing Dcytb in Caco-2 cells, a human cell line model often used to mimic intestinal enterocytes. Iron uptake (59Fe) was significantly higher in Dcytb-transfected Caco-2 cells than in cells transfected with empty vector as a control. Fe(III) reductase activity of Dcytb was measured with ferrozine, a strong chelator of Fe(II) species. Cells expressing Dcytb exhibited enhanced ferric reductase activity as well as increased 59Fe uptake compared with cells transfected with empty vector as a control. Ferrozine blocked iron uptake and preincubation of cells with dehydroascorbate (to increase cellular ascorbate levels) stimulated iron uptake. Cotransfection of Dcytb and DMT1 resulted in an additive increase in iron uptake by the cells. The results confirm Dcytb can act as a ferric reductase that stimulates iron uptake in Caco-2 cells.     

Effects of varying dietary iron on the expression of copper deficiency in the growing rat: anemia, ferroxidase I and II, tissue trace elements, ascorbic acid, and xanthine dehydrogenase

The effect of dietary iron on the development of copper-deficiency anemia in the growing rat was investigated. For up to 80 d, female rats (75 g) were fed purified diets containing adequate, marginal or low levels of iron, and either 0.7 or 10 ppm copper. Hemoglobin levels and factors postulated to affect liver iron mobilization, including ferroxidase (Fox) I and II, ascorbate and liver xanthine dehydrogenase (XDH) were assayed. By d 7, Fox I activity in the copper-deficient groups was 10% that of the copper-sufficient groups; thereafter, Fox I activity remained low, and was not affected by dietary iron. Fox II activity in the copper-deficient groups after d 28 was 50-75% of values from rats adequate in copper. On d 49, hemoglobin levels in the copper-deficient groups were lower than in the copper-sufficient groups fed low and marginal levels of iron, but were similar to those fed adequate iron. Liver iron was similar in both groups fed adequate iron, but was higher in the copper-deficient than in the copper-sufficient rats fed low or marginal levels of iron. Copper deficiency tended to result in slightly lower ascorbate levels on d 80 at all levels of iron. Liver XDH activity tended to be lower in the copper-deficient groups than in the copper-sufficient groups on d 28 and 49. These results show that copper deficiency may impair liver iron mobilization in the growing rat if dietary iron is low. Possible mechanisms include decreased Fox activity and/or decreased iron reduction by ascorbate or XDH.     

Choline Alleviates Parenteral Nutrition–Associated Duodenal Motility Disorder in Infant Rats

Background:Parenteral nutrition (PN) has been found to influence duodenal motility in animals. Choline is an essential nutrient, and its deficiency is related to PN‐associated organ diseases. Therefore, this study was aimed to investigate the role of choline supplementation in an infant rat model of PN‐associated duodenal motility disorder. Materials and Methods: Three‐week‐old Sprague‐Dawley male rats were fed chow and water (controls), PN solution (PN), or PN plus intravenous choline (600 mg/kg) (PN + choline). Rats underwent jugular vein cannulation for infusion of PN solution or 0.9% saline (controls) for 7 days. Duodenal oxidative stress status, concentrations of plasma choline, phosphocholine, and betaine and serum tumor necrosis factor (TNF)–α were assayed. The messenger RNA (mRNA) and protein expression of c‐Kit proto‐oncogene protein (c‐Kit) and membrane‐bound stem cell factor (mSCF) together with the electrophysiological features of slow waves in the duodenum were also evaluated. Results: Rats on PN showed increased reactive oxygen species; decreased total antioxidant capacity in the duodenum; reduced plasma choline, phosphocholine, and betaine; and enhanced serum TNF‐α concentrations, which were reversed by choline intervention. In addition, PN reduced mRNA and protein expression of mSCF and c‐Kit, which were inversed under choline administration. Moreover, choline attenuated depolarized resting membrane potential and declined the frequency and amplitude of slow waves in duodenal smooth muscles of infant rats induced by PN, respectively. Conclusion: The addition of choline to PN may alleviate the progression of duodenal motor disorder through protecting smooth muscle cells from injury, promoting mSCF/c‐Kit signaling, and attenuating impairment of interstitial cells of Cajal in the duodenum during PN feeding.     

The antioxidant BHT normalizes some oxidative effects of iron + ascorbate on lipid metabolism in Caco-2 cells

We showed recently that iron + ascorbate can impair the assembly of intestinal lipoproteins. However, we could not determine whether these changes were caused by iron + ascorbate-mediated lipid peroxidation per se. We therefore conducted studies to evaluate how antioxidants antagonize the iron + ascorbate-induced derangements. To this end, Caco-2 cells, a reliable experimental intestinal model, were incubated with iron + ascorbate (0.2 mmol/L each) alone or with different concentrations of catalase, mannitol, tocopherol or BHT. Exposing Caco-2 cells to iron + ascorbate increased malondialdehyde levels fourfold (P < 0.0001); this effect was decreased markedly (P < 0.02) in the presence of BHT. Furthermore, BHT normalized the abnormal intracellular events involved in fat absorption, i.e., lipid esterification, cholesterol synthesis and apolipoprotein production. On the other hand, it did not fully restore the secretion of lipids and lipoproteins. Thus, our current data imply that iron + ascorbate-catalyzed lipid peroxidation is partially responsible for the disturbances observed in intestinal lipid transport.     

Hepcidin and Iron Deficiency in Women One Year after Sleeve Gastrectomy: A Prospective Cohort Study

Iron deficiency with or without anemia, needing continuous iron supplementation, is very common in obese patients, particularly those requiring bariatric surgery. The aim of this study was to address the impact of weight loss on the rescue of iron balance in patients who underwent sleeve gastrectomy (SG), a procedure that preserves the duodenum, the main site of iron absorption. The cohort included 88 obese women; sampling of blood and duodenal biopsies of 35 patients were performed before and one year after SG. An analysis of the 35 patients consisted in evaluating iron homeostasis including hepcidin, markers of erythroid iron deficiency (soluble transferrin receptor (sTfR) and erythrocyte protoporphyrin (PPIX)), expression of duodenal iron transporters (DMT1 and ferroportin) and inflammatory markers. After surgery, sTfR and PPIX were decreased. Serum hepcidin levels were increased despite the significant reduction in inflammation. DMT1 abundance was negatively correlated with higher level of serum hepcidin. Ferroportin abundance was not modified. This study shed a new light in effective iron recovery pathways after SG involving suppression of inflammation, improvement of iron absorption, iron supply and efficiency of erythropoiesis, and finally beneficial control of iron homeostasis by hepcidin. Thus, recommendations for iron supplementation of patients after SG should take into account these new parameters of iron status assessment.     

Dose-responsive alteration in hepatic lipid peroxidation and retinol metabolism with increasing dietary beta-carotene in iron deficient rats

Phosphatidylcholine hydroperoxide (PCOOH) levels are increased in the iron-deficient rat liver. We investigated the antioxidative effect of dietary beta-carotene and altered retinol metabolism in iron-deficient rats. Experiment 1: Male Wistar-strain rats were divided into six groups and fed a control diet, an iron-deficient diet, and iron-deficient diets with four different levels of dietary beta-carotene. The PCOOH concentration in the iron-deficient rat liver was decreased by supplementation with dietary beta-carotene. However, the beta-carotene dose response was not related to antioxidative potency. Hepatic and plasma beta-carotene concentrations were increased by iron deficiency. The hepatic retinol concentration was increased while the plasma retinol concentration was decreased in iron-deficient rats. Experiment 2: Male Wistar-strain rats were divided into two groups, with one group receiving a control diet with beta-carotene and the other an iron-deficient diet with beta-carotene. Intestinal iron was decreased and intestinal beta-carotene was unchanged in iron-deficient rats. The intestinal beta-carotene conversion ratio and beta-carotene cleavage enzyme activity were decreased in iron-deficient rats. Dietary beta-carotene played the role of an antioxidant in hepatic lipid peroxidation in the iron-deficient state, but there was no dose dependency. Moreover, intestinal beta-carotene cleavage and hepatic retinol release appear to be altered in iron-deficient rats.     

Asymptomatic giardiasis does not affect iron absorption in children with iron deficiency anemia.

Malabsorption of iron has been reported in children with symptomatic giardiasis. The aim of this study was to evaluate intestinal absorption of iron in children with asymptomatic giardiasis and iron deficiency anemia.

Based upon results of blood hemoglobin and stool examination, two groups were established: asymptomatic giardiasis and anemia, and anemia without intestinal parasitosis (control group). Patients were aged 1-6 years. There was no difference in age, weight, height, or iron nutritional status between the asymptomatic giardiasis and control groups on admission to the study.

Intestinal absorption of iron was evaluated using the iron tolerance test and the hemoglobin response to iron therapy. The serum iron tolerance test was based on the increment of iron level 2 hours after administering an iron load of 1 mg/kg of elemental iron in the form of ferrous sulfate, in comparison to the fasting iron level. Hemoglobin response to oral iron therapy was determined by the increment of hemoglobin on day 30 of therapy with ferrous sulfate (5 mg/kg/day of elemental iron).

There was no statistical difference between the asymptomatic giardiasis and control groups with reference to the iron tolerance test (159.1 +/? 73.1 micrograms/dl and 154.5 +/? 76.5 micrograms/dl, respectively) and to the hemoglobin response to iron therapy (1.5 +/? 0.7 g/dl and 1.8 +/? 1.1 g/dl, respectively). The presence or absence of trophozoites of Giardia lamblia on duodenal aspirate did not affect intestinal absorption of iron.

Asymptomatic giardiasis did not affect the intestinal absorption of iron and the hemoglobin response to oral iron therapy in iron-deficient anemic children.     

Iron absorption from ferrous bisglycinate and ferric trisglycinate in whole maize is regulated by iron status

BACKGROUND: There is a need to determine whether iron absorption from iron amino acid chelates is protected from inhibition by dietary phytate and regulated normally by iron status. OBJECTIVE: The objective of this study was to compare iron absorption from ferrous sulfate, ferrous bisglycinate, and ferric trisglycinate in whole-maize meal; to determine whether iron from ferrous bisglycinate and ferrous sulfate exchanges in the intestinal pool; and to assess iron absorption from ferrous bisglycinate and ferric trisglycinate over a range of iron statuses. DESIGN: In study 1A, 10 iron-sufficient men consumed ferrous sulfate-fortified whole-maize meal porridge equilibrated with (59)Fe-sulfate on day 1 and (55)Fe-bisglycinate on day 2. In study 1B, these volunteers consumed ferrous sulfate-fortified porridge equilibrated with (59)Fe-sulfate and (55)Fe-bisglycinate simultaneously. In studies 2A and 2B, iron absorption from 3 mg Fe as (59)Fe-ascorbate, (55)Fe-bisglycinate, or (59)Fe-trisglycinate in water and in porridge was compared in 23 subjects with a range of iron statuses. Iron absorption was determined from blood radioactivity on day 16. RESULTS: In study 1A, geometric mean iron absorption from ferrous bisglycinate was 6.0% (range: 2.6-13.6%), 4 times higher than that from ferrous sulfate (1. 7%; range: 1.0-3.3%; P < 0.05). In study 1B, absorption from neither source was different from that in study 1A. In studies 2A and 2B, absorption from all sources was strongly inversely related to serum ferritin, with geometric means of 32.5% (iron ascorbate), 9.1% (bisglycinate), and 15.3% (trisglycinate). Iron from ferric trisglycinate was poorly absorbed (2.3%; range: 0.5-9.2%) from maize. CONCLUSION: In whole-maize meal, iron from ferrous bisglycinate is better absorbed than is iron from ferrous sulfate and does not exchange with iron from maize or ferrous sulfate in the intestinal pool. Absorption of iron from bisglycinate and trisglycinate is regulated normally by iron status.     

Depressed erythrocyte glutathione reductase activity in sickle cell disease

In a group of 27 sickle cell disease patients ranging in age from 2 yr 3 months to 43 yr, 13 (48%) were found to have depressed erythrocyte glutathione reductase activity suggesting riboflavin deficiency. Glutathione reductase activity coefficients did not correlate with riboflavin intakes which were calculated from 3-day diet records returned by 16 patients. Other causes of riboflavin deficiency including decreased absorption, altered metabolism, or increased excretion of the vitamin must be considered. The potential effect of depressed erythrocyte glutathione reductase activity in the sickle cell disease process is discussed.     

Sugar as a vehicle for iron fortification: further studies.

The data presented confirm the advantages of sugar as a vehicle for iron fortification over other vehicles used in the past. The absorption comparison between ferric and ferrous salts added to sugar demonstrated that Fe(III)-EDTA Complex and ferrous sulfate exhibited the highest absorption, while ferric ammonium citrate was poorly absorbed. It was also found that Fe(III)-EDTA reacts slowly with the tannin contained in tea; the color of the tea changes slightly in the first 2 hr after the addition of the fortified sugar. Iron absorption of sugar fortified with ferrous sulfate was tested in seven beverages. The mean absorption ratio from fortified sugar given with beverages to reference dose of iron ascorbate ranged between 0.42 and 0.70, that is, more than 4 times the absorption from fortified sugar when it is administered with a meal containing one or more vegetals. An absorption of between 0.25 and 0.80 mg of iron/soft drink sugar fortified with 3 mg of iron as ferrous sulfate can be expected in subjects with various degrees of iron deficiency. Thus, two soft drinks per day between meals would be enough to meet the iron requirement in more than 95% of menstruating women, even though the daily iron absorption from the diet is about 0.8 to 1.0 mg.     

Iron supplementation in athletes--first do no harm

Although it generally does not improve performance, iron is often used by elite athletes. The physiologic changes induced by exercise can mimic iron deficiency and decrease hemoglobin and ferritin concentrations. Determination of serum transferrin receptor concentrations may identify true iron deficiency, which occurs particularly in young athletes. In contrast, increased iron stores in the body are a frequent finding in elite athletes who have used long-term iron supplementation. Elite runners have increased intestinal blood loss, but this usually can be compensated by enhanced absorption of dietary iron. The combination of exercise-induced hemolysis with enhanced intestinal blood loss in various endurance sports leads to severe abnormalities of routine tests, and extreme physical activity may be responsible for positive fecal occult blood determinations. Indiscriminate iron supplementation carries the risk of inducing hemochromatosis in individuals homozygous for the widespread C282Y allele of the HFE gene. This polymorphism is common and can be found in about 1% of individuals of Northern European descent; moreover, iron supplementation can modify the presentation of important underlying diseases such as celiac disease or colon carcinoma. In conclusion, iron supplements should be prescribed for athletes with iron-deficiency anemia and carefully monitored if given for prophylaxis; unless a therapeutic response occurs, investigations to establish the cause of iron deficiency should be initiated.     

Duodenal Cytochrome b (DCYTB) in Iron Metabolism: An Update on Function and Regulation

Iron and ascorbate are vital cellular constituents in mammalian systems. The bulk-requirement for iron is during erythropoiesis leading to the generation of hemoglobin-containing erythrocytes. Additionally, both iron and ascorbate are required as co-factors in numerous metabolic reactions. Iron homeostasis is controlled at the level of uptake, rather than excretion. Accumulating evidence strongly suggests that in addition to the known ability of dietary ascorbate to enhance non-heme iron absorption in the gut, ascorbate regulates iron homeostasis. The involvement of ascorbate in dietary iron absorption extends beyond the direct chemical reduction of non-heme iron by dietary ascorbate. Among other activities, intra-enterocyte ascorbate appears to be involved in the provision of electrons to a family of trans-membrane redox enzymes, namely those of the cytochrome b₅₆₁ class. These hemoproteins oxidize a pool of ascorbate on one side of the membrane in order to reduce an electron acceptor (e.g., non-heme iron) on the opposite side of the membrane. One member of this family, duodenal cytochrome b (DCYTB), may play an important role in ascorbate-dependent reduction of non-heme iron in the gut prior to uptake by ferrous-iron transporters. This review discusses the emerging relationship between cellular iron homeostasis, the emergent “IRP1-HIF2α axis”, DCYTB and ascorbate in relation to iron metabolism.     

Highly esterified pectin with low molecular weight enhances intestinal solubility and absorption of ferric iron in rats

The effects of the degree of esterfication (DE) and the molecular weight (MW) of pectins on intestinal solubility and absorption of ferric iron were investigated in vivo and in vitro. The pectins prepared differed in DE and MW, respectively as follows: P-A (73%, 860,000), P-B (75%, 89,000), P-C (22%, 1,260,000), and P-D (24%, 114,000). All the rats were fed a basal diet containing ferric sulfate as an iron source for 7d. Rats were deprived of feed for about 19h and then fed 6g (based on the preliminary experiment) of the basal diet or basal diet containing one of pectins (80 g/kg diet) for 2 h. One hour later, soluble, insoluble and tissue iron concentrations in both stomach and intestinal segments and serum iron concentration in portal blood were determined and the amounts of absorbed iron were calculated. Addition of pectin B to the diet resulted in significantly greater intestinal iron absorption and postprandial iron concentration in portal blood compared to the other groups. There were no significant differences in calculated absorbed iron and serum iron concentration in portal blood between control and other pectin groups except those fed pectin B. When ferric sulfate was added to the pectin solutions and pectin diets in vitro under simulated gastrointestinal condition, more iron was found in the soluble fraction compared to control containing no pectin (P < 0.05). Solubility of ferric sulfate in vitro was also significantly greater in both the solution and diet containing pectin B than in those containing no pectin (P < 0.05). The present findings clearly indicate that iron solubility was influenced by pectin's DE and MW. The pectin with the high DE and low MW enhanced ferric iron solubility and absorption of solubilized iron.     

Effect of zinc and copper deficiency on microsomal NADPH-dependent active oxygen generation in rat lung and liver

Both dietary zinc and copper deficiencies can cause lipid peroxidation in microsomes in rats. The cytochrome P-450 enzyme system can generate active oxygens by uncoupling of the P-450-oxy complex in the catalytic cycle and/or the electron transfer mediated by the NADPH-cytochrome P-450 reductase. The effects of dietary zinc and copper deficiencies on NADPH-dependent H2O2 generation, the catalytic activity of the cytochrome P-450 enzyme with aminopyrine as the substrate and the activity of NADPH-cytochrome P-450 reductase were determined. Zinc deficiency caused increased H2O2 production, increased NADPH-cytochrome P-450 reductase activity, decreased aminopyrine demethylation and two- and fivefold increases in iron concentration in lung and liver microsomes, respectively, compared to Zn-adequate, ad libitum--fed controls. Active oxygen generation by uncoupling of the cytochrome P-450 enzyme system and accumulation of iron are thus possible mechanisms by which zinc deficiency causes microsomal lipid peroxidation. Copper deficiency did not affect H2O2 production; however, it caused two- and fourfold increases in iron concentration in lung and liver microsomes, respectively, compared to Cu-adequate, ad libitum--fed controls. The mechanism by which cooper deficiency causes microsomal lipid peroxidation is still unknown but could be related to the observed accumulation of iron.     

Intestinal iron absorption in chronic alcoholics.

Chronic alcohol misusers frequently accumulate significant amounts of excess iron, but the mechanism of this loading is unknown. In vivo whole-body retention studies demonstrated, on average, a two-fold increase in intestinal iron absorption in six male chronic alcoholics. Degrees of iron loading as assessed by serum ferritin or hepatic iron levels did not correlate with alcohol consumption or liver function tests. In vitro studies of iron uptake at varying medium iron concentrations by duodenal mucosa biopsies showed increased iron uptake by tissue from the chronic alcoholics, particularly at the highest medium iron concentration used. Analysis of the uptake data showed similar Michaelis-Menten kinetic constants for uptake by tissue from control subjects and alcoholics. The analysis showed, in addition, a linear component for 59Fe uptake. This component was five-fold greater for the tissue from the chronic alcoholics compared to the controls at the highest medium iron concentration. 57Co-cyanocobalamin was included in the incubation medium as a tissue extracellular fluid marker (ECF). It was found that the apparent distribution volume of the ECF marker, reflecting tissue permeability, was 75% higher for the biopsies from the alcoholics compared to control subjects. These results, together with the previous reports of enhanced in vitro and in vivo intestinal permeability to 51Cr-EDTA in chronic alcoholics, indicate that unregulated increased iron absorption via the non-carrier-mediated paracellular route contributes to the iron overload in chronic alcoholics.     

Glucose Stimulates Gut Motility in Fasted and Fed Conditions: Potential Involvement of a Nitric Oxide Pathway

(1) Background: Type 2 diabetes (T2D) is associated with a duodenal hypermotility in postprandial conditions that favors hyperglycemia and insulin resistance via the gut-brain axis. Enterosynes, molecules produced within the gut with effects on the enteric nervous system, have been recently discovered and pointed to as potential key modulators of the glycemia. Indeed, targeting the enteric nervous system that controls gut motility is now considered as an innovative therapeutic way in T2D to limit intestinal glucose absorption and restore the gut-brain axis to improve insulin sensitivity. So far, little is known about the role of glucose on duodenal contraction in fasted and fed states in normal and diabetic conditions. The aim of the present study was thus to investigate these effects in adult mice. (2) Methods: Gene-expression level of glucose transporters (SGLT-1 and GLUT2) were quantified in the duodenum and jejunum of normal and diabetic mice fed with an HFD. The effect of glucose at different concentrations on duodenal and jejunal motility was studied ex vivo using an isotonic sensor in fasted and fed conditions in both normal chow and HFD mice. (3) Results: Both SGLT1 and GLUT2 expressions were increased in the duodenum (47 and 300%, respectively) and jejunum (75% for GLUT2) of T2D mice. We observed that glucose stimulates intestinal motility in fasted (200%) and fed (400%) control mice via GLUT2 by decreasing enteric nitric oxide release (by 600%), a neurotransmitter that inhibits gut contractions. This effect was not observed in diabetic mice, suggesting that glucose sensing and mechanosensing are altered during T2D. (4) Conclusions: Glucose acts as an enterosyne to control intestinal motility and glucose absorption through the enteric nervous system. Our data demonstrate that GLUT2 and a reduction of NO production could both be involved in this stimulatory contracting effect.     

Influence of infection/inflammation, thalassemia and nutritional status on iron absorption

Iron balance in human beings is maintained by the control of absorption. Recent observations have demonstrated that a peptide hormone, hepcidin, is the principal regulator of iron homeostasis. It is produced in the liver in response to increasing iron stores. It is also induced by interleukin-6 (IL-6) in infectious and inflammatory diseases. Hepcidin restricts both iron absorption and iron release from stores. Disorders that affect the duodenum or stomach directly, particularly gluten enteropathy and H. pylori infections, also impair iron absorption by damaging enterocytes or reducing gastric acid output. Hepcidin secretion is suppressed by accelerated erythropoiesis even when iron stores are increased. This appears to account for the contribution that excessive absorption makes to the iron overload seen in patients with iron-loading anemias such as thalassemia major. There is some evidence suggesting that two nutritional deficiency disorders (deficiencies of vitamin A and riboflavin) lead to impaired iron absorption or utilization, but further research is needed to reconcile conflicting experimental observations.