Low vitamin a intake affects milk iron level and iron transporters in rat mammary gland and liver

Marginal vitamin A deficiency is common and can result in a secondary iron (Fe) deficiency. A positive correlation between maternal Fe status and milk Fe was observed in lactating women supplemented with both vitamin A and Fe but not with Fe alone, suggesting effects of vitamin A on mammary gland Fe transport. We hypothesized that low vitamin A intake during lactation elicits differential effects on mammary gland and liver Fe transport and storage proteins, thus affecting milk Fe concentration but not maternal Fe status. We fed rats a control (CON, 4 RE/g) or a marginal vitamin A diet (AD, 0.4 RE/g) through midlactation. Effects on plasma, milk, liver and mammary gland Fe and vitamin A concentrations, and divalent metal transporter-1 (DMT1), ferroportin (FPN), ferritin (Ft), and transferrin receptor (TfR) expression were determined. Dams fed AD were not vitamin A or Fe deficient. Milk and liver vitamin A and Fe and mammary gland Fe concentrations were lower in rats fed AD compared with rats fed CON. Liver TfR expression was higher, whereas mammary gland TfR expression was lower in rats fed AD compared with rats fed CON. Liver Ft was unaffected, whereas mammary gland Ft was lower in rats fed AD compared with rats fed CON. Liver and mammary gland DMT1 and FPN protein levels were lower in rats fed AD compared with rats fed CON. Our results indicate that the mammary gland and liver respond differently to marginal vitamin A intake during lactation and that milk Fe is significantly decreased due to effects on mammary gland Fe transporters, putting the nursing offspring at risk for Fe deficiency.     

Iron supplementation during infancy--effects on expression of iron transporters, iron absorption, and iron utilization in rat pups

BACKGROUND: Studies conducted in human infants suggest developmental changes in the regulation of iron absorption; however, little is known about the molecular mechanisms regulating iron absorption during infancy. Two intestinal iron transporters, divalent metal transporter 1 (DMT1) and ferroportin 1 (FPN1), were recently identified. OBJECTIVE: The objective was to investigate at a molecular level the regulation of iron absorption during infancy in a rat pup model. We examined the developmental expression of DMT1 and FPN1 and the effects of iron supplementation on their expression and on iron absorption and utilization during infancy. DESIGN: Rat pups were given daily oral doses of 0, 30, or 150 microg Fe from day 2 to day 20 after birth. On days 10 and 20 after birth, (59)Fe absorption, tissue minerals, and intestinal DMT1, FPN1, and ferritin expression were examined. To assess developmental expression, DMT1 and FPN1 were examined in control rats from days 1 to 50 after birth. RESULTS: Intestinal DMT1 and FPN1 were significantly affected by age; expression increased dramatically by day 40. On day 10, no significant effect of iron supplementation on DMT1 and FPN1 gene expression or on iron absorption was observed. By day 20, DMT1 and FPN1 expression and iron absorption had decreased significantly with iron supplementation. CONCLUSIONS: During early infancy, rat pups are unable to down-regulate intestinal iron transporters or iron absorption in response to iron supplementation, whereas down-regulation occurs during late infancy. The current findings provide evidence of the developmental regulation of iron absorption, which emphasizes the need for caution when giving iron supplements to infants at an early age.     

Obesity Promotes Alterations in Iron Recycling

Hepcidin is a key hormone that induces the degradation of ferroportin (FPN), a protein that exports iron from reticuloendothelial macrophages and enterocytes. The aim of the present study was to experimentally evaluate if the obesity induced by a high-fat diet (HFD) modifies the expression of FPN in macrophages and enterocytes, thus altering the iron bioavailability. In order to directly examine changes associated with iron metabolism in vivo, C57BL/6J mice were fed either a control or a HFD. Serum leptin levels were evaluated. The hepcidin, divalent metal transporter-1 (DMT1), FPN and ferritin genes were analyzed by real-time polymerase chain reaction. The amount of iron present in both the liver and spleen was determined by flame atomic absorption spectrometry. Ferroportin localization within reticuloendothelial macrophages was observed by immunofluorescence microscopy. Obese animals were found to exhibit increased hepcidin gene expression, while iron accumulated in the spleen and liver. They also exhibited changes in the sublocation of splenic cellular FPN and a reduction in the FPN expression in the liver and the spleen, while no changes were observed in enterocytes. Possible explanations for the increased hepcidin expression observed in HFD animals may include: increased leptin levels, the liver iron accumulation or endoplasmic reticulum (ER) stress. Together, the results indicated that obesity promotes changes in iron bioavailability, since it altered the iron recycling function.     

Variable effects of iron status on the concentration of ferritin in rat plasma, liver, and spleen

The present studies were conducted to determine the relationships between iron status and ferritin levels in plasma, liver, and spleen of rats. Rats were fed either iron-adequate or iron-deficient purified diets, and measurements of hemoglobin and plasma and tissue ferritin levels were made at various times during iron depletion and iron repletion. Although mean plasma ferritin concentrations of iron-deficient rats were directionally less than those of iron-adequate rats, these differences were not statistically significant due to high variability among similarly treated animals. During iron repletion plasma ferritin concentrations again were so variable that no significant effect of iron repletion on plasma ferritin concentrations was observed. On the other hand, liver and spleen ferritin concentrations of similarly treated rats were much less variable. Ferritin liver and spleen stores decreased more rapidly than hemoglobin during iron deficiency and were restored more slowly than hemoglobin during iron repletion. There was no evidence of correlation between liver and plasma ferritin concentration. Because of the variable responses of plasma ferritin concentration to iron depletion and repletion and the lack of relationship between plasma and liver ferritin concentrations, it is concluded that plasma ferritin concentration is not a good indicator of iron status in rats.     

Early postnatal iron repletion overcomes lasting effects of gestational iron deficiency in rats

Iron deficiency anemia in early childhood causes developmental delays and, very likely, irreversible alterations in neurological functioning. One primary goal for the present study was to determine whether the effects of late gestational iron deficiency on brain monoamine metabolism, iron content, and behavioral phenotypes could be repaired with iron intervention in early lactation. Young pregnant rats were provided iron-deficient or control diets from mid-gestation (G15). At postnatal d 4 (P4), pups from iron-deficient dams were out-fostered either to other ID dams or control dams while pups of control dams were similarly fostered to other control dams. Dietary treatments continued to adulthood (P65) when brain iron and regional monoamines were evaluated. P4 iron repletion normalized body iron status, brain iron concentrations, monoamine concentrations, and monoamine transporter and receptor densities in most brain regions. Dopamine transporter densities in caudate and substantia nigra were lower in ID rats but were normalized with iron repletion. Serotonin transporter levels in most brain regions and open-field exploration were also normalized with iron repletion. The success of this approach of early postnatal iron intervention following iron deficiency in utero contrasts to a relative lack of success when the intervention is performed at weaning. These data suggest that a window of opportunity exists for reversing the detrimental effects of iron deficiency in utero in rats and provides strong support of intervention approaches in humans with iron deficiency during pregnancy.     

维生素A缺乏对大鼠铁水平及Hepcidin、FP1与DMT1 mRNA表达的影响

目的研究维生素A(VA)缺乏时大鼠铁营养状况的变化,从分子水平探讨VA缺乏对大鼠铁代谢的影响。方法 16只成年健康雄性SD大鼠,随机分为对照组(Con)和VA缺乏组(-VA),每组8只,分别饲以正常饲料和VA缺乏饲料。连续喂养4周,脱椎处死,取血及肝、脾组织检测血红蛋白(Hb)含量、血清铁(SI)、血清铁蛋白(SF)及肝、脾组织铁含量。以β-actin为内参,半定量RT-PCR技术比较两组大鼠铁调素(Hepcidin)、膜铁转运蛋白1(FP1)和二价金属离子转运蛋白(DMT1)肝组织mRNA的表达水平。结果 VA缺乏组SI、SF和Hb显著低于对照组(P0.05),肝、脾铁含量显著高于对照组(P0.05);Hepcidin及DMT1mRNA在肝组织中表达水平显著增加(P0.05)。结论 VA缺乏致使Hepcidin表达水平的升高,导致肝、脾铁储量增加而机体循环铁量和铁营养水平下降。     

Dietary iron supplements and Moringa oleifera leaves influence the liver hepcidin messenger RNA expression and biochemical indices of iron status in rats

In this study, the effects of iron depletion and repletion on biochemical and molecular indices of iron status were investigated in growing male Wistar rats. We hypothesized that iron from Moringa leaves could overcome the effects of iron deficiency and modulate the expression of iron-responsive genes better than conventional iron supplements. Iron deficiency was induced by feeding rats an iron-deficient diet for 10 weeks, whereas control rats were maintained on an iron-sufficient diet (35.0-mg Fe/kg diet). After the depletion period, animals were repleted with different source of iron, in combination with ascorbic acid. Iron deficiency caused a significant (P < .05) decrease in serum iron and ferritin levels by 57% and 40%, respectively, as compared with nondepleted control animals. Significant changes in the expression (0.5- to100-fold) of liver hepcidin (HAMP), transferrin, transferrin receptor-2, hemochromatosis type 2, ferroportin 1, ceruloplasmin, and ferritin-H were recorded in iron-depleted and iron-repleted rats, as compared with nondepleted rats (P < .05). Dietary iron from Moringa leaf was found to be superior compared with ferric citrate in overcoming the effects of iron deficiency in rats. These results suggest that changes in the relative expression of liver hepcidin messenger RNA can be used as a sensitive molecular marker for iron deficiency.     

Regulation of iron metabolism in Hamp −/− mice in response to iron-deficient diet

Background

Hepcidin, the liver-secreted iron regulatory peptide, maintains systemic iron homeostasis in response to several stimuli including dietary iron levels and body iron status. In addition, iron metabolism is controlled by several local regulatory mechanisms including IRP and Hif-2α activities independently of hepcidin. However, the roles of these mechanisms and their interaction particularly in hepcidin-deficient individuals are not yet fully understood. We, therefore, aimed to explore whether Hamp disruption affects iron homeostatic responses to dietary iron deficiency.

Methods

Hepcidin1 knockout (Hamp ^?/?) mice and heterozygous littermates were fed with control or iron-deficient diet for 2 weeks. The expression of iron-related genes and proteins were determined by quantitative PCR and Western blot, respectively.

Results

Two-week iron-deficient diet feeding in Hamp ^?/? mice did not alter serum iron but significantly reduced liver non-heme iron levels. This was also associated with increased ferroportin protein expression in the duodenum and spleen, whereas decreased expression was found in the liver. In addition, significant inductive effects of iron-deficient diet on Dcytb and DMT1 mRNA expression in the duodenum were noted with more pronounced effects in Hamp ^?/? mice compared with controls.

Conclusions

Hamp ^?/? mice exhibited a more dramatic increase in the expression of iron transport machinery, which may be responsible for the unaltered serum iron levels upon iron-deficient diet feeding in these mice. Despite the lack of hepcidin, Hamp ^?/? mice can maintain a degree of iron homeostasis in response to altered dietary iron through several hepcidin-independent mechanisms.     

Vitamin A deficiency increases hepcidin expression and oxidative stress in rat

ObjectiveThe interaction between vitamin A and iron status has been widely reported; however, the exact mechanism involved in this interaction has not been well characterized. The present study investigated the mechanism involved in tissue iron accumulation and changes in the oxidative status in vitamin A–deficient rats.MethodsRats were treated with a control diet, a vitamin A–deficient diet, or a vitamin A/iron-deficient diet for 57 d. The animals were sacrificed; the blood, liver, and spleen were collected for biochemical analysis. Analysis of variance or Mann-Whitney tests were used to compare groups and Pearson's or Spearman's tests to investigate the bivariate correlation.ResultsVitamin A deficiency increased liver hepcidin mRNA and iron spleen concentrations; however, iron deficiency in vitamin A–deficient rats deeply inhibits liver hepcidin mRNA expression and significantly increases divalent metal transporter-1 mRNA levels. Liver ferroportin and hereditary hemochromatosis gene mRNA levels did not change in either treatment. In the vitamin A–deficient groups, liver carbonyl protein increased, whereas catalase and glutathione S-transferase activities decreased, suggesting that vitamin A protects the liver against protein oxidation. A significant positive correlation was found between lipid oxidative damage and iron concentration in the liver and spleen (r = 0.611, P = 0.007; r = 0.558, P = 0.025, respectively).ConclusionThese results suggest that vitamin A maintains iron homeostasis by the modulation of liver hepcidin expression. The increase of lipid peroxidation in vitamin A deficiency seems to be iron dependent, whereas protein oxidation is not.     

铜缺乏对大鼠铁代谢影响的研究

目的研究铜缺乏对大鼠铁营养状况和肝脏转铁蛋白受体mRNA及铁调素mRNA的影响。方法 48只雄性SD大鼠按体重随机分为4组,每组12只,正常对照组(Ⅰ组),铁正常铜完全缺乏组(Ⅱ组),铁正常铜轻度缺乏组(Ⅲ组),铁/铜轻度缺乏组(IV组)。喂饲8周后处死,取大鼠肝脏、脾脏和血清,测定血红蛋白,血清铜、血清铁、血清转铁蛋白受体、血清铁蛋白、肝脏铁和铜含量、脾脏铁和铜含量,并用逆转录-聚合酶链反应(RT-PCR)法检测各组大鼠肝脏转铁蛋白受体(TfR)和铁调素mRNA的表达水平。结果与正常对照组相比,铜缺乏使血清铁、血清铁蛋白含量显著降低(P0.05),血清转铁蛋白受体水平、肝脾脏铁含量显著升高(P0.05),铜缺乏时肝脏TfRmRNA表达增强,而肝脏铁调素mRNA表达下降。结论铜缺乏可能通过影响铁吸收、储存、转运来影响体内铁的营养状况。铜缺乏对大鼠铁调素mRNA的表达与IRE-IRP途径调节的转铁蛋白受体mRNA的表达均产生影响。     

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.     

高铁饲料对大鼠铁水平及铁调素mRNA表达的影响

目的研究不同铁含量的高铁饲料对大鼠体重、体内铁水平及铁调素mRNA的影响。方法60只清洁级雄性SD大鼠随机分为6组,分别喂饲铁含量为35(A组)、100(B组)、200(C组)、500(D组)、1000(E组)、3500(F组)mg/kg的饲料8周后,处死大鼠。取大鼠肝脏、十二指肠和血清,比色法测定大鼠血清铁(SI)、转铁蛋白饱和度(TS)、ELISA法测定血清铁蛋白(SF)水平;逆转录-聚合酶链反应法检测大鼠肝脏铁调素(hepcidin)和十二指肠膜铁转运蛋白(ferroprotin,Fpn1)mRNA表达水平。结果随着饲料铁水平的提高,大鼠体重增重逐渐减少;大鼠血清铁、血清铁蛋白、转铁蛋白饱和度的水平、肝脏铁调素mRNA的表达水平逐渐升高;FPN1mRNA的表达下降。结论高铁饲料对大鼠体重、血清铁水平及肝脏铁调素、十二指肠Fpn1mRNA的表达均产生影响。     

Zinc supplementation reduces iron absorption through age-dependent changes in small intestine iron transporter expression in suckling rat pups

Zinc (Zn) supplementation negatively affects iron (Fe) absorption; however, the molecular mechanisms are not understood. We determined effects of Zn supplementation during mid- and late infancy on intestinal Fe transport mechanisms using a suckling rat model. Suckled rat pups were supplemented with 0 (control), 300 (low), or 750 (high) microg Zn/d until weaning at postnatal day (PN) 20. At mid-(PN10) and late (PN20) infancy, tissue Fe distribution, Fe absorption, intestine DMT1, ferroportin-1 (FPN) and hephaestin expression, and localization and liver hepcidin expression were measured. During early infancy, DMT1 and FPN were localized intracellularly. Negative effects of Zn supplementation on Fe absorption were associated with increased small intestine Fe retention, decreased hephaestin, and increased FPN expression. During late infancy, both DMT1 and FPN were appropriately localized to the apical and basolateral membrane, respectively, and negative effects of Zn supplementation on Fe absorption were absent. Although FPN protein level was lower in Zn-supplemented pups, hephaestin protein level was increased, which may have facilitated enhanced Fe efflux. These results indicate that Zn supplementation reduced Fe absorption during early infancy as a consequence of increased intestinal Fe retention due to reduced hephaestin levels. These effects were age-dependent, further demonstrating that Fe transport regulation is not fully developed until weaning, which may have important implications regarding the safety and efficacy of Zn supplementation programs for infants.     

A postweaning iron-adequate diet following neonatal iron deficiency affects iron homeostasis and growth in young rats

Iron deficiency is among the most prevalent of nutrient-related diseases worldwide, but the long-term consequences of maternal and neonatal iron deficiency on offspring are not well characterized. We investigated the effects of a postweaning iron-adequate diet following neonatal iron deficiency on the expression of genes involved in iron acquisition and homeostasis. Pregnant rats were fed an iron-adequate diet (0.08 g iron/kg diet) until gestational d 15, at which time they were divided into 2 groups: 1) a control group fed an iron-adequate diet, and 2) an iron-deficient group fed an iron-deficient diet (0.005 g iron/kg diet) through postnatal d (P) 23 (weaning). After weaning, pups from both dietary treatment groups were fed an iron-adequate diet until adulthood (P75). Rat pups that were iron deficient during the neonatal period (IDIA) had reduced weight gain and hemoglobin concentrations and decreased levels of serum, liver, and spleen iron on P75 compared with rats that were iron sufficient throughout early life (IA). IDIA rats developed erythrocytosis during postweaning development. Further, hepatic expression of hepcidin in IDIA rats was 1.4-fold greater than in IA rats, which paralleled an upregulation of IL-1 expression in the serum. Our data suggest that an iron-adequate diet following neonatal iron deficiency induced an inflammatory milieu that affected iron homeostasis and early growth and development.     

Iron transporters in rat mammary gland: effects of different stages of lactation and maternal iron status

BACKGROUND: Mechanisms regulating iron transfer from maternal circulation into milk are yet unknown. Whether intestinal iron transporters, divalent metal transporter 1 (DMT1) and ferroportin 1 (FPN1), are present in the mammary gland and are involved in iron transfer into milk are unknown. OBJECTIVE: The objective was to examine DMT1 and FPN1 in rat mammary gland at different stages of lactation and to evaluate the effects of maternal iron status. DESIGN: Rats were fed either 35 mg Fe (control rats) or 8 mg Fe (low-iron rats) per kg diet for 3 wk and were fed the same diet throughout pregnancy and lactation. Mammary gland DMT1, FPN1, transferrin receptor, and ferritin were examined in control rats on days 1, 5, 10, and 20 of lactation and in low-iron rats on days 10 and 20 of lactation. Tissue and milk iron were measured. RESULTS: Milk iron, DMT1, and FPN1 decreased throughout lactation. Iron status was compromised in low-iron rats, whereas milk iron was maintained. On day 10 of lactation, mammary gland iron and ferritin were lower in the low-iron rats. DMT1, FPN1, and transferrin receptor values were unchanged; however, a smaller-size DMT1 protein was observed in the low-iron rats. On day 20, transferrin receptor increased in the low-iron rats, whereas mammary gland iron, ferritin, DMT1, and FPN1 were unchanged. CONCLUSIONS: The results show that DMT1 and FPN1 concentrations are higher during early lactation and are possibly involved in iron transfer into milk. Mammary gland regulation of DMT and FPN1 during low iron status appears to be different from that in the intestine.     

Neonatal iron deficiency results in irreversible changes in dopamine function in rats

Iron deficiency in human infants and in young animal models produces changes in neural functioning that may be related to monoamine metabolism. This study employed both behavioral and biochemical approaches in a design using cross-fostering to examine alterations in dopamine (DA) function when iron deficiency occurs during the neonatal period. We measured brain Fe, dopamine transporters (DAT) and dopamine receptor density in rats made iron deficient, or not, from postnatal day (PND) 4 to PND 14 or 21. Some pups were then weaned to an iron-deficient diet and others to the control diet to examine the reversibility of these effects. Behaviors related to dopamine function were measured. Dopamine D(2) receptor (D(2)R), D(1)R and iron concentrations were approximately 70, 80 and 30% of control values, respectively, in the nucleus accumbens and striatum in iron-deficient rats at PND 14. The DAT density was also reduced to 50% of control density in the nucleus accumbens but was unchanged in the striatum. By PND 21, there was also a significant 50% lowering of DAT, D(1)R and D(2)R densities in the prefrontal cortex (PFC). Iron repletion at PND 21-49 normalized D(1)R, D(2)R, and DAT levels in the nucleus accumbens, PFC and ventral midbrain but not in the striatum. In summary, neonatal iron deficiency is associated with changes in DA biology that vary with duration of iron deficiency, and are not completely normalized despite replenishment of iron status. Changes in DA-related behaviors that were persistent after postweaning iron repletion suggest the existence of a critical neonatal developmental period that is expressed by alterations in DA functioning.     

清华大学教工非酒精性脂肪肝及相关因素分析

  目的  探讨铁超载对高脂膳食诱导非酒精性脂肪肝病（NAFLD）大鼠二价金属离子转运蛋白（DMT1）、膜铁转运蛋白（FPN1）基因表达影响。  方法  采用高脂高铁饮食诱导非酒精性脂肪肝病大鼠模型,测定大鼠血清甘油三酯、总胆固醇、葡萄糖、胰岛素水平,计算胰岛素抵抗指数（HOMA-IR）;苏木素–伊红染色观察大鼠肝脏病理组织改变;逆转录–聚合酶链反应（RT-PCR）法检测大鼠十二指肠 DMT1和FPN1 mRNA表达水平。  结果  高脂高铁组大鼠血清TG含量[（0.61 ± 0.07）μmol/L],明显高于对照组（P < 0.05）,血清胰岛素和HOMA-IR分别为 [（27.73 ± 8.29）mIU/L和（6.06 ± 1.88）],明显高于对照组和高脂组（P < 0.05）;高脂高铁组大鼠肝脏脂肪变性程度较高脂组严重;高脂高铁组大鼠十二指肠DMT1和FPN1 mRNA相对表达量分别为[（0.81 ± 0.03）和（0.69 ± 0.11）],均低于对照组（P < 0.05）。  结论  高脂高铁联合作用可诱发大鼠胰岛素抵抗,加重肝脏脂肪变性程度,并使大鼠肠道铁吸收负反馈调节作用降低。     

Copper and iron deficiencies: effects on serum lipids and tissue minerals

To study the interrelationship between iron and copper on serum lipid concentrations, four diets were fed to growing rats: iron and copper deficient, copper-deficient, iron-deficient, iron and copper adequate. After 18 weeks, concentrations of iron and copper in organs and lipids in sera were determined. Iron deficiency alone or combined with copper deficiency resulted in reduced body weights, hemoglobin concentrations, hematocrits, and iron concentrations in liver, spleen, and heart. Hepatic copper was elevated 8-fold in iron deficiency. Copper deficiency alone or combined with iron deficiency resulted in reduced copper concentrations of liver and spleen and reduced ceruloplasmin. Serum triglycerides and cholesterol did not differ among experimental treatments. No significant effects of the interaction between dietary iron and copper on serum lipid levels were found.