Dietary Iron Restriction Research Articles

Atherosclerosis is aggravated by iron overload and ameliorated by dietary and pharmacological iron restriction.

Whether and how iron affects the progression of atherosclerosis remains highly debated. Here, we investigate susceptibility to atherosclerosis in a mouse model (ApoE-/- FPNwt/C326S), which develops the disease in the context of elevated non-transferrin bound serum iron (NTBI). Compared with normo-ferremic ApoE-/- mice, atherosclerosis is profoundly aggravated in iron-loaded ApoE-/- FPNwt/C326S mice, suggesting a pro-atherogenic role for iron. Iron heavily deposits in the arterial media layer, which correlates with plaque formation, vascular oxidative stress and dysfunction. Atherosclerosis is exacerbated by iron-triggered lipid profile alterations, vascular permeabilization, sustained endothelial activation, elevated pro-atherogenic inflammatory mediators, and reduced nitric oxide availability. NTBI causes iron overload, induces reactive oxygen species production and apoptosis in cultured vascular cells, and stimulates massive MCP-1-mediated monocyte recruitment, well-established mechanisms contributing to atherosclerosis. NTBI-mediated toxicity is prevented by transferrin- or chelator-mediated iron scavenging. Consistently, a low-iron diet and iron chelation therapy strongly improved the course of the disease in ApoE-/- FPNwt/C326S mice. Our results are corroborated by analyses of serum samples of haemochromatosis patients, which show an inverse correlation between the degree of iron depletion and hallmarks of endothelial dysfunction and inflammation. Our data demonstrate that NTBI-triggered iron overload aggravates atherosclerosis and unravel a causal link between NTBI and the progression of atherosclerotic lesions. Our findings support clinical applications of iron restriction in iron-loaded individuals to counteract iron-aggravated vascular dysfunction and atherosclerosis.

Body iron accumulation in obesity, diabetes and its complications, and the possibility of therapeutic application by iron regulation

Iron is an essential trace metal element for maintaining vital functions, and it is involved in hemoglobin synthesis, redox reaction, enzyme activity, cell proliferation and apoptosis in various cells. Iron deficient-related diseases represented anemia are well-known, on the other hand, iron overload disease has attracted little attention. Excessive iron produces hydroxyl radicals via Fenton/Haber-Weiss reaction, causing organ damage in hereditary iron overload diseases. Additionally, it has been clarified that iron accumulation is involved in the pathological conditions even in metabolic diseases thought to be unrelated to iron so far. Therefore, the role of iron in the living body has been raised attention again. Recent studies have reported that body iron content is associated with both obesity and diabetes, and iron might be an aggravating factor of obesity and diabetes. We have revealed that iron chelating agent reduced oxidative stress and inflammation, suppressing the development of adipose hypertrophy in KKAy mice. Dietary iron restriction also diminishes oxidative stress, leading to the inhibition of increased albuminuria excretion and glomerular lesions in db/db mice. In this review, we give an outline of the role of iron on obese and diabetes, and diabetic kidney disease, and present the possibility of application to treatment with iron regulation in those disorders.

Protection of Progression of Nonalcoholic Steatohepatitis in Diet-Induced Obese Mice by a Low-Iron Diet

Iron is a risk factor for type 2 diabetes, but its relationship to other aspects of metabolic syndrome is less clear. To investigate iron’s relation to nonalcoholic steatohepatitis (NASH), mice were fed a “fast food” (FF)-diet (40% energy as fat, 12% SFA, 0.2% cholesterol, and 18.9 g/L glucose and 23.1 g/L fructose in their drinking water) for 10 or 24 weeks. The diets contained either 4 (low iron, LI), 35 (normal iron, NI) or 2000mg iron/kg chow (high iron, HI). HOMA-IR values were 2.2-fold higher (p<0.001) in the FF-groups at 10 weeks, but did not increase further with HI (1.48-fold, p=0.5). At 10 weeks all groups had similar glucose excursions during glucose and pyruvate tolerance testing. There was no significant difference in IP-GTT or IP-PTT area under the glucose curve. Liver triglycerides (TG) were significantly increased by FF (4.2-fold, p<0.01) but not iron at 10 weeks. The effect of FF was greater at 24 weeks (22.4 fold p<0.001), and at 24 weeks iron further increased liver TG (3.3 fold HIFF vs. LIFF, p<0.001). A marker of liver injury (ALT) increased 10.3-fold (p<0.001) on the HIFF diet compared to LI normal chow, but only 3.2-fold (p<0.05) on LIFF. The fibrotic gene collagen1alpha increased 14.3-fold (p<0.001) in HIFF but not at all (0.8-fold) on LIFF compared to LI normal chow. Similar protection was afforded by the LI diet to increases in expression of the inflammatory genes TGF-beta and TNF-alpha seen in HIFF. RNA sequencing data revealed transcriptional regulation of a large family of fat metabolic genes by FF, whereas only the TGF-beta signaling pathway was significantly altered by iron content. We conclude that dietary iron restriction can protect from NASH induced by a high-fat and -carbohydrate diet. A likely candidate mechanism is through a known mediator of hepatic fibrosis, TGF-beta. Disclosure L. Salaye: None. I. Bychkova: None. F. Lorenzo: None. S.T. Sink: None. D. McClain: None.

Influence of dietary iron intake restriction on the development of hypertension in weanling prehypertensive rats.

Hypertension is a major public health problem leading to death. To reduce the morbidity and mortality in patients with hypertension, it is crucial to develop a novel strategy for prevention of hypertension. We have currently reported an attempt at dietary iron intake restriction as non-pharmacological treatment of hypertension in patients with hypertension. However, it remains fully unknown whether dietary iron restriction prevents the development of hypertension. We investigated the influence of dietary iron restriction on the development of hypertension in weanling pre-hypertensive model rats. 3-week-old male stroke-prone spontaneously hypertensive rats (SHR-SP) were randomly divided into two groups and were given an ad libitum normal diet or an iron-restricted diet for 12weeks. Blood pressure was progressively increased in SHR-SP according to growth, while dietary iron restriction attenuated the development of hypertension. Proteinuria was also increased in SHR-SP according to growth, whereas dietary iron restriction suppressed the increment of proteinuria. SHR-SP exhibited glomerulosclerosis and exacerbated renal interstitial fibrosis at 15weeks old, indicating that SHR-SP developed hypertensive nephropathy in the adult stage; however, these changes were attenuated by dietary iron restriction. Gelatin zymography showed dietary iron restriction decreased both renal MMP-2 and MMP-9 activities in SHR-SP at 15weeks old. Of interest, dietary iron restriction suppressed renal TGFβ-RI expression and Smad2 phosphorylation in SHR-SP. Furthermore, dietary iron restriction decreased renal fibrosis, renal MMP-2 and MMP-9 activities, renal TGFβ-RI expression, and Smad2 phosphorylation in rats with unilateral ureteral obstruction. Dietary iron restriction prevented the development of hypertension in weanling pre-hypertensive rats.

Hepcidin deficiency and iron deficiency do not alter tuberculosis susceptibility in a murine M.tb infection model.

Tuberculosis (TB), caused by the macrophage-tropic pathogen Mycobacterium tuberculosis (M.tb) is a highly prevalent infectious disease. Since an immune correlate of protection or effective vaccine have yet to be found, continued research into host-pathogen interactions is important. Previous literature reports links between host iron status and disease outcome for many infections, including TB. For some extracellular bacteria, the iron regulatory hormone hepcidin is essential for protection against infection. Here, we investigated hepcidin (encoded by Hamp1) in the context of murine M.tb infection. Female C57BL/6 mice were infected with M.tb Erdman via aerosol. Hepatic expression of iron-responsive genes was measured by qRT-PCR and bacterial burden determined in organ homogenates. We found that hepatic Hamp1 mRNA levels decreased post-infection, and correlated with a marker of BMP/SMAD signalling pathways. Next, we tested the effect of Hamp1 deletion, and low iron diets, on M.tb infection. Hamp1 knockout mice did not have a significantly altered M.tb mycobacterial load in either the lungs or spleen. Up to 10 weeks of dietary iron restriction did not robustly affect disease outcome despite causing iron deficiency anaemia. Taken together, our data indicate that unlike with many other infections, hepcidin is decreased following M.tb infection, and show that hepcidin ablation does not influence M.tb growth in vivo. Furthermore, because even severe iron deficiency did not affect M.tb mycobacterial load, we suggest that the mechanisms M.tb uses to scavenge iron from the host must be extremely efficient, and may therefore represent potential targets for drugs and vaccines.

Dietary iron restriction alleviates renal tubulointerstitial injury induced by protein overload in mice

Increased proteinuria causes tubulointerstitial injury due to inflammation in chronic kidney disease (CKD). Iron restriction exhibits protective effects against renal dysfunction; however, its effects against protein overload-induced tubulointerstitial damage remain unclear. Here, we investigated dietary iron restriction effect on tubulointerstitial damage in mice with protein-overload tubulointerstitial injury. Renal tubulointerstitial injury in animal model was induced by intraperitoneal injection of an overdose of bovine serum albumin (BSA). We divided mice into three groups: normal saline + normal diet (ND), BSA + ND, and BSA + iron-restricted diet (IRD). BSA overload induced renal tubulointerstitial injury in the ND mice, which was ameliorated in the IRD mice. Inflammatory cytokines and extracellular matrix mRNA expression was upregulated in BSA + ND mice kidneys and was inhibited by IRD. BSA-induced increase in renal superoxide production, NADPH oxidase activity, and p22phox expression was diminished in the IRD mice. IRD suppression increased BSA-induced renal macrophage infiltration. Moreover, BSA mice exhibited nucleotide-binding oligomerisation domain-like receptor pyrin domain-containing protein (NLRP) inflammasome activation, which was inhibited by IRD. Ferrous iron increased in kidneys with BSA overload and was inhibited by IRD. Thus, iron restriction inhibited oxidative stress and inflammatory changes, contributing to the protective effect against BSA overload-induced tubulointerstitial injury.

Iron-Restricted Diet Affects Brain Ferritin Levels, Dopamine Metabolism and Cellular Prion Protein in a Region-Specific Manner.

Iron is an essential micronutrient for several physiological functions, including the regulation of dopaminergic neurotransmission. On the other hand, both iron, and dopamine can affect the folding and aggregation of proteins related with neurodegenerative diseases, such as cellular prion protein (PrPC) and α-synuclein, suggesting that deregulation of iron homeostasis and the consequential disturbance of dopamine metabolism can be a risk factor for conformational diseases. These proteins, in turn, are known to participate in the regulation of iron and dopamine metabolism. In this study, we evaluated the effects of dietary iron restriction on brain ferritin levels, dopamine metabolism, and the expression levels of PrPC and α-synuclein. To achieve this goal, C57BL/6 mice were fed with iron restricted diet (IR) or with normal diet (CTL) for 1 month. IR reduced iron and ferritin levels in liver. Ferritin reduction was also observed in the hippocampus. However, in the striatum of IR group, ferritin level was increased, suggesting that under iron-deficient condition, each brain area might acquire distinct capacity to store iron. Increased lipid peroxidation was observed only in hippocampus of IR group, where ferritin level was reduced. IR also generated discrete results regarding dopamine metabolism of distinct brain regions: in striatum, the level of dopamine metabolites (DOPAC and HVA) was reduced; in prefrontal cortex, only HVA was increased along with the enhanced MAO-A activity; in hippocampus, no alterations were observed. PrPC levels were increased only in the striatum of IR group, where ferritin level was also increased. PrPC is known to play roles in iron uptake. Thus, the increase of PrPC in striatum of IR group might be related to the increased ferritin level. α-synuclein was not altered in any regions. Abnormal accumulation of ferritin, increased MAO-A activity or lipid peroxidation are molecular features observed in several neurological disorders. Our findings show that nutritional iron deficiency produces these molecular alterations in a region-specific manner and provide new insight into the variety of molecular pathways that can lead to distinct neurological symptoms upon iron deficiency. Thus, adequate iron supplementation is essential for brain health and prevention of neurological diseases.

Iron-restricted pair-feeding affects renal damage in rats with chronic kidney disease.

BackgroundWe have previously shown that dietary iron restriction prevents the development of renal damage in a rat model of chronic kidney disease (CKD). However, iron deficiency is associated with appetite loss. In addition, calorie restriction is reported to prevent the development of end-stage renal pathology in CKD rats. Thus, the beneficial effect of iron restriction on renal damage may depend on calorie restriction. Here, we investigate the effect of pair-feeding iron restriction on renal damage in a rat model of CKD.MethodsFirst, to determine the amount of food intake, Sprague-Dawley (SD) rats were randomly given an ad libitum normal diet or an iron-restricted diet, and the food intake was measured. Second, CKD was induced by a 5/6 nephrectomy in SD rats, and CKD rats were given either a pair-feeding normal or iron-restricted diet.ResultsFood intake was reduced in the iron-restricted diet group compared to the normal diet group of SD rats for 16 weeks (mean food intake; normal diet group and iron-restricted diet group: 25 and 20 g/day, respectively). Based on the initial experiments, CKD rats received either a pair-feeding normal or iron-restricted diet (20 g/day) for 16 weeks. Importantly, pair-feeding iron restriction prevented the development of proteinuria, glomerulosclerosis, and tubulointerstitial damage in CKD rats. Interestingly, pair-feeding iron restriction attenuated renal expression of nuclear mineralocorticoid receptor in CKD rats.ConclusionsPair-feeding iron restriction affected renal damage in a rat model of CKD.

Ferroportin Mediated Control of Iron Metabolism and Disease

Ferroportin, the only known mammalian iron export protein, releases iron from the duodenum, reticuloendothelial system and liver, the sites of iron absorption, recycling and storage respectively. By downregulating ferroportin, the liver-derived hormone hepcidin controls systemic iron availability in response to erythroid demand and inflammation. This ferroportin/hepcidin axis has long been recognized as essential for systemic iron homeostasis. However, both ferroportin and hepcidin are found in tissues not recognized for their role in systemic iron control, such as the heart, the kidney, the brain and the placenta. Co-existence within the same tissue suggests a possible function for hepcidin and ferroportin in local iron homeostasis. However, this hypothesis has not been formally explored. Using mouse models with cardiac-specific manipulation of hepcidin and ferroportin, we have uncovered a role for the cardiac hepcidin/ferroportin axis in cell-autonomous iron homeostasis within cardiomyocytes. Disruption of this cardiac pathway leads to fatal cardiac dysfunction, even against a background of normal systemic iron homeostasis. One the one hand, loss of cardiac ferroportin causes by fatal cardiac iron overload that is preventable by dietary iron restriction 1. On the other hand, loss of cardiac hepcidin or of cardiac hepcidin responsiveness causes fatal cardiomyocyte iron deficiency that is preventable by intravenous iron administration. Comparative study of cardiac iron homeostasis and function in cardiac versus systemic models of ferroportin/hepcidin disruption provides insight into the interplay between systemic and cellular iron homeostasis. A role for the hepcidin/ferroportin axis in cell-autonomous iron control, demonstrated here in the context of the heart, has not previously been described in any other tissue. A pertinent question is whether our findings in the heart extend to other tissues that express both hepcidin and ferroportin, such as the kidney, brain and placenta. Disturbances in iron homeostasis are of clinical importance in cardiovascular disease, renal failure, neurodegeneration and developmental defects. Our findings have two clinically relevant implications: a) that disruption of the local hepcidin/ferroportin axis may in itself have a disease-modifying effect, and b) that therapeutic strategies developed to target the systemic hepcidin/ferroportin axis may have off-target effects relating to local iron control within some tissues.Reference1.Lakhal-Littleton S, Wolna M, Carr C, et al. Cardiac ferroportin regulates cellular iron homeostasis and is important for cardiac function. PNAS. 2015; 10;112(10):3164-3169. DisclosuresNo relevant conflicts of interest to declare.

Low Iron Diet Increases Susceptibility to Noise-Induced Hearing Loss in Young Rats.

We evaluated the role of iron deficiency (ID) without anemia on hearing function and cochlear pathophysiology of young rats before and after noise exposure. We used rats at developmental stages as an animal model to induce ID without anemia by dietary iron restriction. We have established this dietary restriction model in the rat that should enable us to study the effects of iron deficiency in the absence of severe anemia on hearing and ribbon synapses. Hearing function was measured on Postnatal Day (PND) 21 after induction of ID using auditory brainstem response (ABR). Then, the young rats were exposed to loud noise on PND 21. After noise exposure, hearing function was again measured. We observed the morphology of ribbon synapses, hair cells and spiral ganglion cells (SGCs), and assessed the expression of myosin VIIa, vesicular glutamate transporter 3 and prestin in the cochlea. ID without anemia did not elevate ABR threshold shifts, but reduced ABR wave I peak amplitude of young rats. At 70, 80, and 90 dB SPL, amplitudes of wave I (3.11 ± 0.96 µV, 3.52 ± 1.31 µV, and 4.37 ± 1.08 µV, respectively) in pups from the ID group were decreased compared to the control (5.92 ± 1.67 µV, 6.53 ± 1.70 µV, and 6.90 ± 1.76 µV, respectively) (p < 0.05). Moreover, ID without anemia did not impair the morphology hair cells and SGCs, but decreased the number of ribbon synapses. Before noise exposure, the mean number of ribbon synapses per inner hair cell (IHC) was significantly lower in the ID group (8.44 ± 1.21) compared to that seen in the control (13.08 ± 1.36) (p < 0.05). In addition, the numbers of ribbon synapses per IHC of young rats in the control (ID group) were 6.61 ± 1.59, 3.07 ± 0.83, 5.85 ± 1.63 and 12.25 ± 1.97 (3.75 ± 1.45, 2.03 ± 1.08, 3.81 ± 1.70 and 4.01 ± 1.65) at 1, 4, 7 and 14 days after noise exposure, respectively. Moreover, ABR thresholds at 4 and 8 kHz in young rats from the ID group were significantly elevated at 7 and 14 days after noise exposure compared to control (p < 0.05). The average number of young rat SGCs from the ID group were significantly decreased in the basal turn of the cochlea compared to the control (p < 0.05). Therefore, ID without anemia delayed the recovery from noise-induced hearing loss and ribbon synapses damage, increased SGCs loss, and upregulated prestin after noise exposure. Thus, the cochleae in rat pups with ID without anemia were potentially susceptible to loud noise exposure, and this deficit may be attributed to the reduction of ribbon synapses and SGCs.

Attenuation of hypertension and renal damage in renovascular hypertensive rats by iron restriction.

Iron is a catalyst in the formation of reactive oxygen species. Oxidative stress is associated with the pathogenesis of both human and experimental animal models of renovascular hypertension. We hypothesized that iron is involved in the pathogenesis of renovascular hypertension and that iron restriction may affect the pathogenesis of renovascular hypertension via the inhibition of oxidative stress. Herein, we investigated the effect of iron restriction on hypertension and renal damage in a rat model of two-kidney one-clip (2K1C) renovascular hypertension. Renovascular hypertension was induced by 2K1C in male Sprague-Dawley rats. At the day of clipping, 2K1C rats were divided into untreated (2K1C) and dietary iron-restricted groups (2K1C+IR). The 2K1C rats showed hypertension after the day of clipping, whereas dietary iron restriction attenuated the development of hypertension. Vascular hypertrophy and the increased fibrotic area were suppressed in the 2K1C+IR group. The clipped kidney developed renal atrophy in both the 2K1C and 2K1C+IR groups after clipping. However, the unclipped kidney showed renal hypertrophy in the 2K1C and 2K1C+IR groups, and the extent was less in the 2K1C+IR group. The 2K1C rats exhibited glomerulosclerosis and tubulointerstitial fibrosis in the unclipped kidney, whereas these changes were attenuated by an iron-restricted diet. Importantly, proteinuria was decreased in the 2K1C+IR group, along with decreased urinary 8-hydroxy-2'-deoxyguanosine excretion and superoxide production of the unclipped kidney. Moreover, the expression of nuclear mineralocorticoid receptor in the unclipped kidney of the 2K1C rats was attenuated by iron restriction. These data indicate a novel effect of iron restriction on hypertension and renal damage in renovascular hypertension.

Deferoxamine ameliorates hepatosteatosis via several mechanisms in ob/ob mice.

Hepatic iron accumulation may be responsible for the pathology of nonalcoholic fatty liver disease (NAFLD), which is both increasingly prevalent in conjunction with obesity and associated with comorbidities. The efficacy of iron reduction therapies, such as phlebotomy or dietary iron restriction, has been demonstrated in patient and animal models, including models of diabetes and obesity; however, the effects on and exact mechanisms responsible for iron depletion in NAFLD have not been clearly elucidated. Our study investigated the role of iron depletion by deferoxamine (DFO) treatment of ob/ob mice with hepatic steatosis. We found that DFO reduced hepatic iron deposition and regulated intracellular iron concentration in a homeostatic process following 15 days of treatment. Compared with vehicle treatment, DFO significantly improved hepatic steatosis by upregulating proteins related to lipid metabolism. Meanwhile, the reduction of free radical formation and proinflammatory cytokines, as well as the increase of hypoxia-inducible factor-1α pathway proteins and Bcl2/Bax ratio, further indicated that DFO was effective for liver protection and hepatic adaptation. These findings show that the intraperitoneal delivery of DFO provides a potential means of both preventing the progression of NAFLD and accelerating healing of hepatic steatosis, with the potential for rapid clinical application.

Iron is associated with the development of hypoxia-induced pulmonary vascular remodeling in mice.

Several recent observations provide the association of iron deficiency with pulmonary hypertension (PH) in human and animal studies. However, it remains completely unknown whether PH leads to iron deficiency or iron deficiency enhances the development of PH.In addition, it is obscure whether iron is associated with the development of pulmonary vascular remodeling in PH. In this study, we investigate the impacts of dietary iron restriction on the development of hypoxia-induced pulmonary vascular remodeling in mice. Eight- to ten-week-old male C57BL/6J mice were exposed to chronic hypoxia for 4weeks. Mice exposed to hypoxia were randomly divided into two groups and were given a normal diet or an iron-restricted diet. Mice maintained in room air served as normoxic controls. Chronic hypoxia induced pulmonary vascular remodeling, while iron restriction led a modest attenuation of this change. In addition, chronic hypoxia exhibited increased RV systolic pressure, which was attenuated by iron restriction. Moreover, the increase in RV cardiomyocyte cross-sectional area and RV interstitial fibrosis was observed in mice exposed to chronic hypoxia. In contrast, iron restriction suppressed these changes. Consistent with these changes, RV weight to left ventricular+interventricular septum weight ratio was increased in mice exposed to chronic hypoxia, while this increment was inhibited by iron restriction. Taken together, these results suggest that iron is associated with the development of hypoxia-induced pulmonary vascular remodeling in mice.

Cardiac remodeling in response to chronic iron deficiency: role of the erythropoietin receptor.

Anemia is a common comorbidity of patients with heart failure, and iron deficiency is known as one of the causes of anemia in heart failure. Recent studies have shown that iron deficiency alone, without overt anemia, is associated with poor outcomes in patients with heart failure. Thus, to minimize the mortality in patients with heart failure, it is important to understand the link between iron deficiency and cardiac function. Chronic untreated iron deficiency results in cardiac remodeling, and we have previously reported that erythropoietin (Epo) and cardiac Epo receptor (EpoR) signaling may be associated with its remodeling. However, the link between EpoR signaling and its remodeling remains to be elucidated. Herein, we investigated the role of EpoR signaling on cardiac remodeling in response to chronic iron deficiency. Wild-type mice and transgene-rescued EpoR-null mutant mice, which express EpoR only in the hematopoietic lineage (EpoR-restricted mice), were fed with either a normal or an iron-restricted diet, and the molecular mechanisms were investigated. Dietary iron restriction gradually induced anemia, Epo secretion, and cardiac hypertrophy in wild-type mice. In contrast, EpoR-restricted mice fed with an iron-restricted diet exhibited anemia, left ventricular dilatation, and cardiac dysfunction compared with wild-type mice. Interestingly, altered cardiac mitochondrial biogenesis was observed in EpoR-restricted mice following iron deficiency. Moreover, cardiac p53 expression was increased in EpoR-restricted mice compared with wild-type mice following iron deficiency. These data indicate that EpoR signaling is associated with cardiac remodeling following chronic iron deficiency.

Iron Deprivation May Enhance Insulin Receptor and Glut4 Transcription in Skeletal Muscle of Adult Rats.

Considering that phenotype related to iron overload associated with pathological conditions differs from that caused by dietary iron excess, our study set out to evaluate the impact of dietary iron restriction and dietary iron supplementation on oxidative stress and functional outcome in adult, healthy rats. adult rats were divided into the three groups and fed diets containing 10, 35 or 350 mg/kg iron (restricted-diet, control-diet and supplemented- diet groups, respectively) for 78 days. Hematological variables, fasting blood glucose, hepatic enzyme activity and C-reactive protein levels were analyzed. Iron and glycogen concentrations in liver and skeletal muscle were determined. The extent of tissue damage caused by either dietary iron restriction or iron supplementation was accessed by measuring malondialdehyde, carbonyl, NADPH oxidase, glutathione peroxidase, glutathione reductase and glutathione-s-transferase in various tissues. The mRNA expression levels of insulin receptor, glucose transporter 4 and p53 were also determined. Fasting blood glucose values trended toward a decrease by dietary iron restriction, moreover, hepatic glycogen content decreased with concomitant increases in skeletal muscle. In addition, dietary iron restriction resulted in a twofold increase in mRNA expression of Insr and fourfold increase in Glut4 expression in skeletal muscle. Although the dietary iron restriction did not affect body iron status, it caused hepatic low oxidative damages. However, high liver NADPH oxidase activity and increased levels of protein oxidation in muscle were observed. Chronic feeding of high iron diet induces iron overload and resulted in elevated levels of stress markers in tissues. Dietary iron deprivation may improve insulin receptor and glucose transporter transcription in muscle; however, our results show that dietary iron restriction can prevent and/or promote oxidative damage in a tissue-specific manner, emphasizing the importance of maintaining optimal iron intake.

Temporary dietary iron restriction affects the process of thrombus resolution in a rat model of deep vein thrombosis.

BackgroundDeep vein thrombosis (DVT) is a major cause of pulmonary thromboembolism and sudden death. Thus, it is important to consider the pathophysiology of DVT. Recently, iron has been reported to be associated with thrombotic diseases. Hence, in this study, we investigate the effects of dietary iron restriction on the process of thrombus resolution in a rat model of DVT.MethodsWe induced DVT in 8-week-old male Sprague-Dawley rats by performing ligations of their inferior venae cavae. The rats were then given either a normal diet (DVT group) or an iron-restricted diet (DVT+IR group). Thrombosed inferior venae cavae were harvested at 5 days after ligation.ResultsThe iron-restricted diet reduced venous thrombus size compared to the normal diet. Intrathrombotic collagen content was diminished in the DVT+IR group compared to the DVT group. In addition, intrathrombotic gene expression and the activity of matrix metalloproteinase-9 were increased in the DVT+IR group compared to the DVT group. Furthermore, the DVT+IR group had greater intrathrombotic neovascularization as well as higher gene expression levels of urokinase-type plasminogen activator and tissue-type plasminogen activator than the DVT group. The iron-restricted diet decreased intrathrombotic superoxide production compared to the normal diet.ConclusionsThese results suggest that dietary iron restriction affects the process of thrombus resolution in DVT.

Aortic Iron Overload With Oxidative Stress and Inflammation in Human and Murine Abdominal Aortic Aneurysm

Although iron is an essential element for maintaining physiological function, excess iron leads to tissue damage caused by oxidative stress and inflammation. Oxidative stress and inflammation play critical roles for the development of abdominal aortic aneurysm (AAA). However, it has not been investigated whether iron plays a role in AAA formation through oxidative stress and inflammation. We, therefore, examined whether iron is involved in the pathophysiology of AAA formation using human AAA walls and murine AAA models. Human aortic walls were collected from 53 patients who underwent cardiovascular surgery (non-AAA=34; AAA=19). Murine AAA was induced by infusion of angiotensin II to apolipoprotein E knockout mice. Iron was accumulated in human and murine AAA walls compared with non-AAA walls. Immunohistochemistry showed that both 8-hydroxy-2'-deoxyguanosine and CD68-positive areas were increased in AAA walls compared with non-AAA walls. The extent of iron accumulated area positively correlated with that of 8-hydroxy-2'-deoxyguanosine expression area and macrophage infiltration area in human and murine AAA walls. We next investigated the effects of dietary iron restriction on AAA formation in mice. Iron restriction reduced the incidence of AAA formation with attenuation of oxidative stress and inflammation. Aortic expression of transferrin receptor 1, intracellular iron transport protein, was increased in human and murine AAA walls, and transferrin receptor 1-positive area was similar to areas where iron accumulated and F4/80 were positive. Iron is involved in the pathophysiology of AAA formation with oxidative stress and inflammation. Dietary iron restriction could be a new therapeutic strategy for AAA progression.

Iron restriction inhibits renal injury in aldosterone/salt-induced hypertensive mice

Excess iron is associated with the pathogenesis of several renal diseases. Aldosterone is reported to have deleterious effects on the kidney, but there have been no reports of the role of iron in aldosterone/salt-induced renal injury. Therefore, we investigated the effects of dietary iron restriction on the development of hypertension and renal injury in aldosterone/salt-induced hypertensive mice. Ten-week-old male C57BL/6J mice were uninephrectomized and infused with aldosterone for four weeks. These were divided into two groups: one fed a high-salt diet (Aldo) and the other fed a high-salt with iron-restricted diet (Aldo-IR). Vehicle-infused mice without a uninephrectomy were also divided into two groups: one fed a normal diet (control) and the other fed an iron-restricted diet (IR) for 4 weeks. As compared with control and IR mice, Aldo mice showed an increase in both systolic blood pressure and urinary albumin/creatinine ratio, but these increases were reduced in the Aldo-IR group. In addition, renal histology revealed that Aldo mice exhibited glomerulosclerosis and tubulointerstitial fibrosis, whereas these changes were attenuated in Aldo-IR mice. Expression of intracellular iron transport protein transferrin receptor 1 was increased in the renal tubules of Aldo mice compared with control mice. Dietary iron restriction attenuated the development of hypertension and renal injury in aldosterone/salt-induced hypertensive mice.

Association of dietary iron restriction with left ventricular remodeling after myocardial infarction in mice

Several epidemiologic studies have reported that body iron status and dietary iron intake are related to an increased risk of acute myocardial infarction (MI). However, it is completely unknown whether dietary iron reduction impacts the development of left ventricular (LV) remodeling after MI. Here, we investigate the effect of dietary iron restriction on the development of LV remodeling after MI in an experimental model. MI was induced in C57BL/6 J mice (9-11 weeks of age) by the permanent ligation of the left anterior descending coronary artery (LAD). At 2 weeks after LAD ligation, mice were randomly divided into two groups and were given a normal diet or an iron-restricted diet for 4 weeks. Sham operation without LAD ligation was also performed as controls. MI mice exhibited increased LV dilatation and impaired LV systolic function that was associated with cardiomyocyte hypertrophy and interstitial fibrosis in the remote area, as compared with the controls at 6 weeks after MI. In contrast, dietary iron restriction attenuated LV dilatation and impaired LV systolic function coupled to cardiomyocyte hypertrophy and interstitial fibrosis in the remote area. Importantly, cardiac expression of cellular iron transport proteins, transferrin receptor 1 and divalent metal transporter 1 was increased in the remote area of MI mice compared with the controls. Dietary iron restriction attenuated the development of LV remodeling after MI in mice. Cellular iron transport might play a role in the pathophysiological mechanism of LV remodeling after MI.

A long-term high-fat diet changes iron distribution in the body, increasing iron accumulation specifically in the mouse spleen.

Although iron is an essential trace metal, its presence in excess causes oxidative stress in the human body. Recent studies have indicated that iron storage is a risk factor for type 2 diabetes mellitus. Dietary iron restriction or iron chelation ameliorates symptoms of type 2 diabetes in mouse models. However, whether iron content in the body changes with the development of diabetes is unknown. Here, we investigated the dynamics of iron accumulation and changes in iron absorption-related genes in mice that developed obesity and diabetes by consuming a high-fat diet (HFD-fed mice). HFD-fed mice (18-20 wk) were compared with control mice for hematologic features, serum ferritin levels, and iron contents in the gastrocnemius muscle, heart, epididymal fat, testis, liver, duodenum, and spleen. In addition, the spleen was examined histologically. Iron absorption-related gene expression in the liver and duodenum was also examined. Hemoglobin and serum ferritin levels were increased in HFD-fed mice. The HFD-fed mice showed iron accumulation in the spleen, but not in the heart or liver. Increased percentages of the splenic red pulp and macrophages were observed in HFD-fed mice and iron accumulation in the spleen was found mainly in the splenic red pulp. The HFD-fed mice also showed decreased iron content in the duodenum. The mRNA expression of divalent metal transporter-1 (DMT-1), an iron absorption-related gene, was elevated in the duodenum of HFD-fed mice. These results indicate that iron accumulation (specifically accumulation in the spleen) is enhanced by the development of type 2 diabetes induced by HFD.