Iron-restricted Diet Research Articles

Consequences of Maternal Iron‐Deficiency: Fetal Anemia, and Hypoxia in Select Tissues of the Conceptus

Iron deficiency (ID) is the most prevalent nutritional deficiency worldwide, and affects populations across the socio‐economic spectrum. The incidence of ID anemia in pregnant women is of chief concern, with rates as high as 80 % in developing countries, and a prevalence of 23 % in Canada. Maternal ID during pregnancy has been shown to impact fetal iron status and growth trajectories, although the precise mechanisms underlying this intrauterine growth restriction are unknown. We hypothesized that maternal ID causes fetal hypoxia, resulting in asymmetric growth restriction in more severe ID. Female rats (initial age 6 weeks for severe (S‐ID), 9 weeks for moderate (M‐ID)) were fed iron restricted diets throughout pregnancy. Dams were treated with pimonidazole on gestational day (GD)20 to assess hypoxia (<10 mm Hg O2). Maternal and fetal hemoglobin (Hb) levels and body weights were assessed on GD21. Pup organs were excised and weighed to assess degree of asymmetric growth restriction. Tissue immunofluorescence was carried out on GD21 pup tissues with fluorescent probes to detect covalent pimonidazole adducts. Both M‐ and S‐ID groups caused fetal anemia ( fetal Hb reduced 30.9 % and 63.8 %, respectively, compared to control of 9.27 g/dL, P<0.01), despite only S‐ID dams becoming anemic (reduction in Hb 10.9 % and 50.3 % for M‐ and S‐ID, respectively, compared to 13.60 g/dL control Hb, P<0.001). Both M‐ and S‐ID caused fetal growth restriction (reduced weight by 9.1 % and 34.9 %, respectively, compared to control weight of 5.45 g, P<0.001). S‐ID, but not M‐ID, caused asymmetrical growth restriction, wherein relative heart and brain weights increased (60 % and 9.8 %, respectively, compared to control weights of 0.005 g/g and 0.041 g/g, P<0.01), and kidney and liver size decreased (20 % and 15 %, respectively, compared to control weights of 0.005 g/g and 0.067 g/g, P<0.01). M‐ID was associated with increased staining for pimonidazole in fetal livers (P=0.01) and kidneys (P=0.01), but not in brains and placentae, versus respective controls; hypoxia staining is S‐ID fetuses is ongoing. Fetal anemia and growth restriction can occur in the absence of maternal anemia, emphasizing the need for new methods of assessment in the clinical setting. Elucidation of the mechanisms through which ID impacts fetal growth and development will enable us to develop a platform with which we can test novel therapeutics.Support or Funding InformationAW held an Alberta Innovates Health Solutions Summer Studentship. SB Holds grants from the Canadian Institute of Health Research (CIHR), and the Women and Children's Health Research Institute (WCHRI).

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.

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.

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.

Abstract 14771: Iron Plays a Role in the Thrombus Formation of a Rat Model of Deep Vein Thrombosis

Introduction: Deep vein thrombosis (DVT) is a major cause of pulmonary thromboembolism. On the other hand, iron is an essential element for maintaining physiological homeostasis in the body, whereas excess iron leads to toxicity and free radical damage. Hence, iron accumulation is associated with the pathogenesis of several cardiovascular diseases. We have previously reported that dietary iron restriction (IR) prevents the development of hypertension and chronic kidney disease (CKD) in animal models of several cardiovascular diseases. In addition, we have shown that intracellular iron transport protein, transferrin receptor 1 (TfR1), is involved in the mechanism of several cardiovascular diseases. In this study, we hypothesized that iron and TfR1 play roles in the pathogenesis of DVT. We investigate the effects of dietary IR on the thrombus formation and resolution in a rat model of DVT. Methods: We performed the ligation of inferior vena cava to induce DVT in 8-week-old male Sprague-Dawley rats. After the ligation, rats were given either a normal diet (DVT group) or iron-restricted diet (DVT+IR group). The thrombus was harvested at 5days after the ligation. Results: At first, TfR1 expression was increased in the thrombus of a rat model of DVT, suggesting that iron and TfR1 play roles in the pathogenesis of DVT. Interestingly, dietary iron restriction reduced the thrombus size compared with the DVT group (180±17 mg/cm vs 135±13 mg/cm, DVT vs DVT+IR ; p < 0.05). Intrathrombotic collagen content was diminished in the DVT+IR group compared with the DVT group. Intrathrombotic gene expression and activity of matrix metalloproteinase (MMP)-9 were increased in the DVT+IR group compared with the DVT group. In addition, the numbers of CD68 positive cells in the thrombus were fewer in the DVT+IR group than in the DVT group. Furthermore, intrathrombotic gene expression of urokinase-type plasminogen activator and tissue-type plasminogen activator was higher in the DVT+IR group than in the DVT group. Conclusions: These results suggest that iron plays a role in the pathogenesis of DVT.

Serum ferritin and liver inflammation: which come first? Chicken or egg?

Beaton MD, et al. For the clinician the serum elevation of ferritin in chronic liver diseases (CLD), with the exception of Emochromatosis, is always assumed to be a non-specific marker of hepatic inflammation and not of iron overload.1 Nonalcoholic fatty liver disease (NAFLD) is now the most common cause of CLD.2 The spectrum of NAFLD is ranging from simple steatosis through nonalcoholic steatohepatitis (NASH).3 Iron is regarded as a putative element that interacts with oxygen radicals, and high rates of hyperferritinemia together with increased hepatic iron stores have been demonstrated in NASH.4 The role of hepatic iron in the progression and in the pathogenesis of NASH remains controversial. We know that phlebotomy or iron-restricted diet reduce hepatic damage as well as insulin resistance (IR) in patients with NAFLD/NASH.5 However, exact mechanisms involved in iron accumulation in NASH remain to be clarified. Genetic factors linked to IR, dysregulation of iron-regulatory molecules, erythrophagocytosis by Kupffer cells may be responsible for hepatic iron overload in NASH.6,7 Chakraberti and Adams, in this Annals of Hepatology issue8 reported a simple clinical study of phlebotomy treatment performed in a series of 56 patients with histology-proven NAFLD. Liver biopsy and liver iron concentration (LIC) was performed at entry and 6 months after phlebotomy. They did not found any significant correlation between LIC and the level of serum ferritin, hepatic inflammation as measured by NAS score, and other generic markers of inflammation, such as ESR and CRP. The authors concluded that elevated serum ferritin is linked to hepatic iron accumulation, but that liver inflammation is not the cause of increased serum ferritin in patients with NAFLD. Other authors7 recently demonstrated that elevated ferritin level reflects iron stores, and not hepatic inflammation, being a predictors of vascular damage in NAFLD. The mechanisms involved in iron accumulation in NAFLD, and in inducing IR, metabolic, hepatic, and vascular damage by iron accumulation are not yet well know and should be further investigated. They may involve up-regulation of hepcidin by increased iron stores in patients not carrying HFE mutations, and iron compartmentalization into macrophages, or the hepatic expressions of the ironexport protein ferroportin-1, and of the iron-sensing molecule hemojuvelin, which have been found to be significantly lower in NAFLD patients.7,9 Whatever the mechanisms should be the only certainty is that an increased serum ferritin level in NAFLD is not a marker or a cause of inflammation, but a consequence of iron accumulation inside the hepatocyte.

Increased Renal Iron Accumulation in Hypertensive Nephropathy of Salt-Loaded Hypertensive Rats

Although iron is reported to be associated with the pathogenesis of chronic kidney disease, it is unknown whether iron participates in the pathophysiology of nephrosclerosis. Here, we investigate whether iron is involved in the development of hypertensive nephropathy and the effects of iron restriction on nephrosclerosis in salt- loaded stroke-prone spontaneously hypertensive rats (SHRSP). SHRSP were given either a normal or high-salt diet for 8 weeks. Another subset of SHRSP were fed a high-salt with iron-restricted diet. SHRSP given a high-salt diet developed severe hypertension and nephrosclerosis. As a result, survival rate was decreased after 8 weeks diet. Importantly, massive iron accumulation and increased iron content were observed in the kidneys of salt-loaded SHRSP, along with increased superoxide production, urinary 8-Hydroxy-2′-deoxyguanosine excretion, and urinary iron excretion; however, these changes were markedly attenuated by iron restriction. Of interest, expression of cellular iron transport proteins, transferrin receptor 1 and divalent metal transporter 1, was increased in the tubules of salt-loaded SHRSP. Notably, iron restriction attenuated the development of severe hypertension and nephrosclerosis, thereby improving survival rate in salt-loaded SHRSP. Taken together, these results suggest a novel mechanism by which iron plays a role in the development of hypertensive nephropathy and establish the effects of iron restriction on salt-induced nephrosclerosis.

Dietary iron restriction prevents further deterioration of renal damage in a chronic kidney disease rat model

Iron accumulation is associated with the pathogenesis of chronic kidney disease (CKD). However, little is known about the effects of isolated iron restriction against CKD. We have recently reported that iron restriction prevents the development of renal damage in the well established 5/6 nephrectomy rat model of CKD. Herein, we investigated the therapeutic effects of iron restriction on preexisting hypertension and renal damage in a rat model of CKD. CKD was induced by 5/6 nephrectomy in Sprague-Dawley rats. After surgery, 5/6 nephrectomized rats were given an iron-restricted diet from 1 day to 16 weeks for prevention protocol or from 8 to 16 weeks for rescue protocol. Other CKD rats were given a normal diet. At 16 weeks after surgery, CKD rats developed hypertension and renal damage. Early intervention with iron restriction prevented the development of hypertension and vascular remodeling. By contrast, late intervention with iron restriction did not remarkably ameliorate preexisting hypertension and vascular remodeling in CKD rats. On the contrary, late intervention with iron restriction prevented further progression of preexisting renal damage in CKD rats. Interestingly, iron restriction led to increased urinary sodium and decreased urinary potassium excretions in CKD rats. Moreover, iron restriction markedly attenuated renal expression of nuclear mineralocorticoid receptor and Rac1 activity in CKD rats. Iron restriction prevented further deterioration of preexisting renal damage. The beneficial effects of iron restriction on renal damage seem to be associated with inhibition of renal mineralocorticoid receptor signaling.

Impact of dietary iron restriction on the development of monocrotaline-induced pulmonary vascular remodeling and right ventricular failure in rats

Pulmonary hypertension (PH) is characterized by pulmonary vascular remodeling leading to right ventricular (RV) failure. Recently, iron deficiency is reported to be prevalent in patients with PH. However, the mechanism by which iron deficiency occurs in patients with PH remains unknown. Here, we investigated the effects of dietary iron restriction on the development of monocrotaline-induced pulmonary vascular remodeling and the involved mechanisms. Male Sprague–Dawley rats were subcutaneously injected with monocrotaline (60mg/kg). Afterwards, monocrotaline-injected rats were randomly divided into two groups and were given a normal diet (n=6) or an iron-restricted diet (n=6) for 4weeks. Saline-injected rats given a normal diet were served as controls (n=6). Monocrotaline-injected rats showed pulmonary vascular remodeling, increased RV pressure, RV hypertrophy, and decreased RV ejection fraction, followed by RV failure after 4weeks. In contrast, iron restriction attenuated the development of pulmonary vascular remodeling and RV failure. Of interest, expression of cellular iron transport protein, transferrin receptor 1 was increased in the pulmonary remodeled artery and the failing right ventricle of monocrotaline-injected rats, as compared with the controls. Moreover, a key regulator of iron homeostasis, hepcidin gene expression was increased in the failing right ventricle of monocrotaline-injected rats. Iron restriction attenuated the development of monocrotaline-induced pulmonary vascular remodeling and RV failure. Cellular iron transport might be involved in the pathophysiology of PH and PH induced RV failure.

Abstract 8857: Iron Restriction Prevents the Progression of Diabetic Nephropathy in db/db Mice

Background: Iron is well-known as an essential trace metal for all living body. On the other hand, excess iron causes oxidative stress through catalyzing highly toxic hydroxyl radical via Fenton/Haber-Weiss reaction. Recent studies have shown that iron reduction ameliorates diabetic condition and prevents development of adipocyte hypertrophy. In the present study, we studied the protective effect of iron reduction against diabetic nephropathy, a diabetic complication. Methods and Results: We employed male db/db mice, a model of diabetic nephropathy and male db/m mice as control mice. db/db mice were divided into 2 groups, normal diet group (ND)(Fe 100mg/kg food) and iron-restricted diet group (ID)(Fe 10mg/kg food). Urinary albumin excretion was increasing in db/db mice with ND compared with db/m mice with ND, which was suppressed in db/db mice with ID. In renal histological analysis, increased glomerular volume and mesangial area expansion in ND-fed db/db mice were reduced in ID-fed db/db mice. Deposition of extracelluar matrix such as collagen IV and fibronectin was decreased in db/db mice with ID. Upregulated glomerular desmin expression in db/db mice with ND was diminished in db/db mice with ID. In terms with oxidative stress, ID diminished increased superoxide production in kidney of ND-fed db/db mice. Renal expression of NADPH oxidase components, p22phox and NOX4, were augmented in db/db mice with ND-fed, which was abolished by ID-fed. In terms with renal iron transporter, there were no differences of expression of transferrin receptor and divalent metal transporter-1 between db/m mice and db/db mice, increased ferroportin expression in db/db mice with ND- was reversed by ID to the same level as db/m mice. Urinary iron excretion was significantly higher in ND-fed db/db mice, which was reduced with ID administration. Conclusion: These findings suggest that dietary iron restriction exert preventive effect against development of diabetic nephropathy via the reduction of oxidative stress.

Effect of iron restriction on renal damage and mineralocorticoid receptor signaling in a rat model of chronic kidney disease

Iron is associated with the pathogenesis of chronic kidney disease (CKD). Activation of mineralocorticoid receptor signaling is implicated in CKD; however, a link between iron and mineralocorticoid receptor signaling in CKD remains unknown. We have previously shown that long-term dietary iron restriction leads to increased sodium and decreased potassium excretions in the rat urine. Herein, we investigated the effect of iron restriction on renal damage and mineralocorticoid receptor signaling in a rat model of CKD. CKD was induced by 5/6 nephrectomy in Sprague-Dawley rats. CKD rats were divided into untreated and dietary iron-restricted groups. CKD rats exhibited proteinuria, glomerulosclerosis, tubulointerstitinal damage, and podocyte injury. In contrast, these changes were attenuated by 16 weeks of iron-restricted diet. Consistent with these findings, iron restriction suppressed increased gene expression of collagen type III, transforming growth factor-β, CD68, and tumor necrosis factor-α in the CKD kidney. Importantly, increased expression of nuclear mineralocorticoid receptor and SGK1, a key downstream effector of mineralocorticoid receptor signaling, in the CKD kidney was markedly attenuated by iron restriction. Of interest, expression of cellular iron transport proteins, transferrin receptor 1, and divalent metal transporter 1 was increased in the CKD renal tubules, along with increased iron accumulation, superoxide production, and urinary iron excretion. CKD rats also developed hypertension, although iron restriction suppressed the development of hypertension. Taken together, these data uncover a novel effect of iron restriction on renal damage and hypertension through the inhibition of renal mineralocorticoid receptor signaling.

Perinatal Iron Deficiency Combined with a High-Fat Diet Causes Obesity and Cardiovascular Dysregulation

Consumption of a high-fat Western diet (WD) and the resultant obesity is linked to a number of chronic pathologies, including cardiovascular dysregulation. The purpose of the present study was to determine whether perinatal iron deficiency (PID) added to the consumption of a WD would precipitate an obese phenotype with exacerbated metabolic and cardiovascular outcomes in adult offspring. Female Sprague Dawley rats were fed either a control (225 mg/kg Fe) or an iron-restricted diet (3-10 mg/kg Fe) prior to and throughout gestation. At birth, all dams were fed an iron-replete diet. At weaning, offspring were fed a normal diet or WD for up to 21 wk. Hemodynamics and locomotor activity were assessed by radiotelemetry starting at 15 wk of age. Iron restriction during pregnancy caused severe anemia in dams and offspring, resulting in 15% lower birth weights in the offspring. PID offspring fed the WD had greater caloric intake and exhibited reduced locomotor activity compared with their normal diet-fed littermates; no such effects were observed in normal iron control offspring. Despite having a similar effect on serum lipid profiles, consumption of the WD had a greater impact on body weight in the PID group, and this weight gain was due largely to visceral adipose tissue accumulation. A significant correlation between visceral adipose tissue weight and mean arterial pressure was observed in the PID offspring but not in controls. These observations demonstrate that PID predisposes offspring to an enhanced response to WD characterized by increased fat accumulation and cardiovascular dysregulation.

Dietary Iron Restriction Prevents Hypertensive Cardiovascular Remodeling in Dahl Salt-Sensitive Rats

Iron accumulation is associated with the pathogenesis of several cardiovascular diseases. However, the preventive effects of iron restriction (IR) against cardiovascular disease remain obscure. We investigated the effects of dietary IR on cardiovascular pathophysiology and the involved mechanism in Dahl salt-sensitive rats. Dahl salt-sensitive rats were provided either a normal or high-salt (HS) diet. Another subset of Dahl salt-sensitive rats were fed an HS with iron-restricted (HS+IR) diet for 11 weeks. Dahl salt-sensitive rats given an HS diet developed hypertension, heart failure, and decreased a survival rate after 11 weeks on the diet. In contrast, IR attenuated the development of hypertension and heart failure, thereby improving survival rate. Dietary IR suppressed cardiovascular hypertrophy, fibrosis, and inflammation in HS rats. The phosphorylation of Akt, AMP-activated protein kinase, and endothelial nitric oxide synthase was decreased in the aorta of HS rats, whereas they were ameliorated by the IR diet. Aortic expression of the cellular iron import protein transferrin receptor 1, and the iron storage protein ferritin H-subunit, was upregulated in HS rats. IR also attenuated proteinuria and increased oxidative stress in the HS group. N(G)-nitro-L-arginine methyl ester abolished the beneficial effects of IR and decreased survival rate in HS+IR rats. Dietary IR had protective effects on salt-induced hypertension, cardiovascular remodeling, and proteinuria through the inhibition of oxidative stress, and maintenance of Akt, AMP-activated protein kinase, and endothelial nitric oxide synthase in the aorta. IR could be an effective strategy for prevention of HS-induced organ damage in salt-sensitive hypertensive patients.

Alopecia in IL‐10‐deficient mouse pups is c‐kit‐dependent and can be triggered by iron deficiency

Hair loss (alopecia) can result from a variety of metabolic, endocrine, immunologic, and environmental causes. This investigation was undertaken to determine the mechanisms underlying the sporadic development of alopecia in litters from C57BL/6 interleukin-10-deficient (Il10(-/-)) mice. All pups in affected litters demonstrated alopecia by postnatal days 17-19, with hair loss from their trunks but not from their head, base of tail, or feet. Histopathology revealed distorted hair follicles containing broken hair shafts and prominent dermal infiltrates containing increased numbers of activated mast cells. Hair re-growth began soon after weaning, suggesting that the alopecia was triggered by factors transmitted during lactation. Milk from Il10(-/-) dams induced macrophage secretion of pro-inflammatory cytokines in vitro regardless of whether or not their pups developed alopecia. Feeding dams a diet containing 3-6 ppm iron increased the percentage of litters with alopecia to 100% for pups with mast cells, with 0% alopecia in mast cell-deficient pups. When dams were fed a diet containing 131 ppm iron, significantly lower haemoglobin and hematocrit values were observed in pups from litters with alopecia (71%; 5 of 7 litters) compared to litters without alopecia. Genetic or pharmacologic inhibition of c-kit that resulted in depletion of mast cells in pups prevented hair loss in at-risk litters. These studies demonstrate that maternal iron-restricted diets enhance the incidence of alopecia in IL-10-deficient mouse pups and suggest mast cells as potential effector cells. Further studies are indicated to further explore the mechanisms involved and to determine how mast cells may contribute to alopecia in humans.

Dietary iron restriction or iron chelation protects from diabetes and loss of β-cell function in the obese (ob/ob lep−/−) mouse

Iron overload can cause insulin deficiency, but in some cases this may be insufficient to result in diabetes. We hypothesized that the protective effects of decreased iron would be more significant with increased beta-cell demand and stress. Therefore, we treated the ob/ob mouse model of type 2 diabetes with an iron-restricted diet (35 mg/kg iron) or with an oral iron chelator. Control mice were fed normal chow containing 500 mg/kg iron. Neither treatment resulted in iron deficiency or anemia. The low-iron diet significantly ameliorated diabetes in the mice. The effect was long lasting and reversible. Ob/ob mice on the low-iron diet exhibited significant increases in insulin sensitivity and beta-cell function, consistent with the phenotype in mouse models of hereditary iron overload. The effects were not accounted for by changes in weight or feeding behavior. Treatment with iron chelation had a more dramatic effect, allowing the ob/ob mice to maintain normal glucose tolerance for at least 10.5 wk despite no effect on weight. Although dietary iron restriction preserved beta-cell function in ob/ob mice fed a high-fat diet, the effects on overall glucose levels were less apparent due to a loss of the beneficial effects of iron on insulin sensitivity. Beneficial effects of iron restriction were minimal in wild-type mice on normal chow but were apparent in mice on high-fat diets. We conclude that, even at "normal" levels, iron exerts detrimental effects on beta-cell function that are reversible with dietary restriction or pharmacotherapy.

Iron Overload Is Associated with Hepatic Oxidative Damage to DNA in Nonalcoholic Steatohepatitis

Several lines of evidence have suggested that oxidative stress plays an important role for the pathogenesis of nonalcoholic steatohepatitis (NASH). Therefore, by using immunohistochemical staining of liver biopsy samples, we measured hepatic 7,8-dihydro-8-oxo-2' deoxyguanosine (8-oxodG), a DNA base-modified product generated by hydroxyl radicals, of 38 NASH patients and compared with 24 simple steatosis and 10 healthy subjects. Relation of hepatic 8-oxodG with clinical, biochemical, and histologic variables and changes after iron reduction therapy (phlebotomy plus iron-restricted diet) were also examined. Hepatic 8-oxodG levels were significantly higher in NASH compared with simple steatosis (17.5 versus 2.0 8-oxodG-positive cells/10(5) microm(2); P < 0.0001). 8-oxodG was significantly related to iron overload condition, glucose-insulin metabolic abnormality, and severities of hepatic steatosis in NASH patients. Logistic regression analysis also showed that hepatic iron deposit and insulin resistance were independent variables associated with elevated hepatic 8-oxodG. After the iron reduction therapy, hepatic 8-oxodG levels were significantly decreased (from 20.7 to 13.8 positive cells/10(5) microm(2); P < 0.01) with concomitant reductions of serum transaminase levels in NASH patients. In conclusion, iron overload may play an important role in the pathogenesis of NASH by generating oxidative DNA damage and iron reduction therapy may reduce hepatocellular carcinoma incidence in patients with NASH.

Perinatal Iron Deficiency Affects Locomotor Behavior and Water Maze Performance in Adult Male and Female Rats ,

Iron deficiency during early growth and development adversely affects multiple facets of cognition and behavior in adult rats. The purpose of this study was to assess the nature of the learning and locomotor behavioral deficits observed in male and female rats in the absence of depressed brain iron levels at the time of testing. Adult female Wistar rats were fed either an iron-enriched diet (>225 mg/kg Fe) or an iron-restricted diet (3 mg/kg Fe) for 2 wk prior to and throughout gestation, and a nonpurified diet (270 mg/kg Fe) thereafter. Open-field (OF) and Morris water maze (MWM) testing began when the offspring reached early adulthood (12 wk). At birth, perinatal iron-deficient (PID) offspring had reduced (P < 0.001) hematocrits (−33%), liver iron stores (−83%), and brain iron concentrations (−38%) compared with controls. Although there were no differences in iron status in adults, the PID males and females exhibited reduced OF exploratory behavior, albeit only PID males had an aversion to the center of the apparatus (2.5 vs. 6.9% in controls, P < 0.001). Additionally, PID males required greater path lengths to reach the hidden platform in the MWM, had reduced spatial bias for the target quadrant, and had a tendency for greater thigmotactic behavior in the probe trials (16.5 vs. 13.0% in controls; P = 0.06). PID females had slower swim speeds in all testing phases (−6.2%; P < 0.001). These results suggest that PID has detrimental programming effects in both male and female rats, although the behaviors suggest different mechanisms may be involved in each sex.

Dietary iron deficiency in strain 40 BXD mice causes diurnal‐dependent alterations in brain iron and activity

There is an existent literature on Restless Legs Syndrome that supports a role for regional brain iron, dopamine and diurnal cycle variations in its pathophysiology. In this study, Strain 40 of the BXD/Ty RI panel was utilized based on its mid‐range ventral midbrain iron levels among 30 BXD/Ty RI strains initially analyzed. Mice were challenged with control or iron restricted diets from postnatal day 21 to 120 and then brain iron, striatal extracellular dopamine, home‐cage locomotor activity and body temperature were assessed across the diurnal cycle. At day 120, mice fed an iron deficient (ID) diet showed reduced hemoglobin, hematocrit and spleen and liver iron compared to controls. Male and female ID mice displayed diurnal dependent decreases in ventral midbrain iron, while in nucleus accumbens, iron content was reduced only in male ID mice. Diurnal monitoring of home cage activity and body temperature revealed that iron deficiency alters the number and duration of activity cycles in the four hours surrounding the light transitions. During these transition periods, activity increased and decreased at a similar rate as extracellular dopamine and body temperature in control mice and ID females, while these transitions were blunted in ID males. These results suggest that dietary iron deficiency in BXD Strain 40 mice produces diurnal‐dependent alterations in brain iron in dopaminergic regions and in activity patterns.

Iron Restriction Negatively Affects Bone in Female Rats and Mineralization of hFOB Osteoblast Cells

We previously reported that severe iron deficiency negatively affects bone microarchitecture. Here we determined whether marginal iron restriction that reflects some human consumption patterns could have similar consequences. Thirty-two weanling female rats were randomly divided into four groups and fed the following diets for 10 weeks: (i) iron-adequate, calcium-adequate (FeA:CaA), (ii) calcium-restricted (FeA:CaR), (iii) iron-restricted (FeR:CaA), and (iv) both calcium- and iron-restricted (FeR:CaR) diets. DEXA analysis revealed that CaR decreased bone mineral density (BMD), and FeR decreased whole-body bone mineral content (BMC). Iron-restricted and calcium-restricted groups had lower BMD than did their adequate counterparts. All treatment-restricted groups had lower BMD in the fourth lumbar (L-4) vertebrae than the FeA:CaA group. Vertebrae BMD was lower in all treatment groups compared to the control group, and for BMC, the CaR groups were lower than the CaA groups and the FeR groups were lower that the FeA groups, and BMC were lower in iron- and calcium-restricted groups. The microarchitecture of the L-4 vertebrae was compromised in FeA:CaR, FeR:CaA, and FeR:CaR: (i) the connectivity density was reduced by FeR and by CaR; and (ii) trabecular number was decreased and trabecular separation was increased by FeR. Cortical thickness of the femur was reduced by both FeR and CaR. Finite element analysis revealed that L-4 vertebrae from the FeR:CaA group had greater internal stress with an applied force than the FeA:CaA group and, thus, would be more likely to break. Chelation of iron in cultured osteoblast cells impaired mineralization but had no impact upon Type I collagen deposition. Iron depletion, similar to that occurring among some human populations, reduced bone strength and microarchitecture based on the in vivo and in vitro results reported here. Impaired mineralization with iron depletion appears to be a possible mechanism for the observed bone abnormalities.