Gut microbiome-short-chain fatty acids interplay in the context of iron deficiency anaemia.

PURPOSE: Anaemia is a global health concern, with iron deficiency anaemia (IDA) causing approximately 50% of cases. Affecting mostly the elderly, pregnant and adult women and children, physiopathology of IDA in relation to the gut microbiome is poorly understood. Therefore, the objective of this study is to analyse, in an animal model, the effect of IDA on the gut microbiome along the gastrointestinal tract, as well as to relate intestinal dysbiosis to changes in microbial metabolites such as short chain fatty acids (SCFA). METHODS: IDA was experimentally induced through an iron deficient diet for a period of 40 days, with twenty weaned male Wistar rats being randomly divided into control or anaemic groups. Blood samples were collected to control haematological parameters, and so were faecal and intestinal content samples to study gut microbial communities and SCFA, using 16S rRNA sequencing and HPLC-UV respectively. RESULTS: An intestinal dysbiosis was observed as a consequence of IDA, especially towards the distal segments of the gastrointestinal tract and the colon. An increase in SCFA was also noticed during IDA, with the major difference appearing in the colon and correlating with changes in the composition of the gut microbiome. Clostridium_sensu_stricto_1 and Clostridium_sensu_stricto_4 showed the greatest correlation with variations in butyric and propionic concentrations in the colon of anaemic animals. CONCLUSIONS: Composition of intestinal microbial communities was affected by the generation of IDA. An enrichment in certain SCFA-producing genera and SCFA concentrations was found in the colon of anaemic animals, suggesting a trade-off mechanism against disease.

Role of Fermented Goat Milk on Liver Gene and Protein Profiles Related to Iron Metabolism during Anemia Recovery.

Despite the crucial role of the liver as the central regulator of iron homeostasis, no studies have directly tested the modulation of liver gene and protein expression patterns during iron deficiency instauration and recovery with fermented milks. Fermented goat milk consumption improves the key proteins of intestinal iron metabolism during iron deficiency recovery, enhancing the digestive and metabolic utilization of iron. The aim of this study was to assess the influence of fermented goat or cow milk consumption on liver iron homeostasis during iron-deficiency anemia recovery with normal or iron-overload diets. Analysis included iron status biomarkers, gene and protein expression in hepatocytes. In general, fermented goat milk consumption either with normal or high iron content up-regulated liver DMT1, FPN1 and FTL1 gene expression and DMT1 and FPN1 protein expression. However, HAMP mRNA expression was lower in all groups of animals fed fermented goat milk. Additionally, hepcidin protein expression decreased in control and anemic animals fed fermented goat milk with normal iron content. In conclusion, fermented goat milk potentiates the up-regulation of key genes coding for proteins involved in iron metabolism, such as DMT1, and FPN1, FTL1 and down-regulation of HAMP, playing a key role in enhanced iron repletion during anemia recovery, inducing a physiological adaptation of the liver key genes and proteins coordinated with the fluctuation of the cellular iron levels, favoring whole-body iron homeostasis.

Fermented Goat Milk Consumption Enhances Brain Molecular Functions during Iron Deficiency Anemia Recovery.

Iron deficiency anemia (IDA) is one of the most prevalent nutritional deficiencies worldwide. Iron plays critical roles in nervous system development and cognition. Despite the known detrimental consequences of IDA on cognition, available studies do not provide molecular mechanisms elucidating the role of iron in brain functions during iron deficiency and recovery with dairy components. In this study, 100 male Wistar rats were placed on a pre-experimental period of 40 days and randomly divided in two groups: a control group receiving a normal-Fe diet, (45 mg/kg), and an Fe-deficient group receiving a low-Fe diet (5 mg/kg). At day 40, 10 rats per group were sacrificed to anemia control, and 80 rats were divided into eight experimental groups fed with fermented goat or cow milk-based diets, with normal Fe content or Fe overload (450 mg/kg) for 30 days. IDA decreased most of the parameters related to brain molecular functions, namely dopamine, irisin, MAO-A, oxytocin, ß-endorphin, and α-MSH, while it increased synaptophysin. These alterations result in an impairment of brain molecular functions. In general, during anemia recovery, fermented goat milk diet consumption increased dopamine, oxytocin, serotonin, synaptophysin, and α-MSH, and decreased MAO-A and MAO-B, suggesting a potential neuroprotective effect in brain functions, which could enhance brain molecular functions.

Publisher Correction: Protective effects of fermented goat milk on genomic stability, oxidative stress and inflammatory signalling in testis during anaemia recovery.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Iron Deficiency and Neuroendocrine Regulators of Basal Metabolism, Body Composition and Energy Expenditure in Rats.

Although dietary iron is a determinant of iron status in animals, body fat mass has been reported to have an inverse association with iron status in human studies. The goal of this study was to determine the relationship between Fe homeostasis, body composition, energy expenditure and neuroendocrine regulators for severe Fe-deficiency anaemia. Forty male Wistar albino rats recently weaned were divided at random into two groups: the control group was fed the basal diet, AIN-93G diet (normal-Fe) and the anaemic group received a low-Fe diet for 40 days. Neuroendocrine parameters that regulate basal metabolism and appetite (thyroid hormones, ghrelin, glucose-dependent insulinotropic polypeptide (GIP), glucagon, insulin, adrenocorticotropic hormone and corticosterone), body composition, respiratory volumes, energy expenditure, haematological and biochemical were assessed. Total body fat was lower, whereas lean mass, free and total water were higher in the anemic group. O2 consumption, CO2 production, energy expenditure (EE) and respiratory quotient (RQ) were lower in the Fe-deficient animals. Triiodothyronine and thyroxine hormones decreased, while thyroid-stimulating hormone increased in the anemic group. Circulating levels of ghrelin were lower in the anemic group, while GIP, glucagon, insulin, corticosterone and adrenocorticotropic hormone levels were higher. Fe-deficiency impairs weight gain in the rats, with marked reductions in lean mass and body fat, indicating lower energy stores.

Protective effects of fermented goat milk on genomic stability, oxidative stress and inflammatory signalling in testis during anaemia recovery.

Oxidative stress is a harmful factor for male reproductive function, and a major cause of infertility. On the other hand, fermented goat milk has positive effects on anemia recovery and mineral metabolism. This study evaluated the effect of feeding rats with fermented milks during anaemia recovery on molecular mechanisms linked to oxidative stress and inflammatory signalling in rats reproductive system. Forty male Wistar rats were placed on a pre-experimental period of 40 days (control group, receiving normal-Fe diet and Fe-deficient group, receiving low-Fe diet). Lately, rats were fed with fermented goat or cow milk-based diets during 30 days. After feeding the fermented milks, Total antioxidant status (TAS) and non-esterified fatty acids (NEFA) increased and 8-hydroxy-2'-deoxyguanosine (8-OHdG), 15-F2t-isoprostanes and thiobarbituric acid reactive substances (TBARS) decreased in testis. DNA oxidative damage in testis germ cells was lower with fermented goat milk. Fermented goat milk reduced IL-6 and TNF-α in control animals, increasing INF-γ in control and anaemic rats. NRF2 and PGC-1α protein levels increased in testis after fermented goat milk consumption in control and anaemic rats. Fermented goat milk also increased TAS and decreased oxidative damage, protecting the main testis cell bioconstituents (lipids, proteins, DNA, prostaglandins) from oxidative damage and reduced inflammatory activity, preventing injuries to testis germinal epithelium. Fermented goat milk enhanced lipolysis, fatty acids degradation and immune response, attenuating inflammatory signalling, representing a positive growth advantage for testicular cells.

Changes in Adiposity and Body Composition during Anemia Recovery with Goat or Cow Fermented Milks.

To date, no studies are available about adipose tissue modifications during anemia recovery; therefore, the aim of this study is to provide detailed information about adipose tissue homeostasis during anemia recovery with fermented milks. Forty male Wistar rats were placed on a pre-experimental period of 40 days, divided in two groups (normal-Fe diet and Fe-deficient diet). Then rats were fed fermented goat or cow milk-based diets with normal-Fe content during 30 days. Ghrelin and adiponectin decreased in both groups of animals fed fermented goat milk, whereas leptin and NEFA increased. UCP-1 decreased in anemic rats fed either fermented milk, and irisin greatly increased in both groups of animals fed fermented goat milk. Fermented goat milk reduces adiposity, inducing leptin elevation and ghrelin reduction. Conversely, plasma adiponectin concentrations decreased in animals fed fermented goat milk, showing an inverse correlation with NEFA, an important marker of lipid mobilization, indicating increased lipolysis. Irisin up-regulation in animals fed fermented goat milk contributes to a favorable metabolic profile and the browning of adipose tissue during anemia recovery.

Fermented goat milk consumption during anaemia recovery: ergogenic effect and improvement of skeletal muscle homeostasis.

PURPOSE: Anaemia is associated with fatigue and diminished muscular oxygenation, which may affect skeletal muscle (SM). No studies are available about the SM modifications during anaemia recovery; therefore, the aim of this study is to study SM homeostasis during anaemia recovery with fermented milks. METHODS: Forty male Wistar rats were placed on a pre-experimental period of 40 days, divided in two groups (control group receiving normal-Fe diet and Fe-deficient group receiving low-Fe diet). Lately, rats were fed with fermented goat or cow milk-based diets, with normal-Fe content during 30 days. After feeding the fermented milks, leptin, adiponectin, non-esterified fatty acids (NEFA) and protein expression (UCP1, PepT1 and irisin) within the SM were assessed. RESULTS: Adiponectin decreased in both groups of animals fed fermented goat milk, while leptin and NEFA increased. UCP1 protein expression increased in control and anaemic animals fed fermented goat milk. UCP1 also increased in both group of anaemic animals fed either fermented cow or goat milk in comparison with their controls. Irisin increased in both group of animals fed fermented goat milk. Finally, PepT1 also showed an increased expression in control and anaemic rats fed fermented goat milk and the anaemia also induced an over-expression of this transporter in animals fed either fermented cow or goat milk. CONCLUSION: Fermented goat milk consumption during anaemia recovery diminishes adiposity depots and enhances lipolysis, increasing UCP1, PepT1 and irisin protein expression, featuring an ergogenic effect in the SM which is an important endocrine regulator of body metabolism.

Fermented Goat's Milk Consumption Improves Duodenal Expression of Iron Homeostasis Genes during Anemia Recovery.

Despite the crucial roles of duodenal cytochrome b (Dcytb), divalent metal transporter 1 (DMT1), ferritin light chain (Ftl1), ferroportin 1 (FPN1), transferrin receptor 1 (TfR1), and hepcidin antimicrobial peptide (Hamp) in Fe metabolism, no studies have investigated the modulations of these genes during Fe repletion with fermented milks. Analysis included Fe status markers and gene and protein expression in enterocytes of control and anemic animals fed fermented milks. Fermented goat's milk up-regulated enterocyte Dcytb, DMT1, FPN1, and Ftl1 and down-regulated TfR1 and Hamp gene expression in control and anemic animals. Anemia decreased Dcytb, DMT1, and Ftl1 in animals fed fermented cow's milk and up-regulated TfR1 and Hamp expression. Fe overload down-regulated Dcytb and TfR1 in animals fed fermented cow's milk and up-regulated DMT1 and FPN1 gene expression. Fermented goat's milk increased expression of duodenal Dcytb, DMT1, and FPN1 and decreased Hamp and TfR1, improving Fe metabolism during anemia recovery.

Production and chemical composition of two dehydrated fermented dairy products based on cow or goat milk.

The aim of this study was to identify the differences between the main macro and micronutrients including proteins, fat, minerals and vitamins in cow and goat dehydrated fermented milks. Fermented goat milk had higher protein and lower ash content. All amino acids (except for Ala), were higher in fermented goat milk than in fermented cow milk. Except for the values of C11:0, C13:0, C16:0, C18:0, C20:5, C22:5 and the total quantity of saturated and monounsaturated fatty acids, all the other fatty acid studied were significantly different in both fermented milks. Ca, Mg, Zn, Fe, Cu and Se were higher in fermented goat milk. Fermented goat milk had lower amounts of folic acid, vitamin E and C, and higher values of vitamin A, D3, B6 and B12. The current study demonstrates the better nutritional characteristics of fermented goat milk, suggesting a potential role of this dairy product as a high nutritional value food.

Folic acid supplemented goat milk has beneficial effects on hepatic physiology, haematological status and antioxidant defence during chronic Fe repletion.

The aim of the current study was to asses the effect of goat or cow milk-based diets, either normal or Fe-overloaded and folic acid supplement on some aspects of hepatic physiology, enzymatic antioxidant defence and lipid peroxidation in liver, brain and erythrocyte of control and anaemic rats after chronic Fe repletion. 160 male Wistar rats were placed on 40 d in two groups, a control group receiving normal-Fe diet and the Fe-deficient group receiving low Fe diet. Lately, the rats were fed with goat and cow milk-based diets during 30 d, with normal-Fe content or Fe-overload and either with normal folic or folic acid supplemented. Fe-overload increased plasma alanine transaminase and aspartate transaminase levels when cow milk was supplied. Dietary folate supplementation reduced plasma transaminases levels in animals fed goat milk with chronic Fe overload. A remarkable increase in the superoxide dismutase activity was observed in the animals fed cow milk. Dietary folate supplement lead to a decrease on the activity of this enzyme in all the tissues studied with both milk-based diets. A concomitant increment in catalase was also observed. The increase in lipid peroxidation products levels in rats fed cow milk with Fe-overload, suggest an imbalance in the functioning of the enzymatic antioxidant defence. In conclusion, dietary folate-supplemented goat milk reduces both plasma transaminases levels, suggesting a hepatoprotective effect and has beneficial effects in situation of Fe-overload, improving the antioxidant enzymes activities and reducing lipid peroxidation.

Goat milk supplemented with folic acid protects cell biomolecules from oxidative stress-mediated damage after anaemia recovery in comparison with cow milk.

BACKGROUND: Fe overload is a common consequence of the anaemia treatment, increasing the oxidative stress and promoting the accumulation of damaged biomolecules, with the subsequently impairment of cell functions. Oxidative stress and the role of folic acid preventing free radical damage have been extensively studied; nevertheless, no studies are available about the influence of folic acid-supplemented goat milk consumption on the oxidative stress-mediated damage. AIM: The objective of the present study was to assess the influence of folic acid supplementation of goat milk- or cow milk-based diets, after Fe-overload treatment to palliate anaemia, on oxidative stress-mediated biomolecular damage in the liver, brain, erythrocytes, duodenal mucosa and plasma. METHODS: Control and anaemic rats were fed goat milk- or cow milk-based diets, either with normal Fe or Fe overload (450 mg/kg), and normal folic acid (2 mg/kg) or folic acid supplemented (40 mg/kg) for 30 days. RESULTS: During chronic Fe repletion, background DNA damage was significantly lower in anaemic rats fed folic acid-supplemented goat milk-based diet, as revealed by tail DNA (%), and folic acid-supplemented goat milk also had a beneficial effect, reducing the extent of lipid peroxidation in liver, plasma, erythrocytes and especially in brain and duodenal mucosa. Furthermore, protein oxidative damage was lower in anaemic rat duodenal mucosa for all goat milk-based diets. CONCLUSIONS: Folic acid supplement in goat milk avoids the undesirable effects of Fe overload during anaemia recovery in all the tissues studied, especially in the liver and duodenal mucosa, which are the tissues with higher exposition to dietary Fe.

Positive influence of a natural product as propolis on antioxidant status and lipid peroxidation in senescent rats

Given the importance of oxidative stress associated to aging, it would be interesting to assess the effect of oral supplementation with antioxidant substances capable of diminishing oxidative aggression and free radicals generation associated to this condition. This study investigated the effects of AIN-93 M diet supplemented either with 2 % of propolis, or with 4 % of a natural product obtained from lyophilizate vegetables, selected by its antioxidant properties, in senescent healthy Wistar rats fed ad libitum over 3 months. Propolis supplementation leads to a lower level of glucose and cholesterol concentrations together with a reduction in protein oxidation. Plasma thiobarbituric acid-reactive substance levels were lower in the rats consuming the natural vegetable product and propolis possibly due to its antioxidant components, neutralizing the free radical produced, and thus preventing cellular damage. The results of the present study suggest a synergic effect of overall propolis compounds reducing the oxidative stress and glucose and cholesterol plasma levels associated with aging (AU)

Influence of several sources and amounts of iron on DNA, lipid and protein oxidative damage during anaemia recovery.

The study was designed to assess the effect of several Fe amounts and sources on haematological parameters, DNA, lipid and protein oxidative damage during the course of Fe-deficiency anaemia recovery. Peripheral DNA damage was assessed using an alkaline comet assay. The brain, liver, erythrocyte and duodenal mucosa lipid peroxidation and protein damage were assessed in control and anaemic rats after Fe repletion with three different sources (FeSO4, haem Fe, and FeSO4 + haem Fe) and amounts (45, 12, and 31 mg Fe/kg diet) of Fe: F diet, H diet or C diet, respectively. After supplying the diets, the haematological parameters studied were recovered; being remarkable is the haemoglobin increase. The DNA damage was lower in rats with the H diet, as revealed by the percentage of DNA in head, tail and Olive tail moment compared in rats with the F (P < 0.001) and C (P < 0.05) diets. Lipid peroxidation was similar in all the tissues, except in the duodenal mucosa which was lower with H and C diets (P < 0.001). The animals fed with C diet showed lower oxidative protein damage in the duodenal mucosa (P < 0.001) and was also lower in the liver and erythrocytes for H and C diets (P < 0.001). No differences were found in the brain under our experimental conditions. In conclusion, Fe supplementation with low doses of haem Fe or combined forms of non-haem and haem Fe (FeSO4 + haem) are efficient in restoring the impaired haematological parameters and prevent the evoked oxidative stress associated with Fe supplements.

Positive influence of a natural product as propolis on antioxidant status and lipid peroxidation in senescent rats.

Given the importance of oxidative stress associated to aging, it would be interesting to assess the effect of oral supplementation with antioxidant substances capable of diminishing oxidative aggression and free radicals generation associated to this condition. This study investigated the effects of AIN-93 M diet supplemented either with 2 % of propolis, or with 4 % of a natural product obtained from lyophilizate vegetables, selected by its antioxidant properties, in senescent healthy Wistar rats fed ad libitum over 3 months. Propolis supplementation leads to a lower level of glucose and cholesterol concentrations together with a reduction in protein oxidation. Plasma thiobarbituric acid-reactive substance levels were lower in the rats consuming the natural vegetable product and propolis possibly due to its antioxidant components, neutralizing the free radical produced, and thus preventing cellular damage. The results of the present study suggest a synergic effect of overall propolis compounds reducing the oxidative stress and glucose and cholesterol plasma levels associated with aging.

Bile composition, plasma lipids and oxidative hepatic damage induced by calcium supplementation; effects of goat or cow milk consumption.

Calcium-fortified foods, especially milk and dairy products are recommended to be consumed daily for groups in risk of nutritional deficiency, including children, young adults, menopausal women, pregnant women and the elderly, however Ca-supplementation promotes gallstone formation because Ca is a nucleating factor. The objective of the current study was to assess the influence of cow or goat milk-based diets, either normal or Ca-supplemented, on bile composition, biochemical parameters and hepatic antioxidant status. Weanling male rats were randomly divided into six groups, fed standard, goat or cow milk-based diets, either with normal Ca content (5.0 g/kg), or Ca-supplemented (10.0 g/kg), for 2 weeks. Bile cholesterol concentration and output was higher in rats fed goat milk in comparison with those fed with standard and cow-milk-based diet. Ca-supplementation increased lithogenic index with the standard and cow-milk based diets, this change was not observed with the goat milk diet. Activities of plasma transaminases were also lower in the animals fed Ca-supplemented goat milk, in comparison with the other diets assayed. In general, Ca-supplement in the diet led to an increase in the hepatic oxidative damage, with an increase in the activities of all the antioxidant enzymes studied in the standard and cow milk diet, but not with goat milk. The habitual consumption of goat milk has positive effects on the plasma lipid profile, biliary composition and hepatic antioxidant defence. In addition, under our experimental conditions, Ca-supplementation of this type of milk does not increase the lithogenic index, or hepatic oxidative damage.

Influence of cow or goat milk consumption on antioxidant defence and lipid peroxidation during chronic iron repletion.

Despite Fe deficiency and overload having been widely studied, no studies are available about the influence of milk consumption on antioxidant defence and lipid peroxidation during the course of these highly prevalent cases. The objective of the present study was to assess the influence of cow or goat milk-based diets, either with normal or Fe-overload, on antioxidant defence and lipid peroxidation in the liver, brain and erythrocytes of control and anaemic rats after chronic Fe repletion. Weanling male rats were randomly divided into two groups: a control group receiving a normal-Fe diet (45 mg/kg) and an anaemic group receiving a low-Fe diet (5 mg/kg) for 40 d. Control and anaemic rats were fed goat or cow milk-based diets, either with normal Fe or Fe-overload (450 mg/kg), for 30 or 50 d. Fe-deficiency anaemia did not have any effect on antioxidant enzymes or lipid peroxidation in the organs studied. During chronic Fe repletion, superoxide dismutase (SOD) activity was higher in the group of animals fed the cow milk diet compared with the group consuming goat milk. The slight modification of catalase and glutathione peroxidise activities in animals fed the cow milk-based diet reveals that these enzymes are unable to neutralise and scavenge the high generation of free radicals produced. The animals fed the cow milk diet showed higher rates of lipid peroxidation compared with those receiving the goat milk diet, which directly correlated with the increase in SOD activity. It was concluded that goat milk has positive effects on antioxidant defence, even in a situation of Fe overload, limiting lipid peroxidation.

Severe nutritional iron-deficiency anaemia has a negative effect on some bone turnover biomarkers in rats.

BACKGROUND: The role of iron (Fe) in bone formation and disease have not received much attention, a fact that is interesting given the known biochemical role that this mineral has upon collagen maturation together with the high prevalence of Fe-deficiency anaemia worldwide. AIM: To investigate the changes in bone formation, resorption and mineral content in developing rats with induced nutritional Fe-deficiency anaemia. METHODS: Thirty male Wistar rats were divided into two groups, a control group receiving AIN-93G diet with normal-Fe content and an anaemic group receiving AIN-93G diet with low-Fe content for 40 days. Both diets were prepared with an adequate calcium (Ca) and phosphorus (P) content. The most representative serum bone turnover biomarkers and femur and sternum calcium and phosphorus content, together with sternum Fe content were determined in both experimental groups. RESULTS: In anaemic rats, bone matrix formation diminished as revealed by the lower amount of procollagen type I N-terminal propeptide. Bone resorption process increased in Fe deficiency as shown by the increase of serum parathyroid hormone, tartrate-resistant acid phosphatase and levels of degradation products from C-terminal telopeptides of type I collagen released to the serum. In addition, mineralization process was affected by Fe deficiency, because Ca and P content in femur decreased markedly. CONCLUSIONS: Fe-deficiency anaemia had a significant impact upon bone, affecting bone mineralization, decreasing the matrix formation and increasing bone resorption, therefore it is of great interest to assess bone status in situation of Fe-deficiency anaemia.

Development of nutritional iron deficiency in growing male rats: haematological parameters, iron bioavailability and oxidative defence.

Despite Fe deficiency having been widely studied, the sequence of events in its development still remains unclear. The aim of the present study was to elucidate the effects of nutritional Fe-deficiency development on haematological parameters, Fe bioavailability and the enzymes involved in oxidative defence in recently weaned male Wistar albino rats. Control (C) and Fe-deficient (ID) groups were fed the AIN-93 G diet with a normal Fe level (45 mg/kg diet) or with a low Fe level (5 mg/kg diet), respectively, for 20, 30 or 40 d. At day 20 serum Fe, serum ferritin and the saturation of transferrin decreased drastically, decreasing further in the course of Fe-deficiency development for the saturation of transferrin. The development of Fe deficiency did not affect plasma thiobarbituric acid-reactive substance production, or catalase (CAT) and glutathione peroxidase (GPx) activities in erythrocyte cytosol. Fe deficiency diminished hepatic Fe content and CAT and GPx activities in hepatic cytosol only at day the 20. However, in spite of the minor Fe deposits in the brain of ID rats, the CAT and GPx activities in the brain cytosolic fraction did not differ in any of the studied periods v. control rats. These results show that brain is a tissue that does not seem to depend on Fe levels for the maintenance of antioxidant defence mechanisms in the course of nutritional Fe deficiency.

Effect of synergic dietary calcium enrichment and induced ferropenic anemia on antioxidant enzymes activity in rats.

OBJECTIVE: The aim of this study was to examine the synergism of dietary calcium enrichment (added to goat's or cow's milk) and induced nutritional ferropenic anemia on oxidative status. METHODS: Control rats and rats with induced nutritional ferropenic anemia were fed for 14 d with diets containing normal (5000 mg/kg) or double (10 000 mg/kg) the recommended calcium content. Thiobarbituric acid-reactive substances in plasma were measured, as were the activities of the antioxidant enzyme catalase, copper/zinc superoxide dismutase, and glutathione peroxidase in erythrocyte cytosol. RESULTS: Dietary calcium enrichment did not affect oxidative stress as assessed by thiobarbituric acid-reactive substances; however, it significantly upregulated the activities of some antioxidant enzymes examined in the erythrocyte cytosol. In particular, adding calcium to standard or milk-based diets significantly increased glutathione peroxidase activity in control and anemic rats and copper/zinc superoxide dismutase activity in control rats. CONCLUSION: The increased activities of glutathione peroxidase and copper/zinc superoxide dismutase induced by dietary calcium enrichment suggest that calcium supplementation may protect against oxidative stress even in nutritionally induced ferropenic anemia.