Iron bioavailability of maize (Zea mays L.) after removing the germ fraction.

Maize is a staple food for many communities with high levels of iron deficiency anemia. Enhancing the iron concentrations and iron bioavailability of maize with traditional breeding practices, especially after cooking and processing, could help alleviate iron deficiency in many of these regions. Previous studies on a small number of maize genotypes and maize flour products indicated that degermination (germ fraction removed with processing) could improve the iron bioavailability of maize. This study expanded upon this research by evaluating the iron bioavailability, mineral concentrations, and phytate concentrations of 52 diverse maize genotypes before (whole kernels) and after degermination. Whole and degerminated maize samples were cooked, dried, and milled to produce corn flour. Iron bioavailability was evaluated with an in vitro digestion Caco2 cell bioassay. In 30 of the maize genotypes, bioavailable iron increased when degerminated, thus indicating a higher fractional iron uptake because the iron concentrations decreased by more than 70% after the germ fraction was removed. The remaining 22 genotypes showed no change or a decrease in iron bioavailability after degermination. These results confirm previous research showing that the germ fraction is a strong inhibitory component for many maize varieties. Phytate concentrations in maize flours were greatly reduced with degermination. However, the relationship between phytate concentrations and the iron bioavailability of processed maize flour is complex, acting as either inhibitor or promoter of iron uptake depending on the color of the maize kernels and processing method used to produce flour. Other factors in the maize endosperm fractions are likely involved in the effects of degermination on iron bioavailability, such as vitreous or floury endosperm compositions and the polyphenol content of the bran. This study demonstrates that iron nutrition from maize can be enhanced by selecting genotypes where the inhibitory effect of the bran color and endosperm fraction are relatively low, especially after processing via degermination.

Investigation of Genotype by Environment Interactions for Seed Zinc and Iron Concentration and Iron Bioavailability in Common Bean.

Iron and zinc malnutrition are global public health concerns afflicting mostly infants, children, and women in low- and middle-income countries with widespread consumption of plant-based diets. Common bean is a widely consumed staple crop around the world and is an excellent source of protein, fiber, and minerals including iron and zinc. The development of nutrient-dense common bean varieties that deliver more bioavailable iron and zinc with a high level of trait stability requires a measurement of the contributions from genotype, environment, and genotype by environment interactions. In this research, we investigated the magnitude of genotype by environment interaction for seed zinc and iron concentration and seed iron bioavailability (FeBIO) using a set of nine test genotypes and three farmers' local check varieties. The research germplasm was evaluated for two field seasons across nine on-farm locations in three agro-ecological zones in Uganda. Seed zinc concentration ranged from 18.0 to 42.0 µg g-1 and was largely controlled by genotype, location, and the interaction between location and season [28.0, 26.2, and 14.7% of phenotypic variability explained (PVE), respectively]. Within a genotype, zinc concentration ranged on average 12 µg g-1 across environments. Seed iron concentration varied from 40.7 to 96.7 µg g-1 and was largely controlled by genotype, location, and the interaction between genotype, location, and season (25.7, 17.4, and 13.7% of PVE, respectively). Within a genotype, iron concentration ranged on average 28 µg g-1 across environments. Seed FeBIO ranged from 8 to 116% of Merlin navy control and was largely controlled by genotype (68.3% of PVE). The red mottled genotypes (Rozi Koko and Chijar) accumulated the most seed zinc and iron concentration, while the yellow (Ervilha and Cebo Cela) and white (Blanco Fanesquero) genotypes had the highest seed FeBIO and performed better than the three farmers' local check genotypes (NABE-4, NABE-15, and Masindi yellow). The genotypes with superior and stable trait performance, especially the Manteca seed class which combine high iron and zinc concentrations with high FeBIO, would serve as valuable parental materials for crop improvement breeding programs aimed at enhancing the nutritional value of the common bean.

Iron Concentrations in Biofortified Beans and Nonbiofortified Marketplace Varieties in East Africa Are Similar.

BACKGROUND: The predominant bean iron (Fe) biofortification approach is to breed for high Fe concentration and assumes the average Fe concentration is 50 µg/g. This approach also assumes that a 40 µg/g increase is sustainable and Fe bioavailability will not decrease to negate the increase in Fe. OBJECTIVE: The overall objective was to determine if bean Fe biofortification via breeding for high Fe is producing beans with higher Fe concentration relative to nonbiofortified lines found in the East Africa marketplace. METHODS: Seventy-six marketplace samples (East Africa Marketplace Collection; EAMC), and 154 genotypes known to be representative of the marketplace were collected from breeders in the Pan-Africa Bean Research Alliance (designated the East Africa Breeder Collection; EABC). Within the EAMC and EABC were 18 and 35 samples, respectively, that were released as biofortified lines. All samples were measured for Fe concentration. The Caco-2 cell bioassay assessed Fe bioavailability of the EAMC. Biofortified versus nonbiofortified samples were compared by the appropriate t-test or ANOVA. RESULTS: The Fe concentration of the 58 nonbiofortified EAMC lines was (mean ± SD [range]) 71 ± 9 µg/g (52-93 µg/g) which did not differ significantly from the 18 biofortified EAMC varieties (71 ± 11 µg/g [55-94 µg/g]). The Fe concentration of the 119 nonbiofortified EABC varieties was 66 ± 7 µg/g (51-90 µg/g) which was significantly different (P < 0.0001) from the 35 EABC biofortified lines (73 ± 9 µg/g [60-91 µg/g]). However, the EABC biofortified lines were not different from the nonbiofortified EAMC samples. In the Caco-2 cell bioassay, biofortified EAMC varieties did not deliver more Fe compared with nonbiofortified EAMC varieties. CONCLUSIONS: The assumptions of the high Fe bean biofortification approach are not met in the East African marketplace. Iron concentration and bioavailability measurement indicate the biofortified bean varieties are providing no additional dietary Fe.

On-farm multi-location evaluation of genotype by environment interactions for seed yield and cooking time in common bean.

Common bean variety choice by farmers in Uganda is driven by seed yield plus end-use quality traits like market class and cooking time. Limited genotype by environment information is available for traits valued by consumers. This research evaluated yield, seed size, hydration properties, and cooking time of 15 common bean genotypes within market classes recognized by consumers along with three farmers' checks at nine on-farm locations in Uganda for two seasons. Yield ranged from 71 to 3,216 kg ha-1 and was largely controlled by location (21.5% of Total Sums of Squares [TSS]), plus the interaction between location and season (48.6% of TSS). Cooking time varied from 19 to 271 minutes with the genotypes Cebo Cela and Ervilha consistently cooking fastest in 24 and 27 minutes respectively. Comparatively, the local checks (NABE-4, NABE-15, and Masindi yellow) took 35 to 45 minutes to cook. Cooking time was largely controlled by genotype (40.6% of TSS). A GGE biplot analysis uncovered the presence of two mega-environments for yield and one mega-environment for cooking time. Identification of mega-environments for these traits will help expedite common bean breeding, evaluation, and variety selection through reduction of number of test environments needed for phenotype evaluations. The high yielding and fast cooking genotypes from this study can be targeted as parental materials to improve existing common bean germplasm for these important traits.

An In Vivo (Gallus gallus) Feeding Trial Demonstrating the Enhanced Iron Bioavailability Properties of the Fast Cooking Manteca Yellow Bean (Phaseolus vulgaris L.).

The common dry bean (Phaseolus vulgaris L.) is a globally produced pulse crop and an important source of micronutrients for millions of people across Latin America and Africa. Many of the preferred black and red seed types in these regions have seed coat polyphenols that inhibit the absorption of iron. Yellow beans are distinct from other market classes because they accumulate the antioxidant kaempferol 3-glucoside in their seed coats. Due to their fast cooking tendencies, yellow beans are often marketed at premium prices in the same geographical regions where dietary iron deficiency is a major health concern. Hence, this study compared the iron bioavailability of three faster cooking yellow beans with contrasting seed coat colors from Africa (Manteca, Amarillo, and Njano) to slower cooking white and red kidney commercial varieties. Iron status and iron bioavailability was assessed by the capacity of a bean based diet to generate and maintain total body hemoglobin iron (Hb-Fe) during a 6 week in vivo (Gallus gallus) feeding trial. Over the course of the experiment, animals fed yellow bean diets had significantly (p ≤ 0.05) higher Hb-Fe than animals fed the white or red kidney bean diet. This study shows that the Manteca yellow bean possess a rare combination of biochemical traits that result in faster cooking times and improved iron bioavailability. The Manteca yellow bean is worthy of germplasm enhancement to address iron deficiency in regions where beans are consumed as a dietary staple.

The Fast Cooking and Enhanced Iron Bioavailability Properties of the Manteca Yellow Bean (Phaseolus vulgaris L.).

The common dry bean (Phaseolus vulgaris L.) is a nutrient-dense pulse crop that is produced globally for direct human consumption and is an important source of protein and micronutrients for millions of people across Latin America, the Caribbean and Sub-Saharan Africa. Dry beans require large amounts of heat energy and time to cook, which can deter consumers worldwide from using beans. In regions where consumers rely on expensive fuelwood for food preparation, the yellow bean is often marketed as fast cooking. This study evaluated the cooking time and health benefits of five major market classes within the yellow bean seed type (Amarillo, Canary, Manteca, Mayocoba, Njano) over two field seasons. This study shows how the Manteca yellow bean possesses a fast cooking phenotype, which could serve as genetic resource for introducing fast cooking properties into a new generation of dry beans with cooking times <20 min when pre-soaked and <80 min unsoaked. Mineral analysis revealed fast cooking yellow beans have high iron retention (>80%) after boiling. An in vitro digestion/Caco-2 cell culture bioassay revealed a strong negative association between cooking time and iron bioavailability in yellow beans with r values = -0.76 when pre-soaked and -0.64 when unsoaked across the two field seasons. When either pre-soaked or left unsoaked, the highest iron bioavailability scores were measured in the fast cooking Manteca genotypes providing evidence that this yellow market class is worthy of germplasm enhancement through the added benefit of improved iron quality after cooking.

The effect of fish oil supplementation on brain DHA and EPA content and fatty acid profile in mice.

Supplementation with omega-3 (n-3) fatty acids may improve cognitive performance and protect against cognitive decline. However, changes in brain phospholipid fatty acid composition after supplementation with n-3 fatty acids are poorly described. The purpose of this study was to feed increasing n-3 fatty acids and characterise the changes in brain phospholipid fatty acid composition and correlate the changes with red blood cells (RBCs) and plasma in mice. Increasing dietary docosahexaenoic (DHA) and eicosapentaenoic acid (EPA) did not alter brain DHA. Brain EPA increased and total n-6 polyunsaturated fatty acids decreased across treatment groups, and correlated with fatty acid changes in the RBC (r > 0.7). Brain cis-monounsaturated fatty acids oleic and nervonic acid (p < .01) and saturated fatty acids arachidic, behenic, and lignoceric acid (p < .05) also increased. These brain fatty acid changes upon increasing n-3 intake should be further investigated to determine their effects on cognition and neurodegenerative disease.

Demonstrating a Nutritional Advantage to the Fast-Cooking Dry Bean (Phaseolus vulgaris L.).

Dry beans (Phaseolus vulgaris L.) are a nutrient-dense food rich in protein and micronutrients. Despite their nutritional benefits, long cooking times limit the consumption of dry beans worldwide, especially in nations where fuelwood for cooking is often expensive or scarce. This study evaluated the nutritive value of 12 dry edible bean lines that vary for cooking time (20-89 min) from four market classes (yellow, cranberry, light red kidney, and red mottled) of economic importance in bean-consuming regions of Africa and the Americas. When compared to their slower cooking counterparts within each market class, fast-cooking dry beans retain more protein and minerals while maintaining similar starch and fiber densities when fully cooked. For example, some of the highest protein and mineral retention values were measured in the fast-cooking yellow bean cultivar Cebo Cela, which offered 20% more protein, 10% more iron, and 10% more zinc with each serving when compared with Canario, a slow-cooking yellow bean that requires twice the cooking time to become palatable. A Caco-2 cell culture model also revealed the bioavailability of iron is significantly higher in faster cooking entries (r = -0.537, P = 0.009) as compared to slower cooking entries in the same market class. These findings suggest that fast-cooking bean varieties have improved nutritive value through greater nutrient retention and improved iron bioavailability.

Genetic diversity and genome-wide association analysis of cooking time in dry bean (Phaseolus vulgaris L.).

KEY MESSAGE: Fivefold diversity for cooking time found in a panel of 206 Phaseolus vulgaris accessions. Fastest accession cooks nearly 20 min faster than average.   SNPs associated with cooking time on Pv02, 03, and 06. Dry beans (Phaseolus vulgaris L.) are a nutrient dense food and a dietary staple in parts of Africa and Latin America. One of the major factors that limits greater utilization of beans is their long cooking times compared to other foods. Cooking time is an important trait with implications for gender equity, nutritional value of diets, and energy utilization. Very little is known about the genetic diversity and genomic regions involved in determining cooking time. The objective of this research was to assess cooking time on a panel of 206 P. vulgaris accessions, use genome- wide association analysis (GWAS) to identify genomic regions influencing this trait, and to test the ability to predict cooking time by raw seed characteristics. In this study 5.5-fold variation for cooking time was found and five bean accessions were identified which cook in less than 27 min across 2 years, where the average cooking time was 37 min. One accession, ADP0367 cooked nearly 20 min faster than average. Four of these five accessions showed close phylogenetic relationship based on a NJ tree developed with ~5000 SNP markers, suggesting a potentially similar underlying genetic mechanism. GWAS revealed regions on chromosomes Pv02, Pv03, and Pv06 associated with cooking time. Vis/NIR scanning of raw seed explained 68 % of the phenotypic variation for cooking time, suggesting with additional experimentation, it may be possible to use this spectroscopy method to non-destructively identify fast cooking lines as part of a breeding program.

Is the omega-3 index a valid marker of intestinal membrane phospholipid EPA+DHA content?

Despite numerous studies investigating n-3 long chain polyunsaturated fatty acid (LCPUFA) supplementation and inflammatory bowel diseases (IBD), the extent to which dietary n-3 LCPUFAs incorporate in gastrointestinal (GI) tissues and correlate with red blood cell (RBC) n-3 LCPUFA content is unknown. In this study, mice were fed three diets with increasing percent of energy (%en) derived from eicosapentaenoic acid (EPA)+docosahexaenoic acid (DHA). Dietary levels reflected recommended intakes of fish/fish oil by the American Heart Association. We analyzed the FA composition of phospholipids extracted from RBCs, plasma, and GI tissues. We observed that the 0.1%en EPA+DHA diet was sufficient to significantly increase the omega-3 index (RBC EPA+DHA) after 5 week feeding. The baseline EPA levels were 0.2-0.6% across all tissues increasing to 1.6-4.3% in the highest EPA+DHA diet; these changes resulted in absolute increases of 1.4-3.9% EPA across tissues. The baseline DHA levels were 2.2-5.9% across all tissues increasing to 5.8-10.5% in the highest EPA+DHA diet; these changes resulted in absolute increases of 3.2-5.7% DHA across tissues. These increases in EPA and DHA across all tissues resulted in strong (r>0.91) and significant (P<0.001) linear correlations between the omega-3 index and plasma/GI tissue EPA+DHA content, suggesting that the omega-3 index reflects the relative amounts of EPA+DHA in GI tissues. These data demonstrate that the GI tissues are highly responsive to dietary LCPUFA supplementation and that the omega-3 index can serve as a valid biomarker for assessing dietary EPA+DHA incorporation into GI tissues.

Altered dopaminergic profile in the putamen and substantia nigra in restless leg syndrome.

Restless leg syndrome (RLS) is a sensorimotor disorder. Clinical studies have implicated the dopaminergic system in RLS, while others have suggested that it is associated with insufficient levels of brain iron. To date, alterations in brain iron status have been demonstrated but, despite suggestions from the clinical literature, there have been no consistent findings documenting a dopaminergic abnormality in RLS brain tissue. In this study, the substantia nigra and putamen were obtained at autopsy from individuals with primary RLS and a neurologically normal control group. A quantitative profile of the dopaminergic system was obtained. Additional assays were performed on a catecholaminergic cell line and animal models of iron deficiency. RLS tissue, compared with controls, showed a significant decrease in D2R in the putamen that correlated with severity of the RLS. RLS also showed significant increases in tyrosine hydroxylase (TH) in the substantia nigra, compared with the controls, but not in the putamen. Both TH and phosphorylated (active) TH were significantly increased in both the substantia nigra and putamen. There were no significant differences in either the putamen or nigra for dopamine receptor 1, dopamine transporters or for VMAT. Significant increases in TH and phosphorylated TH were also seen in both the animal and cell models of iron insufficiency similar to that from the RLS autopsy data. For the first time, a clear indication of dopamine pathology in RLS is revealed in this autopsy study. The results suggest cellular regulation of dopamine production that closely matches the data from cellular and animal iron insufficiency models. The results are consistent with the hypothesis that a primary iron insufficiency produces a dopaminergic abnormality characterized as an overly activated dopaminergic system as part of the RLS pathology.

Dopamine D2 receptor expression is altered by changes in cellular iron levels in PC12 cells and rat brain tissue.

Iron deficiency anemia in early life alters the development and functioning of the dopamine neurotransmitter system, but data regarding the specific effects of brain iron loss on dopamine D(2) receptor regulation are lacking. Cell culture and animal models were employed in this study to determine whether D(2) receptor expression is altered when cellular iron levels are depleted. Endogenous D(2) receptor-expressing PC12 cells exposed to increasing concentrations of the iron chelator desferrioxamine (25-100 micromol/L) exhibited dose-dependent decreases in total D(2) receptor protein concentrations (20-65%), but there were minimal effects on D(2) receptor mRNA levels. When iron-deficient cells were repleted with ferric ammonium citrate for 24 h, D(2) receptor protein densities were similar to control. Dietary iron deficiency for 6 wk in weanling rats also reduced regional iron concentrations by nearly 50% in the ventral midbrain and caudate but did not affect D(2) receptor mRNA levels in the ventral midbrain. Iron deficiency significantly reduced membrane D(2) receptor protein levels by >70% in caudate, whereas cytosolic concentrations showed only 25% losses. D(2) receptor protein densities and regional iron concentrations were restored within 2 wk of dietary iron repletion. These results support the concept that D(2) receptor gene expression is not significantly changed by iron deficiency, whereas dopamine receptor trafficking is affected and is likely related to known dopamine system alterations in iron deficiency.

Down-regulation of dopamine transporter by iron chelation in vitro is mediated by altered trafficking, not synthesis.

Neurological development and functioning of dopamine (DA) neurotransmission is adversely affected by iron deficiency in early life. Iron-deficient rats demonstrate significant elevations in extracellular DA and a reduction in dopamine transporter (DAT) densities in the caudate putamen and nucleus accumbens. To explore possible mechanisms by which cellular iron concentrations control DAT functioning, endogenous DAT-expressing PC12 cells were used to determine the effect of iron chelation on DAT protein and mRNA expression patterns. In addition, we used human DAT (hDAT)-transfected Neuro2a (N2A) cells to examine DAT degradation and trafficking patterns. A 50 microM treatment for 24 h with the iron chelator, desferrioxamine (DFO), significantly decreased dopamine uptake in a dose-dependent manner, with no apparent change in K(m), in both PC12 and N2A cells. Reduced DA uptake was accompanied by concentration- and time-dependent reductions in total DAT protein levels in both cell lines. Exposure to increasing concentrations of DFO did not significantly alter DAT mRNA in either PC12 or N2A cells. However, DAT degradation rates increased three-fivefold in both cell types exposed to 50 microM DFO for 24 h. Biotinylation studies in N2A cells indicate a more dramatic loss of DAT in the membrane fraction, while OptiPrep fractionation experiments revealed an increase in lysosomal DAT with iron chelation. Inhibition of protein kinase C activation with staurosporin prevented the effect of iron chelation on DAT function, suggesting that in vitro iron chelation affects DAT primarily through the effects on trafficking rather than on synthesis.

Cellular iron concentrations directly affect the expression levels of norepinephrine transporter in PC12 cells and rat brain tissue.

Neurological development and functioning are adversely affected by iron deficiency in early life. Iron-deficient rats are known to have elevations in extracellular DA and NE, suggesting alterations in reuptake of these monoamines. To explore possible mechanisms by which cellular iron concentrations may alter NE transporter functioning, we utilized NET expressing PC12 cells and iron-deficient rats to explore the relationship between NET protein and mRNA expression patterns and iron concentrations. Treatment of PC12 with the iron chelator, desferrioxamine mesylate (DFO, 50 microM for 24 h), significantly decreased [3H] NE uptake by more than 35% with no apparent change in Km. PC12 cells exposed to increasing concentrations of DFO (25-100 microM) exhibited a dose response decrease in [3H] NE uptake within 24 h (38-73% of control) that paralleled a decrease in cellular NET protein content. Inhibition of protein synthesis with cycloheximide resulted in NET disappearance rates from DFO-treated cells greatly exceeding the rate of loss from control cells. RT-PCR analysis revealed only a modest decrease in NET mRNA levels. Rat brain locus ceruleus and thalamus NET mRNA levels were also only modestly decreased (10-15%) despite a 40% reduction in regional brain iron. In contrast, NET proteins levels in thalamus and locus ceruleus were strongly affected by regional iron deficiency with high correlations with iron concentrations (r > 0.94 and r > 0.80 respectively). The present findings demonstrate that NET protein concentrations and functioning are dramatically reduced with iron deficiency; the modest effect on mRNA levels suggests a stronger influence on NET trafficking and degradation than on protein synthesis.

Brain iron uptake in hypotransferrinemic mice: influence of systemic iron status.

The relationship of the heterogeneity of iron concentrations in the brain with the regulation of iron uptake into specific brain regions remains unresolved. We used hypotransferrinemic mice and an iron-deficient or control diet to explore whether plasma transferrin (Tf), transferrin saturation, and plasma iron levels influence the uptake of (59)Fe and whether there was brain region specificity. Weaning wild-type (+/+) and heterozygotic mice (+/hpx), were sorted randomly to either a iron-deficient diet or a control iron diet for 8 weeks, whereas homozygous mice (hpx/hpx) ate the control diet for 8 weeks before (59)Fe uptake studies. Iron-deficient heterozygous and wild-type mice both had significantly greater plasma Tf levels (37.5 and 42.5 microM) than control mice had (heterozygous and wild-type controls were 20 and 32.5 microM) and far more than homozygous mice (<0.2 microM) had, thus providing five distinct levels of plasma Tf concentrations. After intravenous injection of (59)Fe, brains of iron-deficient wild-type mice took up significantly more (59)Fe (0.15% dose) compared to control wild-type mice (0.056%) at 2 hr, a treatment effect that persisted through 24 hr. In contrast, diet had no effect in heterozygous mice. Importantly, homozygous mice had equivalent uptake to other groups (0.089% dose) by 24 hr. Early brain radioactivity varied by regions (hypothalamus and prefrontal cortex approximately 10-18% brain uptake > cerebellum, pons, thalamus, and striatum approximately 7-12% > cortex, hippocampus, and substantia nigra approximately 6-8%). This distribution of radioactivity changed over 24 hr in the hypothalamus of heterozygous mice, homozygous mice, and iron-deficient wild-type mice. Homozygous mice also showed higher uptake (13-15%) in some regions (hypothalamus and cerebellum) than in other regions. In wild-type and heterozygous mice, (59)Fe uptake was inversely related to brain Tf and was independent of regional brain iron concentrations and plasma Tf levels or saturation. These experimental data suggest that brain iron uptake may be constitutive and independent of plasma Tf, transferrin saturation, or regional brain iron concentration. The proteins and mechanisms responsible for additional iron uptake into specific regions, or perhaps the redistribution are unclear though the data are supportive of a non-transferrin-bound iron uptake pathway.

Pre- and postweaning iron deficiency alters myelination in Sprague-Dawley rats.

Iron deficiency in early life is associated with hypomyelination; however, the role which iron plays in myelinogenesis is not clearly established. In this study, we examined the effect of preweaning [postnatal days (PND) 4-14 and PND 4-21] and postweaning (PND 21-63) iron deficiency on hindbrain 2',3'-cyclic nucleotide 3'-phosphohydrolase (CNPase) activity (marker of oligodendrocyte metabolic activity) and myelin basic protein (MBP) concentrations. Both CNPase activity and concentrations in the cerebrum and hindbrain were significantly lower in pre- and postweaning iron-deficient rats. Similarly, MBP concentrations were also reduced (25-35%) in all three groups of iron-deficient animals. Iron-deficient animals also had significant alterations in the fatty acid composition of individual phospholipids within the hindbrain as well as changes in cytochrome oxidase activities. These studies show that postnatal iron deficiency, for as little as 10 days, can significantly alter the production of myelin and oligodendrocyte functioning. Importantly, postweaning iron deficiency was still associated with a decrease in CNPase activity and MBP concentrations despite occurring well past a likely key sensitive period of peak myelinogenesis at PND 8-12. This suggests that iron deficiency in later life, as well as during early postnatal growth, can effect the production and maintenance of myelin.

Iron-deficient mice fail to develop autoimmune encephalomyelitis.

Determinations of the effects of iron status on the immune system are complicated by the fact that microorganisms and immune cells both utilize iron. To determine the role of iron in immune function, we utilized a model [experimental autoimmune encephalomyelitis (EAE)] in which a strong antigen-specific CD4+ T-cell response develops in the absence of infection. EAE is an autoimmune disease frequently used as a model for the human disease multiple sclerosis (MS). EAE was induced in B10.PL mice fed low iron (1 mg/kg), normal iron (10 mg/kg) or high iron (160 mg/kg) diets that were replete in all other nutrients. Liver iron measurements verified iron status, i.e., low iron mice had 1.9 micro mol/g tissue, normal iron mice, 3.27 micro mol/g tissue and high iron mice, 5.35 micro mol/g tissue. EAE symptoms were most severe in normal iron mice, and EAE did not develop in low iron mice. The incidence of EAE was 71% in normal iron mice, 62% in iron-overloaded mice and 0% in iron-deficient mice. Two of seven mice in the normal iron group developed severe EAE and were euthanized. None of the iron-overloaded mice developed severe EAE. Other measures of EAE severity were similar in the normal and iron-overloaded mice. The data suggest that iron deficiency provides protection from the development of EAE and that iron excess with its potential contribution to free radical formation was not an important factor. The mechanism of EAE inhibition in iron-deficient mice likely involves the delivery and metabolism of iron for optimal CD4+ T-cell development.

Quantitative genetic analysis of ventral midbrain and liver iron in BXD recombinant inbred mice.

Male and female mice from 15 of the BXD/Ty recombinant inbred strain panel were examined for regional brain and liver iron content. Brain regions included medial prefrontal cortex, nucleus accumbens, caudate-putamen and ventral midbrain. Our focal tissue was the ventral midbrain, containing the ventral tegmentum and substantia nigra. This area contains the perikarya of the dopamine neurons that project to nucleus accumbens and caudate-putamen. Genetic correlations between ventral midbrain and liver iron content were not statistically significant, suggesting that peripheral and central iron regulatory systems are largely independent. Correlations between ventral midbrain iron and iron in the caudate-putamen and nucleus accumbens, but not the prefrontal cortex were moderately high and significant. Ventral midbrain and liver iron contents were subjected to quantitative trait loci analysis to identify associated chromosomal locations. This analysis revealed several suggestive loci for iron content in ventral midbrain but fewer loci for liver. Genetic correlations between ventral midbrain iron and published dopamine functional indices were significant, suggesting a link between ventral midbrain iron status and central dopamine neurobiology. This work shows the value of quantitative genetic analysis in the neurobiology of iron and in showing the close association between ventral midbrain iron and nigrostriatal/mesolimbic dopamine function.