Sharon E. Cox

Maternal nutrition at conception modulates DNA methylation of human metastable epialleles

In experimental animals, maternal diet during the periconceptional period influences the establishment of DNA methylation at metastable epialleles in the offspring, with permanent phenotypic consequences. Pronounced naturally occurring seasonal differences in the diet of rural Gambian women allowed us to test this in humans. We show that significant seasonal variations in methyl-donor nutrient intake of mothers around the time of conception influence 13 relevant plasma biomarkers. The level of several of these maternal biomarkers predicts increased/decreased methylation at metastable epialleles in DNA extracted from lymphocytes and hair follicles in infants postnatally. Our results demonstrate that maternal nutritional status during early pregnancy causes persistent and systemic epigenetic changes at human metastable epialleles.

Mortality in sickle cell anemia in Africa: a prospective cohort study in Tanzania.

Background The World Health Organization has declared Sickle Cell Anemia (SCA) a public health priority. There are 300,000 births/year, over 75% in Africa, with estimates suggesting that 6 million Africans will be living with SCA if average survival reaches half the African norm. Countries such as United States of America and United Kingdom have reduced SCA mortality from 3 to 0.13 per 100 person years of observation (PYO), with interventions such as newborn screening, prevention of infections and comprehensive care, but implementation of interventions in African countries has been hindered by lack of locally appropriate information. The objective of this study was to determine the incidence and factors associated with death from SCA in Dar-es-Salaam. Methods and Findings A hospital-based cohort study was conducted, with prospective surveillance of 1,725 SCA patients recruited from 2004 to 2009, with 209 (12%) lost to follow up, while 86 died. The mortality rate was 1.9 (95%CI 1.5, 2.9) per 100 PYO, highest under 5-years old [7.3 (4.8–11.0)], adjusting for dates of birth and study enrollment. Independent risk factors, at enrollment to the cohort, predicting death were low hemoglobin (<5 g/dL) [3.8 (1.8–8.2); p = 0.001] and high total bilirubin (≥102 µmol/L) [1.7 (1.0–2.9); p = 0.044] as determined by logistic regression. Conclusions Mortality in SCA in Africa is high, with the most vulnerable period being under 5-years old. This is most likely an underestimate, as this was a hospital cohort and may not have captured SCA individuals with severe disease who died in early childhood, those with mild disease who are undiagnosed or do not utilize services at health facilities. Prompt and effective treatment for anemia in SCA is recommended as it is likely to improve survival. Further research is required to determine the etiology, pathophysiology and the most appropriate strategies for management of anemia in SCA.

Maternal nutritional status, C 1 metabolism and offspring DNA methylation: a review of current evidence in human subjects

Evidence is growing for the long-term effects of environmental factors during early-life on later disease susceptibility. It is believed that epigenetic mechanisms (changes in gene function not mediated by DNA sequence alteration), particularly DNA methylation, play a role in these processes. This paper reviews the current state of knowledge of the involvement of C1 metabolism and methyl donors and cofactors in maternal diet-induced DNA methylation changes in utero as an epigenetic mechanism. Methyl groups for DNA methylation are mostly derived from the diet and supplied through C1 metabolism by way of choline, betaine, methionine or folate, with involvement of riboflavin and vitamins B6 and B12 as cofactors. Mouse models have shown that epigenetic features, for example DNA methylation, can be altered by periconceptional nutritional interventions such as folate supplementation, thereby changing offspring phenotype. Evidence of early nutrient-induced epigenetic change in human subjects is scant, but it is known that during pregnancy C1 metabolism has to cope with high fetal demands for folate and choline needed for neural tube closure and normal development. Retrospective studies investigating the effect of famine or season during pregnancy indicate that variation in early environmental exposure in utero leads to differences in DNA methylation of offspring. This may affect gene expression in the offspring. Further research is needed to examine the real impact of maternal nutrient availability on DNA methylation in the developing fetus.

Hepcidin is the major predictor of erythrocyte iron incorporation in anemic African children

Iron supplementation strategies in the developing world remain controversial because of fears of exacerbating prevalent infectious diseases. Understanding the conditions in which iron will be absorbed and incorporated into erythrocytes is therefore important. We studied Gambian children with either postmalarial or nonmalarial anemia, who were given oral iron supplements daily for 30 days. Supplements administered on days 1 and 15 contained the stable iron isotopes 57Fe and 58Fe, respectively, and erythrocyte incorporation was measured in blood samples drawn 14 days later. We investigated how the iron-regulatory hormone hepcidin and other inflammatory/iron-related indices, all measured on the day of isotope administration, correlated with erythrocyte iron incorporation. In univariate analyses, hepcidin, ferritin, C-reactive protein, and soluble transferrin receptor (sTfR) strongly predicted incorporation of 57Fe given on day 1, while hepcidin, ferritin, and sTfR/log ferritin correlated with 58Fe incorporation. In a final multivariate model, the most consistent predictor of erythrocyte isotope incorporation was hepcidin. We conclude that under conditions of competing signals (anemia, iron deficiency, and infection), hepcidin powerfully controls use of dietary iron. We suggest that low-cost point-of-care hepcidin assays would aid iron supplementation programs in the developing world.

DNA methylation potential: dietary intake and blood concentrations of one-carbon metabolites and cofactors in rural African women

Background: Animal models show that periconceptional supplementation with folic acid, vitamin B-12, choline, and betaine can induce differences in offspring phenotype mediated by epigenetic changes in DNA. In humans, altered DNA methylation patterns have been observed in offspring whose mothers were exposed to famine or who conceived in the Gambian rainy season. Objective: The objective was to understand the seasonality of DNA methylation patterns in rural Gambian women. We studied natural variations in dietary intake of nutrients involved in methyl-donor pathways and their effect on the respective metabolic biomarkers. Design: In 30 women of reproductive age (18–45 y), we monitored diets monthly for 1 y by using 48-h weighed records to measure intakes of choline, betaine, folate, methionine, riboflavin, and vitamins B-6 and B-12. Blood biomarkers of these nutrients, S-adenosylhomocysteine (SAH), S-adenosylmethionine (SAM), homocysteine, cysteine, and dimethylglycine were also assessed monthly. Results: Dietary intakes of riboflavin, folate, choline, and betaine varied significantly by season; the most dramatic variation was seen for betaine. All metabolic biomarkers showed significant seasonality, and vitamin B-6 and folate had the highest fluctuations. Correlations between dietary intakes and blood biomarkers were found for riboflavin, vitamin B-6, active vitamin B-12 (holotranscobalamin), and betaine. We observed a seasonal switch between the betaine and folate pathways and a probable limiting role of riboflavin in these processes and a higher SAM/SAH ratio during the rainy season. Conclusions: Naturally occurring seasonal variations in food-consumption patterns have a profound effect on methyl-donor biomarker status. The direction of these changes was consistent with previously reported differences in methylation of metastable epialleles. This trial was registered at www.clinicaltrials.gov as NCT01811641.

Malaria in patients with sickle cell anemia: burden, risk factors, and outcome at the outpatient clinic and during hospitalization

Approximately 280,000 children are born with sickle cell anemia (SCA) in Africa annually, yet few survive beyond childhood. Falciparum malaria is considered a significant cause of this mortality. We conducted a 5-year prospective surveillance study for malaria parasitemia, clinical malaria, and severe malarial anemia (SMA) in Dar-es-Salaam, Tanzania, between 2004 and 2009. We recorded 10,491 visits to the outpatient clinic among 1808 patients with SCA and 773 visits among 679 patients without SCA. Similarly, we recorded 691 hospital admissions among 497 patients with SCA and 2017 in patients without SCA. Overall, the prevalence of parasitemia was lower in patients with SCA than in patients without SCA both at clinic (0.7% vs 1.6%; OR, 0.53; 95% CI, 0.32-0.86; P = .008) and during hospitalization (3.0% vs 5.6%; OR, 0.46; 95% CI, 0.25-0.94; P = .01). Furthermore, patients with SCA had higher rates of malaria during hospitalization than at clinic, the ORs being 4.29 (95% CI, 2.63-7.01; P < .001) for parasitemia, 17.66 (95% CI, 5.92-52.71; P < .001) for clinical malaria, and 21.11 (95% CI, 8.46-52.67; P < .001) for SMA. Although malaria was rare among patients with SCA, parasitemia during hospitalization was associated with both severe anemia and death. Effective treatment for malaria during severe illness episodes and further studies to determine the role chemoprophylaxis are required.

Genetics of fetal hemoglobin in Tanzanian and British patients with sickle cell anemia

Fetal hemoglobin (HbF, α(2)γ(2)) is a major contributor to the remarkable phenotypic heterogeneity of sickle cell anemia (SCA). Genetic variation at 3 principal loci (HBB cluster on chromosome 11p, HBS1L-MYB region on chromosome 6q, and BCL11A on chromosome 2p) have been shown to influence HbF levels and disease severity in β-thalassemia and SCA. Previous studies in SCA, however, have been restricted to populations from the African diaspora, which include multiple genealogies. We have investigated the influence of these 3 loci on HbF levels in sickle cell patients from Tanzania and in a small group of African British sickle patients. All 3 loci have a significant impact on the trait in both patient groups. The results suggest the presence of HBS1L-MYB variants affecting HbF in patients who are not tracked well by European-derived markers, such as rs9399137. Additional loci may be identified through independent genome-wide association studies in African populations.

Expression of the Iron Hormone Hepcidin Distinguishes Different Types of Anemia in African Children

The iron hormone hepcidin correctly identifies African children in whom iron supplementation is most likely to be beneficial. Hepcidin Guides Iron Supplementation in African Children Anemia affects 300 million preschool children worldwide and has multiple causes including iron deficiency or infection. Dietary iron supplementation is used to combat anemia, but substantial concerns exist that iron can be harmful, in part by promoting infection. Iron is likely to preferentially benefit children with iron deficiency anemia, but identification of such children is challenging. Hepcidin is the hormone that regulates body iron levels and inhibits absorption of iron from the diet. Hepcidin concentrations are generally low in iron deficiency but are raised in iron-replete individuals and are also increased by infection. In a new study, Pasricha et al. set out to investigate whether hepcidin measurements would enable targeting of interventions to children who need iron but who are also able to absorb it. They tested this by measuring hepcidin in three cohorts of preschool African children from The Gambia and Tanzania. Single cutoffs of hepcidin concentrations efficiently identified children with iron deficiency, distinguished between iron deficiency anemia and anemia due to infection and inflammation, and predicted which children would incorporate >20% of an oral iron dose into their red blood cells. Thus, hepcidin is a critical determinant of iron homeostasis and may be a useful marker to guide diagnosis of anemia and enable screen-and-treat iron supplementation programs. Childhood anemia is a major global health problem resulting from multiple causes. Iron supplementation addresses iron deficiency anemia but is undesirable for other types of anemia and may exacerbate infections. The peptide hormone hepcidin governs iron absorption; hepcidin transcription is mediated by iron, inflammation, and erythropoietic signals. However, the behavior of hepcidin in populations where anemia is prevalent is not well established. We show that hepcidin measurements in 1313 African children from The Gambia and Tanzania (samples taken in 2001 and 2008, respectively) could be used to identify iron deficiency anemia. A retrospective secondary analysis of published data from 25 Gambian children with either postmalarial or nonmalarial anemia demonstrated that hepcidin measurements identified individuals who incorporated >20% oral iron into their erythrocytes. Modeling showed that this sensitivity of hepcidin expression at the population level could potentially enable simple groupings of individuals with anemia into iron-responsive and non–iron-responsive subtypes and hence could guide iron supplementation for those who would most benefit.

Iron metabolism and malaria.

Recent evidence from a large, randomized, controlled trial has suggested that the universal administration of iron to children in malaria-endemic areas is associated with an increase in adverse health outcomes. The purpose of this paper is to summarize the available ecologic and intervention trials related to iron and malaria in children, and to set these against current knowledge of the biology of host–pathogen interactions involving iron metabolism. We conclude that, although not fully consistent, the balance of evidence confirms that administration of iron (usually in combination with folic acid) increases the incidence of malaria when given without prophylaxis and in the absence of universal access to treatment. The mechanisms by which additional iron can benefit the parasite are far from clear. There is evidence to suggest that the apparent detrimental effect of iron supplementation may vary according to levels of antecedent iron status, the presence of hemoglobinopathies and glucose-6-phosphate dehydrogenase (G6PD) deficiency, and other host genetic variants, such as variants in haptoglobin. The effects of malaria on host iron metabolism are also reviewed and reveal that the key cause of malaria-induced anemia is a maldistribution of iron and suppression of erythropoiesis rather than an exacerbation of gross iron deficiency. We tentatively conclude that, if it is to be recommended, universal iron supplementation in malarious areas should only be considered in conjunction with some form of prophylaxis (e.g., intermittent preventive therapy [IPT]) or in the context of good health services with ready access to facilities for malaria diagnosis and treatment. An alternative approach would be to screen for anemia and target supplementation only to anemic children. With regard to treatment, there is good evidence that iron supplementation should be withheld until the treatment schedule is complete, both because iron may inhibit treatment and because the absorption of oral iron is blocked by the inflammatory response.

Iron incorporation and post-malaria anaemia

Background Iron supplementation is employed to treat post-malarial anaemia in environments where iron deficiency is common. Malaria induces an intense inflammatory reaction that stalls reticulo-endothelial macrophagal iron recycling from haemolysed red blood cells and inhibits oral iron absorption, but the magnitude and duration of these effects are unclear. Methodology/Principal Findings We examined the red blood cell incorporation of oral administered stable isotopes of iron and compared incorporation between age matched 18 to 36 months old children with either anaemia post-malaria (n = 37) or presumed iron deficiency anaemia alone (n = 36). All children were supplemented for 30 days with 2 mg/kg elemental iron as liquid iron sulphate and administered 57Fe and 58Fe on days 1 and 15 of supplementation respectively. 57Fe and58Fe incorporation were significantly reduced (8% vs. 28%: p<0.001 and 14% vs. 26%: p = 0.045) in the malaria vs. non-malaria groups. There was a significantly greater haemoglobin response in the malaria group at both day 15 (p = 0.001) and 30 (p<0.000) with a regression analysis estimated greater change in haemoglobin of 7.2 g/l (s.e. 2.0) and 10.1 g/l (s.e. 2.5) respectively. Conclusion/Significance Post-malaria anaemia is associated with a better haemoglobin recovery despite a significant depressant effect on oral iron incorporation which may indicate that early erythropoetic iron need is met by iron recycling rather than oral iron. Supplemental iron administration is of questionable utility within 2 weeks of clinical malaria in children with mild or moderate anaemia.