BACKGROUND/OBJECTIVES:Liver fibrosis, if left untreated, can lead to cirrhosis and cancer. The current standard liver biopsy for fibrosis staging is invasive and prone to risks of complication. The objective of this study was to develop a new noninvasive method to quantify fibrosis using diamagnetic susceptibility sources generated from multi-echo gradient echo (mGRE) data with both magnitude decay R2* modeling and phase QSM modeling. METHODS:mGRE data of ex vivo liver explants was processed with fat-water separation and then susceptibility source separation. Negative susceptibility was used to measure diamagnetic fibrosis. In 20 formalin-fixed liver explant sections, negative susceptibility maps were compared with other MRI parameters against pathology for fibrosis staging. RESULTS:The correlation between the negative susceptibility sources and the fibrosis stages was evaluated with Spearman coefficients. Negative susceptibility differentiated (i) no or mild fibrosis (stages F0 to F1) from moderate-to-advanced fibrosis (stages F2 to F3; p = 0.0025), (ii) stages F2 to F3 from cirrhosis (stage F4; p = 0.021), and (iii) no-to-moderate fibrosis (stages F0 to F2) from advanced fibrosis or cirrhosis (stages F3 to F4) with a sensitivity of 90%, a specificity of 90%, and a 0.88 Receiver Operating Characteristic Area Under the Curve (AUC) (p = 0.0017). CONCLUSIONS:For staging fibrosis, negative susceptibility was superior to other MRI parameters, including R2*, QSM, and PDFF. Negative susceptibility sources were positively correlated with the fibrosis stage (r = 0.60). Negative susceptibility could be valuable for MRI staging in liver fibrosis.

To compare physiologically based serum hepcidin and ferritin thresholds for iron deficiency in adult female and male blood donors, we analysed laboratory results obtained during screening for a blood donor study. In 907 apparently healthy blood donors, we examined the relationships of hepcidin and ferritin with five indicators of the onset of iron-deficient red blood cell production: haemoglobin (Hb), soluble transferrin receptor (sTfR), erythrocyte zinc protoporphyrin (ZPP), reticulocyte haemoglobin content (Retic-Hb) and mean corpuscular volume (MCV). Serum hepcidin and ferritin were correlated (Pearson's r = 0.69, p = <0.001 females; r = 0.65, p < 0.001, males). At lower serum hepcidin and ferritin concentrations, Hb, Retic-Hb and MCV decreased and sTfR and ZPP increased. Using restricted cubic spline models, the hepcidin thresholds for iron deficiency in adult female donors 18-49 years, 50-75 years and all male blood donors 18-75 years old were 13.9, 24.8 and 28.8 μg/L respectively. The corresponding ferritin thresholds were 25.4, 30.7 and 32.5 μg/L respectively. Among blood donors 18-50 years old, serum hepcidin and ferritin thresholds for iron deficiency were lower in women than in men but were similar after 50 years of age. In conclusion, among adult blood donors, serum hepcidin and ferritin thresholds are similarly effective for detecting iron deficiency, especially among women.

Iron deficiency (ID) is the most predominant nutritional deficiency globally. The current CDC iron guideline for ID screening uses serum ferritin (Ft) ≤15 μg/L based on bone-marrow assessment. We aimed to evaluate physiologically based Ft thresholds for defining ID in children aged 5 to 14 years. Ferritin and hemoglobin (Hb) data were analyzed from 3765 apparently healthy (no inflammation, infection, risk of iron overload or potential liver disease) US children, and an indicator of iron-deficient erythropoiesis such as erythrocyte zinc protoporphyrin (ZPP). Nonlinear ferritin concentration curves (Hb-Ft, ZPP-Ft) and differential equations were applied to identify ferritin thresholds at ID onset with inflection minima and plateau for each iron analyte. Mean age was 10.3 years. The ferritin curve inflection for ID occurred at 23.9 μg/L (95% confidence interval, 21.7–27.3) with Hb and 25.0 μg/L (23.8–27.1) with ZPP. No inter-analyte (P = 0.67), age (P = 0.49 with Hb, P = 0.95 with ZPP), or sex (P = 0.39) differences observed. ID prevalence was 9.0% (7.3–10.7), and ID anemia (IDA) 0.5% (0.3–0.8) using CDC thresholds, versus ID 30.0% (27.4–32.5) and IDA 1.0% (0.6–1.3) % using physiologically based thresholds. Physiologically based ferritin thresholds for ID onset were consistently higher than current CDC thresholds across iron analyte, age, and sex. IDA remained unchanged with physiologically based thresholds, suggesting that anemia is not an adequate proxy for ID screening, and clinicians should consider higher ferritin as a functional threshold for defining ID in children aged 5-14 years.

Aim: Iron supplementation to African weaning infants was associated with increased enteropathogen levels. While cohort studies demonstrated that specific prebiotics inhibit enteropathogens during iron supplementation, their mechanisms remain elusive. Here, we investigated the in vitro impact of galacto-oligosaccharides (GOS) and iron-sequestering bovine lactoferrin (bLF) alone and combined on the gut microbiota of Kenyan infants during low-dose iron supplementation. Methods: Different doses of iron, GOS, and bLF were first screened during batch fermentations (n = 3), and the effect of these factors was studied on microbiota community structure and activity in the new Kenyan infant continuous intestinal PolyFermS model. The impact of different fermentation treatments on barrier integrity, enterotoxigenic Escherichia coli (ETEC) infection, and inflammatory response was assessed using a transwell co-culture of epithelial and immune cells. Results: A dose-dependent increase in short-chain fatty acid (SCFA) production, Bifidobacterium and Lactobacillus /Leuconostoc /Pediococcus (LLP) growth was detected with GOS alone and combined with bLF during iron supplementation in batches. This was confirmed in the continuous PolyFermS model, which also showed a treatment-induced inhibition of opportunistic pathogens C. difficile and C. perfringens . In all tests, supplementation of iron alone and combined with bLF did not have a significant effect on microbiota composition and activity. We observed a strengthening of the epithelial barrier and a decrease in cell death and pro-inflammatory response during ETEC infection with microbiota fermentation supernatants from iron + GOS, iron + bLF, and iron + GOS + bLF treatments compared to iron alone. Conclusion: Overall, beneficial effects on infant gut microbiota were shown using advanced in vitro models for GOS alone and combined with bLF during low-dose iron supplementation.

BACKGROUNDBlood donation increases the risk of iron deficiency, but its effect on brain iron, myelination, and neurocognition remains unclear.METHODSThis ancillary study enrolled 67 iron-deficient blood donors, 19-73 years of age, participating in a double-blind, randomized trial. After donating blood, positive and negative susceptibility were measured using quantitative susceptibility mapping (QSM) MRI to estimate brain iron and myelin levels, respectively. Furthermore, neurocognitive function was evaluated using the NIH Toolbox, and neural network activation patterns were assessed during neurocognitive tasks using functional MRI (fMRI). Donors were randomized to i.v. iron repletion (1 g iron) or placebo, and outcome measures repeated approximately 4 months later.RESULTSIron repletion corrected systemic iron deficiency and led to trends toward increased whole brain iron (P = 0.04) and myelination (P = 0.02), with no change in the placebo group. Although overall cognitive performance did not differ significantly between groups, iron-treated participants showed improved engagement of functional neural networks (e.g., memory pattern activation during speed tasks, P < 0.001). Brain region-specific changes in iron and myelin correlated with cognitive performance: iron in the putamen correlated with working memory scores (P < 0.01), and thalamic myelination correlated with attention and inhibitory control (P < 0.01).CONCLUSIONIron repletion in iron-deficient blood donors may influence brain iron, myelination, and function, with region-specific changes in iron and myelination linked to distinct cognitive domains.REGISTRATIONClinicalTrials.gov NCT02990559FUNDINGThis work was funded by the NIH.

BACKGROUND:Diagnosis of iron deficiency commonly relies on measurement of serum ferritin concentrations. WHO guidelines identify serum ferritin thresholds for iron deficiency among healthy individuals of less than 15 μg/L for women and less than 12 μg/L for children under 5 years, based on expert opinion. We report thresholds for iron deficiency for apparently healthy non-pregnant women and young children based on physiological indicators. METHODS:We performed secondary analyses of cross-sectional data from women (aged 15-49 years) and children (aged 6-59 months) from 12 countries in Africa, Asia, Europe, and central America from available surveys (2007-19). Using haemoglobin and soluble transferrin receptor concentrations as individual-level indicators of iron deficiency, we identified country-specific serum ferritin thresholds. We conducted multivariate meta-analysis using individual participant data to assess multinational heterogeneity and intercountry consistency. FINDINGS:Data were collected from July, 2007 to March, 2019. 18 251 individuals (13 864 women and 4387 children) were included in the final analysis. The thresholds of pooled serum ferritin levels corresponding to the starting point of decline in circulating haemoglobin concentrations were 24·8 μg/L (95% CI 24·4-25·2) for women and 22·1 μg/L (20·8-23·4) for children based on the national survey data from 12 countries. The thresholds were consistent among countries (pheterogeneity: women=0·73, children=0·43) but median serum ferritin concentrations and lower 5% reference ranges differed. In all countries, the prevalence of iron deficiency was higher using physiologically based thresholds than that using WHO current guidelines for women (36·0% [95% CI 25·3-46·8] vs 20·1% [11·5-28·7], p<0·0001) and for children (34·2% [24·3-44·1] vs 16·6% [11·2-22·0], p<0·0001). INTERPRETATION:These results provide evidence that the prevalence of iron deficiency as indicated by physiological measures is substantially higher than those based on current WHO guidelines. The consistency of physiologically based serum ferritin thresholds in apparently healthy women and young children offers a potential means to achieve evidence-informed coordination in thresholds for iron deficiency across populations. The use of physiologically based serum ferritin thresholds could help in detecting the clinical and functional outcomes of iron deficiency. FUNDING:None.

World Health Organization (WHO) and Centers for Disease Control and Prevention (CDC) guidelines recommend a serum ferritin (SF) threshold for iron deficiency (ID) of <15μg/L and ≤ 15μg/L, respectively, for healthy adults aged 15–60+ years. In apparently healthy adults, we used SF at which the circulating hemoglobin (Hb) begins to decrease and the erythrocyte zinc protoporphyrin (eZnPP) begins to increase as a potential physiological indicator of the ID threshold. We analyzed data for 5169 men and 6957 non-pregnant women, aged 15–90 years, from the Third National Health and Nutrition Examination Survey (NHANES III, 1988–1994). Physiologically based SF thresholds for ID were higher than WHO and CDC recommendations (P<0.001 for all). For men and postmenopausal women, SF thresholds corresponding to the initial decline in circulating Hb did not differ significantly, with an overall threshold of 32.6 μg/L (95% CI: (27.4, 37.7)). The SF threshold for premenopausal women was 24.8 μg/L (23.4, 26.9), lower than that for men and postmenopausal women (P<0 .0001). The difference between SF physiologically based thresholds in men and older women, with basal iron losses, compared to younger women, with menstrual/basal iron losses, provides evidence that hepcidin regulation of iron homeostasis controls the onset for ID in healthy adults. Clinically, use of higher physiologically based SF thresholds for ID for adults may lead to earlier diagnosis of increased blood loss.

Serum ferritin (SF) concentration is the most widely used indicator for iron deficiency (ID). During pregnancy, the World Health Organization recently recommended SF thresholds for ID of <15 mu g/L for the first trimester of pregnancy, based on expert opinion, and made no recommendations for the second and third trimesters. We examined the relationship of SF with 2 independent indicators of the onset of iron-deficient erythropoiesis, hemoglobin and soluble transferrin receptor 1, in cross-sectional data from US National Health and Nutrition Examination Survey for 1999 to 2010 and 2015 to 2018. We included 1288 pregnant women aged 15 to 49 years and excluded women with inflammation or potential liver disease. We used restricted cubic spline (RCS) regression analysis to determine SF thresholds for iron-deficient erythropoiesis. SF decreased during pregnancy; geometric mean SF was higher during the first and lower during the second and third trimesters. Using RCS analysis, the SF thresholds identified during pregnancy were <25.8 mu g/L (18.1-28.5) during first trimester, <18.3 mu g/L (16.3-22.9) during second trimester, and <19.0 mu g/L (14.4- 26.1) during third trimester. These SF threshold levels track concentrations of hepcidin, the iron-regulatory hormone controlling the mobilization of iron stores. An SF concentration of <15 mu g/L as the criterion for ID may underestimate the true prevalence of ID throughout pregnancy. In our study, an additional 1 of every 10 pregnant women would be recognized as iron deficient by using the physiologically based thresholds at SF of similar to 25 mu g/L during the first and similar to 20 mu g/L during the second and third trimesters.

SummaryIncreased iron loss may reduce the effectiveness of iron supplementation. The objective of this study was to determine if daily oral iron supplementation increases iron loss, measured using a stable isotope of iron (58Fe). We enrolled and dewormed 24 iron‐depleted Kenyan children, 24–27 months of age, whose body iron was enriched and equilibrated with 58Fe given at least 1 year earlier. Over 3 months of supplementation (6 mg iron/kg body weight [BW]/day), mean (±SD) iron absorption was 1.10 (±0.28) mg/day. During supplementation, 0.55 (±0.36) mg iron/day was lost, equal to half of the amount of absorbed iron. Supplementation did not increase faecal haem/porphyrin or biomarkers of enterocyte damage and gut or systemic inflammation. Using individual patient data, we examined iron dose, absorption and loss among all available long‐term iron isotopic studies of supplementation. Expressed in terms of body weight, daily iron loss was correlated significantly with iron absorption (Pearson's r = 0.66 [95% confidence interval 0.48–0.78]) but not with iron dose (r = 0.16 [95% CI −0.10–0.40]). The results of this study indicate that iron loss is increased with daily oral iron supplementation and may blunt the efficacy of iron supplements in children. This study was registered at ClinicalTrials.gov as NCT04721964.

Background: In otherwise healthy iron-deficient adults, the effects of systemic iron repletion on brain iron and iron-requiring myelination have not been characterized. However, in animal studies, the iron-deficient brain rapidly regains iron following systemic iron repletion. Iron is required for producing vital neurotransmitters in the brain and for synthesizing myelin to facilitate memory and learning by increasing axonal conduction velocity. Myelination is now known to continue throughout adult life. Therefore, we examined the effect of iron repletion on levels of iron and myelin in specific regions of interest in the brain and the correlation of these levels with cognitive performance in iron-deficient blood donors. Methods: This ancillary study enrolled 67 healthy, frequent blood donors, 19-73 years of age and 70% female, who were participating in a parent double-blind, randomized trial. Enrolled donors were initially non-anemic and met all donation standards, but were iron deficient by laboratory criteria (i.e., ferritin <15 ng/mL and zinc protoporphyrin >60 mMol/mol heme). After donating blood, brain iron and myelination were measured using Quantitative Susceptibility Mapping (QSM) MRI with magnetic susceptibility source separation, neurocognitive function was evaluated using the NIH Toolbox, and neural network activation patterns were assessed during neurocognitive tasks using functional MRI (fMRI). Donors were randomized to intravenous iron repletion (i.e., one-gram iron) or placebo, and outcome measures were repeated approximately four months later. The primary null and all secondary hypotheses were tested using linear mixed models for repeated measures to compare differences in the iron repletion and placebo group temporal course at the two defined time points when adjusting for sex, age, and the interaction between sex and age. We used a P-value of P< 0.01 to determine significance. Results: Baseline demographic data did not differ significantly between the iron repletion and placebo groups. After randomization, iron repletion corrected systemic iron deficiency, increasing hemoglobin by a mean of 1.5 g/dL (95% CI 1.1-1.8), and ferritin by a mean of 66 μg/L (49-83). Comparing iron- and placebo-treated subjects, iron repletion produced no significant differences in NIH toolbox measures of overall cognitive performance. Nonetheless, iron repletion improved the utilization of neural network activation patterns in specific domains (e.g., memory pattern score during speed tasks, P<0.001). Iron repletion also produced trends towards increased whole brain iron (P=0.04) and myelination (P=0.02); the placebo had no significant effect. In specific regions of the brain, changes in iron and myelin levels correlated with changes in NIH Toolbox measures of executive function, episodic memory, working memory, and attention and inhibitory control. For example, iron in the putamen, a brain structure regulating movement and various types of learning, correlated with the NIH Toolbox measures of episodic and working memory (Picture Sequencing and List Sorting tests, P<0.01). Myelination in the thalamus, a structure involved in attention, memory, and sensory processing, correlated with the measure of attention and inhibitory control (Flanker Inhibitory Control test, P<0.01). In contrast, changes in cognitive performance were not associated with changes in systemic measures of iron status, such as hemoglobin or ferritin levels. Conclusion: To our knowledge, these are the first studies in otherwise healthy iron-deficient adults to examine the association between measures of neurocognition and the effects of systemic iron repletion on iron and myelin in pre-defined regions of interest in the brain. The results provide evidence that changes in brain iron and myelination in specific brain structures in iron-deficient adult blood donors alter distinct cognitive functions.

In chronic liver disease, liver fibrosis develops as excessive deposition of extracellular matrix macromolecules, predominantly collagens, progressively form fibrous scars that disrupt the hepatic architecture, and fibrosis, iron, and fat are interrelated. Fibrosis is the best predictor of morbidity and mortality in chronic liver disease but liver biopsy, the reference method for diagnosis and staging, is invasive and limited by sampling and interobserver variability and risks of complications. The overall objective of this study was to develop a new non-invasive method to quantify fibrosis using diamagnetic susceptibility sources with histology validation in ex vivo liver explants.

Background Acute chest syndrome (ACS) is an acute complication in SCD but its effects on lung function are not well understood. Inflammation is a key component of SCD pathophysiology but with an unclear association with lung function. We hypothesized that children with ACS had worse lung function than children without ACS and aimed to investigate the association of lung function deficits with inflammatory cytokines. Methods Patients enrolled in a previous 2-year randomized clinical trial who had consented to future data use, were enrolled for the present exploratory study. Patients were categorized into ACS and non-ACS groups. Demographic and clinical information were collected. Serum samples were used for quantification of serum cytokines and leukotriene B4 levels and pulmonary function tests (PFTs) were assessed. Results Children with ACS had lower total lung capacity (TLC) at baseline and at 2 years, with a significant decline in forced expiratory volume in 1 sec (FEV1) and mid-maximal expiratory flow rate (FEF25-75%) in the 2 year period (p = 0.015 and p = 0.039 respectively). For children with ACS, serum cytokines IL-5, and IL-13 were higher at baseline and at 2 years compared to children with no ACS. IP-10 and IL-6 were negatively correlated with PFT markers. In multivariable regression using generalized estimating equation approach for factors predicting lung function, age was significantly associated FEV1 (p = 0.047) and ratio of FEV1 and forced vital capacity (FVC)- FEV1/FVC ratio (p = 0.006); males had lower FEV1/FVC (p = 0.035) and higher TLC (p = 0.031). Asthma status was associated with FEV1 (p = 0.017) and FVC (p = 0.022); history of ACS was significantly associated with TLC (p = 0.027). Conclusion Pulmonary function abnormalities were more common and inflammatory markers were elevated in patients with ACS, compared with those without ACS. These findings suggest airway inflammation is present in children with SCD and ACS, which could be contributing to impaired pulmonary function.

Severe anemia is commonly treated with red blood cell transfusion. Clinical trials have demonstrated that a restrictive transfusion strategy of 7 to 8 g/dL is as safe as a liberal transfusion strategy of 9 to 10 g/dL in many clinical settings. Evidence is lacking for subgroups of patients, including those with preexisting coronary artery disease, acute myocardial infarction, congestive heart failure, and myelodysplastic neoplasms. We present 3 clinical vignettes that highlight the clinical challenges in caring for patients with coronary artery disease with gastrointestinal bleeding, congestive heart failure, or myelodysplastic neoplasms. We emphasize that transfusion practice should be guided by patient symptoms and preferences in conjunction with the patient's hemoglobin concentration. Along with the transfusion decision, evaluation and management of the etiology of the anemia is essential. Iron-restricted erythropoiesis is a common cause of anemia severe enough to be considered for red blood cell transfusion but diagnosis and management of absolute iron deficiency anemia, the anemia of inflammation with functional iron deficiency, or their combination may be problematic. Intravenous iron therapy is generally the treatment of choice for absolute iron deficiency in patients with complex medical disorders, with or without coexisting functional iron deficiency.

Background: Current WHO serum ferritin (SF) thresholds for iron deficiency (ID) in children (<12 mu g/L) and women (<15 mu g/L) are derived from expert opinion based on radiometric assays in use decades ago. Using a contemporary immunoturbidimetry assay, higher thresholds (children, <20 mu g/L; women, <25 mu g/L) were identified from physiologically based analyses.Objective: We examined relationships of SF measured using an immunoradiometric assay from the era of expert opinion with 2 inde-pendently measured indicators of ID, hemoglobin (Hb) and erythrocyte zinc protoporphyrin (eZnPP), using data from the Third National Health and Nutrition Examination Survey (NHANES III, 1988-1994). The SF at which circulating Hb begins to decrease and eZnPP begins to increase provides a physiological basis for identifying the onset of iron-deficient erythropoiesis.Methods: We analyzed NHANES III cross-sectional data from 2616 apparently healthy children, aged 12-59 mo, and 4639 apparently healthy nonpregnant women, aged 15-49 y. We used restricted cubic spline regression models to determine SF thresholds for ID.Results: SF thresholds identified by Hb and eZnPP did not differ significantly in children, 21.2 mu g/L (95% confidence interval: 18.5, 26.5) and 18.7 mu g/L (17.9, 19.7), and, in women, were similar although significantly different, 24.8 mu g/L (23.4, 26.9) and 22.5 mu g/L (21.7, 23.3).Conclusions: These NHANES results suggest that physiologically based SF thresholds are higher than the thresholds from expert opinion established during the same era. SF thresholds found using physiological indicators detect the onset of iron-deficient erythropoiesis, whereas the WHO thresholds identify a later, more severe stage of ID.

Objective To evaluate the frequency of iron status assessment in pediatric heart failure and the prevalence and adverse effects of absolute iron deficiency in dilated cardiomyopathy-induced heart failure. Study design We retrospectively reviewed records of children with chronic heart failure at our center between2010 and 2020. In children with dilated cardiomyopathy, we analyzed baseline cardiac function, hemoglobin level, and subsequent risk of composite adverse events (CAE), including death, heart transplant, ventricular assist device(VAD) placement, and transplant registry listing. Absolute iron deficiency and iron sufficiency were defined as transferrin saturations <20% and(3)30%, respectively; and indeterminant iron status as 20%-29%. Results Of 799 patients with chronic heart failure, 471 (59%) had no iron-related laboratory measurements. Of 68children with dilated cardiomyopathy, baseline transferrin saturation, and quantitative left ventricular ejection fraction (LVEF), 33 (49%) and 14 (21%) were iron deficient and sufficient, respectively, and 21 (31%) indeterminant. LVEF was reduced to 23.6 +/- 12.1% from 32.9 +/- 16.8% in iron deficiency and sufficiency, respectively (P= .04),without a significant difference in hemoglobin. After stratification by New York Heart Association classification, in advanced class IV, hemoglobin was reduced to 10.9 +/- 1.3 g/dL vs 12.7 +/- 2.0 g/dL in iron deficiency and sufficiency, respectively (P= .01), without a significant difference in LVEF. Conclusions In this single-center study, iron deficiency was not monitored in most children with chronic heart failure. In pediatric dilated cardiomyopathy-induced heart failure, absolute iron deficiency was prevalent and associated with clinically consequential and possibly correctable decreases in cardiac function and hemoglobin con-centration.(J Pediatr 2023;263:113721)