Modulation of the HIF2-NCOA4 axis in enterocytes attenuates iron loading in a mouse model of hemochromatosis

Intestinal iron absorption is activated during increased systemic iron demand. The best-studied example is iron-deficiency anemia, which increases intestinal iron absorption. Interestingly, the intestinal response to anemia is very similar to that of iron overload disorders, as both the conditions activate a transcriptional program that leads to a hyperabsorption of iron via the transcription factor hypoxia-inducible factor (HIF)2a. However, pathways to selectively target intestinal-mediated iron overload remain unknown. Nuclear receptor co-activator 4 (NCOA4) is a critical cargo receptor for autophagic breakdown of ferritin (FTN) and subsequent release of iron, in a process termed ferritinophagy. Our work demonstrates that NCOA4-mediated intestinal ferritinophagy is integrated to systemic iron demand via HIF2a. To demonstrate the importance of intestinal HIF2a/ferritinophagy axis in systemic iron homeostasis, whole body and intestine-specific NCOA4-null mouse lines were generated and assessed. These analyses revealed that the intestinal and systemic response to iron deficiency was not altered following disruption of intestinal NCOA4. However, in a mouse model of hemochromatosis, ablation of intestinal NCOA4 was protective against iron overload. Therefore, NCOA4 can be selectively targeted for the management of iron overload disorders without disrupting the physiological processes involved in the response to systemic iron deficiency.

Intestinal ferritinophagy is regulated by HIF-2α and is essential for systemic iron homeostasis

Iron is critical for many processes including oxygen transport and erythropoiesis. Transcriptomic analysis demonstrates that HIF-2α regulates over 90% of all transcripts induced following iron deficiency in the intestine. However, beyond divalent metal transporter 1 (DMT1), ferroportin 1 (Fpn1) and duodenal cytochrome b (Dcytb), no other genes/pathways have been critically assessed with respects to their importance in intestinal iron absorption. Ferritinophagy is associated with cargo specific autophagic breakdown of ferritin and subsequent release of iron. We show here that nuclear receptor co-activator 4 (NCOA4)-mediated intestinal ferritinophagy is integrated to systemic iron demand via HIF-2α. Duodenal NCOA4 expression is regulated by HIF-2α during high systemic iron demands. Moreover, overexpression of intestinal HIF-2α is sufficient to activate NCOA4 and promote lysosomal degradation of ferritin. Promoter analysis revealed NCOA4 as a direct HIF-2α target. To demonstrate the importance of intestinal HIF-2α/ferritinophagy axis in systemic iron homeostasis, whole body and intestine-specific NCOA4-null mouse lines were assessed. These analyses demonstrate an iron sequestration in the enterocytes, and significantly high tissue ferritin levels in the dietary iron deficiency and acute hemolytic anemia models. Together, our data suggests efficient ferritinophagy is critical for intestinal iron absorption and systemic iron homeostasis.

A Review of Nutrients and Compounds, Which Promote or Inhibit Intestinal Iron Absorption: Making a Platform for Dietary Measures That Can Reduce Iron Uptake in Patients with Genetic Haemochromatosis

Objective. To provide an overview of nutrients and compounds, which influence human intestinal iron absorption, thereby making a platform for elaboration of dietary recommendations that can reduce iron uptake in patients with genetic haemochromatosis. Design. Review. Setting. A literature search in PubMed and Google Scholar of papers dealing with iron absorption. Results. The most important promoters of iron absorption in foods are ascorbic acid, lactic acid (produced by fermentation), meat factors in animal meat, the presence of heme iron, and alcohol which stimulate iron uptake by inhibition of hepcidin expression. The most important inhibitors of iron uptake are phytic acid/phytates, polyphenols/tannins, proteins from soya beans, milk, eggs, and calcium. Oxalic acid/oxalate does not seem to influence iron uptake. Turmeric/curcumin may stimulate iron uptake through a decrease in hepcidin expression and inhibit uptake by complex formation with iron, but the net effect has not been clarified. Conclusions. In haemochromatosis, iron absorption is enhanced due to a decreased expression of hepcidin. Dietary modifications that lower iron intake and decrease iron bioavailability may provide additional measures to reduce iron uptake from the foods. This could stimulate the patients’ active cooperation in the treatment of their disorder and reduce the number of phlebotomies.

LIC-R2 Values Predict Severity of SCD: Baseline Data from Licnet-S

Introduction Sickle cell disease (SCD) is a monogenic, yet highly phenotypically variable disease with multisystem pathology. HbS is prone to polymerization upon deoxygenation, and this property underlies all the pathophysiology of SCD. The net result is a chronic hemolytic anemia, with intravascular and extravascular components, and a tendency for microvascular obstruction or vaso-occlusion (VOC). The ability to predict ''severe disease'' depends on the ability to define it specifically and reproducibly. Many genetic, clinical and laboratory modifiers have been suggested as possible predictors of a "severe disease". Despite of this wealth of data the modelling of predictor variable continues to be difficult (Quinn et al, 2016). Considering that the hallmark of SCD is the chronic hemolitic anemia causing increasing of gastro-intestinal iron absorption, our aim was to evaluate if LIC-R2 (Ferriscan) determination may be related with phenotype severity of SCD. Methods This was a retrospective study of patients with SCD attending 7 Italian centres participating in the LICNET-S (Liver Iron Cutino NETwork in Sickle Cell Disease), collected at the Campus of Hematology Franco and Piera Cutino, AOOR Villa Sofia-V. Cervello (Palermo, Italy). The LICNET-S protocol was established on June 2018 by Foundation Franco and Piera Cutino of Palermo and approved by our Ethics Committee on 4, July 2018. This analysis included data from LIC-R2 for those patients presenting between July 2018 and July 2019. The MRI-R2 used protocol was that of St Pierre et al, 2005. Retrieved information included laboratory and clinical findings. These are shown as mean±sd and percentages. A linear regression model (LRM) (Draper & Smith, 1998) was used to study the correlation between LIC-R2 and some potential indicators of phenotype severity. All statistical analyses were performed using Stata 12 (StataCorp, College Station, TX, USA). Results Overall 65 patients (32 (49.3%) females) whom 16 patients with S/S, 15 with S/beta+ tahalseemia and 34 with S/beta0 thalassemia were enrolled in the study. Table 1 shows summary of the main clinical findings included in the study to evaluate the LIC-R2 predictivity based on SCD phenotype severity. Table 2 shows the results of the LRM analysis. The variables statistically significant were the number of the VOC, the Aseptic Avascular Necrosis (AVN) and the Chelation Treatment (CT) (Table 2). However, transfusion regimen was not related with LIC-R2 value, suggesting that chronic hemolitic anemia causing increasing of gastro-intestinal iron absorption rather than transfusion treatment was the main determinant in the LIC-R2 values. This issue is supported by the report of high LIC-R2 in non transfused patients with SCD (Yassim et al, 2017). Table 2 shows even the relationship between the impairment of the single indicator (VOC, AVN, CT) and the LIC-R2 value. All other laboratory and main clinical findings included in Table 2 were not statistically significant. The value of VOC in determining phenotype severity is well known. AVN is a consequence of a chronic ischaemia with inflammatory process evolving at the same time as medullary osteoblastic activity (Mukizi-Musaka et al, 2010). Therefore, it may be related with the chronic hemolitic anemia condition in SCD. Finally, the relationship between CT and LIC-R2 may be explained considering that patients with higher iron body burden are those who received earlier chelation treatment. Conclusions This study suggests, as LIC-R2 value is a strong predictive indicator of phenotype severity in SCD. Probably, this is due because of LIC-R2 is related both with acute and chronic hemolitic anemia of SCD causing an increase, during the years, of gastro-intestinal iron absorption. Therefore, the possibility of preventing VOC, even using new anti-sickling drugs, should be hardly pursued. Disclosures No relevant conflicts of interest to declare.

Iron and Zinc Homeostasis and Interactions: Does Enteric Zinc Excretion Cross-Talk with Intestinal Iron Absorption?

Iron and zinc are essential micronutrients required for growth and health. Deficiencies of these nutrients are highly prevalent among populations, but can be alleviated by supplementation and food fortification. Cross-sectional studies in humans showed positive association of serum zinc levels with hemoglobin and markers of iron status. Dietary restriction of zinc or intestinal specific conditional knock out of ZIP4 (SLC39A4), an intestinal zinc transporter, in experimental animals demonstrated iron deficiency anemia and tissue iron accumulation. Similarly, increased iron accumulation has been observed in cultured cells exposed to zinc deficient media. These results together suggest a potential role of zinc in modulating intestinal iron absorption and mobilization from tissues. Studies in intestinal cell culture models demonstrate that zinc induces iron uptake and transcellular transport via induction of divalent metal iron transporter-1 (DMT1) and ferroportin (FPN1) expression, respectively. It is interesting to note that intestinal cells are exposed to very high levels of zinc through pancreatic secretions, which is a major route of zinc excretion from the body. Therefore, zinc appears to be modulating the iron metabolism possibly via regulating the DMT1 and FPN1 levels. Herein we critically reviewed the available evidence to hypothesize novel mechanism of Zinc-DMT1/FPN1 axis in regulating intestinal iron absorption and tissue iron accumulation to facilitate future research aimed at understanding the yet elusive mechanisms of iron and zinc interactions.