Changes in Hepcidin Levels in an Animal Model of Anemia of Chronic Inflammation: Mechanistic Insights Related to Iron Supplementation and Hepcidin Regulation

Abstract

We examined changes in hepcidin (closely associated with anemia of chronic inflammation (ACI)) and upstream regulatory pathways after intravenous (IV) iron supplementation in an ACI animal model. ACI was induced in male Sprague-Dawley rats by intraperitoneally administering complete Freund’s adjuvant (CFA). Two weeks after starting CFA treatment, ACI rats received IV iron (CFA-iron) or vehicle (CFA-saline). Three days after IV iron treatment, iron profiles, hepcidin levels, and expression of proteins involved in the signaling pathways upstream of hepcidin transcription in the liver were measured. In CFA-treated rats, anemia with a concomitant increase in the levels of serum inflammatory cytokines and reactive oxygen species occurred. In CFA-iron rats, hemoglobin (Hb) concentration was still lower than that in control rats. In CFA-saline rats, hepatic hepcidin and ferritin levels increased compared with those in control rats and were further increased in CFA-iron rats. In CFA-saline rats, NADPH oxidase- (NOX-) 2, NOX-4, and superoxide dismutase levels in the liver were upregulated compared with those in control rats and their levels were further increased in CFA-iron rats. In CFA-saline rats, activities of the IL-6/STAT and BMP/SMAD pathways were enhanced in the liver compared with those in control rats and their levels were further increased in CFA-iron rats, whereas IL-6 expression remained unaffected after IV iron administration. In HepG2 cells, iron caused phosphorylation of STAT-3 and SMAD1/5 and knockdown of STAT-3 and SMAD1/5 using siRNAs reduced iron-induced hepcidin upregulation to levels similar to those in corresponding control cells. Renal erythropoietin expression and serum erythroferrone concentration were lower in CFA-iron rats than those in control rats. In ACI rats, IV iron supplementation did not recover Hb within three days despite an increase in hepatic ferritin levels, which might be attributable to an additional increase in hepcidin levels that was already upregulated under ACI conditions. Both STAT-3 phosphorylation and SMAD1/5 phosphorylation were associated with hepcidin upregulation after IV iron treatment, and this seems to be linked to iron-induced oxidative stress.

Control variables of serum ferritin concentrations in hospitalized newborn infants: an observational study

“…Since incomplete erythropoiesis leads to improper use of iron and destruction of immature erythrocytes and raises serum ferritin levels under anemic conditions [38] , immature response to erythropoietin in newborn infants might explain the observed inverse correlation between hemoglobin and ferritin levels. In addition, in cases when infection and in ammation coincide with anemia, elevated hepcidin expression may restrict iron supply and erythropoiesis, as shown in a rat model of anemia coincided with chronic in ammation [39] .…”

Control variables of serum ferritin concentrations in hospitalized newborn infants: an observational study

“…Since incomplete erythropoiesis leads to improper use of iron and destruction of immature erythrocytes and raises serum ferritin levels under anemic conditions 41 , immature response to erythropoietin in newborn infants might explain the observed inverse correlation between hemoglobin and ferritin levels. In addition, in cases when infection and inflammation coincide with anemia, elevated hepcidin expression may restrict iron supply and erythropoiesis, as shown in a rat model of anemia coincided with chronic inflammation 42 .…”

Critical re‐evaluation of the identification of iron deficiency states and effective iron repletion strategies in patients with chronic heart failure

“…Transcription of the hepcidin gene is highly sensitive to inflammatory states, and increases in proinflammatory cytokines and down-regulation of nutrient deprivation signalling promote increased hepcidin expression. [75][76][77][78] Hepcidin-induced functional iron deficiency is a principal cause of inflammation-induced "anaemia of chronic disease" (Figure 3), 79 as is seen in chronic kidney disease and chronic heart failure, [80][81][82] and hepatic congestion may further increase serum hepcidin. 83 Since hepcidin assays are not well standardized, 84 serum hepcidin is not typically measured in clinical practice.…”

“…Hepcidin-induced impairment of dietary iron absorption and occult gastrointestinal blood loss can cause absolute depletion, which can be superimposed on a functional iron deficiency state. Inflammation and oxidative stress also stimulate the transcription of ferritin and suppress ferritinophagy 76,86,87 (Figure 3); the increases in intracellular ferritin promote the sequestration of cytosolic iron. In most patients, quantification of the relative contributions of absolute and functional iron deficiency is exceedingly difficult.…”

“…The iron deficiency state is functional, since total body iron stores are adequate (even if not normal), but they cannot be fully mobilized. Transcription of the hepcidin gene is highly sensitive to inflammatory states, and increases in proinflammatory cytokines and down‐regulation of nutrient deprivation signalling promote increased hepcidin expression 75–78 . Hepcidin‐induced functional iron deficiency is a principal cause of inflammation‐induced “anaemia of chronic disease” ( Figure ), 79 as is seen in chronic kidney disease and chronic heart failure, 80–82 and hepatic congestion may further increase serum hepcidin 83 …”