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Background: Hereditary hemochromatosis (HH) is an iron overload disorder and can be caused by variants in non-HFE genes in Chinese patients. However, there is still a considerable proportion of patients suffering from unexplained iron overload. In our previous study, we had identified the p.R269Q variant in exon 4 of the Bone morphogenetic protein 4 (BMP4) gene in Chinese patients with unexplained primary iron overload by Whole Exome sequencing, and then the BMP4 p.H251Y variant was identified by Sanger sequencing in a Chinese patient with secondary iron overload. Our study aimed to explore the pathogenicity and underlying mechanism of BMP4 p.H251Y and BMP4 p.R269Q variants in patients with iron overload., Methods: Sanger sequencing was conducted to identify the novel variants in the BMP4 gene of patients with unexplained iron overload. MRI and liver biopsy were used to display iron overload in the liver of the patient harboring the BMP4 p.H251Y variant. The BMP4 and hepcidin levels in BMP4 knockdown and BMP4 variant cells were examined by enzyme-linked immunosorbent assay. The effects of BMP4 p.H251Y and BMP4 p.R269Q variants on the hepcidin-regulation pathway were studied., Results: One of 54 HH patients (1.85%) harbored the BMP4 p.R269Q variant. One of 148 patients (0.68%) with secondary hemochromatosis harbored the BMP4 p.H251Y variant, and these two variants were not found in 100 Chinese general population. For the patient harboring the BMP4 p.H251Y variant, abdominal MRI and Perl's staining of liver tissue displayed iron overload in the liver. Cells transfected with the BMP4 p.H251Y and p.R269Q variants showed down-regulation of hepcidin level and BMP/SMAD pathway compared with cells transfected with the wild-type BMP4 vector., Conclusion: The BMP4 p.H251Y and p.R269Q variants can downregulate hepcidin levels by inhibiting the BMP/SMAD axis, suggesting they may play pathogenic roles in iron overload., Competing Interests: Declarations Ethics approval and consent to participate This study was approved by the Clinical Research Ethics Committee of Beijing Friendship Hospital, Capital Medical University (No. 2016-P2-061–01). Informed and written consent was obtained from all patients. The study was registered with the ClinicalTrials.gov identifier NCT03131427. Consent for publication All authors agreed on the manuscript. Competing interests The authors declared that they have no competing interests., (© 2024. The Author(s).)

Hepcidin belongs to a class of small cationic antimicrobial peptides rich in cysteine. It is synthesized by liver and is widely involved in host antimicrobial, antiviral and other immune responses. We identified and characterized three hepcidin genes (OpHep1, OpHep2 and OpHep3) in spotted knifejaw. All the OpHeps shared high identities with hepcidins in other teleost, containing alpha helix and β-sheets. Three OpHeps were all detected in healthy tissues, with the abundant expression in liver. They were significantly increased after Vibrio harveyi infection in the six immune-relevant tissues (liver, kidney, spleen, gill, skin and intestine). OpHeps knockdown in spotted knifejaw liver cells affected the mRNA levels of inflammation-related genes, including il1β, il6, il8, and nfκb. Further, the recombinant hepcidin proteins were effective in suppressing the growth of both Gram-negative and Gram-positive bacteria. To identify the function of OpHeps in vivo, we performed the overexpression of three OpHeps in zebrafish, and found OpHeps could significantly induce immune-related genes expression in transgenic zebrafish, including myd88, il10, il21, il16, tlr1, tlr3 and lysozyme. When infected with V. harveyi, OpHeps transgenic zebrafishes had a higher survival rate than wild-type zebrafishes. The expression of myd88, il10, il8, il1β, nfκb and lysozyme were all significantly up-regulated in transgenic fishes during bacterial infection. In summary, these results indicated that hepcidin could protect fish fight against pathogen through TLR/NFκB signaling cascade and Lysozyme. Three OpHeps would be potential targets for prevention of bacterial infections in aquaculture industry of spotted knifejaw, which provided a new idea for the molecular breeding of fish disease resistance., Competing Interests: Declaration of competing interest The authors read and approved this version of the final manuscript, confirmed the integrity of this work and declared no conflicts of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Increased iron level is detected in rat kidney and human urine in diabetic condition and implicated in associated nephropathy. However, the biological cue and mechanism of the iron accumulation remain unclear. Here we reveal that glucose increases iron uptake by promoting transferrin receptor 1 (TFRC) in kidney cells by a translational mechanism but does not alter expression of endosomal iron transporter DMT1. Glucose decreases iron exporter ferroportin (FPN) by a protein degradation mechanism. Hepcidin is known to bind at Cys-326 residue in promoting degradation of human ferroportin. When Cys-326 was mutated to Ser in human-FPN-FLAG and expressed in kidney cells, glucose still could degrade FPN-FLAG implicating involvement of hepcidin independent mechanism in glucose induced ferroportin degradation. Chronic hyperglycemia was generated in rats by administering streptozotocin (STZ) with periodic insulin injection to determine the level of iron homeostasis components. Increased TFRC and decreased ferroportin levels were detected in hyperglycemic rat kidney by Western blot and immunohistochemistry analyses. Hepcidin mRNA was not significantly altered in kidney but was marginally decreased in liver. Perls' staining and non-heme iron estimation showed an elevated iron level in hyperglycemic rat kidney. These results suggest that high glucose dysregulates iron transport components resulting iron accumulation in diabetic kidney., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Preterm infants are most at risk of iron deficiency. However, our knowledge of the regulation of iron homeostasis in preterm infants is poor. The main goal of our research was to develop and validate an animal model of human prematurity to assess iron status in preterm infants. We performed a cesarean section on sows on the 109th day of pregnancy, which corresponds to the last trimester of human pregnancy. Preterm piglets showed decreased body weight, red blood cell indices, plasma iron level and transferrin saturation. Interestingly, higher hepatic and splenic non-heme iron content and plasma and hepatic ferritin levels were found in premature piglets compared with term ones. In addition, premature piglets showed higher mRNA levels of iron-regulatory hormone hepcidin in the liver than term animals, which have not been reflected in higher plasma hepcidin-25 levels. We also showed changes in hepcidin regulators, including hepatic bone morphogenetic protein 6, plasma erythroferrone and growth differentiation factor 15 in preterm piglets. Consequently, no difference was observed in iron-exporter ferroportin levels in the spleen and liver. Overall, it seems that premature piglets show a pattern of iron metabolism characteristic of functional iron deficiency and iron accumulation in the tissue.

Heterozygous mutations in SLC40A1, encoding a multi-pass membrane protein of the major facilitator superfamily known as ferroportin 1 (FPN1), are responsible for two distinct hereditary iron-overload diseases: ferroportin disease, which is associated with reduced FPN1 activity (i.e., decrease in cellular iron export), and SLC40A1-related hemochromatosis, which is associated with abnormally high FPN1 activity (i.e., resistance to hepcidin). Here, we report three SLC40A1 missense variants with opposite functional consequences. In cultured cells, the p.Arg40Gln and p.Ser47Phe substitutions partially reduced the ability of FPN1 to export iron and also partially reduced its sensitivity to hepcidin. The p.Ala350Val substitution had more profound effects, resulting in low FPN1 iron egress and weak FPN1/hepcidin interaction. Structural analyses helped to differentiate the first two substitutions, which are predicted to cause local instabilities, and the third, which is thought to prevent critical rigid-body movements that are essential to the iron transport cycle. The phenotypic traits observed in a total of 12 affected individuals are highly suggestive of ferroportin disease. Our findings dismantle the classical dualism of FPN1 loss versus gain of function, highlight some specific and unexpected functions of FPN1 transmembrane helices in the molecular mechanism of iron export and its regulation by hepcidin, and extend the spectrum of rare genetic variants that may cause ferroportin disease., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The regulation of iron (Fe) levels is essential to maintain an adequate supply for erythropoiesis, among other processes, and to avoid possible toxicity. The liver-produced peptide hepcidin is regarded as the main regulator of Fe absorption in enterocytes and release from hepatocytes and macrophages, as it impairs Fe export through ferroportin. The glycoprotein erythropoietin (Epo) drives erythroid progenitor survival and differentiation in the bone marrow, and has been linked to the mobilization of Fe reserves necessary for hemoglobin production. Herein we show that Epo inhibits hepcidin expression directly in the HepG2 hepatic cell line, thus leading to a decrease in intracellular Fe levels. Such inhibition was dependent on the Epo receptor-associated kinase JAK2, as well as on the PI3K/AKT/mTOR pathway, which regulates nutrient homeostasis. Epo was also found to decrease binding of the C/EBP-α transcription factor to the hepcidin promoter, which could be attributed to an increased expression of its inhibitor CHOP. Epo did not only hinder the stimulating effect of C/EBP-α on hepcidin transcription, but also favored hepcidin inhibition by HIF-1α, by increasing is nuclear translocation as well as its protein levels. Moreover, in assays with the inhibitor genistein, this transcription factor was found necessary for Epo-induced hepcidin suppression. Our findings support the involvement of the PI3K/AKT/mTOR pathway in the regulation of Fe levels by Epo, and highlight the contrasting roles of the C/EBP-α and HIF-1α transcription factors as downstream effectors of the cytokine in this process., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Abstract: Ferroportin (Fpn) is the only iron exporter, playing a crucial role in systemic iron homeostasis. Fpn is negatively regulated by its ligand hepcidin, but other potential regulators in physiological and disease conditions remain poorly understood. Diabetes is a metabolic disorder that develops body iron loading with unknown mechanisms. By using diabetic mouse models and human duodenal specimens, we demonstrated that intestinal Fpn expression was increased in diabetes in a hepcidin-independent manner. Protein kinase C (PKC) is hyperactivated in diabetes. We showed that PKCα was required to sustain baseline Fpn expression and diabetes-induced Fpn upregulation in the enterocytes and macrophages. Knockout of PKCα abolished diabetes-associated iron overload. Mechanistically, activation of PKCα increased the exocytotic trafficking of Fpn and decreased the endocytic trafficking of Fpn in the resting state. Hyperactive PKCα also suppressed hepcidin-induced ubiquitination, internalization, and degradation of Fpn. We further observed that iron loading in the enterocytes and macrophages activated PKCα, acting as a novel mechanism to enhance Fpn-dependent iron efflux. Finally, we demonstrated that the loss-of-function of PKCα and pharmacological inhibition of PKC significantly alleviated hereditary hemochromatosis-associated iron overload. Our study has highlighted, to our knowledge, for the first time, that PKCα is an important positive regulator of Fpn and a new target in the control of iron homeostasis.

Abstract: The liver plays a crucial role in maintaining systemic iron homeostasis by secreting hepcidin, which is essential for coordinating iron levels in the body. Imbalances in iron homeostasis are associated with various clinical disorders related to iron deficiency or iron overload. Despite the clinical significance, the mechanisms underlying how hepatocytes sense extracellular iron levels to regulate hepcidin synthesis and iron storage are not fully understood. In this study, we identified Foxo1, a well-known regulator of macronutrient metabolism, which translocates to the nucleus of hepatocytes in response to high-iron feeding, holo-transferrin, and bone morphogenetic protein 6 (BMP6) treatment. Furthermore, Foxo1 plays a crucial role in mediating hepcidin induction in response to both iron and BMP signals by directly interacting with evolutionally conserved Foxo binding sites within the hepcidin promoter region. These binding sites were found to colocalize with Smad-binding sites. To investigate the physiological relevance of Foxo1 in iron metabolism, we generated mice with hepatocyte-specific deletion of Foxo1. These mice exhibited reduced hepatic hepcidin expression and serum hepcidin levels, accompanied by elevated serum iron and liver nonheme iron concentrations. Moreover, high-iron diet further exacerbated these abnormalities in iron metabolism in mice lacking hepatic Foxo1. Conversely, hepatocyte-specific Foxo1 overexpression increased hepatic hepcidin expression and serum hepcidin levels, thereby ameliorating iron overload in a murine model of hereditary hemochromatosis (Hfe-/- mice). In summary, our study identifies Foxo1 as a critical regulator of hepcidin and systemic iron homeostasis. Targeting Foxo1 may offer therapeutic opportunities for managing conditions associated with aberrant iron metabolism., (© 2024 American Society of Hematology. Published by Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

Pathogenic variants in HFE and non-HFE genes have been identified in hemochromatosis in different patient populations, but there are still a certain number of patients with unexplained primary iron overload. We recently identified in Chinese patients a recurrent p.(Arg639Gln) variant in SURP and G-patch domain containing 2 (SUGP2), a potential mRNA splicing-related factor. However, the target gene of SUGP2 and affected iron-regulating pathway remains unknown. We aimed to investigate the pathogenicity and underlying mechanism of this variant in hemochromatosis. RNA-seq analysis revealed that SUGP2 knockdown caused abnormal alternative splicing of CIRBP pre-mRNA, resulting in an increased normal splicing form of CIRBP V1, which in turn increased the expression of BMPER by enhancing its mRNA stability and translation. Furthermore, RNA-protein pull-down and RNA immunoprecipitation assays revealed that SUGP2 inhibited splicing of CIRBP pre-mRNA by a splice site variant at CIRBP c.492 and was more susceptible to CIRBP c.492 C/C genotype. Cells transfected with SUGP2 p.(Arg639Gln) vector showed up-regulation of CIRBP V1 and BMPER expression and down-regulation of pSMAD1/5 and HAMP expression. CRISPR-Cas9 mediated SUGP2 p.(Arg622Gln) knock-in mice showed increased iron accumulation in the liver, higher total serum iron, and decreased serum hepcidin level. A total of 10 of 54 patients with hemochromatosis (18.5%) harbored the SUGP2 p.(Arg639Gln) variant and carried CIRBP c.492 C/C genotype, and had increased BMPER expression in the liver. Altogether, the SUGP2 p.(Arg639Gln) variant down-regulates hepcidin expression through the SUGP2/CIRBP/BMPER axis, which may represent a novel pathogenic factor for hemochromatosis., (© 2024 The Author(s). American Journal of Hematology published by Wiley Periodicals LLC.)

Introduction: Hepcidin, a central regulatory molecule of iron metabolism, is upregulated through the IL-6/STAT3 signaling pathway in inflammatory and infectious states, contributing to the pathogenesis of various diseases. In chronic apical periodontitis (CAP), Porphyromonas gingivalis (P. gingivalis) and its lipopolysaccharides (LPS) activate various immune responses in vivo, contributing to disease progression. This study evaluated the role and mechanism of hepcidin in P. gingivalis-induced bone tissue damage in CAP, focusing on its promotion of macrophage M1 polarization via the IL-6/STAT3 signaling pathway., Methods: We analyzed a GSE77459 dataset from the GEO database, containing data from inflammatory and normal dental pulp tissues. RT-qPCR and immunofluorescence staining were used to detect the expression of hepcidin in human CAP tissues and its relationship with macrophages. Mouse bone marrow derived macrophages (BMDMs) were cultured in vitro and stimulated with P. gingivalis LPS. The effects of Stattic on macrophage hepcidin expression, IL-6 expression, STAT3 phosphorylation, and macrophage polarization were detected by ELISA, western blotting, RT-qPCR, and flow cytometry, respectively., Results: Hepcidin expression in human inflammatory dental pulp tissues was upregulated via the IL-6/STAT3 pathway and correlated with macrophage polarization. Hepcidin-encoding genes were found to be highly expressed and primarily associated with M1 macrophages in CAP tissues. In vitro experiments revealed that P. gingivalis LPS stimulation induced macrophages to express hepcidin through the IL-6/STAT3 pathway and polarize to M1. Additionally, the IL-6/STAT3 pathway inhibitor Stattic suppressed these changes., Conclusions: Our study demonstrates that in CAP, macrophages highly express hepcidin, which subsequently alters macrophage metabolism, regulates M1 polarization, and leads to bone tissue destruction., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Iron is a vital element involved in a plethora of metabolic activities. Mammalian systemic iron homeostasis is mainly modulated by hepcidin, the synthesis of which is regulated by a number of proteins, including the hemochromatosis-associated proteins Hfe and Transferrin Receptor 2 (TfR2). Macrophages play versatile functions in iron homeostasis by storing iron derived from the catabolism of erythrocytes and supplying iron required for erythropoiesis. The absence of Hfe in macrophages causes a mild iron deficiency in aged mice and leads to an overproduction of the iron exporter Ferroportin 1 (Fpn1). Conversely, TfR2 gene silencing in macrophages does not influence systemic iron metabolism but decreases transcription of the macrophage Fpn1 in adult mice and modulates their immune response. This study investigated cellular and systemic iron metabolism in adult and aged male mice with macrophage-specific Hfe and TfR2 silencing (double knock-out, DKO). Serum iron parameters were significantly modified in aged animals, and significant differences were found in hepatic hepcidin transcription at both ages. Interestingly, splenic iron content was low in adult DKOs and splenic Fpn1 transcription was significantly increased in DKO animals at both ages, while the protein amount does not reflect the transcriptional trend. Additionally, DKO macrophages were isolated from mice bone marrow (BMDMs) and showed significant variations in the transcription of iron genes and protein amounts in targeted mice compared to controls. Specifically, Tranferrin Receptor 1 (TfR1) increased in DKO adult mice BMDMs, while the opposite is observed in the cells of aged DKO mice. Fpn1 transcript was significantly decreased in the BMDMs of adult DKO mice, while the protein was reduced at both ages. Lastly, a significant increase in Erythropoietin production was evidenced in aged DKO mice. Overall, our study reveals that Hfe and TfR2 in macrophages regulate hepatic Hepc production and affect iron homeostasis in the spleen and BMDMs, leading to an iron deficiency in aged animals that impairs their erythropoiesis.

Psoriasis is a multifactorial, chronic inflammatory skin disease with unresolved questions on its primary events. Iron overload has been described in the epidermis of psoriasis patients, but its relevance remains unknown. We found that the key iron regulatory hormone hepcidin was highly expressed in the epidermis of psoriasis patients, especially the pustular variants resistant to treatments. In a murine model of acute skin inflammation, keratinocyte-derived hepcidin was required for iron retention in keratinocytes, leading to hyperproliferation of the epidermal layer and neutrophil recruitment, two main features of psoriatic skin lesions. Keratinocytes overexpressing hepcidin were sufficient to elicit these psoriasiform features in a transgenic mouse model. Furthermore, transcriptome analysis of these keratinocytes revealed canonical pathways found in human psoriasis, pointing to a causal role for hepcidin in the pathogenesis of the disease. Altogether, our data suggest that hepcidin could be an actionable target for skin psoriasis treatment, in addition to current therapeutics, or targeted as maintenance therapy during remission to prevent recurrence., (© 2024. The Author(s).)

Subarachnoid hemorrhage (SAH) significantly compromises the blood-brain barrier (BBB) and impairs patient recovery. This study elucidates the critical role of astrocytic Neogenin-1 (NEO1) in BBB integrity post-SAH and examines the regulatory effects of hepcidin on endothelial cell (EC) function amid NEO1-mediated disruptions in iron homeostasis. Proteomic analyses of cerebrospinal fluid (CSF) from SAH patients revealed a substantial decrease in NEO1 expression, identifying it as a key factor in BBB integrity. 111 CSF proteins were significantly reduced in early SAH stages (days 1-3), with NEO1 among the most significantly altered. This dysregulation was linked to poorer patient outcomes, as indicated by a negative correlation between NEO1 levels and Modified Rankin Scale scores six months post-SAH (R = -0.4743, P < 0.0001). Experimental models further highlighted the importance of NEO1: SAH model and NEO1 GFAP-Cre mice exhibited exacerbated EC dysfunction and increased BBB permeability, evidenced by significant Evans Blue retention and dextran leakage in the parietal cortex, effects that were mitigated by hepcidin administration. Our findings highlight the complex interplay between astrocytic signaling and endothelial function in SAH pathophysiology. The loss of astrocytic NEO1 led to increased EC proliferation and altered BBB structure, as confirmed by transmission electron microscopy and immunostaining for PECAM-1, indicating heightened blood vessel density in the affected cortex. Hepcidin treatment effectively reversed the EC dysfunction and BBB disruption in both NEO1-cKO mice and the SAH model, highlighting its potential as a therapeutic agent to enhance recovery and improve prognosis following SAH., (© 2024. The Author(s).)

Impaired self-renewal of Kupffer cells (KCs) leads to inflammation in metabolic dysfunction-associated steatohepatitis (MASH). Here, we identify neutrophil cytosolic factor 1 (NCF1) as a critical regulator of iron homeostasis in KCs. NCF1 is upregulated in liver macrophages and dendritic cells in humans with metabolic dysfunction-associated steatotic liver disease and in MASH mice. Macrophage NCF1, but not dendritic cell NCF1, triggers KC iron overload, ferroptosis, and monocyte-derived macrophage infiltration, thus aggravating MASH progression. Mechanistically, elevated oxidized phospholipids induced by macrophage NCF1 promote Toll-like receptor (TLR4)-dependent hepatocyte hepcidin production, leading to increased KC iron deposition and subsequent KC ferroptosis. Importantly, the human low-functional polymorphic variant NCF1 90H alleviates KC ferroptosis and MASH in mice. In conclusion, macrophage NCF1 impairs iron homeostasis in KCs by oxidizing phospholipids, triggering hepatocyte hepcidin release and KC ferroptosis in MASH, highlighting NCF1 as a therapeutic target for improving KC fate and limiting MASH progression., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Patients with inflammatory bowel diseases (IBD) frequently experience malnutrition and develop nutrients deficiencies such as vitamin D or iron. Collectively, 39 patients were recruited to the study - 32 with IBD and 7 patients constituting the control group (CG). Quantitative real-time polymerase chain reaction was used to assess the expression of vitamin D receptor (VDR), hepcidin (HAMP) and cathelicidin antimicrobial peptide (CAMP) in colon mucosa. The mean expression of VDR and CAMP was higher in patients with ulcerative colitis than other groups (VDR vs. CG, p<0.05). However, the mean HAMP expression reached higher values in CG than in groups with IBD. Further research to understand the relationship between the VDR, HAMP and CAMP may constitute the way for development of future diagnostic and therapeutic options in patients with IBD.

Ferroportin (FPN) is a transmembrane protein and is the only known iron exporter that helps in maintaining iron homeostasis in vertebrates. To maintain stable iron equilibrium in the body, ferroportin works in conjunction with a peptide called hepcidin. In this study, we have identified an alternatively spliced novel isoform of the human SLC40A1 gene, which encodes for the FPN protein and is found to be expressed in different tissues. The novel transcript has an alternate last exon and encodes 31-amino acid long peptide sequence that replaces 104 amino acids at C-terminal in the novel transcript. Molecular modelling and molecular dynamics (MD) simulation studies revealed key structural features of the novel isoform (FPN-N). FPN-N was predicted to have 12 transmembrane domains similar to the reported isoform (FPN), despite being much smaller in size. FPN-N was found to interact with hepcidin, a key regulator of ferroportin activity. Also, the iron-binding sites were retained in the novel isoform as revealed by the MD simulation of FPN-N in bilipid membrane. The novel isoform identified in this study may play important role in iron homeostasis. However, further studies are required to characterize the FPN-N isoform and decipher its role inside the cell., (© 2024 International Union of Biochemistry and Molecular Biology.)

The significance of iron in myocardial mitochondria function cannot be underestimated, because deviations in iron levels within cardiomyocytes may have profound detrimental effects on cardiac function. In this study, we investigated the effects of ferroportin 1 (FPN1) on cardiac iron levels and pathological alterations in mice subjected to chronic intermittent hypoxia (CIH). The cTNT-FPN1 plasmid was administered via tail vein injection to induce the mouse with FPN1 overexpression in the cardiomyocytes. CIH was established by exposing the mice to cycles of 21%-5% FiO 2 for 3 min, 8 h per day. Subsequently, the introduction of hepcidin resulted in a reduction in FPN1 expression, and H9C2 cells were used to establish an IH model to further elucidate the role of FPN1. First, FPN1 overexpression ameliorated CIH-induced cardiac dysfunction, myocardial hypertrophy, mitochondrial damage and apoptosis. Second, FPN1 overexpression attenuated ROS levels during CIH. In addition, FPN1 overexpression mitigated CIH-induced cardiac iron accumulation. Moreover, the administration of hepcidin resulted in a reduction in FPN1 levels, further accelerating the CIH-induced levels of ROS, LIP and apoptosis in H9C2 cells. These findings indicate that the overexpression of FPN1 in cardiomyocytes inhibits CIH-induced cardiac iron accumulation, subsequently reducing ROS levels and mitigating mitochondrial damage. Conversely, the administration of hepcidin suppressed FPN1 expression and worsened cardiomyocyte iron toxicity injury., (© 2024 The Author(s). Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Background: Following the coronavirus disease 2019 outbreak (COVID-19), it became a worrisome health burden worldwide. COVID-19-associated mucormycosis emergence, characterized by dysregulated inflammation and iron metabolism, exacerbated the prognosis of affected patients. Given the significance of hepcidin in regulating inflammation and iron metabolism, this study investigated the significance of hepcidin single nucleotide polymorphisms (SNP) in COVID-19-associated mucormycosis development, along with the association between the clinical and laboratory factors and COVID-19-associated mucormycosis., Methods: From September 2021 to November 2021, COVID-19 patients with and without mucormycosis were enrolled in this cross-sectional study. Their medical records and laboratory results were investigated. SNP genotyping was performed using Sanger sequencing. Hardy-Weinberg Equilibrium, Pearson's Chi square, and student t test were used for analyzing the data using SPSS software version 25. P<0.05 was regarded as statistically significant., Results: Here, 110 COVID-19 patients with and without mucormycosis were investigated. Elevated levels of urea, aspartate aminotransferase, lactate dehydrogenase, and increased ratio of polymorphonuclear neutrophil to lymphocytes were associated with decreased risk of COVID-19-associated mucormycosis in patients (all P<0.05). Moreover, diabetes mellitus increased the risk of mucormycosis (P=0.028). In contrast to patients without mucormycosis, patients with mucormycosis did not display 442 GA and SNP335 GT genotypes. Unlike patients without mucormycosis, none of the patients with mucormycosis had SNP442 GA and SNP335 GT genotypes. Regarding SNP 443 C>T, and the combination of SNPs 582 A>G and 443 C>T, CC genotype and AA+CC genotypes were associated with increased lactate dehydrogenase levels in COVID-19 patients, respectively., Conclusion: Regarding SNP 443 C>T, the CC genotype was associated with increased lactate dehydrogenase levels in COVID-19 patients. In terms of SNP 582 A>G and SNP 443 C>T, COVID-19 patients with AA+CC genotypes had higher levels of LDH. None of the patients with mucormycosis had SNP442 GA and SNP335 GT genotypes., Competing Interests: None declared., (Copyright: © Iranian Journal of Medical Sciences.)

The anemia of inflammation (AI) is characterized by the presence of inflammation and abnormal elevation of hepcidin. Accumulating evidence has proved that Rocaglamide (RocA) was involved in inflammation regulation. Nevertheless, the role of RocA in AI, especially in iron metabolism, has not been investigated, and its underlying mechanism remains elusive. Here, we demonstrated that RocA dramatically suppressed the elevation of hepcidin and ferritin in LPS-treated mice cell line RAW264.7 and peritoneal macrophages. In vivo study showed that RocA can restrain the depletion of serum iron (SI) and transferrin (Tf) saturation caused by LPS. Further investigation showed that RocA suppressed the upregulation of hepcidin mRNA and downregulation of Fpn1 protein expression in the spleen and liver of LPS-treated mice. Mechanistically, this effect was attributed to RocA's ability to inhibit the IL-6/STAT3 pathway, resulting in the suppression of hepcidin mRNA and subsequent increase in Fpn1 and TfR1 expression in LPS-treated macrophages. Moreover, RocA inhibited the elevation of the cellular labile iron pool (LIP) and reactive oxygen species (ROS) induced by LPS in RAW264.7 cells. These findings reveal a pivotal mechanism underlying the roles of RocA in modulating iron homeostasis and also provide a candidate natural product on alleviating AI., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Background: Early studies in cellular models suggested an iron accumulation in Friedreich's ataxia (FA), yet findings from patients are lacking., Objectives: The objective is to characterize systemic iron metabolism, body iron storages, and intracellular iron regulation in FA patients., Methods: In FA patients and matched healthy controls, we assessed serum iron parameters, regulatory hormones as well as the expression of regulatory proteins and iron distribution in peripheral blood mononuclear cells (PBMCs). We applied magnetic resonance imaging with R 2 *-relaxometry to quantify iron storages in the liver, spleen, and pancreas. Across all evaluations, we assessed the influence of the genetic severity as expressed by the length of the shorter GAA-expansion (GAA1)., Results: We recruited 40 FA patients (19 women). Compared to controls, FA patients displayed lower serum iron and transferrin saturation. Serum ferritin, hepcidin, mean corpuscular hemoglobin and mean corpuscular volume in FA inversely correlated with the GAA1-repeat length, indicating iron deficiency and restricted availability for erythropoiesis with increasing genetic severity. R 2 *-relaxometry revealed a reduction of splenic and hepatic iron stores in FA. Liver and spleen R 2 * values inversely correlated with the GAA1-repeat length. FA PBMCs displayed downregulation of ferritin and upregulation of transferrin receptor and divalent metal transporter-1 mRNA, particularly in patients with >500 GAA1-repeats. In FA PBMCs, intracellular iron was not increased, but shifted toward mitochondria., Conclusions: We provide evidence for a previously unrecognized iron starvation signature at systemic and cellular levels in FA patients, which is related to the underlying genetic severity. These findings challenge the use of systemic iron lowering therapies in FA. © 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society., (© 2024 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.)

Background: The sporadic nature of blood transfusion therapy coupled with the alteration of HAMP genes may exacerbate the risk of iron burden in sickle cell anaemia (SCA) patients. The study determined the polymorphic distribution of the HAMP promoter gene rs10421768 and hepcidin levels in SCA patients., Method: Sixty participants aged ≥12years [45 SCA patients and 15 controls (HbA)] were recruited from 15th March, 2023 to 20th July, 2023 for a case-control study at Methodist Hospital Wenchi, Ghana. Complete blood count and hepcidin levels assessment were done using haematology analyzer and ELISA, respectively. Genomic DNA was extracted using the Qiagen Kit, and HAMP gene rs10421768 (c.-582 A>G) was sequenced using the MassARRAY method. Data were analysed using SPSS version 26.0., Results: The frequencies of the HAMP promoter rs10421768 genotypes AA, AG, and GG were 64.4%, 33.3%, and 2.2% in SCA patients, and 86.7%, 13.3%, and 0% in the controls, respectively. Serum hepcidin levels were significantly higher among controls than cases [204.0 (154.1-219.3) vs 150.2 (108.1-195.6)μg/L, p<0.010]. Participants with HAMP rs10421768 homozygous A genotype had higher serum levels of hepcidin compared with those in the wild genotypes (AG/GG) group [(188.7 (130.9-226.9) vs 136.8 (109.7-157.8)μg/L, p<0.016]. Disease severity and blood cell parameters were not associated with the HAMP variants (p>0.05)., Conclusion: The HAMP promoter rs10421768 AA genotype has the highest frequency of distribution and the GG genotype with the least distribution. Participants with HAMP rs10421768 G allele (c.-582A>G) had reduced levels of hepcidin. HAMP rs10421768 genotypes had no association with blood cell parameters and disease severity. The HAMP rs10421768 genotypes may influence serum levels of hepcidin. Further study is required to elucidate the potential effect of the G allele on hepcidin transcription., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Appiah et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Backgrounds & Aims: Given its ability to inhibit HBV replication, Interferon alpha (IFN-α) treatment has been confirmed to be effective in managing Chronic Hepatitis B (CHB). However, its underlying mechanisms are incompletely understood., Methods: Herein, we investigated the antiviral properties of IFN-α by introducing IFN-α expression plasmids into a well-established HBV Hydrodynamic Injection (HDI) mouse model and examined the impact of IFN-α or hepcidin treatment on macrophages derived from THP-1 cells. The cytokine profiles were analyzed using the cytometry microsphere microarray technology, and flow cytometry was used to analyze the polarization of macrophages. Additionally, the IL-6/JAK2/STAT3 signaling pathway and the hepcidin-ferroportin axis were analyzed to better understand the macrophage polarization mechanism., Results: As evidenced by the suppression of HBV replication, injection of an IFN-α expression plasmid and supernatants of IFN-α-treated macrophages exerted anti-HBV effects. The IFN-α treatment up-regulated IL-6 in mice with HBV replication, as well as in IFN-α-treated HepG2 cells and macrophages. Furthermore, JAK2/STAT3 signaling and hepcidin expression was promoted, inducing iron accumulation via the hepcidin-ferroportin axis, which caused the polarization of M1 macrophages. Furthermore, under the effect of IFN-α, IL-6 silencing or blockade downregulated the JAK2/STAT3 signaling pathway and hepcidin, implying that increased hepcidin expression under IFN-α treatment was dependent on the IL-6/JAK2/STAT3 pathway., Conclusion: The IL-6/JAK2/STAT3 signaling pathway is activated by IFN-α which induces hepcidin expression. The resulting iron accumulation then induces the polarization of M1 macrophages via the hepcidin-ferroportin axis, yielding an immune response which exerts antiviral effects against HBV replication., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Abstract: Hemojuvelin (HJV) is a glycosylphosphatidylinositol-anchored protein of the repulsive guidance molecule family acting as a bone morphogenetic protein (BMP) coreceptor to induce the hepatic iron regulatory protein hepcidin. Hepcidin causes ubiquitination and degradation of the sole known iron exporter ferroportin, thereby limiting iron availability. The detailed signaling mechanism of HJV in vivo has yet to be investigated. In the current manuscript, we used an established model of adeno-associated virus (AAV)-mediated liver-specific overexpression of HJV in murine models of hepatocyte-specific deficiency of the BMP type I receptors Alk2 or Alk3. In control mice, HJV overexpression increased hepatic Hamp messenger RNA (mRNA) levels, soluble HJV (sHJV), splenic iron content (SIC), as well as phosphorylated small mothers against decapentaplegic protein (pSMAD1/5/8) levels. In contrast, in Alk2fl/fl;Alb-Cre and Alk3fl/fl;Alb-Cre mice, which present with moderate and severe iron overload, respectively, the administration of AAV-HJV induced HJV and sHJV. However, it did not rescue the iron overload phenotypes of those mice. Serum iron levels were induced in Alk2fl/fl;Alb-Cre mice after HJV overexpression. In phosphate-buffered saline-injected Alk3fl/fl;Alb-Cre mice, serum iron levels and the expression of duodenal ferroportin remained high, whereas Hamp mRNA levels were decreased to 1% to 5% of the levels detected in controls. This was reduced even further by AAV-HJV overexpression. SIC remained low in mice with hepatocyte-specific Alk2 or Alk3 deficiency, reflecting disturbed iron homeostasis with high serum iron levels and transferrin saturation and an inability to induce hepcidin by HJV overexpression. The data indicate that ALK2 and ALK3 are both required in vivo for the HJV-mediated induction of hepcidin., (© 2024 by The American Society of Hematology. Licensed under Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International (CC BY-NC-ND 4.0), permitting only noncommercial, nonderivative use with attribution. All other rights reserved.)

Hepcidin is a crucial regulator of iron homeostasis with protective effects on liver fibrosis. Additionally, gut microbiota can also affect liver fibrosis and iron metabolism. Although the hepatoprotective potential of Akkermansia muciniphila and Faecalibacterium duncaniae, formerly known as F. prausnitzii, has been reported, however, their effects on hepcidin expression remain unknown. We investigated the direct and macrophage stimulation-mediated effects of active, heat-inactivated, and cell-free supernatant (CFS) forms of A. muciniphila and F. duncaniae on hepcidin expression in HepG2 cells by RT-qPCR analysis. Following stimulation of phorbol-12-myristate-13-acetate (PMA) -differentiated THP-1 cells with A. muciniphila and F. duncaniae, IL-6 concentration was assessed via ELISA. Additionally, the resulting supernatant was treated with HepG2 cells to evaluate the effect of macrophage stimulation on hepcidin gene expression. The expression of genes mediating iron absorption and export was also examined in HepG2 and Caco-2 cells via RT-qPCR. All forms of F. duncaniae increased hepcidin expression while active and heat-inactivated/CFS forms of A. muciniphila upregulated and downregulated its expression, respectively. Active, heat-inactivated, and CFS forms of A. muciniphila and F. duncaniae upregulated hepcidin expression, consistent with the elevation of IL-6 released from THP-1-stimulated cells as a macrophage stimulation effect in HepG2 cells. A. muciniphila and F. duncaniae in active, inactive, and CFS forms altered the expression of hepatocyte and intestinal iron-mediated absorption /exporter genes, namely dcytb and dmt1, and fpn in HepG2 and Caco-2 cells, respectively. In conclusion, A. muciniphila and F. duncaniae affect not only directly but also through macrophage stimulation the expression of hepcidin gene in HepG2 cells. These findings underscore the potential of A. muciniphila and F. duncaniae as a potential therapeutic target for liver fibrosis by modulating hepcidin and intestinal and hepatocyte iron metabolism mediated gene expression., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Iron is an essential element for proper cell functioning, but unbalanced levels can cause cell death. Iron metabolism is controlled at the blood-tissue barriers provided by microvascular endothelial cells. Dysregulated iron metabolism at these barriers is a factor in both neurodegenerative and cardiovascular diseases. Mammalian iron efflux is mediated by the iron efflux transporter ferroportin (Fpn). Inflammation is a factor in many diseases and correlates with increased tissue iron accumulation. Evidence suggests treatment with interleukin 6 (IL-6) increases intracellular calcium levels and calcium is known to play an important role in protein trafficking. We have shown that calcium increases plasma membrane localization of the iron uptake proteins ZIP8 and ZIP14, but if and how calcium modulates Fpn trafficking is unknown. In this article, we examined the effects of IL-6 and calcium on Fpn localization to the plasma membrane. In HEK cells expressing a doxycycline-inducible GFP-tagged Fpn, calcium increased Fpn-GFP membrane presence by 2 h, while IL-6 increased membrane-localized Fpn-GFP by 3 h. Calcium pretreatment increased Fpn-GFP mediated 55 Fe efflux from cells. Endoplasmic reticulum calcium stores were shown to be important for Fpn-GFP localization and iron efflux. Use of calmodulin pathway inhibitors showed that calcium signaling is important for IL-6-induced Fpn relocalization. Studies in brain microvascular endothelial cells in transwell culture demonstrated an initial increase in 55 Fe flux with IL-6 that is reduced by 6 h coinciding with upregulation of hepcidin. Overall, this research details one pathway by which inflammatory signaling mediated by calcium can regulate iron metabolism, likely contributing to inflammatory disease mechanisms., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Hepcidin antimicrobial peptide (HAMP) is a small peptide hormone recognized for its role in iron metabolism and cancer treatment. The purpose of this study was to examine the influence of HAMP in NSCLC., Methods: The profile of NSCLC cells and tissues was characterized via HAMP. Gain- or loss-of-function cell models of HAMP were constructed, and CCK8, colony formation, and Transwell analyses were used to confirm the influence of HAMP on NSCLC cells. Upstream and downstream HAMP mechanisms in NSCLC were also analysed. Dual-luciferase reporter and pull-down assays confirmed the associations of miR-873-5p with HAMP, miR-873-5p, and the lncRNA KCNQ1OT1/SNHG14/XIST. Moreover, a xenograft model was established in nude mice for confirming the role of HAMP in NSCLC cell growth., Results: In addition, HAMP expression increased in NSCLC cells and tissues. In terms of cellular functions, the HAMP-overexpressing group exhibited elevated NSCLC cell proliferation, invasion, and migration. HAMP knockdown reversed these changes. Bioinformatics analysis indicated that miR-873-5p targeted HAMP, which affected the nuclear factor kappa B (NF-κB) pathway in NSCLC. HAMP activated the NF-κB pathway, which was negatively modulated by miR-873-5p. NF-κB inhibitor JSH-23 can partly suppress the proliferation, invasion, and migration in HAMP-overexpressed cells. Moreover, miR-873-5p was the target miRNA of long noncoding RNAs (lncRNAs), which included KCNQ1OT1, SNHG14, and XIST, and these three lncRNAs promoted HAMP., Conclusion: Noncoding RNA-mediated HAMP promotes NSCLC cell proliferation, migration, and invasion by initiating the NF-κB pathway.

Over the years, there has been progress in understanding the molecular aspects of iron metabolism and erythropoiesis. However, despite research conducted both in laboratories and living organisms, there are still unanswered questions due to the complex nature of these fields. In this study we investigated the effects of hookworm infection on iron metabolism and how the hosts response to anemia is affected using hamsters infected with Ancylostoma ceylanicum as a model. Our data revealed interesting relationships between infection-induced anemia, erythropoiesis, iron metabolism, and immune modulation, such that the elevated production of erythropoietin (EPO) in renal tissue indicated intensified erythropoiesis in response to anemia. Additionally, the increased expression of the erythroferrone (ERFE) gene in the spleen suggested its involvement in iron regulation and erythropoiesis. Gene expression patterns of genes related to iron metabolism varied in different tissues, indicating tissue-specific adaptations to hypoxia. The modulation of pro-inflammatory and anti-inflammatory cytokines highlighted the delicate balance between immune response and erythropoiesis. Data derived from the investigation of changes induced in iron metabolism and stress erythropoiesis following anemia aid in our understanding of mechanisms related to blood spoliation and anemia, which could potentially be extrapolated or compared to other types or causes of anemia. These findings also contribute to our understanding of the pathophysiology of erythropoiesis in the context of blood loss., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The role of hepcidins, antimicrobial peptides involved in iron metabolism, immunity, and inflammation, is studied. First, gilthead seabream (Sparus aurata L.) head-kidney leucocytes (HKLs) were incubated with λ-carrageenin to study the expression of hepcidin and iron metabolism-related genes. While the expression of most of the genes studied was upregulated, the expression of ferroportin gene (slc40a) was downregulated. In the second part of the study, seabream specimens were injected intramuscularly with λ-carrageenin or buffer (control). The expression of the same genes was evaluated in the head kidney, liver, and skin at different time points after injection. The expression of Hamp1m, ferritin b, and ferroportin genes (hamp1, fthb, and slc40a) was upregulated in the head kidney of fish from the λ-carrageenin-injected group, while the expression of Hamp2C and Hamp2E genes (hamp2.3 and hamp2.7) was downregulated. In the liver, the expression of hamp1, ferritin a (ftha), slc40a, Hamp2J, and Hamp2D (hamp2.5/6) genes was downregulated in the λ-carrageenin-injected group. In the skin, the expression of hamp1 and (Hamp2A Hamp2C) hamp2.1/3/4 genes was upregulated in the λ-carrageenin-injected group. A bioinformatic analysis was performed to predict the presence of transcription factor binding sites in the promoter region of hepcidins. The primary sequence of hepcidin was conserved among the different mature peptides, although changes in specific amino acid residues were identified. These changes affected the charge, hydrophobicity, and probability of hepcidins being antimicrobial peptides. This study sheds light on the poorly understood roles of hepcidins in fish. The results provide insight into the regulatory mechanisms of inflammation in fish and could contribute to the development of new strategies for treat inflammation in farm animals., (© 2024. The Author(s).)

BMP6 is an iron-sensing cytokine whose transcription in liver sinusoidal endothelial cells (LSECs) is enhanced by high iron levels, a step that precedes the induction of the iron-regulatory hormone hepcidin. While several reports suggested a cell-autonomous induction of Bmp6 by iron-triggered signals, likely via sensing of oxidative stress by the transcription factor NRF2, other studies proposed the dominant role of a paracrine yet unidentified signal released by iron-loaded hepatocytes. To further explore the mechanisms of Bmp6 transcriptional regulation, we used female mice aged 10-11 months, which are characterized by hepatocytic but not LSEC iron accumulation, and no evidence of systemic iron overload. We found that LSECs of aged mice exhibit increased Bmp6 mRNA levels as compared to young controls, but do not show a transcriptional signature characteristic of activated NFR2-mediated signaling in FACS-sorted LSECs. We further observed that primary murine LSECs derived from both wild-type and NRF2 knock-out mice induce Bmp6 expression in response to iron exposure. By analyzing transcriptomic data of FACS-sorted LSECs from aged versus young mice, as well as early after iron citrate injections, we identified ETS1 as a candidate transcription factor involved in Bmp6 transcriptional regulation. By performing siRNA-mediated knockdown, small-molecule treatments, and chromatin immunoprecipitation in primary LSECs, we show that Bmp6 transcription is regulated by iron via ETS1 and p38/JNK MAP kinase-mediated signaling, at least in part independently of NRF2. Thereby, these findings identify the new components of LSEC iron sensing machinery broadly associated with cellular stress responses., (© 2024 The Authors. American Journal of Hematology published by Wiley Periodicals LLC.)

Iron resistance iron deficiency anaemia is a rare autosomal recessive disorder characterized by hypochromic microcytic anaemia, low transferrin saturation and inappropriately high hepcidin levels. The aetiology of this condition is rooted in genetic variations within the transmembrane serine protease 6 (TMPRSS6) genes, responsible for encoding matriptase-2, a pivotal negative regulator of hepcidin. We conducted a systematic search across four electronic databases, yielding 538 articles in total out of which 25 were finally included and were preceded further, aiming to prognosticate prevalent single nucleotide polymorphisms (SNPs) and detrimental genetic alterations. This review aims to elucidate the effects of various SNPs and pathogenic mutations on both haematological and biochemical parameters, as well as their potential interethnic correlation. Employing bioinformatics tools, we subjected over 100 SNPs to scrutiny, discerning their potential functional ramifications. We found rs1373272804, rs1430692214 and rs855791 variants to be most frequent and were having a significant impact on haematological and biochemical profile. We found that individuals of European ancestry were more prone to have these variants compared to other ethnic groups. In conclusion, this review not only sheds light on the association of TMPRSS6 polymorphism in iron resistance iron deficiency anaemia (IRIDA), but also highlights the critical need for further investigations involving larger sample size and more diverse ethnic groups around the globe. These future studies will be vital for gaining a stronger and more reliable understanding of how these genetic differences are linked to the development of IRIDA., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

LEAP2 (liver expression antimicrobial peptide 2), is an antimicrobial peptide widely found in vertebrates and mainly expressed in liver. LEAP2 plays a vital role in host innate immunity. In teleosts, a number of LEAP2 homologs have been reported, but their in vivo effects on host defense are still limited. In this study, a LEAP2 homolog (SsLEAP2) was identified from black rockfish, Sebastes schlegelii, and its structure, expression as well as biological functions were analyzed. The results showed that the open reading frame of SsLEAP2 is 300 bp, with a 5'- untranslated region (UTR) of 375 bp and a 3' - UTR of 238 bp. The deduced amino acid sequence of SsLEAP2 shares the highest overall identity (96.97%) with LEAP2 of Sebastes umbrosus. SsLEAP2 possesses conserved LEAP2 features, including a signal peptide sequence, a prodomain and a mature peptide, in which four well-conserved cysteines formed two intrachain disulphide domain. The expression of SsLEAP2 was highest in liver and could be induced by experimental infection with Listonella anguillarum, Edwardsiealla piscicida and Rock bream iridovirus C1 (RBIV-C1). Recombinant SsLEAP2 (rSsLEAP2) purified from Escherichia coli was able to bind with various Gram-positive and Gram-negative bacteria. Further analysis showed that rSsLEAP2 could enhance the respiratory burst activity, and induce the expression of immune genes including interleukin 1-β (IL-1β) and serum amyloid A (SAA) in macrophages; additionally, rSsLEAP2 could also promote the proliferation and chemotactic of peripheral blood lymphocytes (PBLs). In vivo experiments indicated that overexpression of SsLEAP2 could inhibit bacterial infection, and increase the expression level of immune genes including IL-1β, tumor necrosis factor ligand superfamily member 13B (TNF13B) and haptoglobin (HP); conversely, knock down of SsLEAP2 promoted bacterial infection and decreased the expression level of above genes. Taken together, these results suggest that SsLEAP2 is a novel LEAP2 homolog that possesses apparent antibacterial activity and immunoregulatory property, thus plays a critical role in host defense against pathogens invasion., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Abstract: As a functional component of erythrocyte hemoglobin, iron is essential for oxygen delivery to all tissues in the body. The liver-derived peptide hepcidin is the master regulator of iron homeostasis. During anemia, the erythroid hormone erythroferrone regulates hepcidin synthesis to ensure the adequate supply of iron to the bone marrow for red blood cell production. However, mounting evidence suggested that another factor may exert a similar function. We identified the hepatokine fibrinogen-like 1 (FGL1) as a previously undescribed suppressor of hepcidin that is induced in the liver in response to hypoxia during the recovery from anemia, and in thalassemic mice. We demonstrated that FGL1 is a potent suppressor of hepcidin in vitro and in vivo. Deletion of Fgl1 in mice results in higher hepcidin levels at baseline and after bleeding. FGL1 exerts its activity by directly binding to bone morphogenetic protein 6 (BMP6), thereby inhibiting the canonical BMP-SMAD signaling cascade that controls hepcidin transcription., (© 2024 American Society of Hematology. Published by Elsevier Inc. All rights are reserved, including those for text and data mining, AI training, and similar technologies.)

Liver-expressed antimicrobial peptide 2 (LEAP2) is a member of the antimicrobial peptides family and plays a key role in the innate immune system of organisms. LEAP2 orthologs have been identified from a variety of fish species, however, its function in primitive vertebrates has not been clarified. In this study, we cloned and identified Lc-LEAP2 from the primitive jawless vertebrate lamprey (Lethenteron camtschaticum) which includes a 25 amino acids signal peptide and a mature peptide of 47 amino acids. Although sequence similarity was low compared to other species, the mature Lc-LEAP2 possesses four conserved cysteine residues, forming a core structure with two disulfide bonds between the cysteine residues in the relative 1-3 (Cys 58 and Cys 69) and 2-4 (Cys 64 and Cys 74) positions. Lc-LEAP2 was most abundantly expressed in the muscle, supraneural body and buccal gland of lamprey, and was significantly upregulated during LPS and Poly I:C stimulations. The mature peptide was synthesized and characterized for its antibacterial activity against different bacteria. Lc-LEAP2 possessed inhibition of a wide range of bacteria with a dose-dependence, disrupting the integrity of bacterial cell membranes and binding to bacterial genomic DNA, although its inhibitory function is weak compared to that of higher vertebrates. These data suggest that Lc-LEAP2 plays an important role in the innate immunity of lamprey and is of great value in improving resistance to pathogens. In addition, the antimicrobial mechanism of LEAP2 has been highly conserved since its emergence in primitive vertebrates., Competing Interests: Declaration of competing interest The authors declare no conflict of interest., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The Activin A Receptor type I (ALK2) is a critical component of BMP-SMAD signaling that, in the presence of ligands, phosphorylates cytosolic SMAD1/5/8 and modulates important biological processes, including bone formation and iron metabolism. In hepatocytes, the BMP-SMAD pathway controls the expression of hepcidin , the liver peptide hormone that regulates body iron homeostasis via the BMP receptors ALK2 and ALK3, and the hemochromatosis proteins. The main negative regulator of the pathway in the liver is transmembrane serine protease 6 (TMPRSS6), which downregulates hepcidin by cleaving the BMP coreceptor hemojuvelin. ALK2 function is inhibited also by the immunophilin FKBP12, which maintains the receptor in an inactive conformation. FKBP12 sequestration by tacrolimus or its silencing upregulates hepcidin in primary hepatocytes and in vivo in acute but not chronic settings. Interestingly, gain-of-function mutations in ALK2 that impair FKBP12 binding to the receptor and activate the pathway cause a bone phenotype in patients affected by Fibrodysplasia Ossificans Progressiva but not hepcidin and iron metabolism dysfunction. This observation suggests that additional mechanisms are active in the liver to compensate for the increased BMP-SMAD signaling. Here we demonstrate that Fkbp12 downregulation in hepatocytes by antisense oligonucleotide treatment upregulates the expression of the main hepcidin inhibitor Tmprss6, thus counteracting the ALK2-mediated activation of the pathway. Combined downregulation of both Fkbp12 and Tmprss6 blocks this compensatory mechanism. Our findings reveal a previously unrecognized functional cross talk between FKBP12 and TMPRSS6, the main BMP-SMAD pathway inhibitors, in the control of hepcidin transcription. NEW & NOTEWORTHY This study uncovers a previously unrecognized mechanism of hepcidin and BMP-SMAD pathway regulation in hepatocytes mediated by the immunophilin FKBP12 and the transmembrane serine protease TMPRSS6.

Scope: Iron status is regulated via iron absorption as there is no active iron excretion. Divalent metal-ion transporter-1 (DMT1) and ferroportin (FPN) are two key proteins vital for iron absorption, but the regulation of them in suckling mammals differs from that in adults. This study aims to explore regulation of iron transporters under different iron conditions during suckling., Methods and Results: This study developed suckling rats under different iron conditions. Unexpectedly, unchanged FPN at different iron status are detected. Since FPN is the only known iron exporter for mammals, unchanged FPN limits iron exported into blood during suckling. Thus, factors regulating FPN at transcriptional, post-transcriptional, and post-translational levels are detected. Results showed that Fpn mRNA is upregulated, while micro RNA-485(miR-485) which could silence Fpn mRNA is upregulated at low iron status limiting translation of Fpn mRNA. Besides, serum hepcidin and liver Hamp mRNA are upregulated, but ring finger protein 217( Rnf217) mRNA remained unchanged at high iron status leading to FPN not downregulated as adults., Conclusions: Overall, this study indicates that translational regulation limits intestinal FPN protein response to iron deficiency and Rnf217 cannot effectively mediate the degradation of FPN at high iron status, which provides a reference for maintaining iron homeostasis during suckling., (© 2024 Wiley‐VCH GmbH.)

Several studies have shown association of single nucleotide polymorphisms (SNPs) of hepcidin regulatory pathways genes with impaired iron status. The most common is in the TMPRSS6 gene. In Africa, very few studies have been reported. We aimed to investigate the correlation between the common SNPs in the transmembrane protease, serine 6 (TMPRSS6) gene and iron indicators in a sample of Egyptian children for identifying the suitable candidate for iron supplementation.Patients and methods One hundred and sixty children aged 5-13 years were included & classified into iron deficient, iron deficient anemia and normal healthy controls. All were subjected to assessment of serum iron, serum ferritin, total iron binding capacity, complete blood count, reticulocyte count, serum soluble transferrin receptor and serum hepcidin. Molecular study of TMPRSS6 genotyping polymorphisms (rs4820268, rs855791 and rs11704654) were also evaluated.Results There was an association of iron deficiency with AG of rs855791 SNP, (P = 0.01). The minor allele frequency for included children were 0.43, 0.45 & 0.17 for rs4820268, rs855791 & rs11704654 respectively. Genotype GG of rs4820268 expressed the highest hepcidin gene expression fold, the lowest serum ferroportin & iron store compared to AA and AG genotypes (p = 0.05, p = 0.05, p = 0.03 respectively). GG of rs855791 had lower serum ferritin than AA (p = 0.04), lowest iron store & highest serum hepcidin compared to AA and AG genotypes (p = 0.04, p = 0.01 respectively). Children having CC of rs11704654 had lower level of hemoglobin, serum ferritin and serum hepcidin compared with CT genotype (p = 0.01, p = 0.01, p = 0.02) respectively.Conclusion Possible contribution of SNPs (rs855791, rs4820268 and rs11704654) to low iron status., (© 2024. The Author(s).)

Introduction: There is a proven role for hepcidin and the composition of gut microbiota and its derivatives in the pathophysiology of liver fibrosis., Area Covered: This review focuses on the literature search regarding the effect of hepcidin and gut microbiota on regulating liver physiology. We presented the regulating mechanisms of hepcidin expression and discussed the possible interaction between gut microbiota and hepcidin regulation. Furthermore, we investigated the importance of the hepcidin gene in biological processes and bacterial interactions using bioinformatics analysis., Expert Opinion: One of the main features of liver fibrosis is iron accumulation in hepatic cells, including hepatocytes. This accumulation can induce an oxidative stress response, inflammation, and activation of hepatic stellate cells. Hepcidin is a crucial regulator of iron by targeting ferroportin expressed on hepatocytes, macrophages, and enterocytes. Various stimuli, such as iron load and inflammatory signals, control hepcidin regulation. Furthermore, a bidirectional relationship exists between iron and the composition and metabolic activity of gut microbiota. We explored the potential of gut microbiota to influence hepcidin expression and potentially manage liver fibrosis, as the regulation of iron metabolism plays a crucial role in this context., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

Norcantharidin (NCTD) is a demethylated analogue of cantharidin. It was recently demonstrated that NCTD reduces iron contents in the liver and spleen of mice in vivo , indicating that NCTD can affect iron metabolism via hepcidin. Here, we investigated the effects of NCTD on expression of iron storage protein ferritin-light chain (Ft-L), transferrin receptor 1 (TfR1), divalent metal transporter 1 (DMT1), ferroportin 1 (Fpn1), hepcidin, iron regulatory protein 1 (IRP1), IL-6, p-JAK2 and p-STAT3 in lipopolysaccharides (LPS)-treated RAW264.7 cells in vitro via Real-time PCR and Western blotting analysis. We demonstrate that NCTD down-regulates Ft-L, hepcidin, IL-6, pJAK2, pSTAT3 and up-regulates TfR1, DMT1, Fpn1 and IRP1 expression in LPS treated cells, showing that NCTD can inhibit hepcidin via the IL-6/JAK2/STAT3 signalling pathway and also increase TfR1, DMT1 and Fpn1 expression via down-regulating hepcidin and up-regulating IRP1. Our findings provide further evidence in vitro for the role of NCTD in iron metabolism.

Hepcidin is upregulated by increased body iron stores and inflammatory cytokines. It is associated with cardiovascular events, arterial stiffness, and increased iron accumulation in human atheroma with hemorrhage. However, it is unknown whether the expression of hepcidin in human carotid plaques is related to plaque severity and whether hepcidin expression differs between men and women. Carotid samples from 58 patients (38 males and 20 females) were immunostained with hepcidin, macrophages, ferritin, and transferrin receptor. Immunocytochemistry of hepcidin was performed on THP-1 macrophages exposed to iron or 7betahydroxycholesterol. Hepcidin expression significantly increases with the progression of human atherosclerotic plaques. Plaques of male patients have significantly higher levels of hepcidin. Expressions of hepcidin are significantly correlated with the accumulation of CD68-positive macrophages and transferrin receptor 1 (TfR1) and apoptosis. In vitro, hepcidin is significantly increased in macrophages exposed to iron and moderately increased following 7-oxysterol treatment. In the cultured cells, suppression of hepcidin protected against macrophage cell death, lysosomal membrane permeabilization, and oxidative stress. Hepcidin may play a crucial role in the development and progression of atherosclerosis. The differential expression of hepcidin in male and female patients and its significant correlations with plaque severity, highlight the potential of hepcidin as a biomarker for risk stratification and therapeutic targeting in atherosclerosis.

Background: Hepcidin is the master regulator of iron homeostasis. Hepcidin downregulation has been demonstrated in the brains of Alzheimer's disease (AD) patients. However, the mechanism underlying the role of hepcidin downregulation in cognitive impairment has not been elucidated., Methods: In the present study, we generated GFAP-Cre-mediated hepcidin conditional knockout mice (Hamp GFAP cKO) to explore the effect of hepcidin deficiency on hippocampal structure and neurocognition., Results: We found that the Hamp GFAP cKO mice developed AD-like brain atrophy and memory deficits. In particular, the weight of the hippocampus and the number of granule neurons in the dentate gyrus were significantly reduced. Further investigation demonstrated that the morphological change in the hippocampus of Hamp GFAP cKO mice was attributed to impaired neurogenesis caused by decreased proliferation of neural stem cells. Regarding the molecular mechanism, increased iron content after depletion of hepcidin followed by an elevated level of the inflammatory factor tumor necrosis factor-α accounted for the impairment of hippocampal neurogenesis in Hamp GFAP cKO mice. These observations were further verified in GFAP promoter-driven hepcidin knockdown mice and in Nestin-Cre-mediated hepcidin conditional knockout mice., Conclusions: The present findings demonstrated a critical role for hepcidin in hippocampal neurogenesis and validated the importance of iron and associated inflammatory cytokines as key modulators of neurodevelopment, providing insights into the potential pathogenesis of cognitive dysfunction and related treatments., (© 2024. The Author(s).)

RSL3 is a common inhibitor of glutathione peroxidase 4 (GPx4) that can induce ferroptosis. Ferroptosis is an iron ion-dependent, oxidative-type of programmed cell death. In this study, larval/adult zebrafish were stimulated with RSL3 to construct a ferroptosis model, and CYP2R1 -/- zebrafish was used as a 1,25(OH) 2 D 3 knock-down model to explore the regulatory effect and mechanism of 1,25(OH) 2 D 3 /VD 3 on RSL3-induced ferroptosis. The results showed that 1,25(OH) 2 D 3 /VD 3 alleviated RSL3 induced mitochondrial damage in liver of larval/adult zebrafish, reversed the decline of GPx4 activity, and reduced the accumulation of ROS, LPO and MDA. VD 3 also inhibited hepcidin (HEPC) in adult fish liver, promoted the production of ferroportin (FPN), and reduced the aggregation of Fe 2+ . Exogenous 1,25(OH) 2 D 3 increased the CYP2R1 -/- survival and liver GPx4 activity after RSL3 treatment. At the gene level, 1,25(OH) 2 D 3 /VD 3 activated Keap1-Nrf2-GPx4 and inhibited the NFκB-hepcidin axis. In the ferroptosis context, deletion of the cyp2r1 gene resulted in a more severe decline in gpx4 expression, but the exogenous 1,25(OH) 2 D 3 increased the expression of the GPx4 gene and protein in CYP2R1 -/- zebrafish liver after RSL3 treatment. The collective results indicated that 1,25(OH) 2 D 3 /VD 3 can inhibit ferroptosis induced by RSL3 in liver of larval/adult zebrafish by improving the antioxidant capacity and regulating iron ion transport. Exogenous 1,25(OH) 2 D 3 reverses the downregulation of GPx4 in the CYP2R1 -/- zebrafish liver in the ferroptosis state. Compared with the ferroptosis inhibitor Fer-1, the mechanism of action of 1,25(OH) 2 D 3 /VD 3 is diversified and nonspecific. This study demonstrated the resistance of VD 3 to RSL3-induced ferroptosis at different developmental stages in zebrafish., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Mandarin fish (Siniperca chuatsi) is a valuable freshwater fish species widely cultured in China. Its aquaculture production is challenged by bacterial septicaemia, which is one of the most common bacterial diseases. Antimicrobial peptides (AMPs) play a critical role in the innate immune system of fish, exhibiting defensive and inhibitory effects against a wide range of pathogens. This study aimed to identify the antimicrobial peptide genes in mandarin fish using transcriptomes data obtained from 17 tissue in our laboratory. Through nucleotide sequence alignment and protein structural domain analysis, 15 antimicrobial peptide genes (moronecidin, pleurocidin, lysozyme g, thymosin β 12 , hepcidin, leap 2, β-defensin, galectin 8, galectin 9, apoB, apoD, apoE, apoF, apoM, and nk-lysin) were identified, of which 9 antimicrobial peptide genes were identified for the first time. In addition, 15 AMPs were subjected to sequence characterization and protein structure analysis. After injection with Aeromonas hydrophila, the number of red blood cells, hemoglobin concentration, and platelet counts in mandarin fish showed a decreasing trend, indicating partial hemolysis. The expression change patterns of 15 AMP genes in the intestine after A. hydrophila infection were examined by using qRT-PCR. The results revealed, marked up-regulation (approximately 116.04) of the hepcidin gene, down-regulation of the piscidin family genes expression. Moreover, most AMP genes were responded in the early stages after A. hydrophila challenge. This study provides fundamental information for investigating the role of the different antimicrobial peptide genes in mandarin fish in defense against A. hydrophila infection., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Absorption of dietary iron is largely regulated by the liver hormone hepcidin, which is released under conditions of iron overload and inflammation. Although hepcidin-dependent regulation of iron uptake and circulation is well-characterized, recent studies have suggested that the skin may play an important role in iron homeostasis, including transferrin receptor-mediated epidermal iron uptake and direct hepcidin production by keratinocytes. In this study, we characterized direct keratinocyte responses to conditions of high and low iron. We observed potent iron storage capacity by keratinocytes in vitro and in vivo and the effects of iron on epidermal differentiation and gene expression associated with inflammation and barrier function. In mice, systemic iron was observed to be coupled to epidermal iron content. Furthermore, topical inflammation, as opposed to systemic inflammation, resulted in a primary iron-deficiency phenotype associated with low liver hepcidin. These studies suggest a role for keratinocytes and epidermal iron storage as regulators of iron homeostasis with direct contribution by the cutaneous inflammatory state., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: β-thalassemia is an inherited disorder caused by defects in the synthesis of the beta-globin chain. One of the significant clinical complications in β-thalassemia intermedia is iron overload toxicity, which may be attributed to reduced levels of hepcidin. This reduction in hepcidin leads to increased absorption of iron in the intestines, ultimately resulting in iron overload. The objective of this study was to assess the impact of curcumin on the expression of growth differentiating factor-15 (GDF-15) and hepcidin genes in patients with beta-thalassemia intermedia., Methods: This study was designed as a randomized controlled double-blind clinical trial. Prior to and after the intervention period with curcumin, a blood sample of 5 mL was collected from both the placebo and curcumin-treated groups for the assessment of hepcidin and growth differentiating factor-15 gene expression., Results: This study revealed a significant reduction in the expression of growth differentiating factor-15 in the curcumin group compared to the placebo group during the 3-month treatment period. Furthermore, curcumin supplementation led to a remarkable 10.1-fold increase in the levels of hepcidin in the curcumin group compared to the placebo group., Conclusions: The results of this study show that curcumin administration increases the mRNA levels of hepcidin in whole blood of thalassemia intermedia patients and supports the idea that curcumin could be a potential treatment to reduce suppression of hepcidin in thalassemias and other iron-loading anemias., Conclusions: The results of this study show that curcumin administration increases the mRNA levels of hepcidin in whole blood of thalassemia intermedia patients and supports the idea that curcumin could be a potential treatment to reduce suppression of hepcidin in thalassemias and other iron-loading anemias.

Iron accumulation is one of the most essential pathological events after subarachnoid hemorrhage (SAH). Ferroportin1 (FPN1) is the only transmembrane protein responsible for exporting iron. Hepcidin, as the major regulator of FPN1, is responsible for its degradation. Our study investigated how the interaction between FPN1 and hepcidin contributes to iron accumulation after SAH. We found that iron accumulation aggravated after SAH, along with decreased FPN1 in neurons and increased hepcidin in astrocytes. After knocking down hepcidin in astrocytes, the neuronal FPN1 significantly elevated, thus attenuating iron accumulation. After SAH, p-Smad1/5 and Smad4 tended to translocate into the nucleus. Moreover, Smad4 combined more fragments of the promoter region of Hamp after OxyHb stimulation. By knocking down Smad1/5 or Smad4 in astrocytes, FPN1 level restored and iron overload attenuated, leading to alleviated neuronal cell death and improved neurological function. However, the protective role disappeared after recombinant hepcidin administration. Therefore, our study suggests that owing to the nuclear translocation of transcription factors p-Smad1/5 and Smad4, astrocyte-derived hepcidin increased significantly after SAH, leading to a decreased level of neuronal FPN1, aggravation of iron accumulation, and worse neurological outcome., Competing Interests: Declaration of competing interest The authors declare that they have no relevant financial or non-financial interests to disclose., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Infections due to pathogens impact global aquaculture economy, where diseases caused by bacteria should be in particular focus due to antibiotic resistance. Hepcidin and NK-lysin are important innate immune factors having potential to be exploited as alternatives to antibiotics due to their antimicrobial activity and immunomodulatory capacity. In this study, the recombinant proteins of hepcidin 2 and NK-lysin (rPoHep2 and rPoNKL) from flounder (Paralichthys olivaceus) were obtained via a prokaryotic expression system. The results exhibited that rPoHep2 and rPoNKL killed both gram-negative and gram-positive bacteria mainly via attachment and disruption of the membrane. Interestingly, both peptides could bind to bacterial DNA. The antiviral assay showed that both peptides have antiviral activity against hirame nonvirhabdovirus. They exhibited no cytotoxicity to the mammalian and fish cell lines. PoHep2 was found localized in G-CSFR-positive peritoneal cells. Moreover, rPoHep2 significantly enhanced the phagocytosis of flounder leukocytes in vitro. These findings suggested that neutrophils contained rPoHep2 and may respond to the immunoreaction of neutrophils. In summary, both rPoHep2 and rPoNKL possess antimicrobial activities and may be exploited to replace traditional antibiotics. rPoHep2 possess immune regulatory functions, that can be further investigated as an immunostimulant in aquaculture., Competing Interests: Declaration of competing interest The authors declare that this research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Elsevier B.V. All rights reserved.)

Background: Mendelian randomization (MR) studies show iron positively associated with type 2 diabetes (T2D) but included potentially biasing hereditary haemochromatosis variants and did not assess reverse causality., Methods: We assessed the relation of iron homeostasis with T2D and glycaemic traits bidirectionally, using genome-wide association studies (GWAS) of iron homeostasis biomarkers [ferritin, serum iron, total iron-binding capacity (TIBC), transferrin saturation (TSAT) (n ≤ 246 139)], T2D (DIAMANTE n = 933 970 and FinnGen n = 300 483), and glycaemic traits [fasting glucose (FG), 2-h glucose, glycated haemoglobin (HbA1c) and fasting insulin (FI) (n ≤ 209 605)]. Inverse variance weighting (IVW) was the main analysis, supplemented with sensitivity analyses and assessment of mediation by hepcidin., Results: Iron homeostasis biomarkers were largely unrelated to T2D, although serum iron was potentially associated with higher T2D [odds ratio: 1.07 per standard deviation; 95% confidence interval (CI): 0.99 to 1.16; P-value: 0.078) in DIAMANTE only. Higher ferritin, serum iron, TSAT and lower TIBC likely decreased HbA1c, but were not associated with other glycaemic traits. Liability to T2D likely increased TIBC (0.03 per log odds; 95% CI: 0.01 to 0.05; P-value: 0.005), FI likely increased ferritin (0.29 per log pmol/L; 95% CI: 0.12 to 0.47; P-value: 8.72 x 10-4). FG likely increased serum iron (0.06 per mmol/L; 95% CI: 0.001 to 0.12; P-value: 0.046). Hepcidin did not mediate these associations., Conclusion: It is unlikely that ferritin, TSAT and TIBC cause T2D although an association for serum iron could not be excluded. Glycaemic traits and liability to T2D may affect iron homeostasis, but mediation by hepcidin is unlikely. Corresponding mechanistic studies are warranted., (© The Author(s) 2023; all rights reserved. Published by Oxford University Press on behalf of the International Epidemiological Association.)

Background: MicroRNAs (miRNAs) can control several biological processes. Thus, the existence of these molecules plays a significant role in regulating human iron metabolism or homeostasis., Purpose: The study aimed to determine the role of circulating microRNAs and hepcidin in controlling iron homeostasis and evaluating possible anemia among school children., Methods: The study was based on a biochemical and cross-sectional survey study that included three hundred fifty school children aged 12-18 years old. RT-PCR and immunoassay analysis were accomplished to estimate iron concentration, Hgb, serum ferritin (SF), soluble transferrin receptor (sTfR), total body iron stores (TIBs), total oxidative stress (TOS), total antioxidant capacity (TAC), α-1-acid glycoprotein (AGP), high sensitive C-reactive protein (hs-CRP), and miRNAs; miR-146a, miR-129b, and miR-122 in 350 school adolescents., Results: Iron disorders were cross-sectionally predicted in 28.54% of the study population; they were classified into 14.26% with ID, 5.7% with IDA, and 8.6% with iron overload. The overall proportion of iron depletion was significantly higher in girls (20.0%) than in boys (8.6%). MicroRNAs; miR-146a, miR-125b, and miR-122 were significantly upregulated with lower hepcidin expression in adolescence with ID and IDA compared to iron-overloaded subjects, whereas downregulation of these miRNAs was linked with higher hepcidin. Also, a significant correlation was recorded between miRNAs, hepcidin levels, AGP, hs-CRP, TAC, and other iron-related indicators., Conclusion: Molecular microRNAs such as miR-146a, miR-125b, and miR-122 were shown to provide an additional means of controlling or regulating cellular iron uptake or metabolism either via the oxidative stress pathway or regulation of hepcidin expression via activating genes encoding Hfe and Hjv activators, which promote iron regulation. Thus, circulating miRNAs as molecular markers and serum hepcidin could provide an additional means of controlling or regulating cellular iron and be associated as valuable markers in diagnosing and treating cases with different iron deficiencies., (© 2023. The Author(s).)

Iron delivery to the plasma is closely coupled to erythropoiesis, the production of red blood cells, as this process consumes most of the circulating plasma iron. In response to hemorrhage and other erythropoietic stresses, increased erythropoietin stimulates the production of the hormone erythroferrone (ERFE) by erythrocyte precursors (erythroblasts) developing in erythropoietic tissues. ERFE acts on the liver to inhibit bone morphogenetic protein (BMP) signaling and thereby decrease hepcidin production. Decreased circulating hepcidin concentrations then allow the release of iron from stores and increase iron absorption from the diet. Guided by evolutionary analysis and Alphafold2 protein complex modeling, we used targeted ERFE mutations, deletions, and synthetic ERFE segments together with cell-based bioassays and surface plasmon resonance to probe the structural features required for bioactivity and BMP binding. We define the ERFE active domain and multiple structural features that act together to entrap BMP ligands. In particular, the hydrophobic helical segment 81 to 86 and specifically the highly conserved tryptophan W82 in the N-terminal region are essential for ERFE bioactivity and Alphafold2 modeling places W82 between two tryptophans in its ligands BMP2, BMP6, and the BMP2/6 heterodimer, an interaction similar to those that bind BMPs to their cognate receptors. Finally, we identify the cationic region 96-107 and the globular TNFα-like domain 186-354 as structural determinants of ERFE multimerization that increase the avidity of ERFE for BMP ligands. Collectively, our results provide further insight into the ERFE-mediated inhibition of BMP signaling in response to erythropoietic stress., Competing Interests: Conflict of interest T. G. and E. N. are shareholders and scientific advisors of Intrinsic LifeSciences and consultants for Ionis Pharmaceuticals, Disc Medicine, Silence Therapeutics Chugai, and Vifor. E. N. is a consultant for Protagonist, and T. G. is a consultant for Akebia, Dexcel, and Avidity Bio. Other authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)