scholarly journals Response of Monocyte Iron Regulatory Protein Activity to Inflammation: Abnormal Behavior in Genetic Hemochromatosis

Blood ◽  
1998 ◽  
Vol 91 (7) ◽  
pp. 2565-2572
Author(s):  
Stefania Recalcati ◽  
Roberta Pometta ◽  
Sonia Levi ◽  
Dario Conte ◽  
Gaetano Cairo
Keyword(s):  
Iron Homeostasis ◽  
Iron Status ◽  
Regulatory Protein ◽  
Abnormal Behavior ◽  
Iron Regulatory Protein ◽  
Iron Release ◽  
Protein Activity ◽  
Intracellular Iron ◽  
Ifn Γ ◽  
Genetic Hemochromatosis

In genetic hemochromatosis (GH), iron overload affects mainly parenchymal cells, whereas little iron is found in reticuloendothelial (RE) cells. We previously found that RE cells from GH patients had an inappropriately high activity of iron regulatory protein (IRP), the key regulator of intracellular iron homeostasis. Elevated IRP should reflect a reduction of the iron pool, possibly because of a failure to retain iron. A defect in iron handling by RE cells that results in a lack of feedback regulation of intestinal absorption might be the basic abnormality in GH. To further investigate the capacity of iron retention in RE cells of GH patients, we used inflammation as a model system as it is characterized by a block of iron release from macrophages. We analyzed the iron status of RE cells by assaying IRP activity and ferritin content after 4, 8, and 24 hours of incubation with lipopolysaccharide (LPS) and interferon-γ (IFN-γ). RNA-bandshift assays showed that in monocytes and macrophages from 16 control subjects, IRP activity was transiently elevated 4 hours after treatment with LPS and IFN-γ but remarkably downregulated thereafter. Treatment with NO donors produced the same effects whereas an inducible Nitric Oxide Synthase (iNOS) inhibitor prevented them, which suggests that the NO pathway was involved. Decreased IRP activity was also found in monocytes from eight patients with inflammation. Interestingly, no late decrease of IRP activity was detected in cytokine-treated RE cells from 12 GH patients. Ferritin content was increased 24 hours after treatment in monocytes from normal subjects but not in monocytes from GH patients. The lack of downregulation of IRP activity under inflammatory conditions seems to confirm that the control of iron release from RE cells is defective in GH.

scholarly journals Response of Monocyte Iron Regulatory Protein Activity to Inflammation: Abnormal Behavior in Genetic Hemochromatosis

Blood ◽  
1998 ◽  
Vol 91 (7) ◽  
pp. 2565-2572 ◽  
Author(s):  
Stefania Recalcati ◽  
Roberta Pometta ◽  
Sonia Levi ◽  
Dario Conte ◽  
Gaetano Cairo
Keyword(s):  
Iron Homeostasis ◽  
Iron Status ◽  
Regulatory Protein ◽  
Abnormal Behavior ◽  
Iron Regulatory Protein ◽  
Iron Release ◽  
Protein Activity ◽  
Intracellular Iron ◽  
Ifn Γ ◽  
Genetic Hemochromatosis

Abstract In genetic hemochromatosis (GH), iron overload affects mainly parenchymal cells, whereas little iron is found in reticuloendothelial (RE) cells. We previously found that RE cells from GH patients had an inappropriately high activity of iron regulatory protein (IRP), the key regulator of intracellular iron homeostasis. Elevated IRP should reflect a reduction of the iron pool, possibly because of a failure to retain iron. A defect in iron handling by RE cells that results in a lack of feedback regulation of intestinal absorption might be the basic abnormality in GH. To further investigate the capacity of iron retention in RE cells of GH patients, we used inflammation as a model system as it is characterized by a block of iron release from macrophages. We analyzed the iron status of RE cells by assaying IRP activity and ferritin content after 4, 8, and 24 hours of incubation with lipopolysaccharide (LPS) and interferon-γ (IFN-γ). RNA-bandshift assays showed that in monocytes and macrophages from 16 control subjects, IRP activity was transiently elevated 4 hours after treatment with LPS and IFN-γ but remarkably downregulated thereafter. Treatment with NO donors produced the same effects whereas an inducible Nitric Oxide Synthase (iNOS) inhibitor prevented them, which suggests that the NO pathway was involved. Decreased IRP activity was also found in monocytes from eight patients with inflammation. Interestingly, no late decrease of IRP activity was detected in cytokine-treated RE cells from 12 GH patients. Ferritin content was increased 24 hours after treatment in monocytes from normal subjects but not in monocytes from GH patients. The lack of downregulation of IRP activity under inflammatory conditions seems to confirm that the control of iron release from RE cells is defective in GH.

scholarly journals Sensing of Liver Iron Content Requires Cell-Cell Communication between Hepatocytes and Liver Sinusoidal Endothelial Cells

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 432-432
Author(s):  
Silvia Colucci ◽  
Sandro Altamura ◽  
Matthias Hentze ◽  
Martina U. Muckenthaler
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Iron Content ◽  
Iron Homeostasis ◽  
Iron Status ◽  
Molecular Signature ◽  
Iron Release ◽  
Liver Sinusoidal Endothelial Cells ◽  
Intracellular Iron ◽  
Sinusoidal Endothelial Cells

The liver stores iron and senses systemic and tissue iron availability. Hepatocytes control iron homeostasis by producing the peptide hormone hepcidin that controls dietary iron absorption and iron release from intracellular stores. Recent data challenged the exclusive role of hepatocytes in controlling iron levels. Indeed, liver sinusoidal endothelial cells (LSECs) increase BMP2 and BMP6 levels in response to iron, which control hepcidin expression in a paracrine manner. However the molecular mechanism(s) of how BMPs respond to iron levels remain unknown. We established primary murine LSEC cultures and exposed these to iron sources. Unexpectedly, BMP2 mRNA expression is strongly reduced by iron treatment, while BMP6 levels are only mildly increased. This finding suggests that intracellular iron content cannot directly activate BMP2 transcription and only slightly contribute to BMP6 upregulation in LSEC cultures. However, if LSECs are co-cultured with iron-loaded primary hepatocytes the expression of BMP2 and BMP6 is increased and the fold induction of BMP6 is greater compared to LSECs cultured alone, suggesting that the iron status of hepatocytes instructs the LSEC BMP response. These data are supported by findings in a genetic mouse model of iron overload (Slc40a1C326S/C326S). Hepatocytes isolated from Slc40a1C326S/C326S mice display an iron-loaded molecular signature and the expected low mRNA expression of Transferrin Receptor 1 (Tfr1). By contrast, LSECs show high expression of Tfr1, indicating intracellular iron deficiency. Despite this, hepatic BMP levels are increased, suggesting that BMP2 and BMP6 expression are directly related to the intracellular iron content of hepatocytes but not LSECs. RNA-sequencing of isolated hepatic cell populations is ongoing to identify putative hepatocyte regulators involved in the iron-mediated BMP2 and BMP6 regulation. Disclosures Muckenthaler: Silence Therapeutics: Consultancy; Novartis: Research Funding.

scholarly journals Inappropriately High Iron Regulatory Protein Activity in Monocytes of Patients With Genetic Hemochromatosis

Blood ◽  
1997 ◽  
Vol 89 (7) ◽  
pp. 2546-2553 ◽  
Author(s):  
Gaetano Cairo ◽  
Stefania Recalcati ◽  
Giuliana Montosi ◽  
Elisa Castrusini ◽  
Dario Conte ◽  
...  
Keyword(s):  
Iron Homeostasis ◽  
Receptor Gene ◽  
Regulatory Protein ◽  
Mrna Translation ◽  
Iron Regulatory Protein ◽  
Iron Storage ◽  
Control Subjects ◽  
Iron Pool ◽  
Genetic Hemochromatosis

Abstract In genetic hemochromatosis (GH), excess iron is deposited in parenchymal cells, whereas little iron is found in reticuloendothelial (RE) cells until the later stages of the disease. As iron absorption is inversely related to RE cells stores, a failure of RE to retain iron has been proposed as the basic defect in GH. In RE cells of GH subjects, we examined the activity of iron regulatory protein (IRP), a reliable indicator of the elusive regulatory labile iron pool, which modulates cellular iron homeostasis through control of ferritin (Ft) and transferrin receptor gene expression. RNA-bandshift assays showed a significant increase in IRP activity in monocytes from 16 patients with untreated GH compared with 28 control subjects (1.5-fold) and five patients with secondary hemochromatosis (SH) with similar iron burden (fourfold). In 17 phlebotomy-treated GH patients, IRP activity did not differ from that of control subjects. In both GH and SH monocyte-macrophages, Ft content increased by twofold and the L subunit-rich isoferritin profile was unchanged as compared with controls. IRP activity was still upregulated in vitro in monocyte-derived macrophages of GH subjects but, following manipulations of iron levels, was modulated normally. Therefore, the sustained activity of monocyte IRP found in vivo in monocytes of GH patients is not due to an inherent defect of its control, but is rather the expression of a critical abnormality of iron metabolism, eg, a paradoxical contraction of the regulatory iron pool. By preventing Ft mRNA translation, high IRP activity in monocytes may represent a molecular mechanism contributing to the inadequate Ft accumulation and insufficient RE iron storage in GH.

scholarly journals CBIO-10. REDUCED IRON EXPORT FUNCTIONS IN A CELL INTRINSIC MANNER TO DRIVE GLIOBLASTOMA GROWTH

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii17-ii17
Author(s):  
Katie Troike ◽  
Erin Mulkearns-Hubert ◽  
Daniel Silver ◽  
James Connor ◽  
Justin Lathia
Keyword(s):  
Tumor Cells ◽  
Iron Uptake ◽  
Iron Homeostasis ◽  
Iron Status ◽  
Tumor Grade ◽  
Hfe Gene ◽  
Iron Release ◽  
Female Patients ◽  
Cellular Iron

Abstract Glioblastoma (GBM), the most common primary malignant brain tumor in adults, is characterized by invasive growth and poor prognosis. Iron is a critical regulator of many cellular processes, and GBM tumor cells have been shown to modulate expression of iron-associated proteins to enhance iron uptake from the surrounding microenvironment, driving tumor initiation and growth. While iron uptake has been the central focus of previous investigations, additional mechanisms of iron regulation, such as compensatory iron efflux, have not been explored in the context of GBM. The hemochromatosis (HFE) gene encodes a transmembrane glycoprotein that aids in iron homeostasis by limiting cellular iron release, resulting in a sequestration phenotype. We find that HFE is upregulated in GBM tumors compared to non-tumor brain and that expression of HFE increases with tumor grade. Furthermore, HFE mRNA expression is associated with significantly reduced survival specifically in female patients with GBM. Based on these findings, we hypothesize that GBM tumor cells upregulate HFE expression to augment cellular iron loading and drive proliferation, ultimately leading to reduced survival of female patients. To test this hypothesis, we generated Hfe knockdown and overexpressing mouse glioma cell lines. We observed significant alterations in the expression of several iron handling genes with Hfe knockdown or overexpression, suggesting global disruption of iron homeostasis. Additionally, we show that knockdown of Hfe in these cells increases apoptosis and leads to a significant impairment of tumor growth in vivo. These findings support the hypothesis that Hfe is a critical regulator of cellular iron status and contributes to tumor aggression. Future work will include further exploration of the mechanisms that contribute to these phenotypes as well as interactions with the tumor microenvironment. Elucidating the mechanisms by which iron effulx contributes to GBM may inform the development of next-generation targeted therapies.

scholarly journals Glycogen branching enzyme controls cellular iron homeostasis via Iron Regulatory Protein 1 and mitoNEET

2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Nhan Huynh ◽  
Qiuxiang Ou ◽  
Pendleton Cox ◽  
Roland Lill ◽  
Kirst King-Jones
Keyword(s):  
Iron Homeostasis ◽  
Nuclear Translocation ◽  
Regulatory Protein ◽  
Glycogen Metabolism ◽  
Glycogen Storage Disease Type ◽  
Iron Regulatory Protein ◽  
Messenger Rnas ◽  
Branching Enzyme ◽  
Cellular Iron ◽  
Iron Regulatory Protein 1

AbstractIron Regulatory Protein 1 (IRP1) is a bifunctional cytosolic iron sensor. When iron levels are normal, IRP1 harbours an iron-sulphur cluster (holo-IRP1), an enzyme with aconitase activity. When iron levels fall, IRP1 loses the cluster (apo-IRP1) and binds to iron-responsive elements (IREs) in messenger RNAs (mRNAs) encoding proteins involved in cellular iron uptake, distribution, and storage. Here we show that mutations in the Drosophila 1,4-Alpha-Glucan Branching Enzyme (AGBE) gene cause porphyria. AGBE was hitherto only linked to glycogen metabolism and a fatal human disorder known as glycogen storage disease type IV. AGBE binds specifically to holo-IRP1 and to mitoNEET, a protein capable of repairing IRP1 iron-sulphur clusters. This interaction ensures nuclear translocation of holo-IRP1 and downregulation of iron-dependent processes, demonstrating that holo-IRP1 functions not just as an aconitase, but throttles target gene expression in anticipation of declining iron requirements.

scholarly journals Nitric Oxide–Mediated Induction of Ferritin Synthesis in J774 Macrophages by Inflammatory Cytokines: Role of Selective Iron Regulatory Protein-2 Downregulation

Blood ◽  
1998 ◽  
Vol 91 (3) ◽  
pp. 1059-1066 ◽  
Author(s):  
Stefania Recalcati ◽  
Donatella Taramelli ◽  
Dario Conte ◽  
Gaetano Cairo
Keyword(s):  
Nitric Oxide ◽  
Posttranscriptional Regulation ◽  
Iron Homeostasis ◽  
Regulatory Protein ◽  
Intracellular Iron ◽  
Iron Regulatory Proteins ◽  
Ferritin Synthesis ◽  
Direct Demonstration ◽  
J774 Cells

Cytokine-treated macrophages represent a useful model to unravel the molecular basis of reticuloendothelial (RE) iron retention in inflammatory conditions. In the present study, we showed that stimulation of murine macrophage J774 cells with interferon (IFN)-γ/lipopolysaccharide (LPS) resulted in a nitric oxide-dependent modulation of the activity of iron regulatory proteins (IRP)-1 and 2, cytoplasmic proteins which, binding to RNA motifs called iron responsive elements (IRE), control ferritin translation. Stimulation with cytokines caused a small increase of IRP-1 activity and a strong reduction of IRP-2 activity accompanied by increased ferritin synthesis and accumulation. Cytokines induced only a minor increase of H chain ferritin mRNA, thus indicating that IRP-2–mediated posttranscriptional regulation plays a major role in the control of ferritin expression. This was confirmed by direct demonstration that the translational repression function of IRP was impaired in stimulated cells. In fact, translation in cell-free extracts of a reporter transcript under the control of an IRE sequence was repressed less efficiently by IRP-containing lysates from cytokine-treated cells than by lysates from control cells. Our findings throw light on the role of IRP-2 showing that: (1) this protein responds to a stimulus in opposite fashion to IRP-1; (2) when abundantly expressed, as in J774 cells, IRP-2 is sufficient to regulate intracellular iron metabolism in living cells; and (3) by allowing increased ferritin synthesis, IRP-2 may play a role in the regulation of iron homeostasis in RE cells during inflammation.

scholarly journals Iron regulatory protein 1 promotes ferroptosis by sustaining cellular iron homeostasis in melanoma

2021 ◽  
Vol 22 (3) ◽  
Author(s):  
Fengping Yao ◽  
Xiaohong Cui ◽  
Ying Zhang ◽  
Zhuchun Bei ◽  
Hongquan Wang ◽  
...  
Keyword(s):  
Iron Homeostasis ◽  
Regulatory Protein ◽  
Iron Regulatory Protein ◽  
Cellular Iron ◽  
Cellular Iron Homeostasis ◽  
Iron Regulatory Protein 1

scholarly journals Iron-Response Element Binding to Iron Regulatory Protein (IRP) 1 and 2 Are Indispensable for Adipose Tissue Browning

2020 ◽  
Vol 4 (Supplement_2) ◽  
pp. 1283-1283
Author(s):  
Mikyoung You ◽  
Soonkyu Chung
Keyword(s):  
Adipose Tissue ◽  
Iron Metabolism ◽  
Knockout Mice ◽  
Iron Homeostasis ◽  
Regulatory Protein ◽  
Mrna Translation ◽  
Intracellular Iron ◽  
Mobility Shift ◽  
Iron Regulatory Proteins ◽  
Tissue Browning

Abstract Objectives Intracellular iron homeostasis is tightly regulated in posttranscriptional levels via iron regulatory proteins (IRPs). IRPs bind to the iron-responsive elements (IREs), leading to either mRNA translation or stability. Our recent study demonstrated that iron metabolism is intimately linked with adipose tissue browning and thermogenic activation. However, the role of IRP/IRE interactions in the adipose tissue is poorly understood. We aim to characterize the IRP/IRE interactions in the adipose tissue in terms of depot-specificity and thermogenic potential. Methods To induce adipocyte browning, mice were administrated with beta-3 adrenoceptor agonist CL316243 (CL) for 5 days, and different depots of adipose tissue of epididymal (eWAT), inguinal (iWAT), brown (BAT), and liver were collected. Iron metabolism and thermogenesis were evaluated. To investigate the IRP/IRE binding, electrophoretic mobility shift assay (EMSA) was performed in the cytosolic using the fluorescence-labeled IRE (IR-IRE). To distinguish the IRE binding with IRP1 and 2, the cytosolic fraction from IRP1 and 2 knockout mice were used as positive controls. Results In a normal temperature, the constitutive IRP/IRE binding was found in the BAT, but not in the eWAT and iWAT. In response to CL treatment, iron content and transferrin receptor levels significantly increased in the WAT. Accordingly, the IRE/IRPs binding significantly increased in the CL-treated iWAT. Genetic deletion of IRP1 or 2 poses a marginal impact on constitutively active BAT development, suggesting IRP1 and 2 plays a compensatory role. Unlikely to BAT, the deletion of either IRP1 or 2 failed to induce WAT browning in the IRP1 and 2 knockout mice with CL stimulation. Consistently, both IRE binding to IRP1 and 2 were manifest in the CL treated iWAT, implicating that IRP1 and 2 plays a separate and synergistic function for WAT browning. Conclusions Our study defined the depot-specific iron regulatory metabolism in the adipose tissue using an innovative EMSA method. We demonstrated that, for the first time in our knowledge, IRE binding to both IRP1 and IRP2 is indispensable for the thermogenic activation of WAT, which is distinct from the iron regulatory mechanism found in the BAT. We propose that iron metabolism in the WAT is a novel determinant for WAT browning and thermogenic energy expenditure. Funding Sources None.

HFE Downregulates Iron Uptake From Transferrin and Induces Iron-Regulatory Protein Activity in Stably Transfected Cells

Blood ◽  
1999 ◽  
Vol 94 (11) ◽  
pp. 3915-3921 ◽  
Author(s):  
H.D. Riedel ◽  
M.U. Muckenthaler ◽  
S.G. Gehrke ◽  
I. Mohr ◽  
K. Brennan ◽  
...  
Keyword(s):  
Transferrin Receptor ◽  
Iron Uptake ◽  
Iron Homeostasis ◽  
Hereditary Hemochromatosis ◽  
Regulatory Protein ◽  
Intracellular Iron ◽  
Iron Storage ◽  
Iron Regulatory Proteins ◽  
Cellular Iron

Hereditary hemochromatosis (HH) is a common autosomal-recessive disorder of iron metabolism. More than 80% of HH patients are homozygous for a point mutation in a major histocompatibility complex (MHC) class I type protein (HFE), which results in a lack of HFE expression on the cell surface. A previously identified interaction of HFE and the transferrin receptor suggests a possible regulatory role of HFE in cellular iron absorption. Using an HeLa cell line stably transfected with HFE under the control of a tetracycline-sensitive promoter, we investigated the effect of HFE expression on cellular iron uptake. We demonstrate that the overproduction of HFE results in decreased iron uptake from diferric transferrin. Moreover, HFE expression activates the key regulators of intracellular iron homeostasis, the iron-regulatory proteins (IRPs), implying that HFE can affect the intracellular “labile iron pool.” The increase in IRP activity is accompanied by the downregulation of the iron-storage protein, ferritin, and an upregulation of transferrin receptor levels. These findings are discussed in the context of the pathophysiology of HH and a possible role of iron-responsive element (IRE)-containing mRNAs.

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