Iron-related Genes Research Articles

Response of Fe(III)-reducing kinetics, microbial community structure and Fe(III)-related functional genes to Fe(III)-organic matter complexes and ferrihydrite in lake sediment

Microbial Fe(III) reduction significantly influences the fate of various elements and contaminants. Previous research has employed different Fe(III)-OM complexes and ferrihydrite to study Fe(III)-reduction-related biogeochemistry processes. However, the effects of adding specific Fe(III)-OM complexes and ferrihydrite on the Fe(III)-reducing bacterial community, Fe(III)-reducing kinetics, and Fe(III)-related functional genes remain largely unexplored. This study applied microcosm experiments and metagenomic analysis of lake sediments with and without amendments of ferrihydrite, Fe(III)-citrate, or Fe(III)-EDTA. Results showed that sediments amended with Fe(III)-citrate and Fe(III)-EDTA exhibited faster Fe(III) reduction rates and more significant changes in bacterial community structures compared to those amended with ferrihydrite. Geobacter and Clostridium were enriched in the sediments amended with Fe(III)-EDTA and Fe(III)-citrate, respectively. Despite a slower reduction rate and lack of enrichment of specific Fe(III)-reducing bacteria, ferrihydrite still led to an increase in the copy numbers of genes related to Fe(III) reduction and iron assimilation in the metagenomes, suggesting an increase in these capacities. These results suggest that introducing various Fe(III)-OM complexes and ferrihydrite into the environment would result in differences in not only Fe(III) reduction rates and Fe(III)-reducing bacterial communities but also in iron-related functional genes. Meanwhile, variations in Fe(III) reduction rates and Fe(III)-reducing bacterial communities do not necessarily correlate with changes in the abundances of functional genes relevant to Fe(III) reduction and iron assimilation in the metagenomes. These results provide a better understanding of the adaptive mechanisms of Fe(III)-reducing bacteria in different environmental systems.

A bulge uridine in the HIF2α IRE allows IRP1 but not IRP2 to selectively regulate HIF2α expression and ensuing EPO levels

AbstractIron regulatory proteins (IRP1 and IRP2) play a pivotal role in maintaining cellular iron homeostasis by binding to iron-responsive elements (IREs) of target messenger RNAs and regulating the expression of these iron-related genes. Mice and humans who lack functional IRP1 develop erythrocytosis due to erythropoietin (EPO) overproduction, whereas those who lack IRP2 develop microcytic anemia, believed to result from iron deficiency of erythroblasts. Here, we discovered that IRP2 deficiency reduced the expression of hypoxia-inducible factor 2α (HIF2α) and its transcriptional target, EPO, thereby compromising the stress erythropoiesis response to generate red blood cells upon anemia. The distinct consequences of IRP2 and IRP1 on EPO result from the higher binding affinity of the HIF2α IRE for IRP1 than IRP2. This difference in binding affinity arises from a bulge uridine in the upper stem of HIF2α IRE that impairs the ability of IRP2 to bind the IRE. These results reveal that IRP1 and IRP2 play distinct roles in erythropoiesis and unveil an unsuspected IRE binding preference that contributes to the divergent phenotypes observed in IRP1- and IRP2-deficient mammals.

Iron-Status Indicators and HFE Gene Polymorphisms in Individuals with Amyotrophic Lateral Sclerosis: An Umbrella Review of Meta-analyses and Systematic Reviews.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by the progressive loss of motor neurons. Recent meta-analyses and systematic reviews suggest that HFE gene polymorphisms and iron-associated biomarkers may play a key role in the risk and occurrence of ALS. This umbrella study aimed to explore the roles of HFE gene polymorphisms and iron-associated biomarkers in individuals with ALS. A thorough search of three online scientific databases, namely Scopus, Web of Science, and PubMed, was conducted from their inception until September 13, 2024. The screening and selection processes were executed based on the PICO framework and eligibility criteria, followed by two independent reviewers. The Assessment of Multiple Systematic Reviews (AMSTAR)-2 and GRADE tools were utilized to assess the methodological quality and the certainty of evidence. Through an advanced search, 101 records were retrieved, of which eight meta-analyses and systematic reviews were selected for this umbrella review. A significant increase in iron concentrations was found in individuals with ALS compared to healthy controls (SMD, 0.26; 95% CI - 0.05, 0.57). Conversely, selected meta-analyses reported that serum transferrin concentrations in ALS patients were lower compared to healthy controls (SMD, - 0.15; 95% CI - 0.36, 0.05). Furthermore, mutations in H63D polymorphisms resulted in a 13% significant increase in the risk of ALS (OR, 1.13; 95% CI 1.05, 1.22). Our umbrella study of meta-analyses and systematic reviews reveals that individuals with ALS have lower serum concentrations of transferrin compared to healthy controls. Additionally, the H63D polymorphism in the HFE gene is associated with a slight increase in the risk of ALS. Future research should investigate broader aspects of iron-related biomarkers and HFE genes to elucidate their roles in ALS pathogenesis. Registration: Our umbrella study was registered in the International Prospective Register of Systematic Reviews (PROSPERO) with the identification number CRD42024559032 ( https://www.crd.york.ac.uk/prospero/display_record.php?ID=CRD42024559032 ).

Comparison of Molecular Potential for Iron Transfer across the Placenta in Domestic Pigs with Varied Litter Sizes and Wild Boars.

Neonatal iron deficiency anemia is prevalent among domestic pigs but does not occur in the offspring of wild boar. The main causes of this disorder in piglets of modern pig breeds are paucity of hepatic iron stores, high birth weight, and rapid growth. Replenishment of fetal iron stores is a direct result of iron transfer efficiency across the placenta. In this study, we attempted to investigate the molecular potential of iron transfer across the placenta as a possible cause of differences between wild boar and Polish Large White (PLW) offspring. Furthermore, by analyzing placentas from PLW gilts that had litters of different sizes, we aimed to elucidate the impact of the number of fetuses on placental ability to transport iron. Using RNA sequencing, we examined the expression of iron-related genes in the placentas from wild boar and PLW gilts. We did not reveal significant differences in the expression of major iron transporters among all analyzed placentas. However, in wild boar placentas, we found higher expression of copper-dependent ferroxidases such as ceruloplasmin, zyklopen, and hephaestin, which facilitate iron export to the fetal circulation. We also determined a close co-localization of ceruloplasmin and zyklopen with ferroportin, the only iron exporter.

Linalool and Geraniol Defend Neurons from Oxidative Stress, Inflammation, and Iron Accumulation in In Vitro Parkinson's Models.

Parkinson's disease is one of the most prevalent neurological disorders affecting millions of people worldwide. There is a growing demand for novel and natural substances as complementary therapies. Essential oils and their various compounds are highly investigated natural plant-based products as potential treatment options for common human diseases, such as microbial infections, chronic diseases, and neurodegenerative disorders. The present study focuses on the beneficial effects of linalool and geraniol, the major compounds of lavender (Lavandula angustifolia L.) and geranium (Pelargonium graveolens L'Hér. in Aiton) essential oils, on oxidative stress, inflammation, and iron metabolism of the rotenone and 6-hydroxydopamine-induced in vitro Parkinson's models. The experiments were carried out on all-trans retinoic acid differentiated SH-SY5Y cells. The effects of linalool and geraniol were compared to rasagiline, an MAO-B inhibitor. The results revealed that both essential oil compounds reduce the level of reactive oxygen species and alter the antioxidant capacity of the cells. They lower the secretion of IL-6, IL-8, and IL-1β pro-inflammatory cytokines. Moreover, linalool and geraniol change the expression of iron-related genes, such as the iron importer transferrin receptor 1, heme-oxygenase-1, and ferroportin iron exporter, and influence the intracellular iron contents. In addition, it has been unveiled that iron availability is concatenated with the actions of the essential oil compounds. Based on the results, linalool and geraniol are vigorous candidates as an alternative therapy for Parkinson's disease.

Iron-related Genes and Proteins Involved in Iron Homeostasis in Animal Models of Allergic Asthma: A Systematic Review.

The involvement of the immune oxidative stress response in the pathophysiology and pathogenesis of allergic asthma is well documented. However, reports on the role of iron homeostasis in allergic asthma is scarce. Therefore, this study aims to identify iron-related genes and proteins in mouse models of allergic asthma. Related articles were identified from SCOPUS and Web of Science databases. The article search was limited to publications in English, within a 10-year period (2014 - 2023, up to 16 August 2023) and original/research papers. All identified articles were screened for eligibility using the inclusion and exclusion criteria. All eligible articles were quality appraised prior to data extraction. Five studies were selected for data extraction. Based on the extracted data, three themes and seven subthemes related to iron homeostasis were identified. The type of samples and analytical methods used were also identified. In conclusion, our study elucidates that iron-related proteins are regulated in animal models of allergic asthma. However, the currently available data do not allow us to conclude whether the disease model resulted in iron accumulation or depletion. Therefore, further studies with other related markers should be conducted.

Using Biotinylated Iron-Responsive Element to Analyze the Activity of Iron Regulatory Proteins.

Iron regulatory proteins (IRP1 and IRP2) are the master regulators of mammalian iron homeostasis. They bind to the iron-responsive elements (IREs) of the transcripts of iron-related genes to regulate their expression, thereby maintaining cellular iron availability. The primary method to measure the IRE-binding activity of IRPs is the electrophoresis mobility shift assay (EMSA). This method is particularly useful for evaluating IRP1 activity, since IRP1 is a bifunctional enzyme and its protein levels remain similar during conversion between the IRE-binding protein and cytosolic aconitase forms. Here, we exploited a method of using a biotinylated-IRE probe to separate IRE-binding IRPs followed by immunoblotting to analyze the IRE-binding activity. This method allows for the successful measurement of IRP activity in cultured cells and mouse tissues under various iron conditions. By separating IRE-binding IRPs from the rest of the lysates, this method increases the specificity of IRP antibodies and verifies whether a band represents an IRP, thereby revealing some previously unrecognized information about IRPs. With this method, we showed that the S711-phosphorylated IRP1 was found only in the IRE-binding form in PMA-treated Hep3B cells. Second, we found a truncated IRE-binding IRP2 isoform that is generated by proteolytic cleavage on sites in the 73aa insert region of the IRP2 protein. Third, we found that higher levels of SDS, compared to 1-2% SDS in regular loading buffer, could dramatically increase the band intensity of IRPs in immunoblots, especially in HL-60 cells. Fourth, we found that the addition of SDS or LDS to cell lysates activated protein degradation at 37 °C or room temperature, especially in HL-60 cell lysates. As this method is more practical, sensitive, and cost-effective, we believe that its application will enhance future research on iron regulation and metabolism.

MEMO1 binds iron and modulates iron homeostasis in cancer cells.

Mediator of ERBB2-driven cell motility 1 (MEMO1) is an evolutionary conserved protein implicated in many biological processes; however, its primary molecular function remains unknown. Importantly, MEMO1 is overexpressed in many types of cancer and was shown to modulate breast cancer metastasis through altered cell motility. To better understand the function of MEMO1 in cancer cells, we analyzed genetic interactions of MEMO1 using gene essentiality data from 1028 cancer cell lines and found multiple iron-related genes exhibiting genetic relationships with MEMO1. We experimentally confirmed several interactions between MEMO1 and iron-related proteins in living cells, most notably, transferrin receptor 2 (TFR2), mitoferrin-2 (SLC25A28), and the global iron response regulator IRP1 (ACO1). These interactions indicate that cells with high-MEMO1 expression levels are hypersensitive to the disruptions in iron distribution. Our data also indicate that MEMO1 is involved in ferroptosis and is linked to iron supply to mitochondria. We have found that purified MEMO1 binds iron with high affinity under redox conditions mimicking intracellular environment and solved MEMO1 structures in complex with iron and copper. Our work reveals that the iron coordination mode in MEMO1 is very similar to that of iron-containing extradiol dioxygenases, which also display a similar structural fold. We conclude that MEMO1 is an iron-binding protein that modulates iron homeostasis in cancer cells.

Iron-related gene mutations driving global Mycobacterium tuberculosis transmission revealed by whole-genome sequencing

BackgroundIron plays a crucial role in the growth of Mycobacterium tuberculosis (M. tuberculosis). However, the precise regulatory mechanism governing this system requires further elucidation. Additionally, limited studies have examined the impact of gene mutations related to iron on the transmission of M. tuberculosis globally. This research aims to investigate the correlation between mutations in iron-related genes and the worldwide transmission of M. tuberculosis.ResultsA total of 13,532 isolates of M. tuberculosis were included in this study. Among them, 6,104 (45.11%) were identified as genomic clustered isolates, while 8,395 (62.04%) were classified as genomic clade isolates. Our results showed that a total of 12 single nucleotide polymorphisms (SNPs) showed a positive correlation with clustering, such as Rv1469 (ctpD, C758T), Rv3703c (etgB, G1122T), and Rv3743c (ctpJ, G676C). Additionally, seven SNPs, including Rv0104 (T167G, T478G), Rv0211 (pckA, A302C), Rv0283 (eccB3, C423T), Rv1436 (gap, G654T), ctpD C758T, and etgB C578A, demonstrated a positive correlation with transmission clades across different countries. Notably, our findings highlighted the positive association of Rv0104 T167G, pckA A302C, eccB3 C423T, ctpD C758T, and etgB C578A with transmission clades across diverse regions. Furthermore, our analysis identified 78 SNPs that exhibited significant associations with clade size.ConclusionsOur study reveals the link between iron-related gene SNPs and M. tuberculosis transmission, offering insights into crucial factors influencing the pathogenicity of the disease. This research holds promise for targeted strategies in prevention and treatment, advancing research and interventions in this field.

Interplay of Vitamin D, Unfolded Protein Response, and Iron Metabolism in Neuroblastoma Cells: A Therapeutic Approach in Neurodegenerative Conditions

Vitamin D3 (VD) is crucial for various cell functions, including gene regulation, antioxidant defense, and neural health. Neurodegenerative conditions are closely linked to the unfolded protein response (UPR), a mechanism reacting to endoplasmic reticulum (ER) stress. Iron metabolism is intricately associated with UPR and neurodegeneration. This study used SH-SY5Y neuroblastoma cells to investigate the relationship between UPR, iron metabolism, and VD. Different sequences of treatments (pre- and post-treatments) were applied using VD and thapsigargin (Tg), and various methods were used for evaluation, including real-time qPCR, Western blotting, ELISA, and iron content analysis. The findings indicate that VD affects UPR pathways, cytokine release, and iron-related genes, potentially offering anti-inflammatory benefits. It also influences iron transporters and storage proteins, helping to maintain cellular iron balance. Furthermore, pro-inflammatory cytokines like interleukin-6 (IL-6) and tumor necrosis factor alpha (TNFα) were impacting UPR activation in cells. VD also influenced fractalkine (CX3CL1) gene expression and secretion, suggesting its potential as a therapeutic agent for addressing neuroinflammation and iron dysregulation. This research provides insights into the intricate connections among VD, UPR, and iron metabolism in SH-SY5Y neuroblastoma cells, with implications for future investigations and potential therapeutic approaches in neurodegenerative diseases characterized by UPR dysregulation and iron accumulation.

TroR is the primary regulator of the iron homeostasis transcription network in the halophilic archaeon Haloferax volcanii.

Maintaining theintracellular iron concentration within the homeostatic range is vital to meet cellular metabolic needs and reduce oxidative stress. Previous research revealed that the haloarchaeon Halobacterium salinarum encodes four diphtheria toxin repressor (DtxR) family transcription factors (TFs) that together regulate the iron response through an interconnected transcriptional regulatory network (TRN). However, the conservation of the TRN andthe metal specificity of DtxR TFsremained poorly understood. Here we identified and characterized the TRN of Haloferax volcanii for comparison. Genetic analysis demonstrated that Hfx. volcanii relies on three DtxR transcriptional regulators (Idr, SirR, and TroR), with TroR as the primary regulator of iron homeostasis. Bioinformatics and molecular approaches revealed that TroR binds a conserved cis-regulatory motif located ∼100 nt upstream of the start codon of iron-related target genes. Transcriptomics analysis demonstrated that, under conditions of iron sufficiency, TroR repressed iron uptake and induced iron storage mechanisms. TroR repressed the expression of one other DtxR TF, Idr. This reduced DtxR TRN complexity relative to that of Hbt. salinarum appeared correlated with natural variations in iron availability. Based on these data, we hypothesize that variable environmental conditions such as iron availability appear to select forincreasing TRN complexity.

Cross-species comparison illuminates the importance of iron homeostasis for splenic anti-immunosenescence.

Although immunosenescence may result in increased morbidity and mortality, many mammals have evolved effective immune coping strategies to extend their lifespans. Thus, the immune systems of long-lived mammals present unique models to study healthy longevity. To identify the molecular clues of anti-immunosenescence, we first built high-quality reference genome for a long-lived myotis bat, and then compared three long-lived mammals (i.e., bat, naked mole rat, and human) versus the short-lived mammal, mouse, in splenic immune cells at single-cell resolution. A close relationship between B:T cell ratio and immunosenescence was detected, as B:T cell ratio was much higher in mouse than long-lived mammals and significantly increased during aging. Importantly, we identified several iron-related genes that could resist immunosenescence changes, especially the iron chaperon, PCBP1, which was upregulated in long-lived mammals but dramatically downregulated during aging in all splenic immune cell types. Supportively, immune cells of mouse spleens contained more free iron than those of bat spleens, suggesting higher level of ROS-induced damage in mouse. PCBP1 downregulation during aging was also detected in hepatic but not pulmonary immune cells, which is consistent with the crucial roles of spleen and liver in organismal iron recycling. Furthermore, PCBP1 perturbation in immune cell lines would result in cellular iron dyshomeostasis and senescence. Finally, we identified two transcription factors that could regulate PCBP1 during aging. Together, our findings highlight the importance of iron homeostasis in splenic anti-immunosenescence, and provide unique insight for improving human healthspan.

Iron homeostasis in mice: does liver lobe matter?

The liver plays a crucial role in maintaining systemic iron homeostasis through iron storage, sensing of systemic iron needs, and production of the iron-regulatory hormone hepcidin. While mice are commonly used as models for studying human iron homeostasis, their liver structure differs significantly from humans. Since the mouse liver is structured in six separated lobes, often, the analysis of a single defined lobe is preferred due to concerns over data reproducibility between experimental cohorts. In this study, we compared iron-related parameters in distinct liver lobes of C57BL/6 wild-type mice across different ages. We found that the non-heme iron levels, as well as the mRNA and protein expression of iron storage protein Ferritin and the iron importer Transferrin Receptor 1, were similar between liver lobes. Additionally, the mRNA expression of Hepcidin, as well as its regulators, Bmp2 and Bmp6, and iron importers Zip8 and Zip14 were comparable. Minor differences were observed in Ferroportin mRNA levels of 24-wk-old mice; however, this did not correlate with altered iron content. The findings in wild-type mice were reproduced in Hfe knock-out mice - a well-established genetic model of the most prevalent form of hemochromatosis. Overall, our results indicate that C57BL/6 mouse liver lobes can be used interchangeably for assessing iron content and expression of iron-related genes. Understanding if these findings are applicable to other mouse developmental stages, strains, or models of (iron-related) disorders will be key to promote reduction of experimental animal numbers and facilitate resource sharing among research groups studying liver iron homeostasis.NEW & NOTEWORTHY This study reveals that, despite being structurally separated, liver lobes from C57BL/6 wild-type and iron-overloaded mice can be used interchangeably for the evaluation of iron content and expression of iron-related genes.

Aloe-emodin alleviates doxorubicin-induced cardiotoxicity via inhibition of ferroptosis.

Aloe-emodin (AE), a novel ferroptosis inhibitor, alleviates the doxorubicin (DOX)-induced cardiotoxicity in H9c2 rat cardiomyocytes. The inhibition of ferroptosis and the protective effect against cardiotoxicity were evaluated via MTT assay in H9c2 cells. The molecular mechanism of action (MOA) of nuclear factor erythroid 2-related factor 2 (Nrf2) activation, including transactivation of multiple downstream cytoprotective genes, were further assessed by Western blot, luciferase reporter assay and qRT-PCR analyses. Fluorescent imaging was performed to detect the change of intracellular reactive oxygen species, mitochondrial membrane potential and lipid peroxidation. In addition, an infrared spectroscopy was employed to detect the AE-Fe (II) complex. AE, alleviates oxidative stress in DOX-induced H9c2 cells by activating Nrf2 and increasing the expression of Nrf2 downstream antioxidant genes, SLC7A11 and GPX4. Furthermore, AE complexes bivalent iron and regulates the intracellular iron-related genes. In conclusion, the discovery of AE as a novel ferroptosis inhibitor and its MOA provides a new perspective for further exploration of cardio-protective agents in cancer patients during chemotherapy.

Polymorphisms Related to Iron Homeostasis Associate with Liver Disease in Chronic Hepatitis C.

Dysregulation of iron metabolism in chronic hepatitis C (CHC) is a significant risk factor for hepatic cirrhosis and cancer. We studied if known genetic variants related to iron homeostasis associate with liver disease progression in CHC. Retrospective analysis included 249 CHC patients qualified for antiviral therapy between 2004 and 2014. For all patients, nine SNPs within HFE, TFR2, HDAC2, HDAC3, HDAC5, TMPRSS6, and CYBRD1 genes were genotyped. Expression of selected iron-related genes, was determined with qRT-PCR in 124 liver biopsies, and mRNA expression of co-inhibitory receptors (PD-1, Tim3, CTLA4) was measured in 79 liver samples. CYBRD1 rs884409, HDAC5 rs368328, TFR2 rs7385804, and TMPRSS6 rs855791 associated with histopathological changes in liver tissue at baseline. The combination of minor allele in HDAC3 rs976552 and CYBRD1 rs884409 linked with higher prevalence of hepatocellular carcinoma (HCC) during follow up (OR 8.1 CI 2.2-29.2; p = 0.001). Minor allele in HDAC3 rs976552 associated with lower hepatic expression of CTLA4. Tested polymorphisms related to iron homeostasis associate with histopathological changes in the liver. The presence of both HDAC3 rs976552 G and CYBRD1 rs884409 G alleles correlates with HCC occurrence, especially in the group of patients with elevated AST (>129 IU/L). rs976552 in HDAC3 could impact immunological processes associated with carcinogenesis in CHC.

Iron-Related Genes and Proteins in Mesenchymal Stem Cell Detection and Therapy.

Mesenchymal stem cells (MSCs) are located in various tissues of the body. These cells exhibit regenerative and reparative properties, which makes them highly valuable for cell-based therapy. Despite this, majority of MSC-related studies remain to be translated for regular clinical use. This is partly because there are methodical challenges in pre-administration MSC labelling, post-administration detection and tracking of cells, and in retention of maximal therapeutic potential in-vivo. This calls for exploration of alternative or adjunctive approaches that would enable better detection of transplanted MSCs via non-invasive methods and enhance MSC therapeutic potential in-vivo. Interestingly, these attributes have been demonstrated by some iron-related genes and proteins.Accordingly, this unique forward-looking article integrates the apparently distinct fields of iron metabolism and MSC biology, and reviews the utility of iron-related genes and iron-related proteins in facilitating MSC detection and therapy, respectively. Effects of genetic overexpression of the iron-related proteins ferritin, transferrin receptor-1 and MagA in MSCs and their utilisation as reporter genes for improving MSC detection in-vivo are critically evaluated. In addition, the beneficial effects of the iron chelator deferoxamine and the iron-related proteins haem oxygenase-1, lipocalin-2, lactoferrin, bone morphogenetic protein-2 and hepcidin in enhancing MSC therapeutics are highlighted with the consequent intracellular alterations in MSCs. This review aims to inform both regenerative and translational medicine. It can aid in formulating better methodical approaches that will improve, complement, or provide alternatives to the current pre-transplantation MSC labelling procedures, and enhance MSC detection or augment the post-transplantation MSC therapeutic potential.

Cellular and Molecular Processes Are Differently Influenced in Primary Neural Cells by Slight Changes in the Physicochemical Properties of Multicore Magnetic Nanoparticles

Herein, we use two exemplary superparamagnetic iron oxide multicore nanoparticles (SPIONs) to illustrate the significant influence of slightly different physicochemical properties on the cellular and molecular processes that define SPION interplay with primary neural cells. Particularly, we have designed two different SPION structures, NFA (i.e., a denser multicore structure accompanied by a slightly less negative surface charge and a higher magnetic response) and NFD (i.e., a larger surface area and more negatively charged), and identified specific biological responses dependent on SPION type, concentration, exposure time, and magnetic actuation. Interestingly, NFA SPIONs display a higher cell uptake, likely driven by their less negative surface and smaller protein corona, more significantly impacting cell viability and complexity. The tight contact of both SPIONs with neural cell membranes results in the significant augmentation of phosphatidylcholine, phosphatidylserine, and sphingomyelin and the reduction of free fatty acids and triacylglycerides for both SPIONs. Nonetheless, NFD induces greater effects on lipids, especially under magnetic actuation, likely indicating a preferential membranal location and/or a tighter interaction with membrane lipids than NFA, in agreement with their lower cell uptake. From a functional perspective, these lipid changes correlate with an increase in plasma membrane fluidity, again larger for more negatively charged nanoparticles (NFD). Finally, the mRNA expression of iron-related genes such as Ireb-2 and Fth-1 remains unaltered, while TfR-1 is only detected in SPION-treated cells. Taken together, these results demonstrate the substantial impact that minor physicochemical differences of nanomaterials may exert in the specific targeting of cellular and molecular processes. A denser multicore structure generated by autoclave-based production is accompanied by a slight difference in surface charge and magnetic properties that become decisive for the biological impact of these SPIONs. Their capacity to markedly modify the lipidic cell content makes them attractive as lipid-targetable nanomedicines.

Effects of Feeding 5-Aminolevulinic Acid on Iron Status in Weaned Rats from the Female Rats during Gestation and Lactation.

Simple SummaryIn order to develop novel iron supplementation methods for piglets, this study employed female Sprague–Dawley (SD) rats as a model to investigate the effects of 5-aminolevulinic acid (5-ALA) on iron status in weaned rats. The results indicated that fed 5-ALA diets to female rats could improve the blood parameters of weaned rats, increase the concentration of Hepcidin in the liver and serum, and promote the expression of iron-related genes in the liver, suggesting that 5-ALA may be an excellent functional additive to improve the iron status of animals. Nevertheless, these effects still need to be further validated in piglets. Using female Sprague–Dawley (SD) rats as a model, the current study aimed to investigate whether feeding 5-aminolevulinic acid (5-ALA) to female SD rats during gestation and lactation can affect the iron status of weaned rats and provide new ideas for the iron supplementation of piglets. A total of 27 pregnant SD rats were randomly assigned to three treatments in nine replicates, with one rat per litter. Dietary treatments were basal diet (CON), CON + 50 mg/kg 5-ALA (5-ALA50), and CON + 100 mg/kg 5-ALA (5-ALA100). After parturition, ten pups in each litter (a total of 270) were selected for continued feeding by their corresponding mother, and the pregnant rats were fed diets containing 5-ALA (0, 50 and 100 mg/kg diet) until the newborn pups were weaned at 21 days. The results showed that the number of red blood cells (RBCs) in weaned rats in the 5-ALA100 group was significantly higher (p < 0.05) than that in the CON or 5-ALA50 group. The diet with 5-ALA significantly increased (p < 0.05) the hemoglobin (HGB) concentration, hematocrit (HCT) level, serum iron (SI) content, and transferrin saturation (TSAT) level in the blood of weaned rats, as well as the concentration of Hepcidin in the liver and serum of weaned rats and the expression of Hepcidin mRNA in the liver of weaned rats, with the 5-ALA100 group having the highest (p < 0.05) HGB concentration in the weaned rats, and the 5-ALA50 group having the highest (p < 0.05) Hepcidin concentration in serum and in the expression of Hepcidin mRNA in the liver of weaned rats. The other indicators between the 5-ALA groups had no effects. However, the level of total iron binding capacity (TIBC) was significantly decreased (p < 0.05) in the 5-ALA50 group. Moreover, the iron content in the liver of weaned rats fed with 5-ALA showed an upward trend (p = 0.085). In addition, feeding a 5-ALA-supplemented diet could also significantly reduce (p < 0.05) the expression of TfR1 mRNA in the liver of weaning rats (p < 0.05), and the expression of Tfr1 was not affected between 5-ALA groups. In conclusion, dietary supplementation with 5-ALA could improve the blood parameters, increase the concentration of Hepcidin in the liver and serum, and affect the expression of iron-related genes in the liver of weaned rats. Moreover, it is appropriate to add 50 mg/kg 5-ALA to the diet under this condition.

Systemic iron reduction via an iron deficient diet decreases the severity of knee cartilage lesions in the Dunkin-Hartley guinea pig model of osteoarthritis

Iron accumulation is emerging as a player in aging-related disorders due to its propensity for generating reactive oxygen species (ROS). Studies investigating the role of iron in the pathogenesis of primary osteoarthritis (OA) are limited. We designed a proof-of-principle study to determine the effect of systemic iron deficiency, via an iron deficient diet, on knee OA in an animal model. Twelve-week-old male Hartley guinea pigs received the standard diet (n=6) or a diet devoid of iron (n=6) for 19-weeks. Iron levels were determined in the serum, liver, and articular cartilage. Knees were collected to assess structural changes related to OA (microcomputed tomography, histopathology). Immunohistochemistry was performed to evaluate the presence and distribution of a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS4) and ROS-driven 4-hydroxynonenal (4-HNE)-induced protein adducts. Transcript expression was also assessed. Relative to control animals, an iron deficient diet reduced the concentration of this mineral in serum, liver, and articular cartilage. Iron deficient animals had lower histologic OA scores; decreased subchondral bone mineral density was also noted. This reduction in knee joint pathology was accompanied by a decrease in: ADAMTS4 in synovium; and 4-HNE protein adducts from lipid peroxidation in both the menisci and articular cartilage of iron deficient animals. Expression of iron-related genes in these tissues was also altered in treated animals. Results from this study suggest that systemic iron levels may play a role in knee OA pathogenesis, with a short-term deficit in dietary iron reducing the severity of knee cartilage lesions.

PB1748: DIFFERENTIAL IRON HOMEOSTASIS IN CHILDHOOD ACUTE LYMPHOBLASTIC LEUKEMIA IN VITRO MODELS

Background: Iron is an essential element which plays several physiological activities in different human organs. Current studies suggest that dysregulation of iron proteins and subsequent accumulation of iron are associated with the response of leukemia to treatments. In particular, it has been reported that low levels of ferroportin-1 (FPN1), lead to an increased sensitivity to chemotherapy in acute myeloid leukemia. Aims: We aimed at analyzing the gene expression profile of iron-related genes in childhood B acute lymphoblastic leukemia (ALL) and characterize the iron metabolism in vitro models. Methods: Iron related genes (Ceruloplasmin (CP), Ferroportin-1 (SLC40A1), Transferrin Receptor 1 (TFR1) and Ferritin Light Chain (FTL)) were analyzed by qRT-PCR in BM samples of 87 childhood ALL and 8 healthy donors. Cases were selected to be representative of t(12;21), t(4;11), t(9;22) or t(1;19) subgroups, as well as MRD-based standard-risk (SR), medium-risk (MR) and high-risk (HR) response. Furthermore, we evaluated the cellular viability in REH, RS4;11, 697 and TOM-1 cell lines after exposure to iron as ferric ammonium citrate (FAC 100 to 400 µM), deferoxamine (DFO 25 µM) and FAC+DFO (100 µM + 25 µM), using MTT assay at 24-48 and 72h. TFR1 and FTL mRNA levels were assessed at 8h, 16h, 24h, 48h. Labile Iron Pool (LIP) was measured by flow-cytometry using Calcein-AM technique after the exposure to FAC (100-200-400 µM) at 48h. Results:TFR1, SLC40A1 and CP were up-regulated in HR group vs. controls. SLC40A1 and CP were down-regulated in all genetic subgroups, except for CP in t(9;22). TFR1 was slightly expressed in all subgroups. In the t(1;19) subgroup all the iron genes were downregulated, suggesting that this subgroup may not be responsive to iron, as also confirmed by our in vitro data. Iron supplementation by FAC [100 µM] resulted in a boost of viability for TOM-1, REH and RS4;11. TOM-1 and REH viability decreased with FAC 200-400 µM, while RS4;11 proliferation was stimulated by all concentrations. Together with low transcriptional levels of iron genes, 697 viability did not change with different FAC treatments. DFO reduced cell viability in a time-dependent manner. At 48h the viability started to drop for all cell lines (TOM-1 80% - RS4;11 66% - REH 71% - 697 51%) and the co-administration of iron restored the viability to levels similar or even higher than the untreated (TOM-1 100%-RS4;11 80% -REH 92% - 697 81%). FTL expression was significantly up-regulated in all cell lines, while the TFR1 was down-regulated at 8, 16 and 24h vs. untreated. Still, some differences were detected within the cellular models: RS4;11 showed highest level of FTL expression, while in 697 the transcript was slightly upregulated. In 697 cell line, the LIP increased with higher dosages of FAC, suggesting that this cell line was not able to handle high amount of free iron. TOM-1, RS4;11 and REH presented LIP levels higher than the untreated condition, still no significant differences were noticed between the concentrations. Summary/Conclusion:SLC40A1 and CP high levels of HR group potentially suggest that their refractory behavior may be correlated with ability to keep regulate iron intracellular concentration. TOM-1, REH and RS4;11 behave as resistant cell lines, whilst the 697 appear as not sensitive to iron metabolism. In conclusion, leukemia cell lines can be considered as trustable in vitro models for further studies in ALL to investigate whether iron metabolism can be determinant for survival and resistance, and consequently a target for treatment.