A newly identified ferritin L-subunit variant results in increased proteasomal subunit degradation, impaired complex assembly, and severe hypoferritinemia.

Ferritin is a hetero-oligomeric nanocage, composed of 24 subunits of two types, FTH1 and FTL. It protects the cell from excess reactive iron, by storing iron in its cavity. FTH1 is essential for the recruitment of iron into the ferritin nanocage and for cellular ferritin trafficking, whereas FTL contributes to nanocage stability and iron nucleation inside the cavity. Here we describe a female patient with a medical history of severe hypoferritinemia without anemia. Following inadequate heavy IV iron supplementation, the patient developed severe iron overload and musculoskeletal manifestations. However, her serum ferritin levels rose only to normal range. Genetic analyses revealed an undescribed homozygous variant of FTL (c.92A > G), which resulted in a Tyr31Cys substitution (FTLY31C ). Analysis of the FTL structure predicted that the Y31C mutation will reduce the variant's stability. Expression of the FTLY31C variant resulted in significantly lower cellular ferritin levels compared with the expression of wild-type FTL (FTLWT ). Proteasomal inhibition significantly increased the initial levels of FTLY31C , but could not protect FTLY31C subunits from successive degradation. Further, variant subunits successfully incorporated into hetero-polymeric nanocages in the presence of sufficient levels of FTH1. However, FTLY31C subunits poorly assembled into nanocages when FTH1 subunit levels were low. These results indicate an increased susceptibility of unassembled monomeric FTLY31C subunits to proteasomal degradation. The decreased cellular assembly of FTLY31C -rich nanocages may explain the low serum ferritin levels in this patient and emphasize the importance of a broader diagnostic approach of hypoferritinemia without anemia, before IV iron supplementation.

Fusaricide is a Novel Iron Chelator that Induces Apoptosis through Activating Caspase-3.

Nonsmall cell lung cancer (NSCLC) has been a fatal and refractory disease worldwide. Novel therapeutic developments based on fundamental investigations of anticancer mechanisms underlie substantial foundations to win the fight against cancer diseases. In this study, we isolated a natural product fusaricide (FCD) from an endophytic fungus of Lycium barbarum, identified as Epicoccum sp. For the first time, we discovered that FCD potently inhibited proliferation in a variety of human NSCLC cell lines, with relatively less toxicity to normal cells. Our study exhibited that FCD induced apoptosis, caused DNA damage and cell cycle arrest in G0/G1 phase, and activated caspase-3 as well as other apoptosis-related factors in human NSCLC NCI-H460 cells. FCD was proven to be an iron chelator that actively decreased levels of cellular labile iron pool in NCI-H460 cells in our study. FeCl3 supplement reversed FCD-induced apoptosis. The upregulation of transferrin receptor 1 (TfR1) and downregulation of ferritin heavy chain (FTH) expression were observed after FCD treatment. In summary, our study highlighted the potential anticancer effects of FCD against human NSCLCs and demonstrated that the FCD-mediated apoptosis depended on binding to intracellular iron.

Ferritin H deficiency deteriorates cellular iron handling and worsens Salmonella typhimurium infection by triggering hyperinflammation.

Iron is an essential nutrient for mammals as well as for pathogens. Inflammation-driven changes in systemic and cellular iron homeostasis are central for host-mediated antimicrobial strategies. Here, we studied the role of the iron storage protein ferritin H (FTH) for the control of infections with the intracellular pathogen Salmonella enterica serovar Typhimurium by macrophages. Mice lacking FTH in the myeloid lineage (LysM-Cre+/+Fthfl/fl mice) displayed impaired iron storage capacities in the tissue leukocyte compartment, increased levels of labile iron in macrophages, and an accelerated macrophage-mediated iron turnover. While under steady-state conditions, LysM-Cre+/+Fth+/+ and LysM-Cre+/+Fthfl/fl animals showed comparable susceptibility to Salmonella infection, i.v. iron supplementation drastically shortened survival of LysM-Cre+/+Fthfl/fl mice. Mechanistically, these animals displayed increased bacterial burden, which contributed to uncontrolled triggering of NF-κB and inflammasome signaling and development of cytokine storm and death. Importantly, pharmacologic inhibition of the inflammasome and IL-1ß pathways reduced cytokine levels and mortality and partly restored infection control in iron-treated ferritin-deficient mice. These findings uncover incompletely characterized roles of ferritin and cellular iron turnover in myeloid cells in controlling bacterial spread and for modulating NF-κB and inflammasome-mediated cytokine activation, which may be of vital importance in iron-overloaded individuals suffering from severe infections and sepsis.

Unconventional endocytosis and trafficking of transferrin receptor induced by iron.

The posttranslational regulation of transferrin receptor (TfR1) is largely unknown. We investigated whether iron availability affects TfR1 endocytic cycle and protein stability in HepG2 hepatoma cells exposed to ferric ammonium citrate (FAC). NH4Cl and bafilomycin A1, but not the proteasomal inhibitor MG132, prevented the FAC-mediated decrease in TfR1 protein levels, thus indicating lysosomal involvement. Knockdown experiments showed that TfR1 lysosomal degradation is independent of 1) endocytosis mediated by the clathrin adaptor AP2; 2) Tf, which was suggested to facilitate TfR1 internalization; 3) H-ferritin; and 4) MARCH8, previously implicated in TfR1 degradation. Notably, FAC decreased the number of TfR1 molecules at the cell surface and increased the Tf endocytic rate. Colocalization experiments confirmed that, upon FAC treatment, TfR1 was endocytosed in an AP2- and Tf-independent pathway and trafficked to the lysosome for degradation. This unconventional endocytic regulatory mechanism aimed at reducing surface TfR1 may represent an additional posttranslational control to prevent iron overload. Our results show that iron is a key regulator of the trafficking of TfR1, which has been widely used to study endocytosis, often not considering its function in iron homeostasis.

Oligodendrocytes Provide Antioxidant Defense Function for Neurons by Secreting Ferritin Heavy Chain.

An evolutionarily conserved function of glia is to provide metabolic and structural support for neurons. To identify molecules generated by glia and with vital functions for neurons, we used Drosophila melanogaster as a screening tool, and subsequently translated the findings to mice. We found that a cargo receptor operating in the secretory pathway of glia was essential to maintain axonal integrity by regulating iron buffering. Ferritin heavy chain was identified as the critical secretory cargo, required for the protection against iron-mediated ferroptotic axonal damage. In mice, ferritin heavy chain is highly expressed by oligodendrocytes and secreted by employing an unconventional secretion pathway involving extracellular vesicles. Disrupting the release of extracellular vesicles or the expression of ferritin heavy chain in oligodendrocytes causes neuronal loss and oxidative damage in mice. Our data point to a role of oligodendrocytes in providing an antioxidant defense system to support neurons against iron-mediated cytotoxicity.

Identification and functional analysis of an iron-binding protein, ferritin heavy chain subunit, from the swallowtail butterfly, Papilio xuthus.

Ferritin, which is ubiquitous among all living organisms, plays a crucial role in maintaining iron homeostasis, immune response, and detoxification. In the present research, we identified an iron-binding protein, ferritin heavy chain subunit, from Papilio xuthus and named PxFerHCH. The complete complementary DNA of PxFerHCH was 1,252 bp encoding a sequence of 211 amino acids, which includes an iron-responsive element. Phylogenetic analysis showed that PxFerHCH is clustered with Manduca sexta and Galleria mellonella ferritin heavy chain subunits. Expression levels of PxFerHCH in various tissues were analyzed by reverse transcription quantitative polymerase chain reaction, and the results exhibited that PxFerHCH was expressed in all tissues with the highest expression in the fat body. The relative expression level of PxFerHCH in response to bacterial (Escherichia coli and Staphylococcus aureus) challenges sharply increased by about 12 hr postinfection (hpi) and then decreased at 24 hpi. In addition, the iron-binding capacity and antioxidation activity of recombinant PxFerHCH protein were also investigated. These results reveal that PxFerHCH might play an important role in defense against bacterial infection.

Chitosan binding onto the epigallocatechin-loaded ferritin nanocage enhances its transport across Caco-2 cells.

The effect of chitosan decoration on the transport of epigallocatechin (EGC)-encapsulated ferritin cage across the Caco-2 cells was investigated. After the encapsulation of EGC in apo-red bean (adzuki) ferritin (apoRBF), the EGC-loaded apoRBF nanoparticle (ER) was fabricated with an encapsulation ratio of 11.6% (w/w). The results indicated that different chitosan molecules (with molecular weights of 980, 4600, 46 000, and 210 000 Da) could attach onto the apoRBF via electrostatic interactions to form ER-chitosan complexes (ERCs) (ERCs980, ERCs4600, ERCs46000, and ERCs210000). ERCs980 and ERCs4600 retained the typical shell-like morphology of ferritin with a size distribution of 12 nm and showed weak negative zeta-potentials at pH 6.7, while ERCs46000 and ERCs210000 significantly aggregated. Furthermore, the transport of EGC in ERCs980 and ERCs4600 across the Caco-2 cells was enhanced by the transferrin receptor 1 (TfR-1)-mediated absorption pathway, demonstrating that chitosan molecules with low molecular weights of 980 and 4600 Da were beneficial to the absorption of EGC based on the ferritin cage. This study will facilitate the application of ferritin-chitosan materials for fabricating the core-shell platform for encapsulation and bioavailability enhancement of bioactive molecules.

Comparative analysis of two ferritin subunits from blunt snout bream (Megalobrama amblycephala): Characterization, expression, iron depriving and bacteriostatic activity.

Iron is an essential microelement for almost all living organisms, while an excess of iron is toxic, thus maintenance of iron homeostasis is vital. As iron storage protein, ferritin plays an important role in iron metabolism. In the present study, we cloned and characterized the ferritin H subunit from Megalobrama amblycephala, termed as MamFerH. An iron-responsive element (IRE) was predicted in the 5' untranslated region (UTR) of MamFerH, while its bulge structural was different from that of the reported ferritin M subunit (MamFerM). The MamFerH and MamFerM genes exhibited similar expression patterns during early development with specifically high expression post hatching, whereas their tissue expression patterns were different. Specifically, MamFerM was highly expressed in the spleen, liver and kidney, while MamFerH was predominantly expressed in the blood and brain, indicating their different functions. In addition, the expression of the two genes was induced upon Aeromonas hydrophila infection at both transcriptional and translational levels, and MamFerH was more efficient. Immunohistochemistry and immunofluorescence analysis confirmed their significant changes at protein level and distribution in the liver post infection, indicating their participation in host immune response. Furthermore, bacteriostatic experiment revealed that recombinant MamFerH displayed more significant inhibitory effect on the growth of A. hydrophila.

Effect of Systemic Iron Overload and a Chelation Therapy in a Mouse Model of the Neurodegenerative Disease Hereditary Ferritinopathy.

Mutations in the ferritin light chain (FTL) gene cause the neurodegenerative disease neuroferritinopathy or hereditary ferritinopathy (HF). HF is characterized by a severe movement disorder and by the presence of nuclear and cytoplasmic iron-containing ferritin inclusion bodies (IBs) in glia and neurons throughout the central nervous system (CNS) and in tissues of multiple organ systems. Herein, using primary mouse embryonic fibroblasts from a mouse model of HF, we show significant intracellular accumulation of ferritin and an increase in susceptibility to oxidative damage when cells are exposed to iron. Treatment of the cells with the iron chelator deferiprone (DFP) led to a significant improvement in cell viability and a decrease in iron content. In vivo, iron overload and DFP treatment of the mouse model had remarkable effects on systemic iron homeostasis and ferritin deposition, without significantly affecting CNS pathology. Our study highlights the role of iron in modulating ferritin aggregation in vivo in the disease HF. It also puts emphasis on the potential usefulness of a therapy based on chelators that can target the CNS to remove and redistribute iron and to resolubilize or prevent ferritin aggregation while maintaining normal systemic iron stores.

Bioengineered Magnetoferritin Nanoprobes for Single-Dose Nuclear-Magnetic Resonance Tumor Imaging.

Despite all the advances in multimodal imaging, it remains a significant challenge to acquire both magnetic resonance and nuclear imaging in a single dose because of the enormous difference in sensitivity. Indeed, nuclear imaging is almost 10(6)-fold more sensitive than magnetic resonance imaging (MRI); thus, repeated injections are generally required to obtain sufficient MR signals after nuclear imaging. Here, we show that strategically engineered magnetoferritin nanoprobes can image tumors with high sensitivity and specificity using SPECT and MRI in living mice after a single intravenous injection. The magnetoferritin nanoprobes composed of (125)I radionuclide-conjugated human H-ferritin iron nanocages ((125)I-M-HFn) internalize robustly into cancer cells via a novel tumor-specific HFn-TfR1 pathway. In particular, the endocytic recycling characteristic of TfR1 transporters solves the nuclear signal blocking issue caused by the high dose nanoprobes injected for MRI, thus enabling simultaneous functional and morphological tumor imaging without reliance on multi-injections.

Tracking the fear memory engram: discrete populations of neurons within amygdala, hypothalamus, and lateral septum are specifically activated by auditory fear conditioning.

Memory formation is thought to occur via enhanced synaptic connectivity between populations of neurons in the brain. However, it has been difficult to localize and identify the neurons that are directly involved in the formation of any specific memory. We have previously used fos-tau-lacZ (FTL) transgenic mice to identify discrete populations of neurons in amygdala and hypothalamus, which were specifically activated by fear conditioning to a context. Here we have examined neuronal activation due to fear conditioning to a more specific auditory cue. Discrete populations of learning-specific neurons were identified in only a small number of locations in the brain, including those previously found to be activated in amygdala and hypothalamus by context fear conditioning. These populations, each containing only a relatively small number of neurons, may be directly involved in fear learning and memory.

Overexpression of the MRI Reporter Genes Ferritin and Transferrin Receptor Affect Iron Homeostasis and Produce Limited Contrast in Mesenchymal Stem Cells.

Imaging technologies that allow the non-invasive monitoring of stem cells in vivo play a vital role in cell-based regenerative therapies. Recently, much interest has been generated in reporter genes that enable simultaneous monitoring of the anatomical location and viability of cells using magnetic resonance imaging (MRI). Here, we investigate the efficacy of ferritin heavy chain-1 (Fth1) and transferrin receptor-1 (TfR1) as reporters for tracking mesenchymal stem cells. The overexpression of TfR1 was well tolerated by the cells but Fth1 was found to affect the cell's iron homeostasis, leading to phenotypic changes in the absence of iron supplementation and an upregulation in transcript and protein levels of the cell's endogenous transferrin receptor. Neither the sole overexpression of Fth1 nor TfR1 resulted in significant increases in intracellular iron content, although significant differences were seen when the two reporter genes were used in combination, in the presence of high concentrations of iron. The supplementation of the culture medium with iron sources was a more efficient means to obtain contrast than the use of reporter genes, where high levels of intracellular iron were reflected in transverse (T2) relaxation. The feasibility of imaging iron-supplemented cells by MRI is shown using a 3R-compliant chick embryo model.

Involvement of iron depletion in palmitate-induced lipotoxicity of beta cells.

High levels of plasma free fatty acid are thought to contribute to the loss of pancreatic beta-cells in type 2 diabetes. In particular, saturated fatty acid such as palmitate or stearate can induce apoptosis in cultured beta cells (lipotoxicity). Endoplasmic reticulum stress is a critical mediator of free fatty acid-induced lipotoxicity. Recently, disorders in mitochondrial respiratory metabolism have been linked to lipotoxicity. Since iron is a critical component of respiratory metabolism, this study is initiated to determine whether abnormal iron metabolism is involved in palmitate-induced beta cell death. Immunoblotting analysis showed that treatment of INS-1 beta cells with palmitate reduced the level of transferrin receptor 1 (TfR1), but increased the level of heavy chain ferritin (FTH). In addition, palmitate reduced intracellular labile iron pool. Whereas iron depletion through treatment with iron-chelators deferoxamine or deferasirox augmented palmitate-induced cell death, iron supplementation with ferric chloride, ferrous sulfate, or holo-transferrin significantly protected cells against palmitate-induced death. Furthermore, overexpression of TfR1 reduced palmitate-induced cell death, whereas knockdown of TfR1 augmented cell death. In particular, treatment with deferoxamine increased the level of endoplasmic reticulum (ER) stress markers phospho-PERK, phospho-eIF2α, CHOP and phospho-c-Jun N-terminal kinase. Treatment with chemical chaperone significantly protected cells against deferoxamine-induced apoptosis. Iron supplementation also protected cells against palmitate-induced primary islet death. These data suggest that iron depletion plays an important role in palmitate-induced beta cell death through inducing ER stress. Therefore, attempts to block iron depletion might be able to prevent beta cell loss in type 2 diabetes.

Molecular cloning and expression analysis of ferritin, heavy polypeptide 1 gene from duck (Anas platyrhynchos).

H-ferritin is a core subunit of the iron storage protein ferritin, and is related to the pathogenesis of malignant diseases. A differential expressed sequence tag of the ferritin, heavy polypeptide 1 gene (FTH1) was obtained from our previously constructed suppression subtractive cDNA library from 3-day-old ducklings challenged with duck hepatitis virus type I (DHV-1). The expression and function of FTH1 in immune defense against infection remains largely unknown in ducks. In this study, the full-length duFTH1 cDNA was obtained using reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends. It consisted of 153 basepairs (bp) 5'untranslated region (UTR), 183 bp 3'UTR, and 546 bp open reading frame that encodes a single protein of 181 amino acid residues. duFTH1 shares high similarity with FTH1 genes from other vertebrates. The amino acid sequence possesses the conserved domain of typical ferritin H subunits, including seven metal ligands in the ferroxidase center, one iron binding region signature, and a potential bio-mineralization residue (Thy(29)). Moreover, in agreement with a previously reported ferritin H subunit, we identified an iron response element in the 5'UTR. RT-PCR analyses revealed duFTH1 mRNA is widely expressed in various tissues. Real-time quantitative polymerase chain reaction analyses suggested that duFTH1 mRNA is significantly up-regulated in the liver after DHV-1 injection or polyriboinosinic polyribocytidylic acid (polyI:C) treatment, reaching a peak 4 h post-infection, and dropping progressively and returning to normal after 24 h. Our findings suggest that duFTH1 functions as an iron chelating protein subunit in duck and contributes to the innate immune responses against viral infections.

Semi-artificial and bioactive ferroxidase with nanoparticles as the active sites.

Light-chain apoferritin lacks ferroxidase activity, which can be supplemented with Pt-nanoparticles. The hybrid bioinorganic nanoparticle outperforms its heavy-chain pendant in terms of ferroxidase activity, mineralization ability and inhibition resistance. Being active in a cellular environment it regulates the iron homeostasis.

Iron bioavailability in larvae yellow snapper (Lutjanus argentiventris): cloning and expression analysis of ferritin-H.

Ferritin is a major intracellular iron storage protein in higher vertebrates and plays an important role in iron metabolism. In this study, ferritin H subunit was cloned from the larvae of yellow snapper, Lutjanus argentiventris, by rapid amplification of cDNA ends (RACE) following in silico transcriptome analysis. The full-length cDNAs of the LaFeH was 1231 bp in length encoding 177 amino acids with a predicted molecular mass (MW) about 20.82 kDa and theoretical isoelectric point (pI) of 5.79. Amino acid alignment revealed that LaFeH shared high similarity with other known ferritins. It shared high degree identity to the ferritin H subunits of Lates calcarifer (99%), Takifugu rubripes (97%) and Dicentrarchus labrax (97%), and low identity to that of human (82%) and mouse (84%). By real-time PCR assays, the mRNA transcripts of LaFeH was found to be higher expressed in head-kidney, eye, heart and brain. Moreover, mRNA expression levels of LaFeH was measured by real-time PCR in larvae exposed with graded levels of iron (6.8 µg/ml and 13.6 µg/ml (Fe2x and Fe4x, respectively) and an iron chelation assay. Results showed that the expression of the LaFeH mRNA increased gradually with Fe2x in water. The LaFeH gene expression declined with increasing iron exposure levels at Fe4x. Finally, we can observe a high expression of LaFeH gene in larvae exposed to iron chelation therapy at 2 h; however this increase was gradually decreasing over time. In summary, the LaFeH gene expression for larvae yellow snapper showed a dose-depend increase following the iron treatment. These data indicated that iron bioavailability regulates LaFeH at transcriptional level in larvae yellow snapper. Further studies are necessary to ascertain their role in the immune response in teleost fish.

Efficiency of ferritin as an MRI reporter gene in NPC cells is enhanced by iron supplementation.

BACKGROUND: An emerging MRI reporter, ferritin heavy chain (FTH1), is recently applied to enhance the contrast and increase the sensitivity of MRI in the monitoring of solid tumors. However, FTH1-overexpression-related cytotoxicity is required to be explored. METHODS: By using the Tet-Off system, FTH1 overexpression was semi-quantitativiely and dynamicly regulated by doxycycline in a NPC cell line. Effects of FTH1 overexpression on the proliferation, cytotoxicity, apoptosis and migration of NPC cells were investigated in vitro, and MR relaxation rate was measured in vitro and in vivo. RESULTS: In vitro and in vivo overexpression of FTH1 significantly increased the transverse relaxivity (R(2)), which could be enhanced by iron supplementation. In vitro, overexpression of FTH1 reduced cell growth and migration, which were not reduced by iron supplementation. Furthermore, cells were subcutaneously inoculated into the nude mice. Results showed FTH1 overexpression decreased tumor growth in the absence of iron supplementation but not in the presence of iron supplementation. CONCLUSION: To maximize R(2) and minimize the potential adverse effects, supplementation of iron at appropriate dose is recommended during the application of FTH1 as a reporter gene in the monitoring of NPC by MRI.

Postmortem and imaging based analyses reveal CNS decreased myelination in restless legs syndrome.

BACKGROUND: Restless legs syndrome (RLS) is a neurological disorder characterized by a strong urge to move the legs and has been shown in many studies with abnormally low brain iron. Iron deficiency is associated with hypomyelination in brains of animals. Therefore we hypothesized that a myelin deficit should be present in the brains of patients with RLS. METHODS: We performed Western blot analysis on myelin isolated from RLS (n=11) and control (n=11) brain tissue obtained at autopsy for the expression of the integral myelin proteins, myelin basic protein (MBP), and proteolipid protein (PLP) and the oligodendrocyte specific enzyme 3'5'-cyclic nucleotide phosphohydrolase (CNPase). To expand the postmortem findings to in vivo, we analyzed the brains of RLS patients (n=23) and controls (n=23) using voxel-based morphometry (VBM). RESULTS: The expression of MBP, PLP and CNPase in the myelin from RLS was decreased by approximately 25% (p<0.05) compared to controls. The amounts of transferrin (Tf) and H-ferritin (H-Frt) in the myelin fraction were also significantly decreased in RLS compared to controls. The imaging analysis revealed significant small decreases in white matter volume in RLS patients compared to controls in the corpus callosum, anterior cingulum and precentral gyrus. CONCLUSION: A decrease in myelin similar to that reported in animal models of iron deficiency was found in the brains of individuals with RLS. The evidence for less myelin and loss of myelin integrity in RLS brains, coupled with decreased ferritin and transferrin in the myelin fractions, is a compelling argument for brain iron insufficiency in RLS. These data also indicate the need to look beyond the sensorimotor symptoms that typically define the syndrome and its assumed relation to the dopaminergic system. Understanding the full range of RLS pathology may help us better understand the complex, intermittent nature and diversity of the clinical features of RLS and expand our consideration of treatment options for RLS.

Ferritin heavy chain-mediated iron homoeostasis regulates expression of IL-10 in Chlamydia trachomatis-infected HeLa cells.

Chlamydia trachomatis is the leading cause of sexually transmitted infection worldwide, in which disease outcome is determined by the balance between pro- and anti-inflammatory host immune responses. Iron plays important roles in regulation and enhancement of various pro- and anti-inflammatory cytokines. Earlier studies have established essentiality of iron in C. trachomatis infection; however, there is lack of study wherein modulatory effect of iron regulated protein [FHC (ferritin heavy chain)] in regulation of anti-inflammatory cytokine IL (interleukin)-10 has been investigated. In this study, immunoblotting results showed the up-regulation of FHC in C. trachomatis-infected HeLa cells in comparison with mock (in vitro control). Further secretory IL-10 level was significantly increased (P<0.001) or decreased (P<0.001) in response to iron supplementation [FAC (ferric ammonium citrate)] and depletion [DFO (deferoxamine)], respectively. However, in C. trachomatis-infected HeLa cells, levels of IL-10 remain higher, irrespective of availability of iron in comparison with their respective control. These results showed that secretion of IL-10 and expressions of FHC have concordance. Further, to understand interdependence of IL-10 and iron homoeostasis (regulation), the levels of IL-10 were compared with iron-responsive GFP (green fluorescent protein) expression in HeLa-229 cells. The mean fluorescent intensities of GFP were in accordance with levels of IL-10 in C. trachomatis-infected cells. These results showed the association of secreted IL-10, FHC and iron homoeostasis in C. trachomatis-infected HeLa-229 cells. This study provides insight into host-Chlamydia interaction at the crossroad of iron metabolism and immune responses and may help in realizing the potential of iron homoeostasis modulators in treatment of chronic chlamydial infection.

Ferritin as a reporter gene for MRI: chronic liver over expression of H-ferritin during dietary iron supplementation and aging.

The iron storage protein, ferritin, provides an important endogenous MRI contrast that can be used to determine the level of tissue iron. In recent years the impact of modulating ferritin expression on MRI contrast and relaxation rates was evaluated by several groups, using genetically modified cells, viral gene transfer and transgenic animals. This paper reports the follow-up of transgenic mice that chronically over-expressed the heavy chain of ferritin (h-ferritin) in liver hepatocytes (liver-hfer mice) over a period of 2 years, with the aim of investigating the long-term effects of elevated level of h-ferritin on MR signal and on the well-being of the mice. Analysis revealed that aging liver-hfer mice, exposed to chronic elevated expression of h-ferritin, have increased R(2) values compared to WT. As expected for ferritin, R(2) difference was strongly enhanced at high magnetic field. Histological analysis of these mice did not reveal liver changes with prolonged over expression of ferritin, and no differences could be detected in other organs. Furthermore, dietary iron supplementation significantly affected MRI contrast, without affecting animal wellbeing, for both wildtype and ferritin over expressing transgenic mice. These results suggest the safety of ferritin over-expression, and support the use of h-ferritin as a reporter gene for MRI.