Systemic and cellular consequences of macrophage control of iron metabolism.

Iron is necessary for both mammalian cells and microorganisms, which fiercely compete for this essential nutrient. Accordingly, macrophages exploit the denial of iron from microbial pathogens as an important strategy to accomplish their key role in innate immunity and host defense. Macrophages employ multiple mechanisms to accumulate iron and thus contain microbial infections, but this may come at a price. In particular, at the systemic level iron sequestration in the reticuloendothelial cells can lead to the development of anemia of chronic disease. At the local level, iron sequestration in macrophages, which is targeted to extracellular invaders, can in turn favor intracellular pathogens. Moreover, iron accumulation can per se promote pro-inflammatory activation of macrophages and consequently contribute to maintain the process of inflammation, without resolution. Finally, the peculiar iron trafficking that characterizes alternatively polarized macrophages can influence neighboring cells in the microenvironment and impact on the resolution phase of inflammation. In this review, we describe the role of macrophages in iron metabolism in the context of host defense and iron balance.

Localized conventional radiotherapy in the treatment of Mycosis Fungoides: our experience in 100 patients.

BACKGROUND: Radiotherapy (RT) is one of the treatments of choice as skin-directed therapy in Mycosis Fungoides (MF), both in first stages of the disease as total skin electron beam irradiation and in tumoural stage as localized treatment with conventional energies or electrons. OBJECTIVE: Through a retrospective study, to evaluate the results of localized superficial RT in a series of 100 patients affected by MF. METHODS: All the patients, after diagnosis supported by histological and immunophenotyping investigations, have been treated with conventional RT (range 50-150 kV) and a total dose ranging from 9 to 40 Gy. RESULTS: Complete remission of the irradiated lesion has been observed in 88%, partial remission in 6% and non-response in 2%. Four patients were lost to follow-up. Local relapse has been observed in 13 lesions, with a local control rate of 85% after 5 years from the end of RT. Cosmetic results have been good and acceptable in 93% of cases. The treatment has been always well tolerated. The results confirm to be dose dependent, and show that better response is found in the range of higher energies. CONCLUSION: Localized RT is an effective and safe tool in the care and palliation of MF.

Dermatology and COVID-19: The Hidden Pandemic.

Since December 2019, a novel coronavirus, known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has been rapidly spreading across the world, leading to the declared pandemic of COVID-19 [...].

Oral polypodium leucotomos extract photoprotective activity in 57 patients with idiopathic photodermatoses.

AIM: Idiopathic photodermatoses (IP) are a recurrent, acquired sunlight-induced rash of delayed onset, appearing after exposure to ultraviolet radiation in susceptible individuals. The aim of this study was to assess the photoprotective activity of polypodium leucotomos (PL) in IP. METHODS: Fifty-seven patients affected by IP were recruited for the study (53 with polymorphic light eruption and 4 with solar urticaria). The use of UV protection filters or other drugs that could in some way interfere with exposure to light were excluded. All patients exposed themselves to sunlight while consuming 480 mg/day of PL extract orally. A statistical evaluation of the basal clinical conditions compared to those after sunlight exposure with PL was performed. RESULTS: About 73.68% of the patients had a benefit from the administered PL, with a significant reduction of skin reaction and subjective symptoms. No side effects were observed. Results were statistically significant (P<0.05). CONCLUSION: PL complete absence of toxicity combined with its multifactorial protection, makes it an effective and safe treatment for photoprotection in IP.

Effect of reactive oxygen species on iron regulatory protein activity.

Iron may be important in catalyzing excessive production of reactive oxygen species (ROS). Cellular iron homeostasis is regulated by iron regulatory proteins (IRPs), which bind to iron-responsive elements (IRE) of mRNAs for ferritin and transferrin receptor (TfR) modulating iron uptake and sequestration, respectively. Although iron is the main regulator of IRP activity, IRP is also influenced by other factors, including the redox state. Therefore, IRP might be sensitive to pathophysiological alterations of redox state caused by ROS. However, previous studies have produced diverging evidence on the effect of oxidative injury on IRP. Results obtained in an animal model close to a pathophysiological condition, such as ischemia reperfusion of the liver as well as in a cell-free system involving an enzymatic source of O2 and H2O2, indicate that IRP is downregulated by oxidative stress. In fact, IRP activity is inhibited at early times of post-ischemic reperfusion. Moreover, the concerted action of O2 and H2O2 produced by xanthine oxidase in a cell-free system caused a remarkable inhibition of IRP activity. IRP seems a direct target of ROS; in fact, in vivo inhibition can be prevented by the antioxidant N-acetylcysteine and by interleukin-1 receptor antagonist. In addition, modulation of iron levels of the cell-free assay did not affect the downregulation imposed by xanthine oxidase. Conceivably, downregulation of IRP activity by O2 and H2O2 may facilitate iron sequestration into ferritin, thus limiting the pro-oxidant challenge of iron.

Diacerhein blocks iron regulatory protein activation in inflamed human monocytes.

Iron Regulatory Proteins (IRPs), by modulating expression of ferritin, which stores excess iron in a non toxic form, and transferrin receptor, which controls iron uptake, are the main controller of cellular iron metabolism. During inflammation, modification of IRP activity may affect iron availability, free radical generation and cytokine gene response in inflammatory cells. In the present study we tested the effect of inflammatory stimuli on IRP function in a human monocytic-macrophagic cell line and the possibility of interfering with these pathways by using an antiinflammatory compound, diacerhein (DAR). IRP activity was enhanced by interferon gamma/lipopolysaccarhide (IFN/LPS), and this effect was consistently counteracted by increasing concentrations of DAR. No direct effect of DAR on IRP activity was found in vitro. However, in vivo, similar IRP activation was achieved by exposing cells to nitric oxide (NO) donors and the LPS/IFN-induced activation of IRP was reversed by NO inhibitors. Interestingly, NO-induced IRP activation was efficiently blocked by DAR. These data show for the first time that a clinically useful antiinflammatory compound, DAR, interferes with IRP activation by NO in inflammed human cells.

Role of hypoxia-inducible factors in the dexrazoxane-mediated protection of cardiomyocytes from doxorubicin-induced toxicity.

BACKGROUND AND PURPOSE: Iron aggravates the cardiotoxicity of doxorubicin, a widely used anticancer anthracycline, and the iron chelator dexrazoxane is the only agent protecting against doxorubicin cardiotoxicity; however, the mechanisms underlying the role of iron in doxorubicin-mediated cardiotoxicity and the protective role of dexrazoxane remain to be established. As iron is required for the degradation of hypoxia-inducible factors (HIF), which control the expression of antiapoptotic and protective genes, we tested the hypothesis that dexrazoxane-dependent HIF activation may mediate the cardioprotective effect of dexrazoxane. EXPERIMENTAL APPROACH: Cell death, protein levels (by immunoblotting) and HIF-mediated transcription (using reporter constructs) were evaluated in the rat H9c2 cardiomyocyte cell line exposed to low doses of doxorubicin with or without dexrazoxane pretreatment. HIF levels were genetically manipulated by transfecting dominant-negative mutants or short hairpin RNA. KEY RESULTS: Treatment with dexrazoxane induced HIF-1α and HIF-2α protein levels and transactivation capacity in H9c2 cells. It also prevented the induction of cell death and apoptosis by exposure of H9c2 cells to clinically relevant concentrations of doxorubicin. Suppression of HIF activity strongly reduced the protective effect of dexrazoxane. Conversely, HIF-1α overexpression protected against doxorubicin-mediated cell death and apoptosis also in cells not exposed to the chelator. Exposure to dexrazoxane increased the expression of the HIF-regulated, antiapoptotic proteins survivin, Mcl1 and haem oxygenase. CONCLUSIONS AND IMPLICATIONS: Our results showing HIF-dependent prevention of doxorubicin toxicity in dexrazoxane-treated H9c2 cardiomyocytes suggest that HIF activation may be a mechanism contributing to the protective effect of dexrazoxane against anthracycline cardiotoxicity.

New monoclonal antibodies against B-cell antigens: possible new strategies for diagnosis of primary cutaneous B-cell lymphomas.

Reactivities of the monoclonal antibodies (mAbs) of the 9th Human Leukocyte Differentiation Antigen Workshop, in order to define specific antigenic expression of the primary cutaneous B-cell lymphomas (PC-BCL), were analyzed by immunohistology on human tonsil and on PC-BCL, such as follicular centre B-cell lymphomas (FCL), marginal zone lymphomas (MZL) and diffuse large B-cells lymphomas leg-type (DLBL-LT). We identified some subgroups of mAbs that were exclusively or preferentially positive in one lymphoma cell type: the PC-FCL subgroup of mAbs includes PD1/CD279, GCET-1, hFCRL1/CD307a, FCRL2/CD307b, CXCR5/CD185, B7-DC/CD273, MRC/CD200, CD130, CXCR4/CD184, Siglec-5/14, CD150, on the other hand subgroup of mAbs in PC-MZL includes BTLA/CD272, BLIMP-1, hCD38. No specific subgroup of mAbs was found to label PC-DLBCL. This study may be useful to better define specific antigen profile of different PC-BCL entities leading to a correct diagnosis.

A precious metal: Iron, an essential nutrient for all cells.

Iron is an important cofactor required for a number of essential cell functions and hence is a vital nutrient. However, iron can also be dangerous as a catalyst of free radical reactions. Accordingly, intracellular iron homeostasis and body iron balance are tightly regulated. In this review, we presented an overview of the remarkable new insights that over the last years have been gained into the multifaceted and sophisticated molecular mechanisms controlling iron acquisition, storage and release. We also reviewed the data about nutrition-related abnormalities of iron metabolism, such as iron overload and deficiency. Finally, we discussed how pathogenic microorganisms and host cells compete for iron, a battle whose outcome has a relevant role in infectious disease.

Preferential activation of iron regulatory protein-2 in cell lines as a result of higher sensitivity to iron.

Iron regulatory proteins (IRP)-1 and 2 are cytoplasmic mRNA-binding proteins that control intracellular iron homeostasis by regulating the translation of ferritin mRNA and stability of transferrin receptor mRNA in an iron-dependent fashion. Although structurally and functionally similar, the two IRP are different in their mode of regulation, pattern of tissue expression and modulation by multiple factors, such as bioradicals. In the present study RNA bandshift assays demonstrated that IRP-2, but not IRP-1, activity was higher in cultured cells than in tissues. Increased expression of IRP-2 in cell lines was not related to immortalization and differentiation but seemed associated to cell proliferation, although not closely dependent on cell growth rate. As a growing cell consumes more iron than its quiescent counterpart, we assessed the iron status of cell lines and found that ferritin content was lower than in tissues. Analysis of IRP activity in cell lines supplemented with heme or non-heme iron and in livers of iron-loaded and iron-deficient rats indicated that IRP-2 responds more promptly than IRP-1 to modulations of iron content. We propose that enhanced IRP-2 activity in cultured cells could be due to a proliferation-dependent, relative iron deficiency that is sensed first by IRP-2.

Induction of ferritin synthesis in ischemic-reperfused rat liver: analysis of the molecular mechanisms.

BACKGROUND & AIMS: Iron may catalyze the production of reactive oxygen species (ROS) during postischemic reoxygenation. Ferritin, a cellular iron storage protein, can either represent a source of iron or perform a cytoprotective action against ROS. The aim of this study was to address the role of ferritin in postischemic reperfusion. METHODS: Transcriptional and posttranscriptional mechanisms controlling ferritin gene expression were studied in reperfused rat livers. RESULTS: Proteolysis reduced ferritin levels 2 hours after reperfusion, but a concomitant increase of synthesis, accompanied by enhanced transcription and accumulation of H and L ferritin subunit messenger RNAs (mRNAs), almost re-established normal ferritin content at 4 hours. Pretreatment with interleukin 1 receptor antagonist (IL-1RA) did not prevent the rise of ferritin mRNAs. RNA bandshift assays showed that the activity of the iron regulatory proteins (IRPs), which control ferritin mRNA translation, declined early after reperfusion and recovered progressively thereafter. Pretreatment with either the antioxidant N-acetyl cysteine or IL-1RA was sufficient to prevent almost completely down-regulation of IRP activity. CONCLUSIONS: Postischemic reperfusion causes degradation of ferritin, possibly increasing iron levels. However, induction of ferritin gene transcription, possibly mediated by ferritin-derived iron and ROS-mediated inactivation of IRP, which allows translation of ferritin mRNAs, counteracts this effect and concurs to reestablish the amount of ferritin, which may thus act to limit reperfusion damage.

Peroxisomal targeting of mammalian hydroxyacid oxidase 1 requires the C-terminal tripeptide SKI.

Peroxisomal proteins are post-translationally imported into peroxisomes after recognition by specific receptors. The best-defined peroxisomal targeting signal is a C-terminal tripeptide SKL. Different functional variants of this tripeptide have been defined, but mutants with a SKI sequence were recognized as being inefficiently targeted to peroxisomes. Recently, we have cloned a cDNA for the mouse hydroxyacid oxidase 1 (Hao1), a protein that seems to be localized in peroxisomes. Interestingly, the mouse Hao1 sequence comprises a C-terminal SKI tripeptide. We have analyzed the subcellular localization of Hao1 and tested whether its SKI sequence acts as a targeting signal. Ltk(-) and Cos-7 cells were transfected with vectors expressing a fusion protein of green fluorescence protein and Hao1, as well as mutants thereof. Targeting to peroxisomes of the fusion protein with the wild-type SKI sequence was highly selective and as complete as with the peroxisome-specific SKL sequence. By contrast, targeting was lost in a mutant with the sequence CKM. The data show that mammalian Hao1 is a peroxisomal protein and that the C-terminal sequence SKI acts as the targeting signal.

Nitric oxide-mediated induction of ferritin synthesis in J774 macrophages by inflammatory cytokines: role of selective iron regulatory protein-2 downregulation.

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)-gamma/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.

Inappropriately high iron regulatory protein activity in monocytes of patients with genetic hemochromatosis.

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.

Macrophage preconditioning with synthetic malaria pigment reduces cytokine production via heme iron-dependent oxidative stress.

Hemozoin (malaria pigment), a polymer of hematin (ferri-protoporphyrin IX) derived from hemoglobin ingested by intraerythrocytic plasmodia, modulates cytokine production by phagocytes. Mouse peritoneal macrophages (PM) fed with synthetic beta-hematin (BH), structurally identical to native hemozoin, no longer produce tumor necrosis factor alpha (TNFalpha) and nitric oxide (NO) in response to lipopolysaccharide (LPS). Impairment of NO synthesis is due to inhibition of inducible nitric oxide synthase (iNOS) production. BH-mediated inhibition of PM functions cannot be ascribed to iron release from BH because neither prevention by iron chelators nor down-regulation of iron-regulatory protein activity was detected. Inhibition appears to be related to pigment-induced oxidative stress because (a) thiol compounds partially restored PM functions, (b) heme oxygenase (HO-1) and catalase mRNA levels were up-regulated, and (c) free radicals production increased in BH-treated cells. The antioxidant defenses of the cells determine the response to BH: microglia cells, which show a lower extent of induction of HO-1 and catalase mRNAs and lower accumulation of oxygen radicals, are less sensitive to the inhibitory effect of BH on cytokine production. Results indicate that BH is resistant to degradation by HO-1 and that heme-iron mediated oxidative stress may contribute to malaria-induced immunosuppression. This study may help correlate the different clinical manifestations of malaria, ranging from uncomplicated to severe disease, with dysregulation of phagocyte functions and promote better therapeutic strategies to counteract the effects of hemozoin accumulation.