EspH interacts with the host active Bcr related (ABR) protein to suppress RhoGTPases.

Enteropathogenic are bacterial pathogens that colonize the gut and cause severe diarrhea in humans. Upon intimate attachment to the intestinal epithelium, these pathogens translocate via a type III secretion system virulent proteins, termed effectors, into the host cells. These effectors manipulate diverse host cell organelles and functions for the pathogen's benefit.

Enteropathogenic Escherichia coli remodels host endosomes to promote endocytic turnover and breakdown of surface polarity.

Enteropathogenic E. coli (EPEC) is an extracellular diarrheagenic human pathogen which infects the apical plasma membrane of the small intestinal enterocytes. EPEC utilizes a type III secretion system to translocate bacterial effector proteins into its epithelial hosts.

Temporal profiling of redox-dependent heterogeneity in single cells.

Cellular redox status affects diverse cellular functions, including proliferation, protein homeostasis, and aging. Thus, individual differences in redox status can give rise to distinct sub-populations even among cells with identical genetic backgrounds. Here, we have created a novel methodology to track redox status at single cell resolution using the redox-sensitive probe Grx1-roGFP2.

Flow cytometric quantification and characterization of intracellular protein aggregates in yeast.

The sequestration of misfolded proteins into aggregates is an integral pathway of the protein quality control network that becomes particularly prominent during proteotoxic stress and in various pathologies. Methods for systematic analysis of cellular aggregate content are still largely limited to fluorescence microscopy and to separation by biochemical techniques. Here, we describe an alternative approach, using flow cytometric analysis, applied to protein aggregates released from their intracellular milieu by mild lysis of yeast cells.

Ubiquitin conjugation triggers misfolded protein sequestration into quality control foci when Hsp70 chaperone levels are limiting.

Ubiquitin accumulation in amyloid plaques is a pathological marker observed in the vast majority of neurodegenerative diseases, yet ubiquitin function in these inclusions is controversial. It has been suggested that ubiquitylated proteins are directed to inclusion bodies under stress conditions, when both chaperone-mediated refolding and proteasomal degradation are compromised or overwhelmed. Alternatively, ubiquitin and chaperones may be recruited to preformed inclusions to promote their elimination.

The role of endocytic pathways in cellular uptake of plasma non-transferrin iron.

Background: In transfusional siderosis, the iron binding capacity of plasma transferrin is often surpassed, with concomitant generation of non-transferrin-bound iron. Although implicated in tissue siderosis, non-transferrin-bound iron modes of cell ingress remain undefined, largely because of its variable composition and association with macromolecules. Using fluorescent tracing of labile iron in endosomal vesicles and cytosol, we examined the hypothesis that non-transferrin-bound iron fractions detected in iron overloaded patients enter cells via bulk endocytosis.

Non-transferrin bound iron in Thalassemia: differential detection of redox active forms in children and older patients.

Non-transferrin bound iron (NTBI) is commonly detected in patients with systemic iron overload whose serum iron-binding capacity has been surpassed. It has been perceived as an indicator of iron overload, impending organ damage and a chelation target in poly-transfused thalassemia patients. However, NTBI is a heterogeneous entity comprising various iron complexes, including a significant redox-active and readily chelatable fraction, which we have designated as "labile plasma iron" (LPI).

Rescuing iron-overloaded macrophages by conservative relocation of the accumulated metal.

Background And Purpose: Systemic iron deficiency concomitant with macrophage iron retention is characteristic of iron-refractory anaemias associated with chronic disease. The systemic misdistribution of iron, which is further exacerbated by parenteral iron supplementation, is mainly attributable to iron retention exerted on resident macrophages by hepcidin-mediated down-regulation of the iron exporter ferroportin. We aimed at developing an experimental macrophage-based cell model that recapitulates pathophysiological features of iron misdistribution found in chronic disorders and use it as a screening platform for identifying agents with the potential for relocating the accumulated metal and restoring affected functions.

Iron redistribution as a therapeutic strategy for treating diseases of localized iron accumulation.

Defective iron utilization leading to either systemic or regional misdistribution of the metal has been identified as a critical feature of several different disorders. Iron concentrations can rise to toxic levels in mitochondria of excitable cells, often leaving the cytosol iron-depleted, in some forms of neurodegeneration with brain accumulation (NBIA) or following mutations in genes associated with mitochondrial functions, such as ABCB7 in X-linked sideroblastic anemia with ataxia (XLSA/A) or the genes encoding frataxin in Friedreich's ataxia (FRDA). In anemia of chronic disease (ACD), iron is withheld by macrophages, while iron levels in extracellular fluids (e.

Transferrin therapy ameliorates disease in beta-thalassemic mice.

Individuals with beta-thalassemia develop progressive systemic iron overload, resulting in high morbidity and mortality. These complications are caused by labile plasma iron, which is taken up by parenchymal cells in a dysregulated manner; in contrast, erythropoiesis depends on transferrin-bound iron uptake via the transferrin receptor. We hypothesized that the ineffective erythropoiesis and anemia observed in beta-thalassemia might be ameliorated by increasing the amount of circulating transferrin.

Daily labile plasma iron as an indicator of chelator activity in Thalassaemia major patients.

Labile plasma iron (LPI), a non-transferrin-bound component of plasma iron detected in iron overload disorders is a potential source of cellular iron accumulation and ensuing oxidative damage. Periodic monitoring of LPI over a 24 h time-span was used to compare the ability of chelation to control daily LPI levels in 40 Thalassaemia major patients (9-11/group) who had been receiving one of three different chelation protocols for more than a year: Group I. deferrioxamine overnight, Group II.

Exogenous iron increases hemoglobin in beta-thalassemic mice.

Objective: Beta-thalassemia results from beta-globin gene mutations that lead to ineffective erythropoiesis, shortened red cell survival, and anemia. Patients with beta-thalassemia develop iron overload, despite which, hepcidin levels are low. This suggests that hepcidin regulation in beta-thalassemia is more sensitive to factors unrelated to iron state.

Cell functions impaired by frataxin deficiency are restored by drug-mediated iron relocation.

Various human disorders are associated with misdistribution of iron within or across cells. Friedreich ataxia (FRDA), a deficiency in the mitochondrial iron-chaperone frataxin, results in defective use of iron and its misdistribution between mitochondria and cytosol. We assessed the possibility of functionally correcting the cellular properties affected by frataxin deficiency with a siderophore capable of relocating iron and facilitating its metabolic use.

Redistribution of accumulated cell iron: a modality of chelation with therapeutic implications.

Various pathologies are characterized by the accumulation of toxic iron in cell compartments. In anemia of chronic disease, iron is withheld by macrophages, leaving extracellular fluids iron-depleted. In Friedreich ataxia, iron levels rise in the mitochondria of excitable cells but decrease in the cytosol.

Intracellular labile iron.

Cells maintain organellar pools of "labile iron" (LI), despite its propensity for catalyzing the formation of reactive oxygen species. These pools are identifiable by iron-chelating probes and accessible to pharmacological agents. Cytosolic LI has been assumed to have a dual function: providing a rapidly adjustable source of iron for immediate metabolic utilization, and for sensing by iron-regulatory proteins (IRPs) that regulate iron uptake and compartmentalization via transferrin receptors and ferritin.

Ineffective erythropoiesis in beta-thalassemia is characterized by increased iron absorption mediated by down-regulation of hepcidin and up-regulation of ferroportin.

Progressive iron overload is the most salient and ultimately fatal complication of beta-thalassemia. However, little is known about the relationship among ineffective erythropoiesis (IE), the role of iron-regulatory genes, and tissue iron distribution in beta-thalassemia. We analyzed tissue iron content and iron-regulatory gene expression in the liver, duodenum, spleen, bone marrow, kidney, and heart of mice up to 1 year old that exhibit levels of iron overload and anemia consistent with both beta-thalassemia intermedia (th3/+) and major (th3/th3).

Downregulation of hepcidin and haemojuvelin expression in the hepatocyte cell-line HepG2 induced by thalassaemic sera.

Beta-thalassaemia represents a group of diseases, in which ineffective erythropoiesis is accompanied by iron overload. In a mouse model of beta-thalassaemia, we observed that the liver expressed relatively low levels of hepcidin, which is a key factor in the regulation of iron absorption by the gut and of iron recycling by the reticuloendothelial system. It was hypothesised that, despite the overt iron overload, a putative plasma factor found in beta-thalassaemia might suppress liver hepcidin expression.

Action of chelators in iron-loaded cardiac cells: Accessibility to intracellular labile iron and functional consequences.

Labile iron in hemosiderotic plasma and tissue are sources of iron toxicity. We compared the iron chelators deferoxamine, deferiprone, and deferasirox as scavengers of labile iron in plasma and cardiomyocytes at therapeutic concentrations. This comprised chelation of labile plasma iron (LPI) in samples from thalassemia patients; extraction of total cellular iron; accessing labile iron accumulated in organelles and preventing formation of reactive-oxidant species; and restoring impaired cardiac contractility.

Non-transferrin-bound iron and risk of coronary heart disease in postmenopausal women.

Background: Epidemiological studies aimed at correlating coronary heart disease (CHD) with serum ferritin levels have thus far yielded inconsistent results. We hypothesized that a labile iron component associated with non-transferrin-bound iron (NTBI) that appears in individuals with overt or cryptic iron overload might be more suitable for establishing correlations with CHD.

Methods And Results: We investigated the relation of NTBI, serum iron, transferrin saturation, and serum ferritin with risk of CHD and acute myocardial infarction (AMI).

Results of an international round robin for the quantification of serum non-transferrin-bound iron: Need for defining standardization and a clinically relevant isoform.

Non-transferrin-bound iron (NTBI) appears in the circulation of patients with iron overload. Various methods to measure NTBI were comparatively assessed as part of an international interlaboratory study. Six laboratories participated in the study, using methods based on iron mobilization and detection with iron chelators or on reactivity with bleomycin.

LPI-labile plasma iron in iron overload.

Labile plasma iron (LPI) represents a component of non-transferrin-bound iron (NTBI) that is both redox-active and chelatable, capable of permeating into organs and inducing tissue iron overload. It appears in various types of hemosiderosis (transfusional and non-transfusional) and in other iron-overload conditions. Sustained levels of LPI could over time compromise organ (e.

Redox active plasma iron in C282Y/C282Y hemochromatosis.

Labile plasma iron (LPI) represents the redox active component of non-transferrin-bound iron (NTBI). Its presence in thalassemic patients has been recently reported. The aim of the present study was to quantify LPI in HFE genetic hemochromatosis (GH) and to characterize the mechanisms accounting for its appearance.

Labile plasma iron (LPI) as an indicator of chelatable plasma redox activity in iron-overloaded beta-thalassemia/HbE patients treated with an oral chelator.

Persistent levels of plasma nontransferrin bound iron (NTBI) have been associated with tissue iron overload and toxicity. We characterized NTBI's susceptibility to deferoxamine (directly chelatable iron [DCI]) and redox activity (labile plasma iron [LPI]) during the course of long-term, continuous L1 (deferiprone) treatment of patients with hemoglobin E disease and beta-thalassemia (n = 17). In 97% of serum samples (n = 267), the LPI levels were more than 0.

Non-transferrin-bound serum iron (NTBI) in megaloblastic anemia: effect of vitamin B(12) treatment.

Introduction: The abnormalities in iron metabolism associated with megaloblastic anemia are rapidly reversed by B(12) therapy in pernicious anemia (PA). Although non-tranferrin-bound plasma iron (NTBI) was previously shown to be associated with severe iron overload, its origin is unknown.

Methods And Results: Four patients with PA were studied before and after B(12) treatment.