Measurement of total iron in Helicobacter pylori-infected gastric epithelial cells

Flores, Sebastián; Day, Andrew S.; Keenan, Jacqueline I.

doi:10.1007/s10534-014-9810-z

Cited by 17 publications

(9 citation statements)

References 31 publications

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“…The iron concentration of the resultant solution was measured from the absorbance of the Fe 2+ - ferrozine complex at 564 nm (ε 564 = 27.9 mM −1 cm −1 ) using a Varian-Cary 50 Bio UV-vis spectrophotometer, normalized by cell population and reported as Fe atoms/CFU. The colorimetric determination of total intracellular iron using ferrozine offers a sensitive, accurate and low cost analytical technique, 33,34 which has been shown to produce results similar to those obtained by atomic absorption spectrometry. 35 …”

Section: Methodsmentioning

confidence: 95%

Inhibiting the BfrB:Bfd interaction in Pseudomonas aeruginosa causes irreversible iron accumulation in bacterioferritin and iron deficiency in the bacterial cytosol

et al. 2017

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Iron is an essential nutrient for bacteria but the reactivity of Fe2+ and the insolubility of Fe3+ present significant challenges to bacterial cells. Iron storage proteins contribute to ameliorating these challenges by oxidizing Fe2+ using O2 and H2O2 as electron acceptors, and by compartmentalizing Fe3+. Two types of iron-storage proteins coexist in bacteria, the ferritins (Ftn) and the heme-containing bacterioferritins (Bfr), but the reasons for their coexistence are largely unknown. P. aeruginosa cells harbor two iron storage proteins (FtnA and BfrB), but nothing is known about their relative contributions to iron homeostasis. Prior studies in vitro have shown that iron mobilization from BfrB requires specific interactions with a ferredoxin (Bfd), but the relevance of the BfrB:Bfd interaction to iron homeostasis in P. aeruginosa is unknown. In this work we explore the repercussions of (i) deleting the bfrB gene, and (ii) perturbing the BfrB:Bfd interaction in P. aeruginosa cells by either deleting the bfd gene or by replacing the wild type bfrB gene with a L68A/E81A double mutant allele in the P. aeruginosa chromosome. The effects of the mutations were evaluated by following the accumulation of iron in BfrB, analyzing levels of free and total intracellular iron, and by characterizing the ensuing iron homeostasis dysregulation phenotypes. The results reveal that P. aeruginosa accumulate iron mainly in BfrB, and that the nutrient does not accumulate in FtnA to detectable levels, even after deletion of the bfrB gene. Perturbing the BfrB:Bfd interaction causes irreversible flow of iron into BfrB, which leads to the accumulation of unusable intracellular iron while severely depleting the levels of free intracellular iron, which drives the cells to an acute iron starvation response despite harboring “normal” levels of total intracellular iron. These results are discussed in the context of a dynamic equilibrium between free cytosolic Fe2+ and Fe3+ compartmentalized in BfrB, which functions as a buffer to oppose rapid changes of free cytosolic iron. Finally, we also show that P. aeruginosa cells utilize iron stored in BfrB for growth in iron-limiting conditions, and that the utilization of BfrB-iron requires a functional BfrB:Bfd interaction.

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Section: Methodsmentioning

confidence: 95%

Inhibiting the BfrB:Bfd interaction in Pseudomonas aeruginosa causes irreversible iron accumulation in bacterioferritin and iron deficiency in the bacterial cytosol

et al. 2017

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“…H . pylori infection increases the amount of iron in AGS cells, perturbing intracellular iron homeostasis [ 17 ]. In vivo studies have also shown that gastric Helicobacter infection alters iron status in mouse and gerbil models [ 18 – 21 ].…”

Section: Introductionmentioning

confidence: 99%

Helicobacter pylori Infection Induces Anemia, Depletes Serum Iron Storage, and Alters Local Iron-Related and Adult Brain Gene Expression in Male INS-GAS Mice

Burns

Muthupalani

et al. 2015

PLoS ONE

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Iron deficiency anemia (IDA) affects > 500 million people worldwide, and is linked to impaired cognitive development and function in children. Helicobacter pylori, a class 1 carcinogen, infects about half of the world’s population, thus creating a high likelihood of overlapping risk. This study determined the effect of H. pylori infection on iron homeostasis in INS-GAS mice. Two replicates of INS-GAS/FVB male mice (n = 9-12/group) were dosed with H. pylori (Hp) strain SS1 or sham dosed at 6–9 weeks of age, and were necropsied at 27–29 weeks of age. Hematologic and serum iron parameters were evaluated, as was gene expression in gastric and brain tissues. Serum ferritin was lower in Hp SS1-infected mice than uninfected mice (p < 0.0001). Infected mice had a lower red blood cell count (p<0.0001), hematocrit (p < 0.001), and hemoglobin concentration (p <0.0001) than uninfected mice. Relative expression of gastric hepcidin antimicrobial peptide (Hamp) was downregulated in mice infected with Hp SS1 compared to sham-dosed controls (p<0.001). Expression of bone morphogenic protein 4 (Bmp4), a growth factor upstream of hepcidin, was downregulated in gastric tissue of Hp SS1-infected mice (p<0.001). Hp SS1-infected mice had downregulated brain expression of tyrosine hydroxylase (Th) (p = 0.02). Expression of iron-responsive genes involved in myelination (myelin basic protein (Mbp) and proteolipid protein 2 (Plp2)) was downregulated in infected mice (p = 0.001 and p = 0.02). Expression of synaptic plasticity markers (brain derived neurotrophic factor 3 (Bdnf3), Psd95 (a membrane associated guanylate kinase), and insulin-like growth factor 1 (Igf1)) was also downregulated in Hp SS1-infected mice (p = 0.09, p = 0.04, p = 0.02 respectively). Infection of male INS-GAS mice with Hp SS1, without concurrent dietary iron deficiency, depleted serum ferritin, deregulated gastric and hepatic expression of iron regulatory genes, and altered iron-dependent neural processes. The use of Hp SS1-infected INS-GAS mice will be an appropriate animal model for further study of the effects of concurrent H. pylori infection and anemia on iron homeostasis and adult iron-dependent brain gene expression.

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“…Our recent observation of increased total iron in H . pylori -infected AGS gastric epithelial cells that is independent of the extracellular iron concentration supports this hypothesis [ 9 ].…”

Section: Introductionmentioning

confidence: 79%

“…An infection-associated increase in total iron levels seen when H . pylori are added to cultured gastric AGS cells [ 9 ], which was not a result of bacterial association with the cells ( S2 Fig ), suggested that H . pylori may perturb iron uptake and/or storage mechanisms involved in maintaining cellular iron homeostasis.…”

Section: Resultsmentioning

confidence: 99%

Helicobacter pylori infection perturbs iron homeostasis in gastric epithelial cells

et al. 2017

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The iron deficiency anaemia that often accompanies infection with Helicobacter pylori may reflect increased uptake of iron into gastric epithelial cells. Here we show an infection-associated increase in total intracellular iron levels was associated with the redistribution of the transferrin receptor from the cell cytosol to the cell surface, and with increased levels of ferritin, an intracellular iron storage protein that corresponded with a significant increase in lysosomal stores of labile iron. In contrast, the pool of cytosolic labile iron was significantly decreased in infected cells. These changes in intracellular iron distribution were associated with the uptake and trafficking of H. pylori through the cells, and enhanced in strains capable of expressing the cagA virulence gene. We speculate that degradation of lysosomal ferritin may facilitate H. pylori pathogenesis, in addition to contributing to bacterial persistence in the human stomach.

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Measurement of total iron in Helicobacter pylori-infected gastric epithelial cells

Cited by 17 publications

References 31 publications

Inhibiting the BfrB:Bfd interaction in Pseudomonas aeruginosa causes irreversible iron accumulation in bacterioferritin and iron deficiency in the bacterial cytosol

Inhibiting the BfrB:Bfd interaction in Pseudomonas aeruginosa causes irreversible iron accumulation in bacterioferritin and iron deficiency in the bacterial cytosol

Helicobacter pylori Infection Induces Anemia, Depletes Serum Iron Storage, and Alters Local Iron-Related and Adult Brain Gene Expression in Male INS-GAS Mice

Helicobacter pylori infection perturbs iron homeostasis in gastric epithelial cells

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