A study of intracellular iron metabolism using pyridoxal isonicotinoyl hydrazone and other synthetic chelating agents

Ponka, P; Borová, J; Neuwirt, J; Fuchs, Ota; Nečas, E

doi:10.1016/0304-4165(79)90100-4

Cited by 181 publications

(72 citation statements)

References 29 publications

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“…The only supportive evidence for the presence of a LIP are from in vitro experiments showing that membrane permeable chelators are able to bind nonheme iron from cultured cells, including reticulocytes. However, the intracellular source of this Fe is, at best, vaguely identifiable in these studies 17,28,43 ; it is just as likely that chelators strip Fe from organelles or membrane components as from the cytoplasm. Hence, the so-called LIP has been defined by the amount of iron that the experimentor's chelator was able to bind.…”

Section: Discussionmentioning

confidence: 92%

“…59 Fe 2 -Tf and 59 Fe-salicylaldehyde isonicotinoyl hydrazone ( 59 Fe-SIH 2 ) were made from 59 Fe-chloride (Perkin Elmer, Woodbridge, ON) and labeling apo-Tf, as previously described. [27][28][29] Radioactivity measurements were performed using a Cobra II gamma counter (Packard Instruments, Meriden, CT). Confluent dishes of Huh7 cells (hepatocytelike cell line) were generously provided by Dr Kostas Pantopoulos (McGill University, Montréal, Canada).…”

Section: Methodsmentioning

confidence: 99%

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Direct interorganellar transfer of iron from endosome to mitochondrion

Sheftel

Zhang

Brown

et al. 2007

Blood

242

180

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Iron is a transition metal whose physicochemical properties make it the focus of vital biologic processes in virtually all living organisms. Among numerous roles, iron is essential for oxygen transport, cellular respiration, and DNA synthesis. Paradoxically, the same characteristics that biochemistry exploits make iron a potentially lethal substance. In the presence of oxygen, ferrous iron (Fe 2؉ ) will catalyze the production of toxic hydroxyl radicals from hydrogen peroxide. In addition, Fe 3؉ is virtually insoluble at physiologic pH. To protect tissues from deleterious effects of Fe, mammalian physiology has evolved specialized mechanisms for extracellular, intercellular, and intracellular iron handling. Here we show that developing erythroid cells, which are taking up vast amounts of Fe, deliver the metal directly from transferrin-containing endosomes to mitochondria (the site of heme biosynthesis), bypassing the oxygen-rich cytosol. Besides describing a new means of intracellular transport, our finding is important for developing therapies for patients with iron loading disorders. IntroductionAlthough its requirement for life in almost all known organisms has been recognized for decades, some of the most fundamental cell biologic processes of iron (Fe) still elude modern science. Mammalian physiology demands a constant source of bioavailible Fe, which is a functional component of hemoproteins, iron-sulfur cluster containing proteins, and other iron proteins. However, in its reduced form (Fe 2ϩ ), iron catalyzes the production of toxic hydroxyl radicals through Fenton chemistry, while the ferric version (Fe 3ϩ ) is virtually insoluble at physiologic pH. [1][2][3] Nevertheless, the adult human body contains approximately 4 g of Fe, more than 80% of which is in hemoglobin (Hb). 4 Under normal conditions, around 2 million red blood cells (RBCs) are produced per second. Hence, erythropoiesis requires approximately 25 mg of iron, daily, all of which is delivered via transferrin (Tf). The plasma contains approximately 3 M diferric Tf, the Fe of which is concentrated in maturing erythroid tissue to the equivalent of 20 mM iron, in the form of Hb. This exceptionally rapid utilization of the potentially toxic metal requires stringent regulation mechanisms that permit efficient production of hemoglobin, while protecting developing red blood cells and other tissues from iron's harmful properties. 5 Virtually every tissue acquires its iron by receptor mediated endocytosis of Tf (for review, see Richardson and Ponka 6 and Hentze et al 7 ): Diferric Tf binds to its cognate receptor on the cell surface; this binding is succeeded by internalization of the receptorligand complex. The release of Fe from Tf is achieved within the endosome by a lowering of the vesicular pH through the activity of the v-ATPase proton pump. After its liberation from Tf in the acidified endosome, Fe must be reduced (possibly by the recently identified Steap3 8 ) before it is transported across the vesicular membrane by the divalent metal transp...

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Section: Discussionmentioning

confidence: 92%

Section: Methodsmentioning

confidence: 99%

Direct interorganellar transfer of iron from endosome to mitochondrion

Sheftel

Zhang

Brown

et al. 2007

Blood

242

180

View full text Add to dashboard Cite

show abstract

“…Salicylaldehyde isonicotinoyl hydrazone was synthesized by us as described previously (19). 59 Fe-hemin, generated by incubating reticulocytes, was harvested from phenylhydrazinetreated mice (20) with 3 M 59 Fe 2 -transferrin (labeled as reported previously (21)) overnight.…”

Section: Methodsmentioning

confidence: 99%

Non-heme Induction of Heme Oxygenase-1 Does Not Alter Cellular Iron Metabolism

Sheftel

Kim

Ponka

2007

Journal of Biological Chemistry

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The catabolism of heme is carried out by members of the heme oxygenase (HO) family. The products of heme catabolism by HO-1 are ferrous iron, biliverdin (subsequently converted to bilirubin), and carbon monoxide. In addition to its function in the recycling of hemoglobin iron, this microsomal enzyme has been shown to protect cells in various stress models. Implicit in the reports of HO-1 cytoprotection to date are its effects on the cellular handling of heme/iron. However, the limited amount of uncommitted heme in non-erythroid cells brings to question the source of substrate for this enzyme in non-hemolytic circumstances. In the present study, HO-1 was induced by either sodium arsenite (reactive oxygen species producer) or hemin or overexpressed in the murine macrophage-like cell line, RAW 264.7. Both of the inducers elicited an increase in active HO-1; however, only hemin exposure caused an increase in the synthesis rate of the iron storage protein, ferritin. This effect of hemin was the direct result of the liberation of iron from heme by HO. Cells stably overexpressing HO-1, although protected from oxidative stress, did not display elevated basal ferritin synthesis. However, these cells did exhibit an increase in ferritin synthesis, compared with untransfected controls, in response to hemin treatment, suggesting that heme levels, and not HO-1, limit cellular heme catabolism. Our results suggest that the protection of cells from oxidative insult afforded by HO-1 is not due to the catabolism of significant amounts of cellular heme as thought previously.In the average adult, at equilibrium ϳ2 million red blood cells are being turned over every second. In a day, this process requires about 25 mg of iron (Fe) to be recycled from the hemoglobin of effete erythrocytes. Heme oxygenase 1 (HO-1) 2 is the enzyme responsible for catabolizing the heme from senescent red blood cells to release Fe, as well as equimolar amounts of carbon monoxide (CO) and biliverdin, for its eventual reuse in erythropoiesis. Another noninducible isoform of heme oxygenase with heme catabolic properties similar to HO-1, HO-2 is present in substantial levels primarily in the central nervous system and testes (1, 2). Although the heme-degrading function of HO has been known since the 1960s (3, 4), only recently has it been shown that heme oxygenases may be involved in cytoprotection against cellular stresses (for review see Camara and Soares (5), Abraham and Kappas (6), Otterbein et al. (7), and Ryter et al. (8)). This newer discovery of the potential of these enzymes as a tissue defense mechanism has spawned a flurry of studies by numerous groups who have shown by several different approaches that induction or overexpression of heme oxygenase 1 can protect tissues from various insults, including oxidative stress and immune system attack.The precise mechanism by which HO-1 mediates its protective function remains controversial. Breakdown of heme being the only known reaction catalyzed by the enzyme, extensive research has alleged that one or mo...

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“…Condition 4 is FBS-free methylcellulose medium (M3236, MethoCult) supplemented with 10% fetal platelet derived serum (Animal Technologies, Tyler, TX), 5% protein-free hybridoma medium (PFHM-II, Invitrogen), 3 units/ml Epo, 10 ng/ml murine IL3, 10 ng/ml murine IL6, 50 ng/ml murine SCF, 3 ng/ml murine granulocyte-macrophage colony-stimulating factor (PeproTech, Rocky Hill, NJ), as described previously (16), and 100 l of a 1 mM solution of salicylaldehyde isonicotinoyl hydrazone saturated with iron (Fe-SIH) (17), an iron chelate that delivers iron for heme synthesis without involving the TfR/ DMT1 pathway (17). 1 mM Fe-SIH was prepared as follows: 4.7 mg of synthesized SIH (18) was dissolved in 150 l of 1 N NaOH and then diluted with 14.5 ml of PBS; the SIH solution was then saturated with iron by adding 7 ml of 5 mM ferric citrate solution. Colony-forming units-erythroid (CFU-E) were counted at day 2 or 3; burst-forming units-erythroid (BFU-E) were counted at day 7 or 8, and mast cell and granulocyte/macrophage colonies were counted at days 7-10 and multipotential progenitor colonies (CFU-Mix) at days 10 -12 of culture.…”

Section: Mice and Genotyping-hif1␣mentioning

confidence: 99%