1.Iron chelator daphnetin against Pneumocystis carinii in vitro.
Bin YE ; Yu-Qiang ZHENG ; Wei-Hua WU ; Jing ZHANG
Chinese Medical Journal 2004;117(11):1704-1708
BACKGROUNDAlthough there are several drugs and drug combinations for the treatment of Pneumocystis carinii (P. carinii) pneumonia, all drugs have the toxicity as well as low efficacy. Iron chelators have been proposed as a source of new drugs for combating these infections. We hypothesized that iron chelators would suppress the growth of P. carinii by deprivation of the nutritional iron required for growth. In this study, a short-term axenic culture system of P. carinii was established. Daphnetin (7,8-dihydroxycoumarin), a known iron chelator, was demonstrated to exhibit in vitro activity against P. carinii in this system.
METHODSP. carinii organisms were obtained from the lungs of immunosuppressed rats. The culture system consisted of Iscove Dulbecco Eagle's Minimum Essential Medium (IMDM), supplemented with S-adenosyl-L-methionine, N-acetylglucosamine, putrescine, L-cysteine, L-glutamine, 2-mercaptoethanol, and fetal bovine serum, and was maintained at 37 degrees C, in 5% CO(2), 95% O(2), at the optimal pH of 8.0. The culture system was used to assess the effect of daphnetin on the proliferation of P. carinii organisms. The ultrastructures of the treated organisms were observed by transmission electron microscopy.
RESULTSThe number of cysts and trophozoites increased 8- to 9-fold and 11- to 12-fold, respectively, after 10 days of culture. Daphnetin was found to suppress the growth of P. carinii in a dose-dependent manner at concentrations between 1 micromol/L and 20 micromol/L. The inhibitory activity was suppressed by the chelation of daphnetin with ferrous sulfate in a 2:1 molar ratio, but it was not suppressed by mixing the culture medium with magnesium sulfate. Reduction of P. carinii numbers after treatment with daphnetin correlated with morphological changes in the organisms, as determined by transmission electron microscopy.
CONCLUSIONSDaphnetin can suppress the growth of P. carinii in vitro. The efficacy of daphnetin in suppressing the the growth of P. carinii in vitro is related to its ability to chelate iron.
Iron ; physiology ; Iron Chelating Agents ; pharmacology ; Microscopy, Electron ; Pneumocystis carinii ; drug effects ; growth & development ; ultrastructure ; Umbelliferones ; pharmacology
2.Deferoxamine induces apoptosis of K562 cells.
Guo-Cun JIA ; You-Cai TANG ; Feng-Yi LI ; Qing-Kui LIAO
Chinese Journal of Contemporary Pediatrics 2011;13(8):674-676
OBJECTIVETo study the molecular mechanism of apoptosis of leukemic cells (K562 cells) induced by iron chelating agent deferoxamine (DFO).
METHODSThe exponentially growing K562 cells were used (1×10(6)/mL) in this study. The K562 cells were treated with different concentrations of DFO (10, 50 and 100 mmol/L), DFO+FeCl3 (10 μmol/L each) or normal saline (blank control). The cellular labile iron pool was measured with a fluorimetric assay using the metalsensitive probe calcein-AM. The viable count and cell viability were determined by typanblue assay. Cell apoptosis was determined by morphological study and flow cytometry assay. Caspase-3 activity in K562 cells was detected by colorimetry.
RESULTSAfter DFO treatment, the cellular labile iron pool and the viability of K562 cells were reduced and the cell apoptosis increased in a time- and dose-dependent manner compared with the blank control group. The apoptosis rate of K562 cells in the DFO+FeCl3 treatment group was not significantly different from that in the blank control group. The caspase-3 activity in K562 cells increased significantly 24 hrs after 50 and 100 μmmol DFO treatment when compared with the blank control group (P<0.01). There was a negative correlation between cellular labile iron pool and caspase-3 activity of K562 cells (r=-0.894, P<0.05).
CONCLUSIONSDFO induces apoptosis of leukemic cells possibly through decreasing cellular labile iron pool and increasing caspase-3 activity of the cells.
Apoptosis ; drug effects ; Caspase 3 ; metabolism ; Deferoxamine ; pharmacology ; Flow Cytometry ; Humans ; Iron Chelating Agents ; pharmacology ; K562 Cells
3.Effect of Iron-Chelator Deferiprone on the In Vitro Growth of Staphylococci.
Choon Mee KIM ; Sung Heui SHIN
Journal of Korean Medical Science 2009;24(2):289-295
The standard iron-chelator deferoxamine is known to prevent the growth of coagulase-negative staphylococci (CoNS) which are major pathogens in iron-overloaded patients. However, we found that deferoxamine rather promotes the growth of coagulase-positive Staphylococcus aureus. Accordingly, we tested whether deferiprone, a new clinically-available iron-chelator, can prevent the growth of S. aureus strains as well as CoNS. Deferiprone did not at least promote the growth of all S. aureus strains (n=26) and CoNS (n=27) at relatively low doses; moreover, it could significantly inhibit the growth of all staphylococci on non-transferrin-bound-iron and the growth of all CoNS on transferrin-bound iron at relatively high doses. At the same doses, it did not at least promote the growth of all S. aureus strains on transferrin-bound-iron. These findings indicate that deferiprone can be useful to prevent staphylococcal infections, as well as to improve iron overload, in iron-overloaded patients.
Deferoxamine/pharmacology
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Humans
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Iron/metabolism
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Iron Chelating Agents/*pharmacology
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Iron Overload/metabolism
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Microbial Sensitivity Tests
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Pyridones/*pharmacology
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Staphylococcus/*drug effects/growth & development
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Staphylococcus aureus/drug effects/growth & development
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Transferrin/metabolism
4.Detection of the labile iron pool in leukemia cells and its significance.
Guo-Cun JIA ; Ju GAO ; Qing-Kui LIAO ; Feng-Yi LI ; Li-Xing YUAN ; Bin HE
Journal of Experimental Hematology 2006;14(3):468-470
To explore a rapid and easy method to detect labile iron of pool (LIP) in cells, HL-60 and K562 cells were cultured at a concentration 1 x 10(6)/ml in RPMI 1640 containing 10% heat-inactivated fetal bovine serum. The iron deprivation was induced by adding desferrioxamine (DFO) 10 - 100 micromol/L for 0 - 48 hours. The intracellular LIP was measured by probe calcein-AM. Calcein fluorescence was monitored in 1420 multilabel counter. The results indicated that when HL-60 and K562 cells were incubated with different concentrations of DFO, the calcein fluorescence intensity was higher than that of control group at 12, 24 and 48 hours (P < 0.05). Fluorescence value of representing LIP in DFO groups was lower than that in the control group. In conclusion, DFO can decrease LIP in leukemia cells. The approach used in this study may provide a simple and reliable method for detection of intracellular iron homeostasis.
Cation Transport Proteins
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antagonists & inhibitors
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biosynthesis
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metabolism
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Deferoxamine
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pharmacology
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Fluoresceins
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Fluorescent Dyes
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HL-60 Cells
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Humans
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Iron
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metabolism
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Iron Chelating Agents
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analysis
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metabolism
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Iron-Regulatory Proteins
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metabolism
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K562 Cells
5.Use of deferasirox, an iron chelator, to overcome imatinib resistance of chronic myeloid leukemia cells.
Dae Sik KIM ; Yoo Jin NA ; Myoung Hee KANG ; Soo Young YOON ; Chul Won CHOI
The Korean Journal of Internal Medicine 2016;31(2):357-366
BACKGROUND/AIMS: The treatment of chronic myeloid leukemia (CML) has achieved impressive success since the development of the Bcr-Abl tyrosine kinase inhibitor, imatinib mesylate. Nevertheless, resistance to imatinib has been observed, and a substantial number of patients need alternative treatment strategies. METHODS: We have evaluated the effects of deferasirox, an orally active iron chelator, and imatinib on K562 and KU812 human CML cell lines. Imatinib-resistant CML cell lines were created by exposing cells to gradually increasing concentrations of imatinib. RESULTS: Co-treatment of cells with deferasirox and imatinib induced a synergistic dose-dependent inhibition of proliferation of both CML cell lines. Cell cycle analysis showed an accumulation of cells in the subG1 phase. Western blot analysis of apoptotic proteins showed that co-treatment with deferasirox and imatinib induced an increased expression of apoptotic proteins. These tendencies were clearly identified in imatinib-resistant CML cell lines. The results also showed that co-treatment with deferasirox and imatinib reduced the expression of BcrAbl, phosphorylated Bcr-Abl, nuclear factor-kappaB (NF-kappaB) and beta-catenin. CONCLUSIONS: We observed synergistic effects of deferasirox and imatinib on both imatinib-resistant and imatinib-sensitive cell lines. These effects were due to induction of apoptosis and cell cycle arrest by down-regulated expression of NF-kappaB and beta-catenin levels. Based on these results, we suggest that a combination treatment of deferasirox and imatinib could be considered as an alternative treatment option for imatinib-resistant CML.
Antineoplastic Agents/*pharmacology
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Apoptosis/drug effects
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Apoptosis Regulatory Proteins/metabolism
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Benzoates/*pharmacology
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Cell Proliferation/drug effects
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Dose-Response Relationship, Drug
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Drug Resistance, Neoplasm/*drug effects
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G1 Phase Cell Cycle Checkpoints/drug effects
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Humans
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Imatinib Mesylate/*pharmacology
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Iron Chelating Agents/*pharmacology
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K562 Cells
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Leukemia, Myelogenous, Chronic, BCR-ABL Positive/*drug therapy/metabolism
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Protein Kinase Inhibitors/*pharmacology
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Signal Transduction/drug effects
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Triazoles/*pharmacology
6.Bacterial Growth in Amniotic Fluid Is Dependent on the Iron-Availability and the Activity of Bacterial Iron-Uptake System.
Young Joon AHN ; Sang Kee PARK ; Jae Wook OH ; Hui Yu SUN ; Sung Heui SHIN
Journal of Korean Medical Science 2004;19(3):333-340
In the present study, the relationship among iron-availability, antibacterial activity, role of meconium as an iron source and the activity of bacterial iron-uptake system (IUS) for bacterial growth in amniotic fluid (AF) were investigated. Staphylococcus aureus ATCC 6538 and its streptonigrin-resistant (SR) mutant with defective IUS were used as the test strains. The growth of S. aureus in AF was stimulated dosedependently by addition of meconium. Bacterial growth stimulated by meconium was re-inhibited dose-dependently by addition of iron-chelator, dipyridyl and apotransferrin. Iron concentration was correlated with the meconium content in AF (r(2)= 0.989, p=0.001). High-affinity IUS of S. aureus was expressed only in AF but not in AF with meconium. The growth of SR strain was more retarded than that of the parental strain in the iron-deficient brain heart infusion (ID-BHI), clear AF and AF containing apotransferrin. The retarded growth of both strains in the ID-BHI and AF was recovered by addition of holotransferrin, hemoglobin and FeCl3. Taken together, the antibacterial activity of AF is closely related with low iron-availability. Bacterial growth in AF considerably depends on the activity of bacterial IUS. Meconium acts as one of the exogenous iron-sources and thus can stimulate bacterial growth in AF.
Amniotic Fluid/*microbiology
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Antibiotics, Antineoplastic/pharmacology
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Chelating Agents/pharmacology
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Dose-Response Relationship, Drug
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Female
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Ferric Compounds/pharmacology
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Human
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Iron/*metabolism
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Ligands
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Meconium/metabolism
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Mutation
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Pregnancy
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Pregnancy Trimester, Third
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Protein Binding
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Staphylococcus aureus/metabolism
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Streptonigrin/pharmacology
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Support, Non-U.S. Gov't
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Time Factors
7.Iron chelator inducesMIP-3alpha/CCL20 in human intestinal epithelial cells: implication for triggeringmucosal adaptive immunity.
Hyun Ju LEE ; Suck Chei CHOI ; Eun Young CHOI ; Moo Hyung LEE ; Geom Seog SEO ; Eun Cheol KIM ; Bong Joon YANG ; Myeung Su LEE ; Yong Il SHIN ; Kie In PARK ; Chang Duk JUN
Experimental & Molecular Medicine 2005;37(4):297-310
A previous report by this laboratory demonstrated that bacterial iron chelator (siderophore) triggers inflammatory signals, including the production of CXC chemokine IL-8, in human intestinal epithelial cells (IECs). Microarray-based gene expression profiling revealed that iron chelator also induces macrophage inflammatory protein 3 alpha (MIP-3alpha)/ CC chemokine-ligand 20 (CCL20). As CCL20 is chemotactic for the cells involved in host adaptive immunity, this suggests that iron chelator may stimulate IECs to have the capacity to link mucosal innate and adaptive immunity. The basal medium from iron chelator deferoxamine (DFO)-treated HT-29 monolayers was as chemotactic as recombinant human CCL20 at equivalent concentrations to attract CCR6+ cells. The increase of CCL20 protein secretion appeared to correspond to that of CCL20 mRNA levels, as determined by real-time quantitative RT-PCR. The efficacy of DFO at inducing CCL20 mRNA was also observed in human PBMCs and in THP-1 cells, but not in human umbilical vein endothelial cells. Interestingly, unlike other proinflammatory cytokines, such as TNF-alpha and IL-1beta, a time-dependent experiment revealed that DFO slowly induces CCL20, suggesting a novel mechanism of action. A pharmacologic study also revealed that multiple signaling pathways are differentially involved in CCL20 production by DFO, while some of those pathways are not involved in TNF-alpha-induced CCL20 production. Collectively, these results demonstrate that, in addition to some bacterial products known to induce host adaptive immune responses, direct chelation of host iron by infected bacteria may also contribute to the initiation of host adaptive immunity in the intestinal mucosa.
Calcium/metabolism
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Cell Movement/drug effects
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Chemokines, CC/genetics/*metabolism
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Deferoxamine/*pharmacology
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Egtazic Acid/analogs & derivatives/pharmacology
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HT29 Cells
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Humans
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Immunity, Mucosal/*drug effects
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Intestinal Mucosa/*drug effects/immunology/metabolism
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Iron Chelating Agents/*pharmacology
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Macrophage Inflammatory Proteins/genetics/*metabolism
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NF-kappa B/metabolism
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Phosphoprotein Phosphatase/physiology
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Protein Transport/drug effects
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Protein-Serine-Threonine Kinases/physiology
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RNA, Messenger/genetics/metabolism
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Receptors, Chemokine/metabolism
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Research Support, Non-U.S. Gov't