Timosaponin AIII (Tim-AIII), a steroid saponin, exhibits strong anticancer activity in a variety of cancers, especially breast cancer and liver cancer. However, the underlying mechanism of the effects of Tim-AIII-mediated anti-lung cancer effects remain obscure. In this study, we showed that Tim-AIII suppressed cell proliferation and migration, induced G2/M phase arrest and ultimately triggered cell death of non-small cell lung cancer (NSCLC) cell lines accompanied by the release of reactive oxygen species (ROS) and iron accumulation, malondialdehyde (MDA) production, and glutathione (GSH) depletion. Interestingly, we found that Tim-AIII-mediated cell death was reversed by ferroptosis inhibitor ferrostatin-1 (Fer-1). Meanwhile, the heat shock protein 90 (HSP90) was predicted and verified as the direct binding target of Tim-AIII by SwissTargetPrediction (STP) and surface plasmon resonance (SPR) assay. Further study showed that Tim-AIII promoted HSP90 expression and Tim-AIII induced cell death was blocked by the HSP90 inhibitor tanespimycin, indicating that HSP90 was the main target of Tim-AIII to further trigger intracellular events. Mechanical analysis revealed that the Tim-AIII-HSP90 complex further targeted and degraded glutathione peroxidase 4 (GPX4), and promoted the ubiquitination of GPX4, as shown by an immunoprecipitation, degradation and in vitro ubiquitination assay. In addition, Tim-AIII inhibited cell proliferation, induced cell death, led to ROS and iron accumulation, MDA production, GSH depletion, as well as GPX4 ubiquitination and degradation, were markedly abrogated when HSP90 was knockdown by HSP90-shRNA transfection. Importantly, Tim-AIII also showed a strong capacity of preventing tumor growth by promoting ferroptosis in a subcutaneous xenograft tumor model, whether C57BL/6J or BALB/c-nu/nu nude mice. Together, HSP90 was identified as a new target of Tim-AIII. Tim-AIII, by binding and forming a complex with HSP90, further targeted and degraded GPX4, ultimately induced ferroptosis in NSCLC. These findings provided solid evidence that Tim-AIII can serve as a potential candidate for NSCLC treatment.
Palm kernel cake protein was hydrolyzed with different proteases namely papain, bromelain, subtilisin, flavourzyme, trypsin, chymotrypsin, and pepsin to generate different protein hydrolysates. Peptide content and iron-chelating activity of each hydrolysate were evaluated using O-phthaldialdehyde-based spectrophotometric method and ferrozine-based colorimetric assay, respectively. The results revealed a positive correlation between peptide contents and iron-chelating activities of the protein hydrolysates. Protein hydrolysate generated by papain exhibited the highest peptide content of 10.5 mM and highest iron-chelating activity of 64.8% compared with the other hydrolysates. Profiling of the papain-generated hydrolysate by reverse phase high performance liquid chromatography fractionation indicated a direct association between peptide content and iron-chelating activity in most of the fractions. Further fractionation using isoelectric focusing also revealed that protein hydrolysate with basic and neutral isoelectric point (pI) had the highest iron-chelating activity, although a few fractions in the acidic range also exhibited good metal chelating potential. After identification and synthesis of papain-generated peptides, GGIF and YLLLK showed among the highest iron-chelating activities of 56% and 53%, whereas their IC50 were 1.4 and 0.2 μM, respectively.
The Sfp-type 4'-phosphopantetheinyl transferase Ppt1 is required for activation of nonribosomal peptide synthetases, including α-aminoadipate reductase (AAR) for lysine biosynthesis and polyketide synthases, enzymes that biosynthesize peptide and polyketide secondary metabolites, respectively. Deletion of the PPT1 gene, from the maize pathogen Cochliobolus heterostrophus and the rice pathogen Cochliobolus miyabeanus, yielded strains that were significantly reduced in virulence to their hosts. In addition, ppt1 mutants of C. heterostrophus race T and Cochliobolus victoriae were unable to biosynthesize the host-selective toxins (HST) T-toxin and victorin, respectively, as judged by bioassays. Interestingly, ppt1 mutants of C. miyabeanus were shown to produce tenfold higher levels of the sesterterpene-type non-HST ophiobolin A, as compared with the wild-type strain. The ppt1 strains of all species were also reduced in tolerance to oxidative stress and iron depletion; both phenotypes are associated with inability to produce extracellular siderophores biosynthesized by the nonribosomal peptide synthetase Nps6. Colony surfaces were hydrophilic, a trait previously associated with absence of C. heterostrophus Nps4. Mutants were decreased in asexual sporulation and C. heterostrophus strains were female-sterile in sexual crosses; the latter phenotype was observed previously with mutants lacking Nps2, which produces an intracellular siderophore. As expected, mutants were albino, since they cannot produce the polyketide melanin and were auxotrophic for lysine because they lack an AAR.
Conventional techniques to remove Fe impurities in kaolin typically involve high environmental impact and cost. Alternative methods have been focused on the use of bioleaching where Fe in kaolin is reduced with microorganisms. Early results established a noticeable effect of the bacteria on the redox state of Fe, but knowledge gaps persist such as details on the bacterial-kaolin interactions during attachment of bacteria onto kaolin surface, the metabolites produced by bacteria, and changes in Fe(II)/Fe(III) ion equilibria in solution. To bridge these gaps, this study was conducted to determine the detailed physicochemical changes in bacteria and kaolin during bioleaching through surface, structural, and chemical analysis. Bioleaching experiments were conducted for 10 days where each of the three Bacillus sp. was put in contact (at 9 × 108 CFU) with 20 g of kaolin powder using 200 mL of 10 g/L glucose solution. All samples treated with bacteria showed increasing trends in Fe(III) reduction up until day 6 or 8 followed by a slight decrease towards the end of the ten-day period. Examination of scanning electron microscope (SEM) images suggests that bacterial activity damaged the edges of kaolin particles during bioleaching. Ion chromatography (IC) results showed that during bioleaching, Bacillus sp. produced organic acids such as lactic acid, formic acid, malic acid, acetic acid, and succinic acid. EDS analysis of kaolin before and after bioleaching showed Fe removal efficiencies of up to 65.3%. Analyses of color properties of kaolin before and after bioleaching showed an improvement in whiteness index of up to 13.6%. KEY POINTS: • Dissolution of iron oxides by Bacillus species proven with phenanthroline analysis. • Organic acid type and concentration unique to species detected during bioleaching. • Whiteness index of kaolin is improved after bioleaching.
The concentrations of non-haem iron, ferritin and ferritin-iron were measured in the livers of 137 adults and children collected at necropsy. The concentrations of non-haem and ferritin iron were found to be 146.6 +/- 95.2 micrograms/g and 61.6 +/- 32.4 micrograms/g, respectively, in males and 108.0 +/- 61.7 micrograms/g and 60.6 +/- 26.4 micrograms/g, respectively, in females. The values for males in Singapore were lower than those reported in developed Western countries. No correlation was observed between storage iron and age, or ferritin concentration and age. Concentrations of non-haem iron and ferritin were similar for persons dying from accident and coronary heart disease. The non-haem iron concentration in Chinese (187.9 +/- 101.0 micrograms/g) was significantly greater than that in Indians (103.1 +/- 65.8 micrograms/g), while the ferritin concentration in Chinese (6.18 +/- 2.37 mg/g) was significantly greater than either Malays (3.81 +/- 1.8 mg/g) or Indians (3.52 +/- 1.6 mg/g). A significant positive correlation was observed between the non-haem iron and ferritin and also ferritin-iron in Chinese males (r values of 0.678 and 0.598, respectively) and Indian males (r values of 0.576 and 0.612, respectively). However, the correlation between these indices was not significant in the case of Malay males. In premenopausal women the non-haem iron correlated well with ferritin (r = 0.737) and ferritin iron (r = 0.826) while the correlation was lacking in postmenopausal women.
Heavy metals in cigarette tobacco such as iron may cause a serious damage on human health. Surveys showed that the accumulation of certain toxic heavy metals like cadmium, mercury, iron is very often due to the effect of smoking. This work involved 15 volunteers in two randomly divided groups having the habit of cigarette smoking over 15 cigarettes / day. Concentration level of iron in blood and urine before and after treatment using the herbal medicine, widely used in Europe, is analyzed. Determination of Iron concentration in blood and urine was calculated by Inductively Coupled Plasma-Atomic Emission Spectrometry (ICP-AES) according to the procedure DIN EN ISO 11885 ("E22" from April 1998). The analysis shows that the concentration of iron in blood and urine samples in both groups increased in some volunteers instead of decrease. The independent T-test shows that the mean of iron concentration in the group A and group B had no significant difference (p>0.05). The results suggested that the herbal medicine under test does not have significant influence on reduction of iron concentration levels.
This study investigates the twitching ability of 28 clinical and five environmental strains of S. maltophilia grown under iron-depleted condition through in-silico, phenotypic and proteomics approaches. Rapid Annotations using Subsystem Technology (RAST) analysis revealed the presence of 21 targets of type IV pilus shared across S. maltophilia strains K279a, R551-3, D457 and JV3. The macroscopic twitching assay showed that only clinical isolates produced a zone of twitching with a mean of 22.00 mm under normal and 25.00 mm under iron-depleted conditions. (p = 0.002). Environmental isolates did not show any significant twitching activity in both conditions tested. Isobaric Tags for Relative and Absolute Quantification (ITRAQ) analysis showed altered expression of twitching motility protein PilT (99.08-fold change), flagellar biosynthesis protein FliC (20.14-fold change), and fimbrial protein (0.70-fold change) in response to iron-depleted condition. Most of the strains that have the ability to twitch under the normal condition, exhibit enhanced twitching during iron limitation.
Management of Beta (β)-thalassaemia intermedia in contrast to β-thalassaemia major patients has no clear guidelines as to indicators of adequate transfusion. Regular blood transfusion suppresses bone marrow erythropoietic activity. Serum soluble transferrin receptor (sTfR) concentration is a marker for erythropoietic activity, with increased sTfR being associated with functional iron deficiency and increased erythropoietic activity. This study aimed to determine the use of sTfR as an indicator of adequate transfusion in adult β-thalassaemia intermedia patients. A cross-sectional study was conducted at Hospital Ampang, Malaysia, for six months. Patient group included six β-thalassaemia intermedia and 34 HbE-β-thalassaemia transfused patients. None of the patients were on regular monthly blood transfusions as in β-thalassaemia major. The control group comprised of 16 healthy subjects with normal haematological parameters. Haemoglobin (Hb) analysis, sTfR and ferritin assays were performed. Hb and HbA percentages (%) were found to be significantly lower in patients compared to the controls, while HbE%, HbF%, sTfR and ferritin were significantly higher in patients. An inverse relationship was found in the controls between HbF% with Hb (r = -0.515, p < 0.05) and HbA% (r = -0.534, p < 0.05). In patients, sTfR showed an inverse relationship with HbA% (r = -0.618, p = 0.000) and a positive correlation with HbE% (r = 0.418, p = 0.007) and HbF% (r = 0.469, p = 0.002). Multivariate analysis showed that HbA% (r = 2.875, p = 0.048), HbE% (r = 2.872, p = 0.020) and HbF% (r = 2.436, p = 0.013) best predicted sTfR independently in patients. Thus, sTfR is a useful marker for erythropoiesis. The elevated sTfR in these patients indicate that the transfusion regimen used was inadequate to suppress ineffective erythropoiesis. Hb levels may not be the best target for monitoring transfusion treatment in β-thalassaemia intermedia patients, but the use of sTfR is helpful in individualising transfusion regimens.
Iron is an essential micronutrient for most living beings since it participates as a redox active cofactor in many biological processes including cellular respiration, lipid biosynthesis, DNA replication and repair, and ribosome biogenesis and recycling. However, when present in excess, iron can participate in Fenton reactions and generate reactive oxygen species that damage cells at the level of proteins, lipids and nucleic acids. Organisms have developed different molecular strategies to protect themselves against the harmful effects of high concentrations of iron. In the case of fungi and plants, detoxification mainly occurs by importing cytosolic iron into the vacuole through the Ccc1/VIT1 iron transporter. New sequenced genomes and bioinformatic tools are facilitating the functional characterization, evolution and ecological relevance of metabolic pathways and homeostatic networks across the Tree of Life. Sequence analysis shows that Ccc1/VIT1 homologs are widely distributed among organisms with the exception of animals. The recent elucidation of the crystal structure of a Ccc1/VIT1 plant ortholog has enabled the identification of both conserved and species-specific motifs required for its metal transport mechanism. Moreover, recent studies in the yeast Saccharomyces cerevisiae have also revealed that multiple transcription factors including Yap5 and Msn2/Msn4 contribute to the expression of CCC1 in high-iron conditions. Interestingly, Malaysian S. cerevisiae strains express a partially functional Ccc1 protein that renders them sensitive to iron. Different regulatory mechanisms have been described for non-Saccharomycetaceae Ccc1 homologs. The characterization of Ccc1/VIT1 proteins is of high interest in the development of biofortified crops and the protection against microbial-derived diseases.
Iron is a crucial element required for the proper functioning of the body. For instance, hemoglobin is the vital component in the blood that delivers oxygen to various parts of the body. The heme protein present in hemoglobin comprises iron in the form of a ferrous state which regulates oxygen delivery. Excess iron in the body is stored as ferritin and would be utilized under iron-deficient conditions. Surprisingly, cancer cells as well as cancer stem cells have elevated ferritin levels suggesting that iron plays a vital role in protecting these cells. However, apart from the cytoprotective role iron also has the potential to induce cell death via ferroptosis which is a non-apoptotic cell death dependent on iron reserves. Apoptosis a caspase-dependent cell death mechanism is effective on cancer cells however little is known about its impact on cancer stem cell death. This paper focuses on the molecular characteristics of apoptosis and ferroptosis and the importance of switching to ferroptosis to target cancer stem cells death thereby preventing cancer relapse. To the best of our knowledge, this is the first review to demonstrate the importance of intracellular iron in regulating the switching of tumor cells and therapy resistant CSCs from apoptosis to ferroptosis.
Iron deficiency is common in the developing world; consequently, programmes of presumptive therapy and mass supplementation have been introduced in several countries. In this article Stephen Oppenheimer suggests caution, as recent evidence suggests that these practices may actually increase the likelihood of the subject developing patent malaria in endemic areas. This may be especially significant in infants, who are less likely to be immune to malaria, and in pregnant women, who are often routinely given iron supplements and in whom malaria may damage the foetus.
Terrestrial plants as potential phytoremediators for remediation of surface soil contaminated with toxic metals have gained attention in clean-up technologies. The potential of kenaf (Hibiscus cannabinus L.) to offer a cost-effective mechanism to remediate Fe and As from landfill leachate-contaminated soil was investigated. Pot experiment employing soil polluted with treatments of Jeram landfill leachate was conducted for 120 days. Plants were harvested after 8th, 12th, and 16th weeks of growth. Accumulation of Fe and As was assessed based on Bioconcentration Factor and Translocation Factor. Results showed sequestration of 0.06-0.58 mg As and 66.82-461.71 mg Fe per g plant dry weight in kenaf root, which implies that kenaf root can be an bioavailable sink for toxic metals. Insignificant amount of Fe and As was observed in the aerial plant parts (< 12% of total bioavailable metals). The ability of kenaf to tolerate these metals and avoid phytotoxicity could be attributed to the stabilization of the metals in the roots and hence reduction of toxic metal mobility (TF < 1). With the application of leachate, kenaf was also found to have higher biomass and subsequently recorded 11% higher bioaccumulation capacity, indicating its suitability for phytoextraction of leachate contaminated sites.
Scurrula ferruginea (Jack) Danser is one of the mistletoe species belonging to Loranthaceae family, which grows on the branches of many deciduous trees in tropical countries. This study evaluated the antioxidant activities of S. ferruginea extracts. The cytotoxic activity of the selected extracts, which showed potent antioxidant activities, and high phenolic and flavonoid contents, were investigated in human breast cancer cell line (MDA-MB-231) and non-cancer human skin fibroblast cells (HSF-1184). The activities and characteristics varied depending on the different parts of S. ferruginea, solvent polarity, and concentrations of extracts. The stem methanol extract showed the highest amount of both phenolic (273.51 ± 4.84 mg gallic acid/g extract) and flavonoid contents (163.41 ± 4.62 mg catechin/g extract) and strong DPPH• radical scavenging (IC50 = 27.81 μg/mL) and metal chelation activity (IC50 = 80.20 μg/mL). The stem aqueous extract showed the highest ABTS•+ scavenging ability. The stem methanol and aqueous extracts exhibited dose-dependent cytotoxic activity against MDA-MB-231 cells with IC50 of 19.27 and 50.35 μg/mL, respectively. Furthermore, the extracts inhibited the migration and colony formation of MDA-MB-231 cells in a concentration-dependent manner. Morphological observations revealed hallmark properties of apoptosis in treated cells. The methanol extract induced an increase in ROS generation and mitochondrial depolarization in MDA-MB-231 cells, suggesting its potent apoptotic activity. The present study demonstrated that the S. ferruginea methanol extract mediated MDA-MB-231 cell growth inhibition via induction of apoptosis which was confirmed by Western blot analysis. It may be a potential anticancer agent; however, its in vivo anticancer activity needs to be investigated.
Comparative genomic studies have reported widespread variation in levels of gene expression within and between species. Using these data to infer organism-level trait divergence has proven to be a key challenge in the field. We have used a wild Malaysian population of S. cerevisiae as a test bed in the search to predict and validate trait differences based on observations of regulatory variation. Malaysian yeast, when cultured in standard medium, activated regulatory programs that protect cells from the toxic effects of high iron. Malaysian yeast also showed a hyperactive regulatory response during culture in the presence of excess iron and had a unique growth defect in conditions of high iron. Molecular validation experiments pinpointed the iron metabolism factors AFT1, CCC1, and YAP5 as contributors to these molecular and cellular phenotypes; in genome-scale sequence analyses, a suite of iron toxicity response genes showed evidence for rapid protein evolution in Malaysian yeast. Our findings support a model in which iron metabolism has diverged in Malaysian yeast as a consequence of a change in selective pressure, with Malaysian alleles shifting the dynamic range of iron response to low-iron concentrations and weakening resistance to extreme iron toxicity. By dissecting the iron scarcity specialist behavior of Malaysian yeast, our work highlights the power of expression divergence as a signpost for biologically and evolutionarily relevant variation at the organismal level. Interpreting the phenotypic relevance of gene expression variation is one of the primary challenges of modern genomics.
Anaemia is a common condition in Malaysia, and is mostly due to iron deficiency. In many cases, allogeneic blood transfusion (ABT) is administered unnecessarily to treat anaemia. Patient blood management (PBM) is a concept whereby a patient becomes his or her "own blood bank", instead of receiving ABT. The concept encompasses three pillars namely optimising erythropoiesis, minimising blood loss and harnessing human physiological reserve. We present a safe and fruitful outcome of managing severe anaemia without utilising any ABT, made possible with the PBM approach including administration of intravenous iron.
Stenotrophomonas maltophilia is a multi-drug-resistant global opportunistic nosocomial pathogen, which possesses a huge number of virulence factors and antibiotics resistance characteristics. Iron has a crucial contribution toward growth and development, cell growth and proliferation, and pathogenicity. The bacterium found to acquire iron for its cellular process through the expression of two iron acquisition systems. Two distinct pathways for iron acquisition are encoded by the S. maltophilia genome-a siderophore-and heme-mediated iron uptake system. The entAFDBEC operon directs the production of the enterobactin siderophore of catecholate in nature, while heme uptake relies on hgbBC and potentially hmuRSTUV operon. Fur and sigma factors are regulators of S. maltophilia under iron-limited condition. Iron potentially act as a signal which plays an important role in biofilm formation, extracellular polymeric substances (EPS), extracellular enzymes production, oxidative stress response, diffusible signal factor (DSF) and siderophore production in S. maltophilia. This review summarizes the current knowledge of iron acquisition in S. maltophilia and the critical role of iron in relation to its pathogenicity.
Recently, microbial-based iron reduction has been considered as a viable alternative to typical chemical-based treatments. The iron reduction is an important process in kaolin refining, where iron-bearing impurities in kaolin clay affects the whiteness, refractory properties, and its commercial value. In recent years, Gram-negative bacteria has been in the center stage of iron reduction research, whereas little is known about the potential use of Gram-positive bacteria to refine kaolin clay. In this study, we investigated the ferric reducing capabilities of five microbes by manipulating the microbial growth conditions. Out of the five, we discovered that Bacillus cereus and Staphylococcus aureus outperformed the other microbes under nitrogen-rich media. Through the biochemical changes and the microbial behavior, we mapped the hypothetical pathway leading to the iron reduction cellular properties, and found that the iron reduction properties of these Gram-positive bacteria rely heavily on the media composition. The media composition results in increased basification of the media that is a prerequisite for the cellular reduction of ferric ions. Further, these changes impact the formation of biofilm, suggesting that the cellular interaction for the iron(III)oxide reduction is not solely reliant on the formation of biofilms. This article reveals the potential development of Gram-positive microbes in facilitating the microbial-based removal of metal contaminants from clays or ores. Further studies to elucidate the corresponding pathways would be crucial for the further development of the field.
Pasteurella multocida causes respiratory infectious diseases in a multitude of birds and mammals. A number of virulence-associated genes were reported across different strains of P. multocida, including those involved in the iron transport and metabolism. Comparative iron-associated genes of P. multocida among different animal hosts towards their interaction networks have not been fully revealed. Therefore, this study aimed to identify the iron-associated genes from core- and pan-genomes of fourteen P. multocida strains and to construct iron-associated protein interaction networks using genome-scale network analysis which might be associated with the virulence. Results showed that these fourteen strains had 1587 genes in the core-genome and 3400 genes constituting their pan-genome. Out of these, 2651 genes associated with iron transport and metabolism were selected to construct the protein interaction networks and 361 genes were incorporated into the iron-associated protein interaction network (iPIN) consisting of nine different iron-associated functional modules. After comparing with the virulence factor database (VFDB), 21 virulence-associated proteins were determined and 11 of these belonged to the heme biosynthesis module. From this study, the core heme biosynthesis module and the core outer membrane hemoglobin receptor HgbA were proposed as candidate targets to design novel antibiotics and vaccines for preventing pasteurellosis across the serotypes or animal hosts for enhanced precision agriculture to ensure sustainability in food security.
Although more than 100 genome sequences of Pasteurella multocida are available, comprehensive and complete genome sequence analysis is limited. This study describes the analysis of complete genome sequence and pathogenomics of P. multocida strain PMTB2.1. The genome of PMTB2.1 has 2176 genes with more than 40 coding sequences associated with iron regulation and 140 virulence genes including the complete tad locus. The tad locus includes several previously uncharacterized genes such as flp2, rcpC and tadV genes. A transposable phage resembling to Mu phages was identified in P. multocida that has not been identified in any other serotype yet. The multi-locus sequence typing analysis assigned the PMTB2.1 genome sequence as type ST101, while the comparative genome analysis showed that PMTB2.1 is closely related to other P. multocida strains with the genomic distance of less than 0.13. The expression profiling of iron regulating-genes of PMTB2.1 was characterized under iron-limited environment. Results showed significant changes in the expression profiles of iron-regulating genes (p < 0.05) whereas the highest expression of fecE gene (281 fold) at 30 min suggests utilization of the outer-membrane proteins system in iron acquisition at an early stage of growth. This study showed the phylogenomic relatedness of P. multocida and improved annotation of important genes and functional characterization of iron-regulating genes of importance to the bacterial growth.