Transferrin Binding Research Articles

The effect of age on intestinal iron uptake

Background: The pro-oxidant role of iron in neurodegenerative and chronic disease has led to concern over the danger of excessive levels posing a health risk (Kell, 2009). As the body has limited capacity to excrete iron, absorption acts as the main homeostatic mechanism (Anderson et al., 2009). Although iron stores increase with age (Liu et al., 2003), the effect of advancing age on iron regulation has not been extensively studied and age related misregulation has been implied (Levenson et al., 2004). The aim of this study was to investigate the effect of age on intestinal iron uptake in adults. Method: Thirty-five participants (29 females and six males subdivided into three age groups) provided fasted blood samples. A second sample was taken three hours following a standard breakfast (low in iron inhibitors and enhancers) and a 200 mg iron sulphate supplement as used by Hutchinson et al. (2004). Total plasma iron, total iron binding capacity (TIBC), transferrin saturation (TSAT) and non-transferrin bound iron (NTBI) were measured. Participants completed a 52 item food frequency questionnaire, adapted from the validated EPIC-Norfolk study to be iron specific. Data was statistically analysed using anova and Pearson regression. Approval was granted by the University of Plymouth Ethical Committee. Results: Total plasma iron absorption and post supplement increases in TSAT and NTBI were greatest in the 35–55 years age group (Table 1). TIBC and iron intake were similar for all groups. A weak correlation was found between iron uptake and change in NTBI (r = 0.405, P < 0.022). Table 1. Mean (SD) plasma analysis, standard deviation and significance between groups Variable 18–34 years 35–55 years 56+ years Significance Δ = change, PP = postprandial, TIBC = total iron binding capacity, TSAT = transferrin saturation. Δ In Iron μM 11.9 (17.4) 20.8 (10) 6.2 (7.4) P < 0.051 Δ In NTBI μM 0.24 (0.36) 1.09 (1.48) 0.29 (0.5) P < 0.060 TIBC μM 102 (23) 92 (23) 97 (11) P < 0.525 % TSAT (PP) 49 (13) 60 (9) 41 (9) P < 0.014 Iron Intake (mg) 66 (29) 67 (21) 73 (1) P < 0.747 Discussion: This study does not support the suggestion that iron absorption is misregulated in older age (Levenson et al., 2004) but does support the ‘set point’ theory that iron uptake increases with age until a certain point (Garry et al., 2000). In healthy individuals the plasma concentration of NTBI is small; the elevated amount found in the 35–55 year age group is of some concern. This study examined a small, predominantly female sample and the results may have been affected by factors that were not considered such as menstruation, menopause and contraceptive use. Conclusion: It is possible that iron absorption is greater in the middle years of life and that this increases both stores and pro-oxidant damage, which in later years manifests into chronic disease. This study may provide new insight into iron regulation and warrants further research.

Transcriptome Analysis of Neisseria meningitidis in Human Whole Blood and Mutagenesis Studies Identify Virulence Factors Involved in Blood Survival

During infection Neisseria meningitidis (Nm) encounters multiple environments within the host, which makes rapid adaptation a crucial factor for meningococcal survival. Despite the importance of invasion into the bloodstream in the meningococcal disease process, little is known about how Nm adapts to permit survival and growth in blood. To address this, we performed a time-course transcriptome analysis using an ex vivo model of human whole blood infection. We observed that Nm alters the expression of ≈30% of ORFs of the genome and major dynamic changes were observed in the expression of transcriptional regulators, transport and binding proteins, energy metabolism, and surface-exposed virulence factors. In particular, we found that the gene encoding the regulator Fur, as well as all genes encoding iron uptake systems, were significantly up-regulated. Analysis of regulated genes encoding for surface-exposed proteins involved in Nm pathogenesis allowed us to better understand mechanisms used to circumvent host defenses. During blood infection, Nm activates genes encoding for the factor H binding proteins, fHbp and NspA, genes encoding for detoxifying enzymes such as SodC, Kat and AniA, as well as several less characterized surface-exposed proteins that might have a role in blood survival. Through mutagenesis studies of a subset of up-regulated genes we were able to identify new proteins important for survival in human blood and also to identify additional roles of previously known virulence factors in aiding survival in blood. Nm mutant strains lacking the genes encoding the hypothetical protein NMB1483 and the surface-exposed proteins NalP, Mip and NspA, the Fur regulator, the transferrin binding protein TbpB, and the L-lactate permease LctP were sensitive to killing by human blood. This increased knowledge of how Nm responds to adaptation in blood could also be helpful to develop diagnostic and therapeutic strategies to control the devastating disease cause by this microorganism.

Structural Variations within the Transferrin Binding Site on Transferrin-binding Protein B, TbpB

Pathogenic bacteria acquire the essential element iron through specialized uptake pathways that are necessary in the iron-limiting environments of the host. Members of the Gram-negative Neisseriaceae and Pasteurellaceae families have adapted to acquire iron from the host iron binding glycoprotein, transferrin (Tf), through a receptor complex comprised of transferring-binding protein (Tbp) A and B. Because of the critical role they play in the host, these surface-exposed proteins are invariably present in clinical isolates and thus are considered prime vaccine targets. The specific interactions between TbpB and Tf are essential and ultimately might be exploited to create a broad-spectrum vaccine. In this study, we report the structure of TbpBs from two porcine pathogens, Actinobacillus pleuropneumoniae and suis. Paradoxically, despite a common Tf target, these swine related TbpBs show substantial sequence variation in their Tf-binding site. The TbpB structures, supported by docking simulations, surface plasmon resonance and hydrogen/deuterium exchange experiments with wild-type and mutant TbpBs, explain why there are structurally conserved elements within TbpB homologs despite major sequence variation that are required for binding Tf.

Regulation of cellular iron metabolism

Iron is an essential but potentially hazardous biometal. Mammalian cells require sufficient amounts of iron to satisfy metabolic needs or to accomplish specialized functions. Iron is delivered to tissues by circulating transferrin, a transporter that captures iron released into the plasma mainly from intestinal enterocytes or reticuloendothelial macrophages. The binding of iron-laden transferrin to the cell-surface transferrin receptor 1 results in endocytosis and uptake of the metal cargo. Internalized iron is transported to mitochondria for the synthesis of haem or iron–sulfur clusters, which are integral parts of several metalloproteins, and excess iron is stored and detoxified in cytosolic ferritin. Iron metabolism is controlled at different levels and by diverse mechanisms. The present review summarizes basic concepts of iron transport, use and storage and focuses on the IRE (iron-responsive element)/IRP (iron-regulatory protein) system, a well known post-transcriptional regulatory circuit that not only maintains iron homoeostasis in various cell types, but also contributes to systemic iron balance.

Urinary hepcidin level as an early predictor of iron deficiency in children: A case control study

BackgroundThe ideal screening test would be capable of identifying iron deficiency in the absence of anemia. We tried to detect role of urinary hepcidin-25 level in early prediction of iron deficiency in children.MethodsThis is a case control study performed on 100 children in Hematology Unit of Pediatric Department, Zagazig University Hospital, Egypt. Our study included 25 cases of iron deficiency (ID) stage-1 (iron depletion), 25 cases ID stage-2 (iron-deficient erythropoiesis), 25 cases ID stage-3 (iron deficiency anemia) and 25 healthy children as a control group. Estimation of iron status parameters was done. Urinary hepcidin-25 level was detected.ResultsUrinary hepcidin-25 level was significantly lower in all stages of iron deficiency than in control group, more significant reduction in its level was observed with the progress in severity of iron deficiency. Urinary hepcidin showed significant positive correlation with hemoglobin, mean corpuscular volume, hematocrit value, serum iron and ferritin and transferrin saturation. In contrary, it showed significant negative correlation with serum transferrin and total iron binding capacity.Urinary hepcidin at cutoff point ≤0.94 nmol/mmol Cr could Predict ID stage-1 with sensitivity 88% and specificity 88%. Cutoff point ≤0.42 nmol/mmol Cr could predict ID stage-2 with sensitivity 96% and specificity 92%. Cutoff point ≤0.08 nmol/mmol Cr could Predict ID stage-3 with Sensitivity 96% and specificity 100%.ConclusionsWe can conclude that detection of urinary hepcidin-25 level was a simple and non invasive test and could predict iron deficiency very early, before appearance of hematological affections.

The influence of iron status and genetic polymorphisms in the HFE gene on the risk for postoperative complications after bariatric surgery: a prospective cohort study in 1,064 patients

BackgroundGastric bypass surgery is a highly effective therapy for long-term weight loss in severely obese patients, but carries significant perioperative risks including infection, wound dehiscence, and leaks from staple breakdown. Iron status can affect immune function and wound healing, thus may influence peri-operative complications. Common mutations in the HFE gene, the gene responsible for the iron overload disorder hereditary hemochromatosis, may impact iron status.MethodsWe analyzed 1064 extremely obese Caucasian individuals who underwent open and laparoscopic Roux-n-Y gastric bypass surgery at the Geisinger Clinic. Serum iron, ferritin, transferrin, and iron binding capacity were measured pre-operatively. All patients had intra-operative liver biopsies and were genotyped for the C282Y and H63D mutations in the HFE gene. Associations between surgical complications and serum iron measures, HFE gene status, and liver iron histology were determined.ResultsWe found that increased serum iron and transferrin saturation were present in patients with any post-operative complication, and that increased serum ferritin was also increased in patients with major complications. Increased serum transferrin saturation was also associated with wound complications in open RYGB, and transferrin saturation and ferritin with prolonged lengths of stay. The presence of 2 or more HFE mutations was associated with overall complications as well as wound complications in open RYGB. No differences were found in complication rates between those with stainable liver iron and those without.ConclusionSerum iron status and HFE genotype may be associated with complications following RYGB surgery in the extremely obese.

Immune response to dna vaccine expressing transferrin binding protein a gene of Pasteurella multocida

Haemorrhagic Septicaemia (HS), an acute and fatal disease of cattle and buffalo is primarily caused by serotype B:2 or E:2 of Pasteurella multocida. The transferrin binding protein A (TbpA) has been found to act as immunogen and potent vaccine candidate in various Gram negative bacteria including P. multocida. The present study was carried out to evaluate the potential of this antigen as a DNA vaccine against HS in mice model. The tbpA gene of P. multocida serotype B:2 was cloned in a mammalian expression vector alone and along with murine IL2 gene as immunological adjuvant to produce monocistronic and bicistronic DNA vaccine constructs, respectively. The immune response to DNA vaccines was evaluated based on serum antibody titres and lymphocyte proliferation assay. A significant increase in humoral and cell mediated immune responses was observed in mice vaccinated with DNA vaccines as compared to non immunized group. Additionally, the bicistronic DNA vaccine provided superior immune response and protection level following challenge as compared to monocistronic construct. The study revealed that DNA vaccine presents a promising approach for the prevention of HS.

Molecular Pathogenesis of Neisseria Gonorrhoeae

Molecular Pathogenesis of Neisseria Gonorrhoeae

Risk Factors Associated with Iron Deficiency Among Children Admitted to Paediatric Emergency Unit in Erbil, Iraq

Background and objective: Iron deficiency is the most prevalent and common micronutrient deficiency in the developing world today. The aim is to determine the iron status among 1-5 years old children admitted to emergency department for different reasons and to determine the risk factors associated with iron deficiency in these children. Methods: a cross sectional study was carried out on a total of 106 children aged 1-5 years attended the emergency department of Raparin teaching hospital for children in Erbil from 16th of May to 16th of August 2007 were invited to participate in the study, parent were interviewed for potential risk factors of iron deficiency, blood were taken for analysis of hemoglobin, serum iron, total iron binding capacity and transferrin saturation after a verbal consent. Iron deficiency was defined as serum iron &lt;50 microgram/dl and/or transferrin saturation (TS)&lt;16%. Results: iron deficiency was noted in 51.9% and 48.1% according to serum iron and transferrin saturation respectively. Male have more risk to have lower TS&lt;16% as compared with female ( OR 0.400, 95% CI 0.182-0.879). Age and tea ingestion were significant independent predictor of iron deficiency, family income associated significantly with low serum iron while pica, frequency of meat ingestion, number of sibling and weight percentile have non significant relation with neither low serum iron nor low transferrin saturation Conclusion: iron deficiency is important health problem in this population of children, prevalence of iron deficiency found in 1-2 year age group in this study was alarming, this suggest the need for greater efforts of prevention.

Intraendosomal Transferrin Saturation Governs Interorganellar Association for Iron Delivery In Erythroid Cells

AbstractAbstract 4253Delivery of iron (Fe) to most cells occurs following the binding of diferric transferrin (Tf) to its cognate receptors on the cell membrane. The Tf-receptor complexes are then internalized via endocytosis, and Fe is released from Tf by a process involving endosomal acidification. Iron, following its reduction to Fe2+ by Steap3 (six-transmembrane epithelial antigen of the prostate 3), is then transported across the endosomal membrane by the divalent metal transporter, DMT1. Unfortunately, the post-endosomal path of Fe within cells remains elusive or is, at best, controversial. It has been commonly accepted that a low molecular weight intermediate chaperones iron in transit from endosomes to mitochondria and other sites of utilization; however, this much sought iron binding intermediate has never been identified. In erythroid cells, more than 90% of iron must enter mitochondria since ferrochelatase, the final enzyme in the heme biosynthetic pathway that inserts Fe2+ into protoporphyrin IX, resides in the inner part of the inner mitochondrial membrane. In erythroid cells, strong evidence exists for specific targeting of Fe toward mitochondria: iron acquired from Tf continues to flow into mitochondria even when the synthesis of protoporphyrin IX is suppressed. Based on this, we have formulated the hypothesis that in erythroid cells a transient mitochondrion-endosome interaction is involved in Fe translocation to its final destination and have collected experimental support for this proposition (Zhang et al. Blood 105:368, 2005; Sheftel et al. Blood 110: 125, 2007). In the present study, we developed two additional experimental strategies to seek further support for the above hypothesis. First, instead of using two-dimensional (2D) confocal microscopy (Sheftel et al. 2007), we generated 3D confocal images using a Quorum WaveFX Spinning Disc Confocal System: a laser-based system using optimized Nipkow Spinning Disc Technology capable of acquiring confocal images at video rates, allowing us to capture images of endosomes interacting with mitochondria in space, excluding the possibility of “pseudo-interactions” caused by coincidental overlapping. This technique clearly revealed contact of endosomes with mitochondria in 3D space. Moreover, we developed a novel method using a flow subcytometry to analyze lysates obtained from reticulocytes with fluorescently labeled endosomes (Alexa Green Transferrin) and mitochondria (MitoTracker Deep Red). Using this strategy, we have identified three distinct flow particulate populations: endosomes, mitochondria, and a population double labeled with both fluorescent markers. The size of the double-labeled population (presumably representing endosomes associated with mitochondria) increases with incubation time and reaches a plateau in ∼20 min. Reticulocyte re-incubation with unlabeled Fe2-Tf leads to a time-dependent decrease of the dual-labeled population, indicating a reversible nature of mitochondria-endosome interactions. Bafilomycin A1, which blocks endosome acidification and Fe release from Tf, slows down the generation of the double-labeled population. One likely explanation for this observation is that bafilomycin A1 blocks Fe2+ export from endosomes already associated with mitochondria, extending their residence time on these organelles, thus causing a “traffic jam” that decelerates the movement of endosomes containing fluorescent Tf towards mitochondria. Similar effects were obtained using heme, which was shown to feedback inhibit the release of iron from Tf within reticulocytes endosomes (Ponka et al. Biochem J 251:105, 1988). The presence of heme during the re-incubation of double-labeled reticulocytes with unlabeled Fe2-Tf slowed down the disappearance of the double population, suggesting that the efficient dissociation of endosomes requires them to be iron-free. Taken together, our new data in support of this “kiss-and-run” behavior of organelles in erythroid cells suggest that a molecular mechanism is in place which coordinates the iron status of intraorganellar Tf with endosomal trafficking. Disclosures:No relevant conflicts of interest to declare.

Delineating the regions of human transferrin involved in interactions with transferrin binding protein B from Neisseria meningitidis

Pathogenic bacteria in the Neisseriaceae possess a surface receptor mediating iron acquisition from human transferrin (hTf) that consists of a transmembrane iron transporter (TbpA) and a surface-exposed lipoprotein (TbpB). In this study, we used hydrogen/deuterium exchange coupled to mass spectrometry (H/DX-MS) to elucidate the effects on hTf by interaction with TbpB or derivatives of TbpB. An overall conserved interaction was observed between hTf and full-length or N-lobe TbpB from Neisseria meningitidis strains B16B6 or M982 that represent two distinct subtypes of TbpB. Changes were observed exclusively in the C-lobe of hTf and were caused by the interaction with the N-lobe of TbpB. Regions localized to the 'lip' of the C1 and C2 domains that flank the interdomain cleft represent sites of direct contact with TbpB whereas the peptides within the interdomain cleft that encompass iron binding ligands are inaccessible in the closed (holo) conformation. Although substantial domain separation upon binding TbpB cannot be excluded by the H/DX-MS data, the preferred model of interaction involves binding hTf C-lobe in the closed conformation. Alternate explanations are provided for the substantial protection from deuteration of the peptides encompassing iron binding ligands within the interdomain cleft but cannot be differentiated by the H/DX-MS data.

Cadmium suppresses the proliferation of piglet Sertoli cells and causes their DNA damage, cell apoptosis and aberrant ultrastructure

ObjectiveVery little information is known about the toxic effects of cadmium on somatic cells in mammalian testis. The objective of this study is to explore the toxicity of cadmium on piglet Sertoli cells.MethodsSertoli cells were isolated from piglet testes using a two-step enzyme digestion and followed by differential plating. Piglet Sertoli cells were identified by oil red O staining and Fas ligand (FasL) expression as assayed by immunocytochemistry and expression of transferrin and androgen binding protein by RT-PCR. Sertoli cells were cultured in DMEM/F12 supplemented with 10% fetal calf serum in the absence or presence of various concentrations of cadmium chloride, or treatment with p38 MAPK inhibitor SB202190 and with cadmium chloride exposure. Apoptotic cells in seminiferous tubules of piglets were also performed using TUNEL assay in vivo.ResultsCadmium chloride inhibited the proliferation of Piglet Sertoli cells as shown by MTT assay, and it increased malondialdehyde (MDA) but reduced superoxide dismutase (SOD) and Glutathione peroxidase (GSH-Px) activity. Inhibitor SB202190 alleviated the proliferation inhibition of cadmium on piglet Sertoli cells. Comet assay revealed that cadmium chloride caused DNA damage of Piglet Sertoli cells and resulted in cell apoptosis as assayed by flow cytometry. The in vivo study confirmed that cadmium induced cell apoptosis in seminiferous tubules of piglets. Transmission electronic microscopy showed abnormal and apoptotic ultrastructure in Piglet Sertoli cells treated with cadmium chloride compared to the control.Conclusioncadmium has obvious adverse effects on the proliferation of piglet Sertoli cells and causes their DNA damage, cell apoptosis, and aberrant morphology. This study thus offers novel insights into the toxicology of cadmium on male reproduction.

N‐Glycosylation is important for the correct intracellular localization of HFE and its ability to decrease cell surface transferrin binding

HFE is a type 1 transmembrane protein that becomes N-glycosylated during transport to the cell membrane. It influences cellular iron concentrations through multiple mechanisms, including regulation of transferrin binding to transferrin receptors. The importance of glycosylation in HFE localization and function has not yet been studied. Here we employed bioinformatics to identify putative N-glycosylation sites at residues N110, N130 and N234 of the human HFE protein, and used site-directed mutagenesis to create combinations of single, double or triple mutants. Compared with the wild-type protein, which co-localizes with the type 1 transferrin receptor in the endosomal recycling compartment and on distributed punctae, the triple mutant co-localized with BiP in the endoplasmic reticulum. This was similar to the localization pattern described previously for the misfolding HFE-C282Y mutant that causes type 1 hereditary haemachromatosis. We also observed that the triple mutant was functionally deficient in beta2-microglobulin interactions and incapable of regulating transferrin binding, once again, reminiscent of the HFE-C282Y variant. Single and double mutants that undergo limited glycosylation appeared to have a mixed phenotype, with characteristics primarily of the wild-type, but also some from the glycosylation-deficient protein. Therefore, although they displayed an endosomal recycling compartment/punctate localization like the wild-type protein, many cells simultaneously displayed additional reticular localization. Furthermore, although the majority of cells expressing these single and double mutants showed decreased surface binding of transferrin, a number appeared to have lost this ability. We conclude that glycosylation is important for the normal intracellular trafficking and functional activity of HFE.

Abstract 4456: Molecular events required for the induction of lethal iron deprivation in malignant hematopoietic cells via an antibody-avidin fusion protein specific for human transferrin receptor 1

Abstract Iron is an essential nutrient for cell growth and division. The transferrin receptor 1 (TfR1 or CD71) is a major gateway for iron import, and as such is expressed on a variety of cells; it is over-expressed in a number of tumors including hematopoietic malignancies. We have previously shown that an antibody-avidin fusion protein (ch128.1Av) specific for TfR1 is toxic to cancer cells, capable of inducing lethal iron deprivation in some malignant B-cells such as IM-9, but not in others such as U266. Importantly, ch128.1Av induces iron deprivation without directly preventing the binding of transferrin (Tf) to TfR1. In an attempt to identify the molecular events behind the iron deprivation induced by ch128.1Av, we have found that like Tf, internalization of ch128.1Av bound to TfR1 is clathrin, dynamin, and temperature dependent. Additionally, recycling of TfR1-bound ch128.1Av at short timescales (30min) is also similar to that of Tf and does not significantly alter the rate of Tf uptake. Long-term incubation (&amp;gt;6h) of IM-9 cells with ch128.1Av precludes their uptake of Tf and induces degradation of TfR1. This is not the case with U266 cells, in which Tf uptake remains unaffected and degradation of TfR1 is negligible in the presence of ch128.1Av. Clustering of cell surface TfR1 by ch128.1Av, given its multivalent nature, was previously thought to cause degradation of the receptor in sensitive cells such as IM-9. We now show that clustering of TfR1 occurs both in cells that are sensitive and resistant to treatment with the antibody (IM-9 and U266), indicating that clustering of TfR1 induced by ch128.1Av is not sufficient to induce receptor degradation and subsequent iron deprivation. Instead, resistance to treatment with ch128.1Av may be attributed to elements downstream of receptor binding/clustering; possibly related to those involved with TfR1 recycling and degradation. Further studies are needed to identify these resistance factors and potentially enhance the efficacy of ch128.1Av as well as other antibodies and/or small molecules that similarly target TfR1 for the treatment of hematologic malignancies. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4456.

Transferrin Receptor 2 Is Frequently and Highly Expressed in Glioblastomas

Under physiological conditions, transferrin receptor 2 (TfR2) is expressed in the liver and its balance is related to the cell cycle rather than to intracellular iron levels. We recently showed that TfR2 is highly expressed in glioblastoma cell lines. Here, we demonstrate that, in these cells, TfR2 appears to localize in lipid rafts, induces extracellular signal-regulated kinase 1/2 phosphorylation after transferrin binding, and contributes to cell proliferation, as shown by RNA silencing experiments. In vitro hypoxic conditions induce a significant TfR2 up-regulation, suggesting a role in tumor angiogenesis. As assessed by immunohistochemistry, the level of TfR2 expression in astrocytic tumors is related to histologic grade, with the highest expression observed in glioblastomas. The level of TfR2 expression represents a favorable prognostic factor, which is associated with the higher sensitivity to temozolomide of TfR2-positive tumor cells in vitro. The endothelial cells of glioblastoma vasculature also stain for TfR2, whereas those of the normal brain vessels do not. Importantly, TfR2 is expressed by the subpopulation of glioblastoma cells with properties of cancer-initiating cells. TfR2-positive glioblastoma cells retain their TfR2 expression on xenografting in immunodeficient mice. In conclusion, our observations demonstrate that TfR2 is a neoantigen for astrocytomas that seems attractive for developing target therapies.

Clinical observation on the treatment of 60 patients with iron deficiency anemia with Jianpi Shengxue Granule

Objective To compare the effects of Jianpi Shengxue Granule and Hexue Capsule in treating patients with iron deficiency anemia. Methods All 120 patients with iron deficiency anemia were randomly recruited into group A (treated with Jianpi Shengxue Granule) and group B (treated with Hexue Capsule), with 60 patients in each group. The levels of serum iron (Fe), total iron binding capacity (T IBC), transferrin saturation (TS), transferrin (Tf), ferritin (SF), hemoglobin (Hb) and serum soluble transferrin receptor(sTIR) were observed in both groups. Results Jianpi Shengxue Granule showed better results in treating patients with iron deficiency anemia than Hexue Capsule with significant difference (P<0.05). Conclusion Jianpi Shengxue Granule is effective in treating iron deficiency anemia. Key words: Jianpi Sheng Granule; Hexue Capsule; Iron deficiency anemia

1-Cinnamoyl-3,11-dihydroxymeliacarpin delays glycoprotein transport restraining virus multiplication without cytotoxicity

The 1-cinnamoyl-3,11-dihydroxymeliacarpin (CDM), isolated from extracts of Melia azedarach L., displays antiviral and immunomodulating properties. CDM is the first reported tetranortriterpenoid responsible for the alkalinization of intracellular compartments affecting both, viral endocytic and exocytic pathways. Considering that viral glycoprotein synthesis is completely dependent upon cellular membrane trafficking, we questioned whether CDM might also interfere with the normal transport of cellular glycoproteins. This study demonstrates that CDM promoted a transient block in the transport of two cellular glycoproteins, the transferrin receptor (TfR) and TNF-α. Nevertheless, CDM did not affect the transferrin binding ability of TfR and did not impede the TNF-α secretion. On the other hand, CDM disturbed the intracellular localization of capsid, glycoprotein and tegument proteins simultaneously in the same HSV-1 infected cells. Besides, we show that concanamycin A and monensin provoke a permanent blockage of viral and cellular glycoproteins, in contrast to the delay observed after CDM treatment. Thus, the delay on glycoprotein transport caused by CDM would account for the strong inhibition on virus multiplication without interfering with the bioactivity of cellular glycoproteins.

Characterization of the Interaction between Diferric Transferrin and Transferrin Receptor 2 by Functional Assays and Atomic Force Microscopy

Transferrin receptor 2 (TfR2), a homologue of the classical transferrin receptor 1 (TfR1), is found in two isoforms, α and β. Like TfR1, TfR2α is a type II membrane protein, but the β form lacks transmembrane portions and therefore is likely to be an intracellular protein. To investigate the functional properties of TfR2α, we expressed the protein with FLAG tagging in transferrin-receptor-deficient Chinese hamster ovary cells. The association constant for the binding of diferric transferrin (Tf) to TfR2α is 5.6×106 M−1, which is about 50 times lower than that for the binding of Tf to TfR1, with correspondingly reduced rates of iron uptake. Evidence for Tf internalization and recycling via TfR2α without degradation, as in the TfR1 pathway, was also found. The interaction of TfR2α with Tf was further investigated using atomic force microscopy, a powerful tool used for investigating the interaction between a ligand and its receptor at the single-molecule level on the living cell surface. Dynamic force microscopy reveals a difference in the interactions of Tf with TfR2α and TfR1, with Tf–TfR1 unbinding characterized by two energy barriers, while only one is present for Tf–TfR2. We speculate that this difference may reflect Tf binding to TfR2α by a single lobe, whereas two lobes of Tf participate in binding to TfR1. The difference in the binding properties of Tf to TfR1 and TfR2α may help account for the different physiological roles of the two receptors.

Effect of Calcium and Phosphorus on Nonhaeme Iron Absorption and Haematogenic Characteristics in Rats

The objectives of this study were to use the dry thyme leaves as source of nonhaeme iron and evaluate the effects of calcium, phosphorus and calcium + phosphorus on nonhaeme iron absorption and haematogenic characteristics in rats. Thirty adult male albino rats, weighing 150 ± 5 g were divided into five groups. The first group fed basal diet, the second group fed thyme diet, the third group fed thyme diet + calcium, the fourth group fed thyme diet + phosphorus and the fifth group fed thyme diet + calcium + phosphorus. All groups fed experimental diets for six weeks. Hemoglobin (Hb), haematocrit (Ht), red blood cell (RBC), mean corpuscular volume (MCV), serum iron (SI), serum ferritin (SF), total iron–binding capacity and transferrin saturation were determined at the beginning and the at end of the experiment. Iron in diet, Fe intake, Fe feces and Fe absorption were also evaluated. The results indicated that the lowest Fe absorption was observed in rats fed the thyme diet + calcium and thyme diet + calcium + phosphorus. Supplementation the thyme diet with calcium or calcium + phosphorus decreased the values of Hb, Ht, RBC, SI and SF. However, supplementation the thyme diet with phosphorus did not affect in Ht, RBC and MCV but Hb, SI and SF increased. The results suggest that supplementation the diet with calcium or calcium + phosphorus interfere with iron absorption.

Luminescent quantum clusters of gold in transferrin family protein, lactoferrin exhibiting FRET

We report the synthesis of highly luminescent, water soluble quantum clusters (QCs) of gold, which are stabilized by an iron binding transferrin family protein, lactoferrin (Lf). The synthesized AuQC@Lf clusters were characterized using UV-Visible spectroscopy, X-ray photoelectron spectroscopy (XPS), transmission electron microscopy (TEM), photoluminescence (PL), matrix assisted laser desorption ionization mass spectrometry (MALDI-MS), FTIR spectroscopy and circular dichroism (CD) spectroscopy along with picosecond-resolved lifetime measurements. Detailed investigations with FTIR and CD spectroscopy have revealed changes in the secondary structure of the protein in the cluster. We have also studied Förster resonance energy transfer (FRET) occurring between the protein and the cluster. The ability of the clusters to sense cupric ions selectively at ppm concentrations was tested. The stability of clusters in widely varying pH conditions and their continued luminescence make it feasible for them to be used for intracellular imaging and molecular delivery, particularly in view of Lf protection.