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PubMed1989

Transformation-induced changes in transferrin and iron metabolism in myogenic cells.

The uptake of transferrin and iron by cultured myogenic cells transformed with a temperature-sensitive strain of the Rous sarcoma virus (tsLA24) was compared with that of normal developing myogenic cells which were proliferating at the same rate as the transformed cells. The mechanism of transferrin and iron uptake was the same in the transformed cells as in normal myogenic cells and involved receptor-mediated endocytosis of transferrin. However, there were differences in transferrin receptor numbers and receptor function. The number of receptors in transformed cells was more than twice as great as in the normal cells largely due to increased surface receptor numbers. Despite this, the rate of iron uptake increased by only 20% in the transformed cells due to less efficient cycling of the transferrin receptors and less efficient release of iron from transferrin to intracellular sites. Some internalized iron was released from the transformed cells still bound to transferrin. A fast and a slow rate of transferrin exocytosis were identified in transformed cells, as in normal cells, indicating that there were at least two intracellular pathways for transferrin. The fast pathway predominated in the transformed cells, compared with an equal importance of the two pathways in the normal cells.

Animals↗

PubMed1989

Effects of transferrin-indium on cellular proliferation of a human leukemia cell line.

In previous studies, we have demonstrated that transferrin-gallium inhibits cellular proliferation by a mechanism whereby cellular iron utilization is impaired. Since indium, a similar class 3A metal, has not been well studied, we examined its effects on cellular iron uptake and cellular proliferation. In these studies, we provide evidence that indium, when bound to transferrin, has a 50-fold higher effect on inhibition of cellular proliferation than indium added as indium salt. Cells exposed to relatively low concentrations of transferrin-indium exhibit markedly increased transferrin receptor expression but, as with transferrin-gallium, these cells incorporate an inappropriately low amount of iron, suggesting that there is a defect in the release of internalized iron from transferrin. In further studies, we utilize a monoclonal antibody against transferrin receptor that inhibits transferrin-mediated iron uptake. This antibody exhibits a dose-related inhibition of cellular proliferation, and when both transferrin-indium and monoclonal antibody are added to media, there is a more than additive effect on inhibition of cellular proliferation.

Antibodies, Monoclonal↗

PubMed1989

[Anemia in malignant tumor diseases. I. Tumor-induced loss of transferrin and its relation to tumor size and degree of malignancy in the rat model].

Cellular uptake of 67Ga-labelled transferrin by the tumor tissue was studied in rats with tumors of different malignancy and different tumor mass using the slowly growing Morris hepatoma 5123C, the moderately growing Novikoff hepatoma and the very fast and aggressive Yoshida hepatoma AH130. The cellular accumulation of 67Ga-transferrin was found to correlate with the proliferation activity of the tumor. The 67Ga-transferrin concentration in the very fast growing Yoshida hepatoma was 4.8 times higher than the concentration in the slowly growing Morris hepatoma. The uptake of 67Ga-transferrin by the tumors resulted in a faster disappearance of circulating 67Ga-transferrin from the blood. The rate of disappearance correlated with the proliferation activity and the spread of the tumors. Using tumors of identical size the elimination of 67Ga-transferrin from the blood was much faster in the rats with Yoshida hepatoma than in those with the slowly growing Morris hepatoma. On the other hand, using tumors of different tumor size it could be demonstrated that the rate of disappearance of 67Ga-transferrin from the blood correlated directly with tumor mass. It is concluded that cellular incorporation of transferrin within the tumor cells results in a loss of circulating transferrin, which correlates with tumor mass and proliferation of tumor. This mechanism is supposed to be the cause for the hypotransferrinemia seen in patients with malignant tumors.

Anemia↗

PubMed1985

Mechanism of phorbol diester-induced regulation of surface transferrin receptor involves the action of activated protein kinase C and an intact cytoskeleton.

Phorbol diesters are tumor-promoting agents that cause differentiation of HL60 human leukemic cells and concomitantly regulate surface transferrin receptors. Regulation of transferrin receptors by phorbol diesters involves receptor internalization in association with increased receptor phosphorylation (hyperphosphorylation). The intracellular mechanism of action of phorbol diester involves binding to and activation of the Ca2+-phospholipid-dependent protein kinase (protein kinase C). Present studies comparing results obtained with whole cells and those from a cell-free system reconstituted from purified protein kinase C and transferrin receptor components have revealed that the transferrin receptor is phosphorylated by protein kinase C activated by phorbol esters. Following tryptic digestion and two-dimensional separation of phosphopeptides of phosphorylated transferrin receptors, two major and several minor phosphoserine-containing fragments are resolved. These fragments are identical whether transferrin receptor is phosphorylated in whole cells incubated with phorbol diesters or following phosphorylation of affinity immobilized transferrin receptor in the in vitro reconstitution system. Phosphoamino acid analysis of these fragments indicates that serine is the only amino acid phosphorylated in whole cells or in the cell-free system. In addition, colchicine is shown to inhibit in a dose-dependent manner phorbol diester-induced internalization but not hyperphosphorylation of the surface transferrin receptor in whole cells. This inhibition is specific for colchicine since inactive beta- and gamma-Lumicolchicine have no such effect, while taxol reverses the inhibition. These results indicate that the phorbol diester-mediated process of down-regulation of the surface transferrin receptor is associated with phosphorylation of the receptor by activated protein kinase C and requires an intact cytoskeleton to affect receptor internalization.

Cell Line↗

PubMed1986

Transferrin binding to two human colon carcinoma cell lines: characterization and effect of 60-Hz electromagnetic fields.

125I-Labeled human transferrin was used to study the binding of transferrin to Colo 320 DM and Colo 205 human cell lines derived from adenocarcinomas of the colon. Although transferrin uptake was greater in both cases at 37 degrees than at 4 degrees it was found that slightly greater than two-thirds of the transferrin associated with the cells at 37 degrees was not bound to surface receptors but rather had been internalized by the cells. Subsequent analysis of true surface binding at 4 degrees by Scatchard analysis allowed determination of the number of transferrin receptors as well as association constants for the interaction. The number of transferrin receptors per cell was found to be inversely related to the cell density of the cultures from which cells were removed for study. Association constants were unaffected by cell density, with average values of 1.2 and 5.4 X 10(8) M-1 obtained for Colo 320 DM and Colo 205, respectively. Additionally, maximum theoretical numbers of receptors of 1.05 X 10(5)/cell for Colo 320 DM and 1.39 X 10(5)/cell for Colo 205 were determined. Furthermore, exposure of Colo 205 cells to three different experimental situations, i.e., 60 Hz-generated electric field only (E+, 300 mA/m2rms), magnetic field only (M+, 1.0 gauss rms), and combined electric + magnetic fields at these intensities (E+M+), altered the expression of transferrin receptors as compared to a concurrently run unexposed control population of cells (E-M-). In three separate experiments the number of transferrin receptors quantitated on both M+ and E+M+ cells was independent of cell culture density and was close to or exceeded the maximum theoretical number of receptors determined for this cell line. In contrast, E+ cells expressed fewer transferrin receptors than was predicted on the basis of cell culture density.

Adenocarcinoma↗

PubMed1986

The p97 antigen is mapped to the q24-qter region of chromosome 3; the same region as the transferrin receptor.

Since the p97 antigen, a membrane-associated iron-binding protein, has extensive amino acid sequence with homology with transferrin, is functionally related to the transferrin receptor, and has been previously mapped to chromosome 3, we have performed additional studies for regional mapping of the gene expressing p97 antigen. In these experiments, Chinese hamster-human cell lines were chosen that contained a large spectrum of autosomal human chromosomes, but mainly consisted of clones expressing all or a part of chromosome 3. These cell lines included a clone that previously allowed for mapping of human transferrin receptor to q22-qter region. Human p97 expression was assessed by specific binding of [125I]monoclonal antibody 96.5, and human transferrin receptor expression was tested by specific [125I]human transferrin binding and [125I]monoclonal antibody OKT-9 specific for human transferrin receptor. Based on these analyses, both human p97 antigenic expression and human transferrin receptor are mapped concordantly to the q24-qter region. These data and previous reports, therefore, suggest that the related iron-transport proteins are closely linked and may be under coordinate regulation. However, studies of several cell lines that exhibit up-regulation of human transferrin receptor expression with cellular proliferation, and down-regulation of receptor with increased transferrin-iron in the media, showed no change in expression of p97 antigen. p97 antigenic expression increased when melanocyte-stimulating hormone was added to a human melanoma cell line in tissue culture. These latter studies suggest that in mammalian cells the two proteins do not show coordinate regulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1984

Effect of iron chelators on the transferrin receptor in K562 cells.

Delivery of iron to K562 cells by diferric transferrin involves a cycle of binding to surface receptors, internalization into an acidic compartment, transfer of iron to ferritin, and release of apotransferrin from the cell. To evaluate potential feedback effects of iron on this system, we exposed cells to iron chelators and monitored the activity of the transferrin receptor. In the present study, we found that chelation of extracellular iron by the hydrophilic chelators desferrioxamine B, diethylenetriaminepentaacetic acid, or apolactoferrin enhanced the release from the cells of previously internalized 125I-transferrin. Presaturation of these compounds with iron blocked this effect. These chelators did not affect the uptake of iron from transferrin. In contrast, the hydrophobic chelator 2,2-bipyridine, which partitions into cell membranes, completely blocked iron uptake by chelating the iron during its transfer across the membrane. The 2,2-bipyridine did not, however, enhance the release of 125I-transferrin from the cells, indicating that extracellular iron chelation is the key to this effect. Desferrioxamine, unlike the other hydrophilic chelators, can enter the cell and chelate an intracellular pool of iron. This produced a parallel increase in surface and intracellular transferrin receptors, reaching 2-fold at 24 h and 3-fold at 48 h. This increase in receptor number required ongoing protein synthesis and could be blocked by cycloheximide. Diethylenetriaminepentaacetic acid or desferrioxamine presaturated with iron did not induce new transferrin receptors. The new receptors were functionally active and produced an increase in 59Fe uptake from 59Fe-transferrin. We conclude that the transferrin receptor in the K562 cell is regulated in part by chelatable iron: chelation of extracellular iron enhances the release of apotransferrin from the cell, while chelation of an intracellular iron pool results in the biosynthesis of new receptors.

Apoproteins↗

PubMed1984

Heme regulation of HeLa cell transferrin receptor number.

The number of diferic transferrin receptors on HeLa cells decreases when cells are grown in iron-supplemented media. The experiments reported here suggest that heme is the iron-containing compound which serves as the signal for receptor number regulation. When HeLa cells were grown in the presence of hemin, transferrin receptor number decreased to a greater degree than when cells were grown in equivalent amounts of iron supplied as ferric ammonium citrate. Incubation of cells in conditions which increased cellular heme content resulted in a decrease in cellular transferrin receptors. Incubating cells with 5-aminolevulinic acid (thus bypassing the rate-limiting step in heme biosynthesis, 5-aminolevulinic acid synthase) led to a decrease in transferrin receptor number. Incubation of cells with an inhibitor of heme oxygenase, Sn-protoporphyrin IX, also led to a decrease in transferrin receptor number. When cellular heme content was decreased by inhibiting heme synthesis with succinylacetone (an inhibitor of 5-aminolevulinic acid dehydratase), or by depriving cells of iron with deferoxamine, an increase in HeLa cell transferrin receptor number was seen. When HeLa cells were incubated with inducers of heme oxygenase (CoCl2, SnCl2, Co-protoporphyrin IX), transferrin receptor number also increased. The effects of all compounds which alter transferrin receptor number were dependent on the concentration of the supplement, as well as the duration of the supplementation. These experiments suggest that intracellular heme content may be an important signal controlling transferrin receptor number.

Aminolevulinic Acid↗

PubMed1982

Regulation of HeLa cell transferrin receptors.

HeLa cells were found to have a single class of non-interacting receptors specific for transferrin. Both apotransferrin and diferric transferrin competed equally with 125I-diferric transferrin for receptor binding. Transferrin binding was temperature-dependent and reversible. Binding of transferrin to cells exhibited a KD of 27 nM with a maximum binding capacity of 1.8-3.7 x 10(6) molecules/cell. Cells grown in the presence of diferric transferrin or in the presence of ferric ammonium citrate exhibited a concentration- and time-dependent decrease in 125I-diferric transferrin binding. The decrease in binding activity reflected a reduction in receptor number rather than an alteration in ligand receptor affinity. Growth of cells in saturating concentrations of apotransferrin did not cause a decrease in receptor number. When iron-treated cells were removed to media free of ferric ammonium citrate, the receptor number returned to control values by 40 h. When receptors were removed with trypsin, cells grown and maintained in ferric ammonium citrate-supplemented media demonstrated a rate of receptor reappearance 47% that of control cells grown in ferric ammonium citrate-free media. Cells grown in media supplemented with diferric transferrin or ferric ammonium citrate exhibited an increase in cytosolic iron content. The transferrin receptor number returned to normal after cells were removed to unsupplemented media, despite persistent elevation of cytosolic iron content. Increased iron content did not appear to be the sole factor determining receptor number.

Binding, Competitive↗

PubMed1981

Studies of the interaction between human transferrin and specific receptors on the trophoblast membrane.

The concept of specific receptors for maternal transferrin on the human syncytiotrophoblast membrane has been generally accepted for many years, but definitive evidence of their existence has been established only recently. In this study, experiments were performed to characterize transferrin receptors further, both on intact membranes and after solubilization. Intact, isolated membrane fragments were examined for transferrin binding, both qualitatively by immunofluorescence and quantitatively by radiobinding. The amounts of ligand bound varied inversely with the quantities of residual maternal transferrin remaining at the time of testing. Scatchard plots revealed that the calculated affinity (K alpha) and the number of receptors increased substantially when transferrin was removed by initial washing with chaotropic agents. After dissociation from the membrane by detergents, occupied receptors largely retained bound transferrin, and transferrin binding by unoccupied receptors was also preserved. The stability of such ligand:receptor complexes was considerably enhanced at pH 5.0, and was apparently unaffected by prior treatment with chaotropic agents. Specific immunoprecipitation of solubilized radioiodinated membrane with rabbit antiserum to human transferrin, followed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate under reducing conditions, indicated a relative molecular mass for the unoccupied receptor of 90,000. These results confirm the existence of high-affinity receptors for transferrin on the trophoblast, and also delineate certain potential pitfalls in studies of their interactions with ligand.

Chemical Precipitation↗

PubMed1984

A highly cytotoxic human transferrin-ricin A chain conjugate used to select receptor-modified cells.

The toxic A chain of ricin was linked to human transferrin via a disulfide bond and the resulting conjugate was shown to bind to cell membrane transferrin receptors. Surface-localized transferrin A chain (TF-A chain) gained access to the cytoplasm and inactivated ribosomes as witnessed by a rapid curtailment of cellular protein synthesis (t1/2 = 6 h) and subsequent cytolysis. The intact conjugate produced potent cytotoxic effects on human leukemia CEM cells, (ID50 = 3 X 10(-11) M), while a 10,000-fold higher concentration of uncoupled transferrin plus A chain was required for comparable action. TF-A chain cytotoxicity was totally blocked by native transferrin or by antibodies directed against ricin A chain and human transferrin. CEM cells gradually acclimated to grow in the presence of TF-A chain displayed 1000-fold resistance to the conjugate while their sensitivity to whole ricin was undiminished. The level of transferrin receptor expressed by these cells was 1/20 the amount present on the parent line and their capacity to bind human transferrin was below the limits of detectability using fluorescent probes. This receptor-deficient cell line was unresponsive to the low levels of transferrin which stimulated proliferation of control CEM cells, but their growth was supported by greatly elevated concentrations of ligand. Receptor variant CEM cells, together with the specific TF-A chain toxin, will be useful for studying the mechanisms for transmembrane delivery of both Fe3+ and ricin A chain into cells and will aid in understanding the growth regulatory functions of the receptor-ligand interaction.

Cell Line↗

PubMed2000

Alternative splicing prevents transferrin secretion during differentiation of a human oligodendrocyte cell line.

Transferrin, the iron-transport protein of vertebrate serum, is synthesized mainly in the liver, from which it is secreted into the blood. Transferrin is also synthesized in oligodendrocytes and is an early marker of their differentiation. We have analyzed the regulation of transferrin expression in HOG cells, a human oligodendrocyte cell line. Transferrin expression was correlated with the appearance of oligodendrocyte differentiation markers when cells were exposed to differentiation medium. In contrast to the protein expressed in hepatocytes or in Sertoli cells, transferrin was secreted by neither HOG cells nor immature rat primary oligodendrocytes in vitro. Moreover, transferrin appears to be localized in the cytosol and not in the secretory compartment, as is expected for secreted proteins. This transferrin localization was correlated with the synthesis of a specific transcript, resulting from an alternative splicing, which leads to the elimination of the signal peptide sequence. These results suggest the existence of a functional difference between transferrin synthesized in the brain and in other organs such as liver and testis. They are in accordance with the hypothesis that transferrin plays a specific role, other than iron transport, in oligodendrocyte maturation and in the myelination process.

Alternative Splicing↗

PubMed1994

Laser desorption time-of-flight mass spectrometric analysis of transferrin precipitated with antiserum: a unique simple method to identify molecular weight variants.

Serum transferrin precipitated with anti-transferrin serum was analysed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOFMS). The transferrin-antibody complex in the immunoprecipitates was separated into transferrin and IgG in an acidic pH, which is the usual condition of loading on MALDI-TOFMS. Ions of IgG and other minor components were not superimposed on the transferrin ions. Transferrin isoforms with different carbohydrate contents could be identified by this simple method easier than by affinity chromatography requiring the time-consuming preparation of an insolubilized specific antibody. The transferrin isoform with a molecular weight of approximately 2.2 kDa smaller than normal transferrin, which is contained in the serum from patients with carbohydrate-deficient glycoprotein (CDG) syndrome, was identified by this method. In addition to the M(1+) ion detected using sinapinic acid as a matrix, the M(2+) and M(3+) ions of transferrin were clearly detected using alpha-cyano-4-hydroxycinnamic acid as matrix and the molecular weight heterogeneity was identified more clearly in multivalent ions than that in the M(1+) ion. The MALDI-TOF analysis of immunoprecipitates may serve as a simple and sensitive method to identify the molecular weight heterogeneity of various biological materials.

Carbohydrate Metabolism, Inborn Errors↗

PubMed1998

Optimized determination of carbohydrate-deficient transferrin isoforms in serum by capillary zone electrophoresis.

Carbohydrate deficient transferrin (CDT) is one of the most reliable markers of chronic alcohol abuse. It consists of a group of minor isoforms of human transferrin (the main iron transport serum protein) deficient in sialic acid groups (asialo, monosialo and disialo) with a pI > 5.7, while the main isotransferrin (tetrasialo) has a pI of 5.4. The aim of the present work was to develop a capillary electrophoretic method to determine CDT in serum, suitable for routine use as a confirmatory technique of the current screening methods based on immunoassays. Serum samples (0.5 mL) were saturated with iron by incubation with 10 mM FeCl3 (9 microL) and 500 mM NaHCO3 (12 microL) for 30 min, then diluted 1/10 in water and injected by positive pressure (0.5 psi for 10 s). Separation was performed with a capillary zone electrophoretic method using bare fused-silica capillaries (20 microm ID, 37 cm in length) and a buffer composed of 100 mM sodium tetraborate adjusted with boric acid to pH 8.3. Applied voltage was 10 kV and temperature 25 degrees C. Detection was by UV absorption at 200 nm wavelength. Under the described conditions, asialo-, monosialo-, disialo-, trisialo- and tetrasialo-transferrin were separated in human serum. The limit of detection (signal-to-noise ratio of 2) was about 0.3% for disialo-transferrin, and 0.4% of trisialo-transferrin, expressed as percentages of the terasialo-transferrin peak area. Relative standard deviations (RSD) of absolute migration times were < 1%, while RSD of relative migration times (on the basis of tetrasialo-transferrin) were < 0.1%. Intra-day and day-to-day peak quantitation precision studies showed RDS ranging from 4 to 9% and from 13 to 24% for disialo- and trisialo-transferrin, respectively. The results from 30 control subjects, including social drinkers, and 13 alcoholics showed disialo- and trisialo-transferrin significantly increased in patients by a factor of about 4.5 (P < 0.0001).

Adult↗

PubMed1994

Reconstitution of a surface transferrin binding complex in insect form Trypanosoma brucei.

In the bloodstream of the mammalian host, Trypanosoma brucei takes up host transferrin by means of a high-affinity uptake system, presumably a transferrin receptor. Transferrin-binding activity is seen in the flagellar pocket and is absent in insect form trypanosomes. By transfection we have reconstituted a transferrin-binding complex in insect form trypanosomes. Formation of this complex requires the products of two genes that are part of a variant surface glycoprotein expression site, expression site-associated gene (ESAG) 6 (encoding a protein with GPI-anchor) and ESAG 7 (encoding a protein without any obvious membrane attachment). This complex can be precipitated by transferrin-Sepharose and by an antibody directed only against the ESAG 6 protein. Transfection of ESAG 6 or 7 alone did not result in transferrin binding. In the transfected trypanosomes, the products of ESAG 6 alone and the combination of ESAG 6 and 7 did not exclusively localize to the flagellar pocket, but were present all over the surface of the trypanosome. The reconstituted transferrin-binding complex also did not result in the uptake of transferrin. Additional proteins present in bloodstream trypanosomes, but not in sufficient amounts in insect form trypanosomes, may therefore be required for the correct routing of the transferrin-binding complex to the flagellar pocket, and for its rapid internalization after ligand binding.

Amino Acid Sequence↗

PubMed1991

Lung-derived growth factor that stimulates the growth of lung-metastasizing tumor cells: identification as transferrin.

We have previously shown that culture medium conditioned by lung fragments contains mitogenic activity for lung-metastasizing tumor cells but not for their non-metastatic counterparts. The growth-promoting component from media conditioned by rat and porcine lungs has been purified and partially characterized as a Mr approximately 66,000 (unreduced) or Mr approximately 72,000 (reduced) glycoprotein [Cancer Res 49:3928, 1989; J Cell Biochem 43:127, 1990]. Here we report that this factor is the iron transport protein transferrin. Migration distances in sodium dodecyl sulfate and native gel polyacrylamide electrophoresis systems were similar, as were the specific activities and spectrum of mitogenic activities of the lung-derived growth factor and transferrin. Electrophoretically separated holo-rat transferrin and rat lung-derived growth factor displayed similar positive stains for iron. A polyclonal antibody generated against the lung-derived growth factor cross-reacted with human and rat transferrin in Western blots, and anti-human transferrin cross-reacted with rat lung-derived growth factor. All of the mitogenic activity contained in crude lung conditioned media could be removed by antibody-mediated transferrin depletion. The putative cell receptor molecular weights for the lung-derived growth factor and transferrin were similar as were the molecular weights of polypeptides produced by partial trypsin cleavage of the two. Finally, the amino acid sequence of certain regions of rat lung-derived growth factor demonstrated a high degree of homology to human transferrin. The physical and biochemical properties, antigenicity, and mitogenic activity of a previously unidentified lung-derived growth factor for lung-metastasizing tumor cells indicate that it is transferrin.

Amino Acid Sequence↗

PubMed1980

Effect of changes in the ionic environment of reticulocytes on the uptake of transferrin-bound iron.

Rabbit reticulocytes were incubated with rabbit transferrin labelled with 59Fe and 125I in media in which the NaCl was replaced by other electrolytes or sucrose. Iron and transferrin uptake by the cells was affected by changes in the pH, ionic strength, ionic composition, and the osmolarity of the medium. Uptake was maximal at pH 7.4. A reduction in ionic strength produced by replacing NaCl with sucrose inhibited the uptake in a concentration-dependant manner, greatest inhibition occurring at lowest salt concentration. Similar results were obtained when KCl, LiCl, RbCl, Na2SO4, or K2SO4 were used instead of NaCl. Low ionic strength was found to inhibit the endocytotic uptake of transferrin labelled with colloidal gold, but had only a small effect on transferrin binding to cell membrane receptors. It was concluded that low ionic strength inhibits iron uptake primarily by blocking the endocytosis of transferrin. Three salts, NH4Cl, CaCl2, and MgCl2, produced different results from the above. NH4Cl inhibited iron uptake at all concentrations used. This action was due to an effect on the release of iron from transferrin, which appeared to be taken up by the cells in a normal manner. When the ionic strength of the sucrose medium was increased by adding low concentrations of CaCl2, iron uptake was greater than with equivalent concentrations of NaCl. However, with CaCl2 concentrations above 10 mM, iron uptake was inhibited, due to inhibition of transferrin uptake, possibly by blocking endocytosis. By contrast, MgCl2 stimulated iron uptake at all concentrations used. The results are discussed in terms of the possible effects of ionic strength, pH, and ionic composition of the extracellular fluid on the three main steps involved in iron uptake by immature erythroid cells: transferrin-receptor interaction, endocytosis, and iron release from transferrin.

Ammonium Chloride↗

PubMed1987

Iron-transferrin-induced increase in protein kinase C activity in CCRF-CEM cells.

Iron transferrin has been found to induce a mean 10-fold increase in the activity of protein kinase C in CCRF-CEM cells. This increase was not detectable up to 45 min after treatment of cells with iron transferrin, although after 60 min, a maximal increase in enzyme activity was observed. Similarly, iron transferrin at concentrations of 0.1-0.5 microgram/ml did not alter protein kinase C activity, while concentrations of iron transferrin of 1-100 micrograms/ml induced a maximal increase in enzyme activity. Apotransferrin and iron in the form of ferric citrate, as well as complexes of transferrin with copper, nickel, zinc, manganese, and cobalt did not increase protein kinase C activity. Additionally, CCRF-CEM cells pretreated with either actinomycin D or cycloheximide and then incubated with iron transferrin did not exhibit increased enzyme activity. Treatment with iron transferrin was found to have no effect on protein kinase C activity in normal human peripheral blood lymphocytes and in HL60, Daudi, and U937 cells. However, normal lymphocytes stimulated with phytohemagglutinin for 48 hr exhibited a 2-fold increase in protein kinase C activity following treatment with iron transferrin. These results indicate a specific effect of iron transferrin on protein kinase C activity in CCRF-CEM cells and in mitogen-stimulated human lymphocytes that may occur through increased synthesis of the enzyme.

Cell Line↗