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H G van Eijk

Publications and source records attributed to H G van Eijk.

At least 37 records · Page 2Linked to original sources
PubMed1996

Analytical and clinical implications of soluble transferrin receptors in serum.

In this study the analytical performances of two recently introduced assays for soluble transferrin receptors in serum were tested. The Ramco transferrin assay was compared with the Eurogenetics assay. In a small clinical study serum samples from patients with anaemia of chronic disease, iron deficiency and myelodysplastic syndrome were analysed, as well as sera from healthy volunteers. The analytical performances of the Ramco assay were found to be acceptable. In the Eurogenetics test however, inter-assay imprecision and the end of run drift were unacceptably high. We were able to confirm that in patients with uncomplicated iron deficiency the concentration of soluble transferrin receptors is higher than in healthy volunteers. In cases of anaemia of chronic and inflammatory disease, the levels of soluble transferrin receptors in serum are slightly, but not significantly, higher than in normal subjects. Measurement of soluble transferrin receptors in serum provides a good differentiation between anaemia of chronic disease and iron deficiency.

Anemia, Iron-Deficiency↗
PubMed1995

Expression of haemopexin receptors by cultured human cytotrophoblast.

The expression of cell-surface haemopexin (Hx) receptors on human cytotrophoblasts was assessed by using four different Hx species purified from plasma: human Hx isolated by wheatgerm-affinity chromatography, human Hx isolated by haem-agarose-affinity chromatography and rabbit and rat Hx, also isolated by haem-agarose-affinity chromatography. About 3500-7000 high-affinity (Kd 0.34-0.85 nM) receptors per cell were measured by Scatchard-type analysis at 4 degrees C using human (species obtained by both methods) or rabbit 125I-labelled haem-Hx. Measured simultaneously, transferrin receptor number and affinity were 40,000/cell and 0.83 nM respectively. In contrast with transferrin receptors, the number of Hx receptors did not increase during 24 h in cytotrophoblast culture. Rat Hx showed no specific binding to human Hx receptors in cytotrophoblast cultures.

Animals↗
PubMed1995

An in vitro study on the binding of Al(III) to human serum transferrin with the isoelectric focusing technique.

Transferrin saturated with Al3+ subjected to isoelectric focusing (IEF) in a pH gradient can be separated into four fractions, representing the apotransferrin, transferrin with aluminum at the metal binding site in the C- or N-terminal lobe, or both. The electrophoretic mobilities of these four fractions are identical to those of the iron-transferrin counterparts. Simultaneous binding of aluminum and iron to transferrin can also be demonstrated. The decreased saturation after IEF indicates that the affinity of transferrin for aluminum is low compared with its affinity for iron. This effect is particularly evident when bicarbonate is used as the synergistic anion in the loading procedure. In contrast, loading of transferrin with aluminum in the presence of oxalate produces a di-aluminum-transferrin complex that is stable during IEF.

Aluminum↗
PubMed1995

Isolation, purification and characterization of porcine serum transferrin and hemopexin.

Two techniques are described for the isolation of porcine serum transferrin and hemopexin, respectively, yielding nearly pure proteins (> 99%) as tested with crossed immunoelectrophoresis. Porcine transferrin has an estimated molecular weight of 79 kDa and porcine hemopexin a molecular weight of 62 kDa. Both purified proteins were subjected to amino acid and carbohydrate analyses. Based on carbohydrate and sialic acid analyses, it is proposed that transferrin contains one bi-antennary glycan chain, whereas hemopexin contains two bi-antennary and one tri-antennary glycan chains.

Amino Acids↗
PubMed1995

Optimized bacterial production of nonglycosylated human transferrin and its half-molecules.

Transferrin is a glycoprotein functioning in iron transport in higher eukaryotes, and consists of two highly homologous domains. To study the function of the glycan residues attached exclusively to the C-terminal domain, we have constructed a plasmid allowing production of nonglycosylated human transferrin in Escherichia coli. By molecular biological and genetic techniques, production was stepped up to 60 mg/l. Similar plasmids were constructed for production of the two half-transferrins. The recombinant proteins accumulate in inclusion-body-like aggregates, where they appear to bind iron without causing bacteriostasis. Proteins active in iron binding have been purified from these inclusion bodies.

Amino Acid Sequence↗
PubMed1995

Quantification of different transferrin receptor pools in primary cultures of porcine blood-brain barrier endothelial cells.

Distribution of iron in the brain varies with region, cell type, and age. Furthermore, some neurological diseases are accompanied by an abnormal accumulation of iron in specific areas of the CNS. These findings implicate a mobile intracerebral iron pool; however, transport of iron across the blood-brain barrier and its regulation are largely unknown. In an extensive series of experiments in primary cultures of porcine blood-brain barrier endothelial cells, we separately quantified surface-bound and total cellular transferrin receptor pools. Although 90% of all transferrin receptors were located inside the cell, only 10% of these intracellular receptors actively took part in the endocytic cycle. This large "inactive" intracellular transferrin receptor pool could either function as a storage site for spare receptors or be activated by the cell to increase its capacity for iron transport. Data were corrected for nonspecific binding by a separate biochemical assessment using a 100-fold excess of unlabeled ligand. Data were also analyzed in a nonlinear curve-fit program. This resulted in a less elaborate and less biased estimate of nonspecific binding.

Animals↗
PubMed1994

CCC.UGA: a new site of ribosomal frameshifting in Escherichia coli.

To activate expression of a human transferrin (Tf)-encoding cDNA in Escherichia coli by translational coupling, it was placed in an expression plasmid downstream from a 5'-terminal fragment from the replicase (R)-encoding gene of bacteriophage MS2. The resulting construct was found to produce, besides the desired Tf, a protein with the mobility of a fusion product (RTf) of the N-terminal R fragment and Tf. Analysis of available mutants showed that this fusion results from +1 ribosomal frameshifting at the end of the R reading frame. This region contains the sequence, CCC.UGA, suggesting that before termination occurs, tRNA(Pro) may dislodge from the CCC codon and reassociate with the +1 triplet CCU. By further site-directed mutagenesis, we demonstrate that both the CCC codon and the termination codon are indeed required for the observed 2-4% frameshifting. When either triplet is changed, the frequency of frameshifting drops to 0.3% or less. These results classify CCC.UGA as a new '+1 shifty stop'.

Bacterial Proteins↗
PubMed1994

Purification of isotransferrins by concanavalin A sepharose chromatography and preparative isoelectric focusing.

1. From pooled serum containing genetically homogeneous transferrin C1, transferrin was purified and separated in three fractions (tri-tri, bi-tri- and bi-bi-antennary transferrin C1), using Concanavalin A-Sepharose. 2. Each of these fractions was separated into its sialic acid-dependent subfractions by preparative isoelectric focusing. Sixteen iso-transferrin C1 fractions were obtained, which differed in their degree of glycan branching and/or their sialic acid content. 3. Preliminary carbohydrate analyses suggest that in some iso-transferrins the N-acetylglucosamine and the galactose content is lower than expected.

Adult↗
PubMed1993

Regulation of transferrin receptor expression and distribution in in vitro cultured human cytotrophoblasts.

During gestation the transplacental iron transport is very important to the fetus. Iron uptake by the placenta can be studied in cultured cytotrophoblasts. The influence of culture time and human differic transferrin on the number and distribution of transferrin receptors (TfRs) was investigated in human cytotrophoblasts. Cytotrophoblasts cultured for 2.5 h had few TfRs (0.28 pmol/mg protein). With time, total TfR amounts increase (4.14 pmol/mg protein at 70 h). They increase to a higher level in cells cultured in iron-poor medium, indicating that iron has an effect on the TfR synthesis/breakdown ratio. TfRs were distributed between two 'active' (located at the cell surface and intracellularly) and one 'inactive' (located intracellularly) receptor pools. TfR distribution among these pools was modulated by culture time and iron. Trophoblasts regulated iron uptake by variation of number of surface TfRs via changes in total TfRs and redistribution of TfRs among the receptor pools.

Cell Differentiation↗
PubMed1993

The effect of desferrioxamine on iron metabolism and lipid peroxidation in hepatocytes of C57BL/10 mice in experimental uroporphyria.

The effects of the iron chelator desferrioxamine (DFx) on liver iron accumulation, malondialdehyde (MDA) production, porphyrin accumulation and uroporphyrinogen decarboxylase (URO-D; EC 4.1.1.37) activity were investigated over a period of 14 weeks in C57BL/10 mice, made porphyric by the administration of hexachlorobenzene (HCB) and iron-dextran (Imferon, IMF) or IMF alone. In addition, we measured the amount of low molecular weight (LMW) iron in liver tissue to determine a possible correlation with MDA production. These experiments showed that combined treatment with HCB + IMF, as well as IMF alone, resulted in porphyrin accumulation, increased MDA production and reduced URO-D activity, whereas HCB alone had no effect. DFx caused a reduction in hepatic porphyrins, this reduction being more distinct in the IMF group than in the HCB + IMF group. The effect of DFx on MDA production and URO-D activity was in agreement with the results on porphyrin accumulation. LMW iron pool measurements at 11 weeks correlated well with data on MDA production in all treated groups in that period (r2 = 0.84), suggesting both variables are interdependent. In conclusion, these results suggest an important role for iron in porphyrin accumulation, probably through its catalytic role in the generation of oxygen-related free radicals, resulting in direct damage to URO-D. The effectiveness of DFx in reducing porphyrin accumulation is probably the result of a reduction in LMW iron, thus diminishing the amount of iron available for a catalytic role in the generation of oxygen-related free radicals.

Animals↗
PubMed1993

Decreased affinity and number of transferrin receptors on erythroblasts in the anemia of rheumatoid arthritis.

In anemia of chronic disease (ACD) in rheumatoid arthritis (RA) a decreased iron uptake and transferrin binding by erythroblasts are postulated to play a pathophysiological role. To examine whether this is related to changes in transferrin receptor expression by erythroblasts, we studied bone marrow from 5 healthy controls, 5 nonanemic RA patients, and 9 RA patients with ACD. Bone marrow mononuclear cells were incubated with increasing concentrations of 125I-transferrin and specific binding data were analyzed by the method of Scatchard. The number of transferrin receptors on erythroblasts from RA patients with ACD was significantly lower as compared to nonanemic RA patients (P < .05) and controls (P < .02). The affinity of the transferrin receptor tended to be lower in ACD. These preliminary data may indicate that transferrin receptor expression by erythroblasts is impaired in ACD. Since the rate of erythroid iron uptake is mainly determined by the number of transferrin binding sites, this may explain a decrease in erythroblast iron availability in ACD in RA.

Aged↗
PubMed1993

Ferritin accumulation and uroporphyrin crystal formation in hepatocytes of C57BL/10 mice: a time-course study.

To establish the time-sequence relationship between ferritin accumulation and uroporphyrin crystal formation in livers of C57BL/10 mice, a biochemical, morphological and morphometrical study was performed. Uroporphyria was induced by the intraperitoneal administration of hexachlorobenzene plus iron dextran and of iron dextran alone. Uroporphyrin crystal formation started in hepatocytes of mice treated with hexachlorobenzene plus iron dextran at 2 weeks and in mice treated with iron dextran alone a 9 weeks. In the course of time, uroporphyrin crystals gradually increased in size. Uroporphyrin crystals were initially formed in hepatocytes in the periportal areas of the liver, in which also ferric iron staining was first detected. The amount and the distribution of the main storage form of iron in hepatocytes, ferritin, did not differ between the two treatment groups. Ferritin accumulation preceded the formation of uroporphyrin crystals in hepatocytes in both treatment groups. Moreover, uroporphyrin crystals were nearly always found close to ferritin iron. We conclude that uroporphyrin crystals are only formed in hepatocytes in which also iron (ferritin) accumulates. Hexachlorobenzene accelerates the effects of iron in porphyrin metabolism, but does not influence the accumulation of iron into the liver.

Animals↗
PubMed1993

Iron chelators may enhance erythropoiesis by increasing iron delivery to haematopoietic tissue and erythropoietin response in iron-loading anaemia.

Based on the mode of action of iron chelators, one might expect a decrease in bone marrow iron availability, resulting in worsening of the anaemia in certain types of iron-loading anaemia. However, improvement of anaemia or reduction in transfusion requirements during chelation treatment has been reported in various types of iron-loading anaemia. It is suggested that iron chelators act as mediators facilitating iron release from storage sites and its delivery to haematopoietic tissues. In addition, a reduction of iron stores may upregulate erythropoietin response and bring about a decrease of disease activity in inflammatory disorders, resulting in a haemoglobin rise. Large trials with (oral) iron chelators are required to verify these possible effects.

Anemia↗