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Biomedical subjects

J Lindemans

Publications and source records attributed to J Lindemans.

At least 91 records · Page 5Linked to original sources
PubMed1986

Uptake of transcobalamin II-bound cobalamin by isolated rat kidney tubule cells.

The uptake and intracellular processing of transcobalamin II-bound cobalamin by isolated rat kidney tubule cells were studied. The cells absorbed the complex in a temperature-and calcium-dependent process, which could be inhibited by monensin, an inhibitor of endocytosis. Cells, loaded with a mixture of 125I- and 57Co-labelled transcobalamin II-vitamin B12, released 125I-labelled protein-degradation products, while keeping the 57Co-labelled vitamin. Protein degradation was inhibited by chloroquine and monensin, which is further evidence for a process of endocytosis, followed by intralysosomal hydrolysis of the transport protein. Transcobalamin II-vitamin B12 uptake was not fully saturable and other proteins, for example, haemoglobin, inhibited the uptake in a concentration-dependent way. Apparently the uptake proceeds through relatively unspecific protein-binding sites, probably involved in the reabsorption of filtrated proteins, although the affinity for transcobalamin II seems relatively high. Consequently, elevated urinary excretion of cobalamin is expected in patients with overflow proteinuria, and was indeed found in a patient with paroxysmal nocturnal haemoglobinuria.

Animals↗
PubMed1984

Nitrous oxide reduces growth of experimental rat leukemia.

The ability of nitrous oxide to inhibit the in vivo growth of hematological neoplasms was investigated in a rat model for acute myeloid leukemia (BNML). Nitrous oxide, administered in a concentration of 67% with 33% oxygen, resulted in a reduction of spleen and liver weights of approx. 30%, as compared with leukemic rats kept in ambient air. Peripheral white cell counts were also considerably lower in the treated rats. Plasma levels of vitamin B12 were found to be elevated in untreated leukemia, but fell to about normal levels after nitrous oxide exposure. On the contrary, folic acid levels were low in untreated leukemic rats, and significantly higher in animals exposed to nitrous oxide. The observed effects of nitrous oxide appeared to be dose-dependent. The deoxyuridine suppression test performed with leukemic cells became abnormal after nitrous oxide inhalation, in accordance with the effect on normal bone marrow. These results indicate that the interference of nitrous oxide with vitamin B12-related metabolism, which leads to impairment of de novo thymidine synthesis, has the potency to reduce leukemic proliferation in vivo.

Animals↗
PubMed1984

Synergistic growth inhibiting effect of nitrous oxide and cycloleucine in experimental rat leukaemia.

Nitrous oxide (N2O) inactivates the vitamin B12-dependent enzyme methionine synthetase with subsequent impairment of folate metabolism and a reduction of cellular proliferation. Indications exist that this effect is antagonized by S-adenosylmethionine (SAM), and it was investigated whether combination with an inhibitor of SAM synthesis, cycloleucine, would result in increased inhibition of growth in rat leukaemia model (BNML). Leukaemic growth was compared in untreated rats, in rats treated with either nitrous oxide/oxygen (1:1) or cycloleucine (50 mg kg-1 i.p.), and in rats receiving both agents. Combined treatment resulted in the strongest reduction of leukaemic infiltration in spleen and liver, and this reduction often was more than the added effects of single treatments. Peripheral leukocyte counts were also lowest after combined treatment. The deoxyuridine suppression test, measuring folate-dependent de novo synthesis of thymidine, was more severely disturbed with combined treatment. Levels of vitamin B12 in plasma were reduced in rats receiving N2O, but an increase in plasma folate occurred in all treated rats. These results indicate that a reduction of SAM synthesis by cycloleucine can increase the disturbance of folate metabolism that is caused by nitrous oxide, with a potentiation of the effects on leukaemic growth.

Amino Acids↗
PubMed1984

Vitamin B12 absorption in cystic fibrosis.

Vitamin B12 absorption was measured in 30 patients with cystic fibrosis by means of the urinary excretion method and found to be impaired, i.e. less than 10%, in 25. The mean urinary excretion amounted to 4.7 +/- 0.8%. In all patients vitamin B12 absorption improved by the addition of trypsin (18.9 +/- 2.1%). Addition of the vitamin B12 analogue cobinamide, which prevents vitamin B12-binding by R-binders, raised the vitamin B12 absorption to 15.0 +/- 2.2%. A further improvement was obtained by the simultaneous addition of cobinamide and trypsin, 18.2 +/- 2.6%, the same value as with trypsin alone. Assuming that cobinamide addition was effective in suppressing all R-binder activity, the additional effect of trypsin suggests a second, stimulatory function of trypsin on vitamin B12 absorption, separate from R-binder-inactivation. In 5 patients only marginal improvement of vitamin B12 absorption was gained by the addition of either trypsin or cobinamide. The deficient serum vitamin B12 (110 pmol/l) in one of them indicates that the normal pancreas-substitution therapy not always implies sufficient restoration of vitamin B12 absorption.

Adolescent↗
PubMed1984

Elevated serum vitamin B12 in cystic fibrosis.

In 62 patients with cystic fibrosis the serum vitamin B12 concentration ranged from 160-2600 pmol/l with a mean of 1 105 pmol/l. Both vitamin B12-binding proteins in the serum, transcobalamin II and R-binders, carried increased amounts of vitamin B12, but showed relatively normal levels of unsaturated vitamin B12-binding capacity. This combination is rather typical for hepatic dysfunction, although the recurrent pulmonary infections might exert an upward effect on plasma R-binder concentration through increased turnover of myeloid cells. A significant positive correlation between transcobalamin II-vitamin B12 and serum alkaline phosphatase suggests that transcobalamin II-bound vitamin B12 might be an early indicator of focal biliary cirrhosis, which is known to occur in these patients.

Adolescent↗
PubMed1983

Application of a simple immunoadsorption assay for the measurement of saturated and unsaturated transcobalamin II and R-binders.

In this paper an immunoadsorption method for the selective measurement of the concentrations of saturated and unsaturated transcobalamin II and R-binders in human plasma is presented. With this assay the concentrations of saturated transcobalamin II found in the plasma of 70 normal individuals ranged from 20-220 pmol per litre, with a mean of 70 pmol/l. The concentration of R-binders, carrying cobalamin, ranged from 87-491 pmol/l (mean 195 pmol/l). The 33-203 pmol/l (mean 111 pmol/l) of cobalamin analogues were found to be almost exclusively bound to the R-binder fraction. The level of saturated binding proteins is not, or only marginally, influenced by the absorption of ingested cobalamin, even with an oral dose of 15 micrograms. The concentration of transcobalamin II-bound cobalamin is apparently not determined by the availability of unsaturated binding protein. On the contrary, a positive correlation was found between the R-binder-cobalamin concentration and total R-binder level. These observations suggest that the concentration of transcobalamin II-bound cobalamin is primarily determined by the concentration of cobalamin in the tissues.

Humans↗
PubMed1983

Improved distribution analysis of cobalamins and cobalamin analogues in human plasma in which the use of thiol-blocking agents is a prerequisite.

In this paper the quantitative analysis of cobalamins and cobalamin analogues in human plasma by means of high performance liquid chromatography (HPLC) and radioisotope dilution assay (RIDA) is described. Current methods for the extraction of cobalamins from plasma proved impracticable due to the selective loss of hydroxo- and sulphitocobalamin, caused by concomitant reduction to the Co2+ form and tight binding to thiol groups of denatured plasma proteins. Although this process can be prevented by exchange of the hydroxyl group with sulphite, azide or nitrite ions, further separation of the respective cobalamin forms from the other endogenous cobalamins by HPLC proved to be impossible. Efficient extraction of hydroxo- and sulphitocobalamin has been obtained in the presence of the thiol-blocking agent N-ethylmaleimide, which does not interfere with the subsequent chromatographic separation of any of the plasma cobalamin forms. The use of R-binder free Intrinsic Factor and salivary R-binder separately as binding substances in RIDA made it possible to analyse the distribution of biologically active cobalamins as well as cobalamin analogues over the various HPLC fractions. Analyses are reported of 15 normal human plasma samples, in which the major components were found to be methylcobalamin (46.9 +/- 4.5%, mean +/- SD) and hydroxocobalamin (40.4 +/- 7.1%, mean +/- SD). Analyses of plasma extracts in the absence of N-ethylmaleimide showed a gross relative overestimation of the amount of methylcobalamin, due to loss of hydroxocobalamin during the extraction procedure. Cobalamin analogues appeared to be evenly distributed over the cobalamin fractions in a pattern almost similar to that of the biologically active forms.

Blood Proteins↗
PubMed1982

The uptake of R-type cobalamin-binding protein by isolated rat liver cells.

The uptake of R-type cobalamin-binding protein from human granulocytes and plasma by isolated parenchymal rat liver cells has been studied. When [57Co] cyanocobalamin-saturated granulocyte-binding protein or transcobalamin III was incubated with the liver cells in a concentration of 500 pM, more than 80% of the vitamin was taken up in 1 h. Vitamin B-12 bound to plasma transcobalamin I, however, was not taken up unless the protein was desialylated by neuraminidase from Vibrio cholerae. The uptake of iodinated pure granulocyte-binding protein, saturated with cobalamin, reached 100% and was accompanied by increasing intracellular proteolytic degradation of the binding protein. EGTA and asialo-orosomucoid completely inhibited this process of uptake and degradation, whereas partial inhibition was caused by chloroquine and colchicine. These observations provide evidence that these (asialo)-R-type cobalamin-binding proteins are taken up by the cell through the plasma membrane receptor for asialoglycoproteins by means of endocytosis followed by proteolysis of the binding protein in the lysosomes.

Animals↗
PubMed1981

An improved method for large scale purification of human holo-transcobalamin II.

25 mg of human holo-transcobalamin II with a specific cobalamin-binding capacity of 0.95 mol cobalamin/mol TC II was purified from 122 kg Cohn fraction III with a yield of 73% and a purification factor of 9.34 . 10(5). Consecutive purification steps comprised CM-Sephadex batchwise ion-exchange chromatography, affinity chromatography, using cyanocobalamin as a ligand, thermolability attached to 3.3'-diaminodipropylamine-substituted CH-Sepharose, and gel filtration. The high yield of the purification procedure was achieved by improving the stability of apo-transcobalamin II in the eluate of the CM-Sephadex, and by a few other modifications of a former procedure. In the latter, rapid denaturation of apo-transcobalamin II prohibited the use of long term affinity chromatography, which is obligatory for processing large amounts of Crohn fraction. In addition, subfractionation of transcobalamin II into smaller fragments which occurred in SDS-polyacrylamide gel electrophoresis in previous studies, was now reduced, indicating that proteolysis in the CM-Sephadex eluate had been prevented effectively.

Animals↗
PubMed1981

Plasma lysozyme level and reticuloendothelial system function in human liver disease.

Plasma lysozyme levels have been reported to reflect the functional status of the reticuloendothelial system (RES). We measured plasma lysozyme levels in 22 patients with acute hepatitis and 21 patients with cirrhosis and a mesocaval shunt. In 17 of these patients RES function was assessed by measuring the disappearance rate (t/2) of radio-labelled sulphur colloid. In acute hepatitis plasma lysozyme levels and colloid t/2 were significantly lower than in health controls and cirrhotics. In the acute hepatitis patients, the plasma lysozyme levels rose significantly two weeks after admission as the hepatitis improved. The colloid t/2 of the 17 patients with liver disease was significantly correlated with the plasma lysozyme level (r = +0.66, p = 0.005). These results suggest that in human liver disease, in comparison with animal experiments, plasma lysozyme is dependent on RES functional status in the sense that a more active RES will result in a lower lysozyme level.

Acute Disease↗
PubMed1980

Plasma lysozyme levels and decay of neutrophilic granulocytes in patients with Crohn's disease.

In patients with Crohn's disease, the lysozyme concentration in plasma was compared with the Crohn's disease activity index, the granulocyte lysozyme content, the number of circulating neutrophilic granulocytes in peripheral blood and the unsaturated vitamin B12-binding capacity of the plasma cobalophilins as an index for the granulocyte decay. There was no difference in the lysozyme content of granulocytes from patients and controls. Patients with increased plasma lysozyme levels suffered from a more active disease and had larger numbers of circulating granulocytes in their blood than those with normal plasma lysozyme levels. The plasma lysozyme levels correlated well with the cobalophilin levels, in patients with increased levels even more markedly than in those with normal levels. The results suggest that in patients with Crohn's disease increased plasma lysozyme levels are due to an increase in granulocyte decay.

Adult↗
PubMed1979

Purification of human transcobalamin II-cyanocobalamin by affinity chromatography using thermolabile immobilization of cyanocobalamin.

Transcobalamin II-cyanocobalamin was isolated from Cohn fraction III of pooled human plasma by affinity chromatography on cyanocobalamin-Sepharose and some conventional separation methods. The affinity ligand cyanocobalamin was coupled to AH-Sepharose by a thermolabile linkage. The unsaturated binding protein was absorbed at 4 degrees C and eluted from the column at 37 degrees C as transcobalamin II-cyanocobalamin complex. The final preparation had a specific cyanocobalamin-binding capacity of 0.98 mol cyanocobalamin/mol transcobalamin II, the yield was 55% and the purification index amounted to 1.1 . 10(6). In dodecyl sulphate polyacrylamide gel electrophoresis one major protein band was observed at a molecular weight of 37 000 and a faint band at a molecular weight of 29 000. In polyacrylamide gel isolectric focusing the pure preparation turned out to be heterogeneous with isoelectric points ranging from pH 6.2 to 6.8, possibly by the occurrence of isoproteins.

Blood Proteins↗
PubMed1979

Preparation of vitamin B-12-free serum for the use in vitamin B-12 radioassays.

In radioassays for serum vitamin B-12, the separation of free and bound vitamin is usually made with charcoal absorption. The specificity of this separation depends on the amount of charcoal and the protein content and constitution of the medium. The large difference in protein concentration between the samples for the dilution curve and the serum samples introduces an uncontrolled variable in the test. In order to equalize the experimental circumstances, the standard dilutions were made in serum freed from vitamin B-12 after boiling the serum for 20 min in a 4-fold dilution with glutamic acid buffer at pH 3.3 and subsequent passage over a CH-Sepharose 4-B column complexed with hog intrinsic factor (IF). The vitamin B-12-binding capacity of such an affinity column prepared from 1 g CH-Sepharose and 20 mg IF, suffices for the absorption of vitamin B-12 in 3000 ml serum from which 300 series of vitamin B-12 standard solutions can be made. Our first results with this method confirm that the charcoal absorption radioassay has become more accurate by the use of vitamin B-12-free serum in the standard dilutions.

Charcoal↗
PubMed1975

Evaluation of a radioassay for serum folate and the effects of ascorbate and methotrexate.

In this study a competitive binding assay for serum folate is evaluated with special attention to storage conditions of the serum samples and the effect of methotrexate on the assay results. Average serum folate levels were 31 nmol/1 +/- 8 (S.D.) in 59 nromal sera and 19 nmol/1 +/- 12 (S.D.) in 354 sera from hospitalized patients of which 43 values were lower than 9 nmol/1. Serum folate appeared to be stable at - 20 degrees C for at least 5 weeks. The addition of ascorbate, which is said to stabilize serum folate, caused a non-specific increase of the measured folate. Methotrexate acted competitively with the tracer folate in the assay sytem when added in vitro and elevated the folate readings. The same occurred in the serum of a patient after intravenous methotrexate.

Ascorbic Acid↗
PubMed1975

Detection of water-to-blood leakages in heat exchangers for cardiopulmonary bypass.

In the construction of heat exchangers, it is of great importance that when they are in use it shall be impossible for water to leak into the blood, as this might lead to hemolysis, intoxications, air emboli, or infection by microorganisms. In an investigation the aim of which was to discover water-to-blood leakages in the heat exchangers that are generally used during cardiopulmonary bypass, tests were carried out on 17 heat exchangers that are used all over the world and are supplied by five different makers. They included both disposable and nondisposable models. In six stainless steel heat exchangers, water leakages to the blood compartment were detected with the aid of helium. Some of these heat exchangers were brand new and had never been used. The size of the leakages was found to be sufficient to enable them to act as channels for the passage of foreign microorganisms.

Blood↗