PubMed Health⌕ Search

Biomedical subjects

D K Meijer

Publications and source records attributed to D K Meijer.

At least 109 records · Page 6Linked to original sources
PubMed1995

Antiviral effects of plasma and milk proteins: lactoferrin shows potent activity against both human immunodeficiency virus and human cytomegalovirus replication in vitro.

Native and chemically derivatized proteins purified from serum and milk were assayed in vitro to assess their inhibiting capacity on the cytopathic effect of human immunodeficiency virus (HIV)-1 and human cytomegalovirus (HCMV) on MT4 cells and fibroblasts, respectively. Only native and conformationally intact lactoferrin from bovine or human milk, colostrum, or serum could completely block HCMV infection (IC50 = 35-100 micrograms/mL). Moreover, native lactoferrin also inhibited the HIV-1-induced cytopathic effect (IC50 = 40 micrograms/mL). When negatively charged groups were added to lactoferrin by succinylation, there was a 4-fold stronger antiviral effect on HIV-1, but the antiviral potency for HCMV infection was mostly decreased. Lactoferrin likely exerts its effect at the level of virus adsorption or penetration (or both), because after HCMV penetrated fibroblasts, the ongoing infection could not be further inhibited.

Animals↗
PubMed1994

Characterization of transport in isolated human hepatocytes. A study with the bile acid taurocholic acid, the uncharged ouabain and the organic cations vecuronium and rocuronium.

The uptake and efflux of three categories of substrates were measured in isolated human hepatocytes and compared to those in rat hepatocytes. In addition, the extent to which the in vitro experiments quantitatively reflect liver function in vivo in both species was investigated. The anionic bile acid taurocholic acid was taken up by isolated human hepatocytes at a considerably lower rate than observed in isolated rat hepatocytes. Taurocholic acid uptake both in human hepatocytes and in liver plasma membrane vesicles showed sodium dependency. The uptake rate of taurocholic acid in isolated hepatocytes of both species was quantitatively compatible with the reported liver clearance of the bile acid in vivo. Ouabain uptake rate in isolated human hepatocytes was lower than in rat hepatocytes. This species difference was in accordance with pharmacokinetic studies in vivo on hepatic clearance of ouabain in man and rat. Uptake of vecuronium into human hepatocytes was about a factor of 10 lower than that in rat hepatocytes. Uptake into and efflux from human hepatocytes was comparable for the two short acting muscle relaxants vecuronium and rocuronium. Since distribution to the liver is considered to be a major factor in termination of action of vecuronium and rocuronium these observations were in line with the human pharmacokinetic profiles. In conclusion, the uptake rate of the studied model compounds in human hepatocytes appeared to be lower than that in rat hepatocytes. These observed transport rates reflected the relative hepatic transport rates observed in these species in the intact organism, but the absolute values in both species for some substrates may have been somewhat lower than calculated from in vivo data. It is concluded that transport studies in isolated hepatocytes are suitable for comparative drug transport studies, but are less precise in the prediction of quantitative membrane transport.

Androstanols↗
PubMed1994

Effect of ethanol on hepatobiliary transport of cationic drugs. A study in the isolated perfused rat liver, rat hepatocytes and rat mitochondria.

The effect of ethanol on the hepatic uptake of various cationic drugs was studied in isolated perfused rat livers, isolated rat hepatocytes and isolated rat liver mitochondria. In isolated rat hepatocytes and in isolated perfused rat livers, the uptake of the model organic cation tri-n-butylmethylammonium was found to be markedly stimulated by ethanol in a concentration-dependent fashion. The uptake of tri-n-butylmethylammonium at 1 microM was increased to 120% and 137% at 0.5% (v/v, (= 87 mM)) and 1% (v/v. (= 174 mM)) ethanol, respectively. At 25 microM, tri-n-butylmethylammonium uptake was increased to 124% and 152% at 0.5% (v/v) and 1% (v/v) of ethanol, respectively. The uptake of the organic cations azidoprocainamide methoiodide, vecuronium, ORG 9426 and ORG 6368, the anionic compound taurocholate and the uncharged compound ouabain was not markedly increased at these ethanol concentrations. The mechanism of action of ethanol on the uptake of tri-n-butylmethylammonium was further studied. Competitive inhibitors for the type I organic cation uptake system, procainamide ethobromide and verapamil, almost completely blocked uptake of tri-n-butylmethylammonium (1 microM) in the presence of 1% (v/v) ethanol, indicating that carrier-mediated uptake is still involved and that additional passive diffusion is unlikely. Neither the plasma membrane potential nor the accumulation of the cation in mitochondria was altered after ethanol treatment, suggesting that potential driving forces for uptake and sequestration were not affected.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetaldehyde↗
PubMed1994

The sinusoidal efflux of dibromosulfophthalein from rat liver is stimulated by albumin, ligandin and fatty acid binding protein but not by other dibromosulfophthalein binding proteins.

Organic anions can be excreted from the liver into the bile or back into the general circulation (sinusoidal efflux). It has previously been shown that the net sinusoidal efflux rate of dibromosulfophthalein from the perfused liver into the perfusate is the result of actual efflux from and reuptake into the liver, and can be strongly influenced by the presence of bovine serum albumin in the perfusion medium. The present study investigated whether the influence of albumin on the net sinusoidal efflux process is albumin-specific or whether other binding proteins could have a similar effect on the sinusoidal efflux. Using a single-pass liver perfusion technique and short-lasting (pulse) protein infusions, the stimulatory effect of a wide range of dibromosulfophthalein binding proteins on the sinusoidal efflux process were determined. These experiments showed that all the serum albumins tested as well as the liver cytosolic binding proteins fatty acid binding protein and ligandin (glutathione S-transferase) stimulated this process. The other proteins tested, bovine beta lactoglobulin-b, human gamma globulin and chicken egg lysozyme showed no stimulatory effect, despite relatively high equilibrium binding of dibromosulfophthalein. No clear-cut relationship was found between the equilibrium unbound ligand concentration as measured in perfusate and the stimulatory effect, suggesting absence of equilibrium binding in the sinusoids. Equilibrium binding of dibromosulfophthalein to chicken serum albumin and ligandin as well as the dissociation rate constants were determined in vitro with rapid filtration techniques.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Disposition of glycosidase inhibitors in the isolated perfused rat liver: hepatobiliary and subcellular concentration gradients of 1-deoxymannojirimycin and N-methyl-1-deoxynojirimycin.

The hepatic disposition of two glycosidase inhibitors was studied in the isolated perfused rat liver and after subcellular fractionation. The mannosidase inhibitor 1-deoxymannojirimycin (dMM) and the glucosidase inhibitor N-methyl-1-deoxynojirimycin (MedNM) exhibited minimal binding to albumin and reached liver concentrations that approximately equaled their medium concentrations, after 30 min (MedNM) or 90 min (dMM). Within 2 hr 0.5% of the dose of MedNM and 2.9% of dMM were excreted in bile. No metabolites were found for MedNM, whereas minor (bio)degradation was inferred for dMM. After subcellular fractionation, dMM and MedNM were found predominantly in the cytosolic fraction. Compared to the other particulate fractions, MedNM was elevated in the microsomal fraction, and both compounds were slightly enriched in the lysosomal fraction. We conclude that dMM and MedNM will likely inhibit liver enzymes when sufficiently high plasma levels are reached.

1-Deoxynojirimycin↗
PubMed1994

Metabolism of three pharmacologically active drugs in isolated human and rat hepatocytes: analysis of interspecies variability and comparison with metabolism in vivo.

1. The metabolism of the three drugs (Org GB 94, Org 3770 and Org OD 14) was studied in isolated human and rat hepatocytes. The metabolic profiles in rat and human hepatocytes were compared with the available in vivo data in both species. 2. All three drugs were metabolized extensively under the conditions used, both in human and rat hepatocytes, showing both extensive phase I and II metabolism. 3. During 3-h incubation with rat hepatocytes the three compounds were metabolized completely, whereas incubation with human hepatocytes only resulted in partial metabolism, amounting for 58% (Org GB 94), 36% (Org 3770) and 94% (Org OD 14) of the dose. In addition, rat hepatocytes excreted relatively more of the formed metabolites than human hepatocytes. 4. For both species, the metabolites formed in the isolated cells were quite similar to those found in vivo. With respect to Org GB 94 and Org 3770, metabolites were detected in man in vivo and in isolated human hepatocytes that were not found in any of the animal species studied previously. 5. The reflection of interspecies differences in isolated hepatocytes, with respect to both metabolite profiles and human-specific metabolites, renders isolated human hepatocytes a very valuable tool during preclinical drug development.

Anabolic Agents↗
PubMed1993

Maintenance of viability and transport function after preservation of isolated rat hepatocytes in various simplified University of Wisconsin solutions.

Rat hepatocytes were preserved for 24 hr with high recovery and good maintenance of viability and transport function both in University of Wisconsin (UW) solution and in various simplified UW solutions. Cell quality is somewhat affected after 48 hr of preservation in both the original UW solution and the simplified solutions. ATP content and uptake rate of taurocholic acid are more sensitive markers of cell viability than Trypan blue exclusion or the MTT test. A much less expensive solution than UW, containing only K(+)-lactobionate, KH2PO4, MgSO4 and raffinose, can be used successfully for preservation of rat hepatocytes for 24 hr for drug transport studies.

Adenosine Triphosphate↗
PubMed1993

Hepatic and intrahepatic targeting of an anti-inflammatory agent with human serum albumin and neoglycoproteins as carrier molecules.

The anti-inflammatory agent naproxen (Nap) was covalently coupled to human serum albumin (HSA) and to the neoglycoproteins, galactose and mannose terminated HSA, to deliver this drug selectively to different cell types of the liver. Disposition of Nap20-HSA was studied in rats and compared to that of equivalent doses of mixtures of uncoupled drug and protein. The liver to kidney ratios of the drug (L/K-Nap) and the protein (L/K-prot.) were increased, indicating an improved delivery of both protein and drug to the target site. After injection of 10 micrograms Nap20-HSA the L/K-prot. was increased 15.0 +/- 0.21-fold as measured 1 hr after injection. Even after injection of 5 mg of the conjugate, the L/K-prot. was enhanced 5.6 +/- 0.34-fold and the L/K-Nap 4.6 +/- 0.23-fold as measured 1 hr after injection. Immunohistochemical staining of liver slices revealed that the endothelial cells were the main sites for hepatic uptake. Further pharmacokinetic studies of Nap20-HSA in isolated perfused rat livers showed a saturable uptake process (Vmax = 2.46 micrograms/min/10.0 g liver and Km = 4.27 x 10(-6) M). The uptake in the liver could be inhibited by various polyanionic probes, indicating the major involvement of a scavenger receptor system in the internalization mechanism of Nap20-HSA. This endothelial uptake via the scavenger receptor system is likely to be related to the increased negative charge of the Nap-albumin conjugate as was revealed by anion exchange chromatography. Studies in the intact organ and in purified liver lysosomal lysates indicate that after internalization of Nap20-HSA the conjugate is proteolytically degraded leading to the formation of the lysine conjugate of Nap. This amino acid conjugate of Nap was shown in a previous study by us to be equipotent to Nap itself with regard to prostaglandin-E2 synthesis inhibition. A pronounced altered intrahepatic distribution was observed when Nap was coupled to lactosaminated and mannosylated HSA (Lact-HSA and Man-HSA, respectively). Coupling of Nap to Lact27-HSA and Man10-HSA resulted in a major shift in intrahepatic distribution from endothelial cells to the hepatocytes and Kupffer cells, respectively. We conclude that conjugation of Nap to HSA itself results in a selective delivery to endothelial cells and that the local proteolysis of the conjugate produces an active catabolite. Selective delivery to other cell types of the liver can be achieved by attaching naproxen to neoglycoproteins with an appropriate type and number of sugar groups.

Animals↗
PubMed1993

Differential effects of metabolic inhibitors on cellular and mitochondrial uptake of organic cations in rat liver.

The effects of several metabolic inhibitors on the uptake of tri-n-butylmethylammonium (TBuMA) were studied in isolated rat liver mitochondria, isolated rat hepatocytes and isolated perfused rat livers, in order to characterize further the mechanisms for carrier-mediated uptake and cellular accumulation of organic cations in the liver. Treatment of isolated hepatocytes with valinomycin, carbonylcyanide-m-chlorophenyl-hydrazone (CCCP), dinitrophenol, oligomycin or antimycin resulted in a rapid decrease in cellular ATP within 3 min of addition. The initial uptake rate of TBuMA was generally largely affected by these treatments. However, fructose at 10 mM had no effect at all on the uptake rate of the cation whereas cellular ATP was decreased to an extent comparable to that after treatment with the metabolic inhibitors. Consequently it was hypothesized that the metabolic inhibitors affected the initial cellular uptake rate of organic cations due to either altered intracellular sequestration (e.g. mitochondria) or alternatively to direct effects on the plasma membrane rather than by decreasing cellular ATP. Isolated rat mitochondria were shown to take up organic cations very efficiently. Accumulation in this organelle is probably driven by the negative membrane potential as measured by the uptake of the lipophilic cation [3H]tetraphenylphosphonium. Treatment of the isolated mitochondria with various metabolic inhibitors decreased the membrane potential in parallel to the effects on the uptake of TBuMA. Since mitochondria constitute a considerable intracellular volume, they may contribute largely to the storage of the organic cation in the hepatocyte. In isolated perfused livers, preloaded with either TBuMA or tetraphenylphosphonium (TPP+), the addition of valinomycin or CCCP leads to a marked backflux of the cations from the liver into the perfusion medium. This suggests strongly that a large part of the intracellular storage capacity is lost after metabolic inhibitor treatment, probably as the consequence of dissipation of the mitochondrial membrane potential. Since the metabolic inhibitors in contrast to TBuMA uptake did not decrease the initial uptake rate of TPP+ into isolated hepatocytes, it was concluded that mitochondrial uptake (mitochondria are the major storage sites for TPP+) is not an essential determinant of the initial uptake rate in intact hepatocytes.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenosine Triphosphate↗
PubMed1993

Identification with liquid chromatography-ionspray mass spectrometry of the metabolites of the enantiomers N-methyl dextrorphan and N-methyl levorphanol after rat liver perfusion.

To gather more information on stereochemical factors in the hepatic disposition of organic cations, mass spectrometry coupled to liquid chromatography was used to determine the identity of the metabolites excreted in bile after isolated rat liver perfusions with the quaternary ammonium derivatives of the enantiomeric drugs dextrorphan and levorphanol. Ionspray mass spectrometry was chosen for its soft ionization and absence of thermal degradation of labile compounds. The drugs were labelled with a stable (2H) isotope and mixed with unlabelled drugs to create an artificial isotope pattern in the mass spectrum and facilitate the recognition of unknown metabolites. In mass spectra that were recorded under normal conditions, fragmentation was absent and metabolites of N-methyl dextrorphan and N-methyl levorphanol were visible as parent-ion 'doublets'. Collision-induced fragmentation studies were performed to support the identification of the metabolites. For N-methyl dextrorphan the glucuronide, the glutathione conjugate and the glucuronide of the N-demethylated metabolite were found in bile. For N-methyl levorphanol the glucuronide, the glutathione conjugate, the sulphate conjugate and the glucuronide of a hydroxylated N-methyl levorphanol were excreted in bile. Thus a remarkable stereoselectivity occurs in the metabolism of these quaternary ammonium compounds in the rat liver.

Animals↗
PubMed1993

Pharmacokinetic modeling of the sinusoidal efflux of anionic ligands from the isolated perfused rat liver: the influence of albumin.

This study contains a pharmacokinetic analysis on the efflux of organic anions from the liver into the bloodstream (sinusoidal efflux) with specific reference to the influence of albumin. The net sinusoidal efflux rate of dibromosulfophthalein (DBSP) from preloaded livers, being the resultant of sinusoidal efflux and reuptake of ligand by hepatocytes downstream the sinusoid, can be strongly increased by the presence of bovine serum albumin (BSA), a protein having multiple binding sites for DBSP. We previously attributed this effect to a reduction of reuptake through extracellular binding of the organic anion to the protein, rather than to an intrinsic stimulatory effect on the actual membrane transport process from the cells. In the present study we tested this hypothesis using a pharmacokinetic multicompartment liver model. This model resembles the parallel tube model in that the liver is described by several compartments placed in series instead of a single well-stirred compartment and it takes into account rates of dissociation and association in binding to proteins in the sinusoidal space. The model parameters were fitted from the sinusoidal efflux and biliary excretion data from efflux experiments measuring the stimulatory effect of various concentrations of BSA. Equilibrium binding of DBSP to albumin as well as the dissociation rate constant (koff) were determined in vitro with rapid filtration techniques. The experimental data could not be fitted satisfactorily when using the experimentally obtained values of the protein association and dissociation rate constants (kon and koff). However, they could be simulated accurately assuming 16 times higher values for the association and dissociation rate constant compared to those determined in vitro. Time constants of the perfusate flow, liver (re)uptake, and protein association and dissociation indicate that binding equilibrium does not exist within the sinusoids and that, in particular at low protein concentrations, the net sinusoidal efflux rate is association rate-limited: A large fraction of the ligand effluxed from the cell into the median is taken up by the hepatocyte before binding to the proteins occurs. Higher kon and koff values predicted by the model might indicate altered DBSP-albumin binding characteristics upon passage through the liver but alternatively can be explained by an intrinsic effect of albumin on the carrier-mediated efflux process. Efflux experiments showed a marked stimulatory effect of the protein on sinusoidal efflux but only a moderate effect on biliary excretion, despite a strong decrease in liver content. These patterns indicate that sinusoidal efflux and biliary excretion occur from two different intracellular compartments that equilibrate slowly.

Albumins↗
PubMed1993

The isolated heart-lung preparation in the cat. An in situ model to study the role of the lungs in the disposition of drugs.

In the search for drugs with an extreme short time course of action, compounds should be developed that are rapidly distributed to and temporarily stored in well-perfused organs. Since the lungs receive the complete cardiac output and have the ability to temporarily store drugs, we have developed an in situ, isolated lung preparation in the cat to study the contribution of the lungs to the disposition of drugs. The cat's own heart perfuses the lung in situ with autologous blood. The circulation between the left ventricle and the right atrium is short-circuited via an aorta-caval shunt. The right forelimb is added to study pharmacodynamics simultaneously (only for muscle relaxants). Validation of the model for 180 min of perfusion showed complete isolation of the organs without major biochemical changes or edema and a stable muscle response. In pilot experiments with two structurally related muscle relaxants, initial muscle relaxation was followed by spontaneous recovery of neuromuscular function and a gradually decreasing plasma concentration, indicating partial disposition by the lungs. This was confirmed by direct concentration measurements in the lung. The present model may provide a powerful experimental tool to elucidate the role of the lungs in the disposition of drugs.

Albumins↗
PubMed1993

Low molecular weight proteins as carriers for renal drug targeting: naproxen coupled to lysozyme via the spacer L-lactic acid.

Low molecular weight proteins (LMWPs) are potential carriers for targeting drugs to the kidney. To test whether ester bonds are suitable for the reversible drug conjugation, the antiinflammatory drug naproxen (Nap) was conjugated to the LMWP lysozyme (LYSO) via an ester bond using an L-lactic acid spacer (Nap-lact-LYSO, 1:1:1). The distribution and degradation of the conjugate in rats were compared to those of an equimolar mixture of free drug and LMWP and of a directly coupled conjugate without spacer (Nap-LYSO). The plasma clearance of Nap-lact-LYSO closely resembled that of Nap-LYSO and LYSO itself. Its major accumulation site appeared to be the kidney as demonstrated by extracorporal gamma-camera counting of the LMWP. Renally released naproxen was excreted in the urine as 6-desmethyl-naproxen-sulfate (6-DMN-S). Apparently the kidneys represent the main sites of demethylation and sulfation after administration of the LMWP-coupled drug. In addition, the renal excretion of naproxen (including its metabolites) was significantly delayed and sustained as compared to that after injection of uncoupled naproxen. Using the L-lactic acid spacer LMWP conjugation, the renal selectivity of Nap was increased 5.6 +/- 0.41-fold. Additional in vitro studies with Nap-lact-LYSO revealed that renal generation of the parent drug coincided with formation of low molecular weight catabolites, mainly as naproxen-L-lactic acid-lysine (Nap-lact-Lys). This indicated that in vitro the rate of cleavage of the ester bond is significantly slower than digestion of the carrier backbone itself.(ABSTRACT TRUNCATED AT 250 WORDS)

Anesthesia↗
PubMed1993

Pharmacokinetic analysis and cellular distribution of the anti-HIV compound succinylated human serum albumin (Suc-HSA) in vivo and in the isolated perfused rat liver.

After intravenous injection of a low dose (25 micrograms/kg) in rats, the anti HIV-1 compound succinylated human serum albumin (Suc-HSA) is taken up mainly in the liver and spleen and is proteolytically degraded. Ten minutes after injection of 125I-Suc-HSA, 72 and 14% of the dose were found in the liver and spleen, respectively. With immunohistochemistry we demonstrated that in both organs, Suc-HSA was specifically endocytosed in endothelial cells. In the isolated perfused rat liver preparation, liver uptake was shown to be saturable, with a Km of 2.9 10(-8) M and a Vmax of 2.4 micrograms/min/100 g body weight. The apparent Km and Vmax in vivo were 2.2 10(-7) M and 10.3 micrograms/min/100 g, respectively. Uptake in liver and spleen was inhibited by preadministration of an excess of formaldehyde-treated albumin and with polyinosinic acid, indicating the involvement of the scavenger receptor, as anticipated for such polyanionic compounds. Suc-HSA is not absorbed intact from the colon and the ileum. After injecting (i.v.) rats with a high dose of Suc-HSA (10 mg/kg), the elimination t1/2 was 3 hr, and therefore, sustained plasma levels above the concentration needed for in vitro anti-HIV-1 activity can be achieved.

Animals↗
PubMed1993

Coupling of the antiviral drug ara-AMP to lactosaminated albumin leads to specific uptake in rat and human hepatocytes.

We covalently coupled 9-beta-D-arabinofuranosyladenine 5'-monophosphate (ara-AMP) to the carrier molecule lactosaminated human serum albumin using a water-soluble carbodiimide with a two-step conjugation method (pH 4.5 and pH 7.5) instead of the commonly used single-step conjugation at pH 7.5. This resulted in a predominantly monomeric conjugate (lac27-HSA-ara-AMP9). The conjugate was stable in buffer (pH 7.4) and blood plasma. After in vivo injection, the carrier and the monomeric conjugate were subjected to selective endocytosis in rat hepatocytes, as shown on immunohistochemical study and cell-separation techniques using 125I-labeled material. In competition experiments with other ligands for the asialoglycoprotein receptor N-acetylgalactosamine and asialofetuin, we showed that both lactosaminated human serum albumin and lac27-HSA-ara-AMP9 are subject to endocytosis by this receptor system. Although the coupling of ara-AMP significantly increased the net negative charge of the conjugate compared with the native carrier, liver uptake was not affected by coadministration of an excess of succinylated human serum albumin (suc-HSA), a negatively charged ligand for the scavenger receptor. Incubation studies with purified rat liver lysosomes showed that in this acidic and proteolytic environment, mainly ara-AMP and, to a much lesser extent, ara-A itself were released from the carrier. After injection into the rat in vivo and in isolated perfused rat liver, no free ara-AMP or 9-B-D-arabinofuranosyladenine (ara-A) could be detected in plasma and perfusate, respectively, indicating proper retention of the virally active components in hepatocytes.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗