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P Meijer

Publications and source records attributed to P Meijer.

At least 37 records · Page 2Linked to original sources
PubMed1995

Insulin suppresses bile acid synthesis in cultured rat hepatocytes by down-regulation of cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase gene transcription.

Evidence from in vivo studies indicates that the bile acid pool and bile acid excretion are increased in humans with diabetes mellitus and in experimental diabetic animals, and that both parameters return to normal levels after administration of insulin. To investigate the biochemical background of these changes, the effects of insulin on bile acid synthesis and cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase, two key enzymes in routing of cholesterol toward bile acids, were studied in cultured rat hepatocytes. Mass production of bile acids was dose dependently diminished, showing significant reduction (-33% to -53%) at physiological concentrations of the hormone (1.4 to 14 nmol/L) and a maximal decrease at 140 nmol/L (-65%). The decrease of bile acid synthesis correlated well with the suppression of cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase activity. The enzyme activity for cholesterol 7 alpha-hydroxylase, examined in more detail, was dose dependently diminished on incubation of hepatocytes with various concentrations of insulin, reaching maximal reduction at 14 nmol/L of insulin. Maximal decrease of the enzyme activity was seen after 8 hours of incubation (-70%). Insulin strongly reduced the rise in cholesterol 7 alpha-hydroxylase activity induced by incubation with dexamethasone. Sterol 27-hydroxylase activity was inhibited up to -58% after 24 hours of incubation with 140 nmol/L insulin. To study the mechanism of suppression of cholesterol 7 alpha-hydroxylase and sterol 27-hydroxylase activity, the effects of insulin on their respective levels of messenger RNA (mRNA) and gene transcription were assessed. The decrease in enzyme activities could be explained by a concomitant reduction in the cholesterol 7 alpha-hydroxylase (-76%) and sterol 27-hydroxylase (-62%) mRNA level. Transcriptional activity, as assessed by nuclear runoff assays, was decreased to the same extent, i.e., -60% for cholesterol 7 alpha-hydroxylase and -75% for sterol 27-hydroxylase. Transient expression experiments using a construct containing the proximal 348 basepairs of the cholesterol 7 alpha-hydroxylase promoter fused to the chloramphenicol acetyltransferase (CAT) gene (-348Rcat) showed a significant reduction of transcriptional activity (-64%) with insulin, indicating that a sequence important for an insulin-induced transcriptional response is located within the first 348 basepairs, preceding the transcription start of the cholesterol 7 alpha-hydroxylase promoter.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1994

Specificity of antigen assays of plasminogen activator inhibitor in plasma: Innotest PAI-1 immunoassay evaluated.

We evaluate a new commercial enzyme immunoassay (EIA) of plasminogen activator inhibitor-1 (PAI-1) in plasma, the Innotest PAI-1. Because we wanted to measure PAI-1 in blood samples, we developed a procedure for evaluating the specificity of the assay for different PAI-1 forms in their natural environment. All molecular forms were prepared from a plasma that contained only active PAI-1. The recovery of the different molecular forms of PAI-1, relative to active PAI-1 (100%), was 99% +/- 7% for PAI-1 complexed with recombinant tissue-type plasminogen activator (t-PA), 104% +/- 4% for PAI-1 complexed with melanoma t-PA, 94% +/- 11% for PAI-1 complexed with high-M(r) urokinase, and 113% +/- 3% for latent PAI-1. The parallelism between the calibration curve of the EIA and the serial dilutions of the different PAI-1 forms was considered acceptable for clinical purposes. In selected clinical plasma samples, the PAI-1 values obtained with the Innotest PAI-1 EIA correlated well with those of the TintElize PAI-1 EIA (r = 0.913, n = 106); the observed correlation of the Innotest measurements with PAI activity was r = 0.795 (n = 79). The Innotest PAI-1 antigen assay appears to detect all molecular forms of PAI-1 to a similar degree, and comes close to being the so-called grand total assay for detecting the total molecular concentration of PAI-1 in plasma.

Humans↗
PubMed1993

Effect of isotretinoin on endogenous tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor 1 (PAI-1) in humans.

The effect of isotretinoin on fibrinolysis was investigated in 10 healthy, male volunteers in a randomized, double-blind, crossover-designed study. Isotretinoin (40 mg) was administered in the morning and in the evening for 5 days. t-PA, u-PA and PAI-1 antigen and activity in plasma were measured every morning at 9 a.m. on days 1 to 4 and every 3 hours over 24 hours on day 5. Isotretinoin treatment had no significant stimulatory effect on endogenous t-PA antigen and activity in morning plasma samples nor on their circadian variation. Also, u-PA antigen levels did not change after isotretinoin treatment. Mean PAI-1 antigen and PAI activity in 9 a.m. plasma samples were non-significantly higher during isotretinoin than during placebo treatment. After treatment with isotretinoin a significant rise of fasting triglyceride plasma levels was observed as compared to placebo. The study shows that isotretinoin has no clinically significant effect on endogenous fibrinolysis.

Adult↗
PubMed1993

Thrombin generation induced by the intrinsic or extrinsic coagulation pathway is accelerated by streptokinase, independently of plasminogen.

Thrombolytic therapy paradoxically induces the formation of fibrinopeptide A, fibrin degradation products and thrombin-antithrombin complexes, indicating thrombin generation. Part of the mechanism of this thrombin generation under the influence of thrombolytic agents was unraveled in this study. We measured thrombin with a chromogenic substrate at several time intervals after recalcification of citrated plasma which had been preincubated with urokinase, streptokinase, recombinant tissue plasminogen activator (rt-PA) or recombinant single-chain urokinase-type plasminogen activator (rscu-PA). Thrombin generation induced by the addition of thromboplastin together with calcium (extrinsic pathway) was greatly accelerated in the presence of streptokinase (from about 7 to 2 min), and to a lesser extent in the presence of urokinase, rt-PA or rscu-PA. Similar effects were seen after the addition of calcium to the plasma containing the thrombolytic agent and preincubated with partial thromboplastin (intrinsic pathway). Hirudin quenched the conversion of the chromogenic substrate completely, confirming that thrombin was the active enzyme. Aprotinine did not affect the results, and the effect of streptokinase was also observed in plasminogen-depleted plasma. We conclude that streptokinase, and to a lesser extent other thrombolytic agents, activate the prothrombinase complex directly or indirectly through a calcium-dependent mechanism, independently of plasminogen, with a resulting acceleration of thrombin generation.

Blood Coagulation↗
PubMed1993

A self-rating scale for the assessment of posttraumatic stress disorder in Dutch Resistance veterans of World War II.

The present study reports on the development of a Dutch PTSD scale based on the DSM-III criteria for PTSD. Test-retest reliability was .91. The scale showed an internal consistency with a coefficient alpha of .88. Factor analysis on a large sample of Resistance veterans (N = 967) yielded six factors, which represent intrusive thoughts, physiological reactions, detachment, rage, active confrontation, and guilt.

Aged↗
PubMed1991

Heterogeneity of rat liver parenchyma in cholesterol 7 alpha-hydroxylase and bile acid synthesis.

Periportal and perivenous hepatocytes were isolated from rat liver by digitonin/collagenase perfusion for investigating the acinar distribution of bile acid synthesis. The specific activity of cholesterol 7 alpha-hydroxylase (EC 1.14.13.17) was 7.9-fold higher in perivenous cells than in periportal hepatocytes. Mass production of bile acids differed 4.4-fold between cultured perivenous and periportal hepatocytes. In contrast, the levels of free cholesterol in homogenates and microsomes derived from both subfractions were similar. Feeding of rats with the bile-acid-sequestering anion-exchange resin colestid resulted in a pronounced stimulation of cholesterol 7 alpha-hydroxylase activity and bile acid mass production, but decreased the perivenous/periportal ratio of both parameters. These results demonstrate that bile acid mass production, but decreased the perivenous hepatocytes, possibly owing to feedback suppression by bile acids from the enterohepatic circulation. Furthermore, the opposite acinar localization of cholesterol and bile acid biosynthesis provides an interesting alternative to current views of the regulation of their metabolic pathways.

Animals↗
PubMed1991

Cyclosporin A blocks bile acid synthesis in cultured hepatocytes by specific inhibition of chenodeoxycholic acid synthesis.

Bile acid synthesis, determined by conversion of [4-14C]cholesterol into bile acids in rat and human hepatocytes and by measurement of mass production of bile acids in rat hepatocytes, was dose-dependently decreased by cyclosporin A, with 52% (rat) and 45% (human) inhibition of 10 microM. The decreased bile acid production in rat hepatocytes was due only to a fall in the synthesis of beta-muricholic and chenodeoxycholic acids (-64% at 10 microM-cyclosporin A), with no change in the formation of cholic acid. In isolated rat liver mitochondria, 26-hydroxylation of cholesterol was potently inhibited by the drug (concn. giving half-maximal inhibition = 4 microM). These results suggest that cyclosporin A blocks the alternative pathway in bile acid synthesis, which leads preferentially to the formation of chenodeoxycholic acid.

Animals↗
PubMed1990

[Canine scabies in man].

Parasitic prurigo caused by scabies mites from recently acquired puppies is described in two patients and their family members. Because Sarcoptes scabiei var. canis (dog) occasionally survives in man, patients with persisting complaints despite treatment of their dogs should be treated with the topical scabicide lindane for one night.

Adult↗
PubMed1990

Maintenance of bile acid synthesis and cholesterol 7 alpha-hydroxylase activity in cultured rat hepatocytes.

Addition of foetal-bovine serum to rat hepatocytes cultured in Williams E medium resulted in improved maintenance of bile-acid-synthetic capacity and cholesterol 7 alpha-hydroxylase activity as compared with cultures supplemented with rat or newborn-bovine serum or cultures in a hormonally defined serum-free medium. Minimally, 5% (v/v) foetal-bovine serum was necessary to maintain these liver-specific functions. Serum factor(s) responsible for these effects were not dialysable or associated with lipoproteins, but were removed by charcoal extraction.

Animals↗
PubMed1990

[Contact allergy for Alstrumeria (inca lily)].

We report 6 patients with occupational contact allergy to Alstroemeria cultivars. Four of them presented with the clinical picture of 'tulip fingers'. They all reacted to parts of fresh plants and to tuliposide A. The literature on Alstroemeria allergy is reviewed.

Adolescent↗
PubMed1989

Dexamethasone regulates bile acid synthesis in monolayer cultures of rat hepatocytes by induction of cholesterol 7 alpha-hydroxylase.

To study the effect of steroid hormones on bile acid synthesis by cultured rat hepatocytes, cells were incubated with various amounts of these compounds during 72 h and conversion of [4-14C]cholesterol into bile acids was measured. Bile acid synthesis was stimulated in a dose-dependent way by glucocorticoids, but not by sex steroid hormones, pregnenolone or the mineralocorticoid aldosterone in concentrations up to 10 microM. Dexamethasone proved to be the most efficacious inducer, giving 3-fold and 7-fold increases in bile acid synthesis during the second and third 24 h incubation periods respectively, at a concentration of 50 nM. Mass production of bile acids as measured by g.l.c. during the second day of culture (28-52 h) was 2.2-fold enhanced by 1 microM-dexamethasone. No change in the ratio of bile acids produced was observed during this period in the presence of dexamethasone. Conversion of [4-14C]7 alpha-hydroxycholesterol, an intermediate of the bile acid pathway, to bile acids was not affected by dexamethasone. Measurement of cholesterol 7 alpha-hydroxylase activity in homogenates of hepatocytes, incubated with 1 microM-dexamethasone, showed 10-fold and 90-fold increases after 48 and 72 h respectively, as compared with control cells. As with bile acid synthesis from [14C]cholesterol, no change in enzyme activity was found in hepatocytes cultured in the presence of 10 microM steroid hormones other than glucocorticoids. Addition of inhibitors of protein and mRNA synthesis lowered bile acid production and cholesterol 7 alpha-hydroxylase activity and prevented the rise of both parameters with dexamethasone, suggesting regulation at the mRNA level. We conclude that glucocorticoids regulate bile acid synthesis in rat hepatocytes by induction of enzyme activity of cholesterol 7 alpha-hydroxylase.

Animals↗
PubMed1989

Comparison of taurocholate accumulation in cultured hepatocytes of pig, rat and man.

Intracellular accumulation at 37 degrees C of 50 microM [14C]taurocholic acid by hepatocytes of pig and rat, cultured for 24 hours, and by human hepatocytes, cultured for 12 hours, reached equilibrium after an incubation time of 1 to 2 hours. Maximum capacity to accumulate taurocholate intracellularly was assessed in 3-hour incubations with increasing extracellular taurocholate concentrations. Accumulation capacity of pig and rat hepatocytes was saturated at 100 microM, while uptake by human hepatocytes slightly increased even further above this concentration. At extracellular concentrations of 100 to 500 microM, hepatocytes of these three species concentrated taurocholic acid intracellularly to between 13 and 17 nmol per mg cell protein, corresponding to an intracellular concentration which was 10-70 times higher than the added extracellular concentration. With proceeding culture age, accumulation capacity of rat and human hepatocytes declined steeply (-80% and -60%, respectively between the first and second culture day). In contrast, in cultured pig hepatocytes, this capacity was only 40% lower on the third day compared to the first day of culture. It is concluded that in cultured pig hepatocytes, the capacity to accumulate bile acids is retained for a longer time than in cultured rat and human hepatocytes.

Animals↗
PubMed1988

Hydroxylation, conjugation and sulfation of bile acids in primary monolayer cultures of rat hepatocytes.

Hydroxylation of lithocholic, chenodeoxycholic, deoxycholic and cholic acids was studied in monolayers of rat hepatocytes cultured for 76 h. The majority of added lithocholic and chenodeoxycholic acids was metabolized to beta-muricholic acid (56-76%). A small part of these bile acids (9%), however, and a considerable amount of deoxycholic and cholic acids (21%) were converted into metabolites more polar than cholic acid in the first culture period. Formation of these compounds decreased during the last day of culture. Bile acids synthesized after addition of [4-14C]-cholesterol were almost entirely (97%) sulfated and/or conjugated, predominantly with taurine (54-66%), during culture. Sulfated bile acids were mainly composed of free bile acids. The ability of hepatocytes to sulfurylate bile acids declined with culture age. Thus, rat hepatocytes in primary monolayer culture are capable to sulfurylate bile acids and to hydroxylate trihydroxylated bile acids, suggesting formation of polyhydroxylated metabolites.

Animals↗
PubMed1988

Assay of cholesterol 7 alpha-hydroxylase activity in rat hepatocytes in primary monolayer culture.

A sensitive and precise method is described to assay cholesterol 7 alpha-hydroxylase activity in homogenates of rat hepatocytes cultured in monolayers for up to 76 h. The assay is based on measurement of the amount of radioactive cholesterol converted into 7 alpha-[14C]-hydroxycholesterol. Since no subcellular fractionation was applied to measure enzyme activity, this method is rapid and can be performed with cell protein, corresponding to as little as 1 to 2 million hepatocytes. Optimal assay conditions were determined and the reproducibility of this cholesterol 7 alpha-hydroxylase determination was established. Exogenous cholesterol (105 microM), solubilized in Tween 80, was added to saturate the enzyme, giving an apparent Km of 56 microM. Under these conditions, 70% of the cholesterol present in the homogenates is directly accessible to the cholesterol 7 alpha-hydroxylase. The detection limit of the assay was found to be about 10 pmol per incubation. A time course of the cholesterol 7 alpha-hydroxylase activity in cultured hepatocytes revealed that after an initial loss of approximately 60% of the activity as compared with 287 pmol/h/mg for freshly isolated cells, the enzyme activity was increased to the initial level in hepatocytes cultured for 52 h. This result and the finding that the cholesterol 7 alpha-hydroxylase activity was diminished by 94% after a 24-h incubation with 5 microM cycloheximide suggest that the enzyme activity is associated with de novo protein synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗