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Biomedical subjects

H Jansen

Publications and source records attributed to H Jansen.

At least 91 records · Page 5Linked to original sources
PubMed1994

Quantitative flow cytometric analysis of opsonophagocytosis and killing of nonencapsulated Haemophilus influenzae by human polymorphonuclear leukocytes.

Since nonencapsulated Haemophilus influenzae persists in the lower respiratory tracts of patients with chronic bronchitis despite the presence of specific antibodies, complement, and polymorphonuclear leukocytes (PMNs), opsonophagocytosis of H. influenzae was analyzed. Nonencapsulated H. influenzae isolated from the sputa of chronic bronchitis patients was labeled with fluorescein isothiocyanate and incubated with human PMNs in the presence of complement and antibodies for 30 min at 37 degrees C. Candida albicans was added to each sample as an internal standard, and the reduction of the number of bacteria was determined by flow cytometry. Fluorescence quenching with ethidium bromide was used to discriminate between intracellular and extracellular bacteria. Opsonophagocytosis of viable H. influenzae d1 was 17% +/- 29% in the presence of complement and human pooled sera containing high titers of strain-specific antibodies. Opsonophagocytosis of six other H. influenzae strains was also poor. Under the same conditions, opsonophagocytosis of Staphylococcus aureus was 90% +/- 5%, and opsonophagocytosis of C. albicans was 55% +/- 23%. About half of the number of H. influenzae bacteria associated with PMNs was internalized. Opsonophagocytosis of heat-killed H. influenzae d1 (41% +/- 20%) was higher than that of viable bacteria of the same strain (P < 0.05). This result suggests that the accessibility of epitopes on H. influenzae for opsonizing antibodies is better on killed than on viable bacteria. We conclude that viable nonencapsulated H. influenzae is poorly opsonophagocytized in the presence of strain-specific antibodies and complement.

Bacterial Capsules↗
PubMed1994

Opsonic antibodies to outer membrane protein P2 of nonencapsulated Haemophilus influenza are strain specific.

The ability of monoclonal antibodies (MAbs) specific for variable and conserved epitopes of outer membrane protein (OMP) P2 (b,c) of nonencapsulated Haemophilus influenza to promote opsonophagocytosis of this bacterium by human polymorphonuclear leucocytes (PMNs) was determined by flow cytometry. MAbs rendering PMNs fluorescent because of association with fluorescein isothiocyanate-labelled bacteria were defined as stimulating opsonophagocytosis. Opsonophagocytosis was dependent on the presence of both antibodies and complement. Of the 14 MAbs directed to the variable parts of OMP P2 (L. van Alphen, P. Eijk, L. Geelen-van den Broek, and J. Dankert, Infect. Immun. 59:247-252, 1991), 9 stimulated opsonophagocytosis. Four of the five nonopsonophagocytic MAbs that were immunoglobulin G1 were unable to cause complement activation. The MAbs promoting opsonophagocytosis included MAbs specific for one or more OMP P2 antigenic variants of H. influenzae strains isolated from patients with chronic bronchitis during persistent infection. MAbs cross-reacting in enzyme-linked immunosorbent assays with nonrelated H. influenzae did not promote opsonophagocytosis of strains from other patients. Opsonophagocytosis was not observed in the presence of three MAbs reacting with OMP P2 epitopes common in H. influenzae. These results indicate that OMP P2-dependent opsonophagocytosis of nonencapsulated H. influenzae is strictly strain specific.

Animals↗
PubMed1994

Hepatic lipase gene is transcribed in rat adrenals into a truncated mRNA.

Rat adrenals contain a lipase activity that is indistinguishable from hepatic lipase (HL) present in liver. Expression of HL mRNA in adrenals was studied using the method of reverse transcription-polymerase chain reaction (RT-PCR). A 596-bp fragment of HL cDNA spanning exons 5 to 8 was amplified when using total RNA from rat adrenals and liver, but not from heart or kidney. The abundance of HL mRNA was quantified by competitive RT-PCR using a standard RNA that was generated in vitro by transcription from a deleted HL cDNA construct. Adrenals contained 0.4 attomoles of HL mRNA per microgram of total RNA, compared to 16 attomoles in liver. In hypertrophic adrenals isolated from corticotrophin-treated rats, the abundance also amounted to 0.4 attomoles of mRNA per microgram of total RNA. However, amplification of full-length cDNA from either control or hypertrophic adrenals was never observed. Detailed analysis by PCR using different combinations of primers indicated that exons 3 to 9 including the 3'-untranslated region were expressed in adrenal RNA, but not the first two coding exons. The upstream part of the adrenal lipase mRNA was cloned after rapid amplification of cDNA ends (RACE). The resulting clones showed a unique 126-bp sequence 5' of the exon 2-exon 3 junction. This sequence contained multiple termination codons in all three reading frames but lacked a potential start codon. RT-PCR using an HL-specific primer and an oligonucleotide directed against this 5'-sequence showed that it is not only expressed in RNA from adrenals but also in liver. Pulse-labeling of freshly isolated adrenocortical cells with [35S]methionine followed by immunoprecipitation with anti-HL antibodies failed to show synthesis of mature HL, but indicated the synthesis of immunoreactive proteins in the 40-45 kDa range that remained mainly intracellular. Hence, the HL gene is transcribed in adrenals but results in an mRNA species with a unique 5'-end. Translation from an internal start site may produce an intracellular HL isoform that differs markedly from the liver-type lipase previously identified in adrenals.

Adrenal Cortex↗
PubMed1994

A compound heterozygote for lipoprotein lipase deficiency, Val69-->Leu and Gly188-->Glu: correlation between in vitro LPL activity and clinical expression.

We analyzed the molecular defects in the lipoprotein lipase gene of a patient with type I hyperlipidemia suffering from recurrent pancreatitis, indicative for lipoprotein lipase deficiency. Postheparin lipoprotein lipase activity in the patient was decreased by 70%. Direct genomic sequencing revealed compound heterozygosity for two mutation: the well-known Gly188-->Glu and a new Val69-->Leu substitution. Val69 is situated in a conserved hydrophobic region of the lipoprotein lipase protein, and the substitution with leucine gives rise to a 80% decrease in specific catalytic activity, as supported by site-directed mutagenesis experiments, followed by expression in COS-cells. The combination of both defects in the lipoprotein lipase gene was incidentally associated with severe clinical expression of disease, and triglyceride levels of more than 30 mmol/l were measured. In our patient, triglyceride levels wer usually below 10 mmol/l. We, therefore, postulate that the residual LPL activity in our patient is usually sufficient to keep the triglyceride level within bounds and expression of disease occurred only when conditions such as alcohol abuse or poor compliance to diet were present.

Adult↗
PubMed1993

Low density lipoprotein receptor internalizes low density and very low density lipoproteins that are bound to heparan sulfate proteoglycans via lipoprotein lipase.

It has previously been shown that lipoprotein lipase (LPL) enhances the binding of low density lipoproteins (LDL) and very low density lipoproteins (VLDL) to HepG2 cells and fibroblasts, up to 80-fold. This increase in binding is LDL receptor-independent and is due to a bridging of LPL between extracellular heparan sulfate proteoglycans (HSPG) and the lipoproteins. In the present paper, we show that preincubation of the cells with LPL, followed by washing prior to the binding experiment, increased binding to the same extent as occurs when the binding is performed in the presence of LPL. This indicates that the formation of a complex of LPL with the lipoproteins is not a prerequisite of binding. Binding curves and Scatchard analyses reveal that both the number of binding sites and the affinity of the binding are increased 20-30-fold by the addition of 3.4 micrograms/ml LPL. The addition of LPL also resulted in an enhanced uptake and subsequent lysosomal degradation of both LDL and VLDL when compared with binding, although to a lesser extent (up to 25-fold when measured after 5 h at 37 degrees C). Strikingly, enhanced uptake did not occur in LDL receptor-negative fibroblasts. In addition, down-regulation of the LDL receptor activity by preincubation of the cells for 48 h with either LDL or beta-VLDL resulted in a parallel decrease in the uptake of LPL-mediated HSPG-bound LDL, whereas the LPL-mediated binding itself was not diminished. These observations indicate that the uptake of LPL-mediated HSPG-bound LDL and VLDL mainly proceeds via the LDL receptor. Binding of labeled LDL to the cells at 4 degrees C for 2 h followed by a chase period at 37 degrees C revealed that in absolute terms, the initial rate of internalization of HSPG-bound LDL is comparable with that of LDL receptor-bound LDL (0.58 and 0.44 ng/min/mg of cell protein, respectively). We conclude that in LDL receptor-positive cells, the LPL-mediated binding of LDL and VLDL to HSPG is followed by internalization of the lipoproteins mainly through the rapid process of the classical LDL receptor recycling system, whereas only a minor portion is internalized via the much slower process of HSPG uptake.

Animals↗
PubMed1993

Simvastatin improves chylomicron remnant removal in familial combined hyperlipidemia without changing chylomicron conversion.

It is unknown whether the clearance of atherogenic chylomicron remnants and the postprandial lipoprotein metabolism in general can be improved by 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors in subjects with familial combined hyperlipidemia (FCH). Therefore, the postprandial chylomicron remnant clearance was studied in nine normolipidemic untreated controls and seven FCH patients before and after treatment with simvastatin using an oral vitamin A-fat load (24 hours, 50 g/m2). Treatment with simvastatin reduced plasma cholesterol level by 16% (mean +/- SEM, 8.1 +/- 0.8 v 6.8 +/- 0.8 mmol/L; P < .05) and plasma apolipoprotein (apo) B level by 19% (1.6 +/- 0.2 v 1.3 +/- 0.2 g/L; P < .05). Plasma apo E level (89.6 +/- 21.0 mg/L) was reduced by 29% (63.5 +/- 14.1 mg/L; P < .05). High-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) levels did not change; consequently, the reductions seen had been due to a decrease in very-low-density lipoprotein (VLDL) levels. Fasting plasma triglyceride (30% reduction) and plasma apo C-II (31% reduction) levels did not change significantly. Mean postheparin plasma lipoprotein lipase (LPL) activity increased by 13% after treatment (90.4 +/- 19.8 v 102.6 +/- 20.3 mU/mL; P < .05), but hepatic lipase (HL) activity was not altered. The clearance of chylomicrons (Sf > 1,000), expressed as the area under the 24-hour retinyl palmitate curve, did not change with simvastatin (52.8 +/- 12.9 v 51.8 +/- 13.4 h.mg-1/L).(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

Growth hormone normalizes hepatic lipase in hypothyroid rat liver.

The effects of growth hormone (GH) administration on hepatic lipase (HL) activity and mRNA levels were studied in pair-fed hypothyroid rats. In the hypothyroid state, liver HL mRNA levels and liver and postheparin plasma HL activities were decreased. In hypothyroid rats, GH normalized HL mRNA levels and HL activity in postheparin plasma; GH also increased the activity of HL in the liver, which remained lower than that in controls. These data indicate that at least part of the decrease of HL activity during hypothyroidism is due to the concomitant GH deficiency, and that GH rather than thyroid hormone may regulate HL mRNA levels.

Analysis of Variance↗
PubMed1993

Lipoprotein(a) plasma concentrations associated with lipolytic activities in eight kindreds with familial combined hyperlipidemia and normolipidemic subjects.

The relationship between lipoprotein(a) [Lp(a)] and metabolism of triglyceride-rich lipoproteins (TRL) was studied in 58 untreated patients with familial combined hyperlipidemia (FCH) from eight different kindreds, 17 spouse controls, and 17 unrelated controls. Lp(a) plasma concentrations were not significantly different between FCH subjects (343 +/- 61 mg/L, mean +/- SEM) and controls (249 +/- 52 mg/L). In FCH, log-transformed Lp(a) levels correlated positively with postheparin lipoprotein lipase ([LPL] r = .61, P = .0002) and hepatic lipase ([HL] r = .46, P = .008) activities and total plasma cholesterol level (r = .30, P = .03). In controls, Lp(a) correlated with LPL (r = .50, P = .04) and total plasma cholesterol level (r = .51, P = .003). In eight FCH patients, treatment with the 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor simvastatin resulted in significantly increased mean LPL activities and plasma Lp(a) concentrations. In three of these FCH patients, repeated measurements during 1 year demonstrated that changes in Lp(a) concentrations were paralleled by similar changes in LPL activity, but not HL activity. The observed correlation between postheparin plasma lipolytic activities and Lp(a) plasma concentrations suggests a connection between the metabolism of TRL and Lp(a).

Adult↗
PubMed1993

Different clearance of intravenously administered olive oil and soybean-oil emulsions: role of hepatic lipase.

The elimination of two intravenously administered fat emulsions consisting of either 20% (wt:vol) soybean oil or 17% olive oil plus 3% soybean oil was studied in six normolipidemic young men according to a randomized crossover protocol. Slower elimination was found with the olive oil emulsion. A significantly lower maximal removal capacity (K1) and fractional catabolic rate (K2) were measured with olive oil emulsion (P < 0.05). Removal of olive oil emulsion was inversely related to hepatic lipase activity (r = -0.85; P < 0.05). Removal of soybean-oil emulsion was related to the initial plasma triglyceride concentration (r = -0.84; P < 0.05) but not to lipolytic activity. In vivo apolipoprotein C-II binding was similar for both emulsions. Therefore, hepatic lipase activity is more important in the elimination of olive oil emulsions than soybean-oil emulsions. The faster elimination of soybean-oil emulsions suggests an additional elimination pathway, such as the reticuloendothelial system.

Adult↗
PubMed1993

Different postprandial metabolism of olive oil and soybean oil: a possible mechanism of the high-density lipoprotein conserving effect of olive oil.

The postprandial lipoprotein metabolism of two orally administered vitamin A-fat loads consisting of either 20% (wt:vol) soybean oil or 17% olive oil plus 3% soybean oil was studied in six normolipidemic young men according to a randomized crossover design. Mean (+/- SEM) retinyl palmitate concentrations (area under the 24-h curve) were higher in olive oil chylomicrons (97.3 +/- 5.5 mmol.L-1 x h-1), than in soybean-oil chylomicrons (84.0 +/- 10.5 mmol.L-1 x h-1; P < 0.02). Apolipoprotein B-48 concentrations were higher in the olive oil chylomicron remnants with densities (d) of 1.006-1.019 compared with soybean-oil remnants. The slower removal of olive oil chylomicron remnants was correlated to hepatic lipase activity (r = 0.84, P < 0.02). The initial HDL-cholesterol concentration (0.87 +/- 0.17 mmol/L--relatively low but within the normal range for young Dutch men) decreased significantly after ingestion of soybean oil to 0.66 +/- 0.10 mmol/L after 5 and 7 h, but no significant decrease was observed after olive oil ingestion. Soybean oil induced decreases in HDLs correlated inversely with hepatic lipase (r = -0.88, P < 0.02). The results suggested that competition between olive oil chylomicron remnants and HDL for hepatic lipase may have been the underlying mechanism that prevented the postprandial decrease in HDL cholesterol.

Adult↗
PubMed1993

Impaired chylomicron remnant clearance in familial combined hyperlipidemia.

Postprandial chylomicron remnant clearance was studied in six patients with familial combined hyperlipidemia (FCH) and seven control subjects by using an oral retinyl palmitate (RP) fat-loading test. The chylomicron remnant clearance (Sf < 1,000 fraction), expressed as the area under the RP curve (AUC-RP), was delayed in FCH subjects (65.05 +/- 12.84 hours x [mg/L]) compared with control subjects (25.1 +/- 5.4 hours x [mg/L]; p = 0.01). Postprandial lipoprotein particle size and composition in the Sf > 1,000 fraction were different between FCH and control subjects as analyzed by molecular-sieve chromatography. Fasting high density lipoprotein cholesterol was lower in FCH patients (0.54 +/- 0.09 mmol/L) than in control subjects (0.89 +/- 0.05 mmol/L; p < 0.01). Mean plasma postheparin lipoprotein lipase and hepatic lipase activities were similar between FCH patients (94 +/- 25 and 427 +/- 57 milliunits/mL, respectively) and control subjects (126 +/- 16 and 362 +/- 33 milliunits/mL, respectively). In FCH, a 54% reduction (p < 0.05) of plasma triglycerides to 2.63 +/- 0.41 mmol/L by drug treatment resulted in an enhanced, but not normalized, clearance of chylomicron remnants (39.4 +/- 6.0 hours x [mg/L]). Univariate regression analysis revealed that in FCH subjects the changes in fasting plasma apolipoprotein C-III concentrations after therapy were significantly associated with the changes in chylomicron remnant AUC-RP (r = 0.87; p = 0.02). Delayed elimination of atherogenic chylomicron remnants may contribute to the increased risk of premature atherosclerosis in FCH.

Adult↗
PubMed1993

Impaired fatty acid metabolism in familial combined hyperlipidemia. A mechanism associating hepatic apolipoprotein B overproduction and insulin resistance.

To establish whether insulin resistance and/or postprandial fatty acid metabolism might contribute to familial combined hyperlipidemia (FCH) we have examined parameters of insulin resistance and lipid metabolism in six FCH kindreds. Probands and relatives (n = 56) were divided into three tertiles on the basis of fasting plasma triglycerides (TG). Individuals in the highest tertile (TG > 2.5 mM; n = 14) were older and had increased body mass index, systolic blood pressure, and fasting plasma insulin concentrations compared with individuals in the lowest tertile (n = 24). The former also presented with decreased HDL cholesterol and increased total plasma cholesterol, HDL-TG, and apoprotein B, E, and CIII concentrations. Insulin concentrations were positively correlated with plasma apo B, apo CIII, apo E, and TG, and inversely with HDL cholesterol. Fasting nonesterified fatty acids (NEFA) were elevated in FCH subjects compared to six unrelated controls and five subjects with familial hypertriglyceridemia. Prolonged and exaggerated postprandial plasma NEFA concentrations were found in five hypertriglyceridemic FCH probands. In FCH the X2 minor allele of the AI-CIII-AIV gene cluster was associated with increased fasting plasma TG, apo CIII, apo AI, and NEFA concentrations and decreased postheparin lipolytic activities. The clustering of risk factors associated with insulin resistance in FCH indicates a common metabolic basis for the FCH phenotype and the syndrome of insulin resistance probably mediated by an impaired fatty acid metabolism.

Apolipoproteins B↗
PubMed1993

Functional role of N-linked glycosylation in human hepatic lipase: asparagine-56 is important for both enzyme activity and secretion.

Hepatic lipase (HL) and lipoprotein lipase (LPL) are evolutionarily related enzymes that are essential for normal lipoprotein metabolism. While much has been published on the structure-function relationship of LPL, little is known concerning the structural basis of HL action and secretion. Human HL is a glycoprotein and its predicted amino acid sequence contains four putative N-linked glycosylation sites at Asn residues 20, 56, 340, and 375. We studied the role of these residues in the secretion and catalytic activity of hHL by analysis of hHL expressed in stable CHO cell lines. Using site-specific mutagenesis, the wild-type human HL and substitution mutants of each of the four Asn residues were expressed in vitro. The relative sizes of these site-specific mutants indicate that all four putative sites are utilized for glycosylation in CHO cells. Abolition of N-linked glycosylation of three (residues 20, 340, and 375) of the four sites did not affect enzyme secretion or activity. Mutations of Asn-56 to either Gln or Ala resulted in the production of a totally inactive HL which accumulated intracellularly but was not secreted into the culture medium. Therefore, Asn-56 is required for both HL enzyme activity and secretion. The fact that the homologous N-linked glycosylation site (Asn-43) is required for both enzyme activity and secretion for human LPL (Semenkovich et al. 1990. J. Biol. Chem. 265: 5429-5433) indicates that carbohydrate chains at this site are essential for the active conformation and correct folding for secretion of these evolutionarily related lipases. Our observations provide insight into the structural basis of lipase action and secretion.

Animals↗
PubMed1992

Opposite regulation of hepatic lipase and lecithin: cholesterol acyltransferase by glucocorticoids in rats.

Rats were treated with hydrocortisone, dexamethasone or triamcinolone for 4 days. The effect of treatment on hepatic lipase and lecithin:cholesterol acyltransferase (LCAT) mRNA levels and catalytic activities was determined. Hepatic lipase mRNA was not affected by hydrocortisone, but was decreased after dexamethasone (-28%) and triamcinolone (-54%). Hepatic lipase activity followed the same pattern, it was not affected by hydrocortisone and lowered by dexamethasone (-38%) and triamcinolone (-70%). The LCAT mRNA level in the liver was also not affected by hydrocortisone, but increased upon treatment with dexamethasone (+22%) and triamcinolone (+72%). Plasma LCAT, determined with an excess exogenous substrate (designated LCAT-II), tended to decrease after hydrocortisone treatment (-11%) and was higher after dexamethasone (+21%) and triamcinolone (+22%). The plasma cholesterol esterification rate (designated LCAT-I), determined by incubation of the plasma at 37 degrees C, followed the same pattern. The activity ratio of hepatic lipase/LCAT-II decreased from 1 in the controls to 0.51 after dexamethasone and 0.25 in the triamcinolone-treated animals. The plasma HDL cholesterol concentration in the different groups changed oppositely to the hepatic lipase/LCAT activity ratio. It is concluded that HDL cholesterol is raised by synthetic glucocorticoids due, among other factors, to a lowered hepatic lipase and an increased plasma LCAT activity. The influence of glucocorticoids on these enzymes is, at least partly, explained by the effects on the hepatic mRNA contents.

Animals↗
PubMed1992

Heparan sulphate proteoglycans are involved in the lipoprotein lipase-mediated enhancement of the cellular binding of very low density and low density lipoproteins.

We found that LPL enhances the binding to HepG2 cells and fibroblasts of both VLDL and apoE free LDL. In the presence of 1.7 micrograms/ml of purified bovine LPL, the binding of LDL and VLDL was up to 60 fold increased as compared to the control binding. In addition, LPL enhances the binding in LDL-receptor negative fibroblasts to the same extent as it does in normal fibroblasts. The presence of 10 mM of EGTA could not prevent the LPL-mediated enhancement of the binding of both LDL and VLDL to fibroblasts, indicating that the binding is calcium independent. Furthermore, up- and down regulation of the LDL receptor did not influence the binding of these lipoproteins in the presence of LPL. Strikingly, we found that the enhancing effect of LPL on the binding of LDL and VLDL to HepG2 cells could be abolished by preincubation of the cells with heparinase, suggesting that heparan sulphate proteoglycans are involved in the LPL-mediated stimulation. We hypothesize that the enhancement of the cellular binding of LDL and VLDL in the presence of LPL is caused by an LPL-bridging between proteoglycans present on the plasma membrane and the lipoproteins, and that the LDL receptor and LRP are not involved.

Cell Membrane↗