Lambert Dikkeschei

Background: In patients with acute myocardial infarction, estimation of infarct size by cumulative lactate dehydrogenase release at 72 h (LDHQ72) is a simple and widely used method. Our objective was to study the value of estimating infarct size, by the cumulative release of LDH over 72, 60, 48 and 36 h in predicting left ventricular ejection fraction (LVef) and cardiac death at 1 year. Methods: In the Zwolle Infarction Study infarct size estimated as LDHQ was calculated in 1224 patients treated with primary percutaneous coronary intervention for acute myocardial infarction between December 1993 and June 2001. Patients were categorized as having small (LDHQ72<800 U/L), medium (LDHQ72 800-2500 U/L) or large (LDHQ72>2500 U/L) myocardial infarction. Results: LDHQ72 was closely correlated with LDHQ60, LDHQ48 and LDHQ36 ( r = 0.998, 0.993 and 0.987, respectively, P <0.0001). The relations between LDHQ infarct size classification and mean LVef (51% vs 45% vs 35%, P <0.001) or ...

The mass fragmentographic determination of medroxyprogesterone acetate (MPA) in serum, using as internal standard medroxyprogesterone propionate (MPP) synthesized from MPA, is described. After addition of MPP, the sera are extracted on Sep-Pak C18 cartridges and MPA and MPP are detected as their respective 3-enol trifluoroacyl esters. Serum samples from 84 patients with breast cancer, daily receiving MPA orally, were determined showing a large variation in MPA concentrations (4-349 ng/ml). Our proposed gas chromatographic-mass spectrometric (GC-MS) method, which can be considered as a reference, was compared with a radioimmunoassay (RIA) method showing a correlation coefficient of 0.73 (n = 69; p much less than 0.001). The assay was also used to determine sequential serum levels of patients receiving a single oral dose of MPA. With only minor adjustments, the GC-MS method allows the determination of serum concentrations of related steroids such as megestrol acetate and cyproterone a...

A reversed-phase high-performance liquid chromatographic method with ultraviolet detection of megestrol acetate and cyproterone acetate in human sera is described. The proposed assay is linear up to 1400 ng/ml (r = 0.999) and has a detection limit of 5 ng/ml. Recoveries of both compounds in spiked sera were ca. 95%; inter-assay coefficients of variation were 4.0 and 3.1% and intra-assay values were 1.3 and 1.4%, respectively. For validation of the method we also developed a gas chromatographic-mass spectrometric method for both steroids. The results obtained by the two methods showed good correlation: for megestrol acetate r = 0.98, n = 31, p less than 0.0001, and for cyproterone acetate r = 0.94, n = 0, p less than 0.0001. Large inter-individual differences in the serum concentrations of both substances were found in groups of patients with metastatic breast cancer receiving the same oral load of either steroid.

The tritium water release assay, originally described for the analysis of aromatase activity in placental tissue, was used to estimate aromatase activity in breast tissue samples. The lower activity in this tissue necessitates longer incubation times and thus optimization of the assay conditions. To prevent oxidative and proteolytic inactivation of aromatase, dithiothreitol and albumin were added to the incubation mixture. Extra NADPH, cofactor in the aromatase reaction, also improved reaction rate in placental incubations, but after approximately 120 min activity rapidly decreased. Inhibitors gradually produced during the incubation could explain this phenomenon. Quantitative gas chromatography-mass spectrometry (GC-MS) analyses of testosterone, oestradiol, oestrone and androstenedione after incubation with non-labelled androstenedione proved that a substantial amount of the substrate is converted into testosterone. Qualitative GC-MS steroid profiling of the incubation mixture demonstrated the presence of hydroxylated oestradiol and hydroxylated testosterone, produced during incubation, which could have caused partial aromatase inhibition. The adjusted assay was used to analyse 84 breast tissue samples, histologically classified as normal, adenoma or carcinoma. Aromatase activity was found in 56% of all samples and ranged from 0.6 to 26 pmol oestrogen/g protein per hour. Aromatase positivity was found in 80% of the normal samples, 56% of the adenoma samples and 48% of the carcinoma samples. Although carcinoma samples were less often aromatase positive than normal tissue samples (chi2 = 4.80; P < 0.050) there was no difference in absolute aromatase activity. Because no less than approximately 50% of the carcinomas contained aromatase activity and because of the non-routine character of the assay we conclude that it is justified to start aromatase inhibition therapy without previous knowledge of the aromatase status.

A phase II study with cyproterone acetate (CPA) was done as the primary treatment in female breast cancer patients. Twenty-three patients, mean age 64 years, range 52-75 years, were entered and treated with CPA 400 mg daily. Twenty patients were evaluable and responses were sparse. There was one partial and one complete remission, 17 patients were stable and one patient progressed within 3 months. Side-effects were frequent: five patients complained of nausea, three had severe weight loss, one suffered from depression and seven showed disturbed liver function tests. Six patients had to stop treatment for side-effects, while two other patients were taken off treatment because they developed an acute necrotizing hepatitis. The hepatitis recovered after drug withdrawal in both patients. The serum levels of CPA, cortisol, androstenedione, DHAS, LH, FSH and prolactin were measured during CPA treatment. The levels of cortisol and androstenedione did not change, while LH, FSH and DHAS were suppressed. The DHAS showed an inverse relation to serum CPA concentrations. The prolactin levels rose uniformly. The therapeutic effect of CPA in postmenopausal patients with advanced breast cancer is disappointing, and inferior to that of other progestins. Side-effects are frequent, possibly as a result of the high dosage used in this study. The hormonal changes are different from those of other progestins, which may explain the different efficacies.

We developed an assay for delta-5-androstenediol (adiol) and delta-5-androstenediol-3-sulphate (adiol-3S) in serum and adiol and total adiol sulphate (adiol-S) in urine. An analytical procedure using HPLC and gas chromatography-mass spectrometry was devised and tested for its reliability. After addition of deuterated androstenediol as internal standard, serum and urine samples were extracted. Steroid sulphates were hydrolysed. The extracts and hydrolysates were purified on HPLC, adiol was derivatized using heptafluorobutyric anhydride and finally quantified by gas chromatography-mass spectrometry. The assay is accurate and reproducible. The coefficient of variation (CV) for the determination of adiol in serum samples is 4% (intra-assay) and 9% (interassay) and for urine samples 3 and 8% respectively. The intra-assay CV for the adiol-3S analyses is 5% for serum and 2% for urine samples while the interassay CV values for adiol-3S are 10% for serum and 7% for urine samples. The recovery of adiol and adiol-3S from serum and urine samples is 97%. The developed assay meets the analytical demands needed for clinical applications.

Wolfram syndrome patients are mainly characterised by juvenile onset diabetes mellitus and optic atrophy. A synonym is the acronym DIDMOAD: diabetes insipidus, diabetes mellitus, optic atrophy, deafness. Diabetes insipidus and sensorineural high-frequency hearing impairment are important additional features. This rare autosomal recessively inherited neurodegenerative syndrome is caused by mainly inactivating mutations in the WFS1 gene. It is located at chromosome 4p16 and encodes wolframin, a transmembrane protein. No function has yet been ascribed to this protein.

Dyslipidemia contributes to increased cardiovascular risk in nephrotic syndrome. We questioned whether reduction in proteinuria not only lowers low-density lipoprotein cholesterol (LDL-C), but also high-density lipoprotein cholesterol (HDL-C) and cholesteryl ester transfer protein (CETP) mass and whether changes in HDL-C were related to changes in plasma adiponectin. Thirty-two non-diabetic proteinuric patients (12 on statin therapy), were followed during two double blind 6-week periods of placebo and treatment (low sodium + 100mg losartan + 25 mg hydrochlorothiazide). With placebo HDL-C was lower but LDL-C and CETP were not different in proteinuric patients compared with matched controls. LDL-C, HDL-C and CETP decreased upon proteinuria reduction. The decrease in LDL-C correlated with the drop in CETP and the degree of proteinuria reduction. HDL-C also decreased in proportion to proteinuria lowering. Individual changes in HDL-C were correlated with changes in adiponectin. LDL-C lowering upon robust reduction of proteinuria may be affected by changes in plasma CETP mass, but this treatment also decreases HDL-C in relation to the degree of proteinuria reduction. This adverse effect on HDL-C may in part be attributable to changes in adiponectin.

Plasma cholesteryl ester transfer (CET) from high density lipoproteins (HDL) to very low and low density lipoproteins (VLDL+LDL) may predict (subclinical) atherosclerosis. We tested the extent to which plasma CET and cholesterol esterification (EST) are decreased by statin and fibrate combination therapy compared to statin and fibrate administration alone in type 2 diabetic patients. Plasma CET and EST were measured by isotope assays in 14 type 2 diabetic patients, in whom a randomized placebo-controlled crossover study was carried out (8 weeks treatment with simvastatin (40 mg daily), bezafibrate (400 mg daily) and their combination). Plasma CET and EST from diabetic patients were compared with 42 non-diabetic control subjects with similar triglyceride levels. Plasma CET and EST were elevated in diabetic patients at baseline compared to control subjects (p < 0.01), and were correlated positively with non-HDL cholesterol and triglycerides in non-diabetic subjects and in diabetic patients at baseline (p < 0.01). Decreases in CET during combined treatment (p < 0.05) were not greater than the changes during simvastatin and bezafibrate monotherapy (p > 0.20). EST only decreased during bezafibrate therapy (p < 0.05). Changes in CET during treatment were correlated positively with changes in non-HDL cholesterol (p < 0.05) and triglycerides (p < 0.001). Changes in HDL cholesterol were related inversely to changes in CET (p < 0.05). Diabetes-associated plasma CET elevations are ameliorated by statin and fibrate monotherapy, but combined lipid lowering drug treatment does not additively lower CET. CET lowering likely contributes to HDL cholesterol changes during statin and fibrate administration.

This study examined the audiovestibular profile of 11 Wolfram syndrome patients (4 males, 7 females) from 7 families, with identified WFS1 mutations, and the audiometric profile of 17 related heterozygous carriers of WFS1 mutations. Patients with Wolfram syndrome showed a downsloping audiogram and progressive hearing impairment. None of the carriers had sensorineural hearing loss. Two patients with missense (non-inactivating) mutations in WFS1 had normal hearing and mild symptoms of Wolfram syndrome and were excluded from the analyses. Of the identified patients with inactivating WFS1 mutations, 5 female patients were significantly more hearing impaired than four male patients (p < 0.05). Female patients showed hearing impairment progressing by 1.5-2.0 dB HL per year for the low frequencies and 4.0-4.5 dB HL per year for the mid and high frequencies. The age of onset (90% phoneme recognition score) was 21 years and the onset level 78 dB HL. The deterioration rate was 4.0% per year and the deterioration gradient 1.4% per dB HL. One of the 6 examined patients had vestibular areflexia.

The usefulness of peanut specific IgE levels for diagnosing peanut allergy has not been studied in primary and secondary care where most cases of suspected peanut allergy are being evaluated. We aimed to determine the relationship between peanut-specific IgE levels and clinical peanut allergy in peanut-sensitized children and how this was influenced by eczema, asthma and clinical setting (primary or secondary care). We enrolled 280 children (0-18 years) who tested positive for peanut-specific IgE (> 0.35 kU/L) requested by primary and secondary physicians. We used predefined criteria to classify participants into three groups: peanut allergy, no peanut allergy, or possible peanut allergy, based on responses to a validated questionnaire, a detailed food history, and results of oral food challenges.Fifty-two participants (18.6%) were classified as peanut allergy, 190 (67.9%) as no peanut allergy, and 38 (13.6%) as possible peanut allergy. The association between peanut-specific IgE...

To describe changes over time in the number of allergy tests for specific IgE ordered and outcomes in children, to help address the question whether the increase in allergies is due to an actual increase in sensitisation or an increase in diagnostic awareness of allergies among physicians. Retrospective and descriptive. We reviewed the results of all specific IgE tests performed in our hospital's laboratory for children 0-18 years of age in the period 1985-2003. This included tests ordered by both general practitioners and hospital-based specialists. We analysed trends over time in the number of tests ordered (as an indicator ofdiagnostic awareness) and test results (as an indicator ofsensitisation). Between 1989 and 1995, the annual number of tests ordered increased from 1 per 10,000 children to 95 per 10,000 children and remained stable thereafter. Before 1990, more than 90% of tests were ordered by hospital-based specialists; after 1990, approximately 70% of the tests were or...

BACKGROUND.: Although techniques for percutaneous coronary intervention (PCI) have improved, patients with PCI of more vessels may still have an increased risk. We performed a prospective observational study evaluating the differences between multivessel and single-vessel procedures according to postprocedural troponin T (TnT) elevation and events during follow-up. METHODS.: The study included 713 patients without elevated TnT (<0.05 ng/ml) before PCI. Primary endpoint was the combined endpoint of death, myocardial infarction, stroke, repeat coronary angiography and readmission for anginal symptoms during the mean follow-up of 10.9 months. RESULTS.: TnT after PCI was elevated in 150 patients (21%) and was significantly associated with an increased incidence of the primary endpoint (RR 1.55, 95% CI 1.01 to 2.38). PCI of more than one vessel was performed in 146 patients (20%). These patients more often had increased TnT levels after the procedure (31.5 vs. 18.3%, p=0.001) and an i...

To evaluate the consequences of the new Dutch College of General Practitioners (NHG) protocol 'Diabetes mellitus type 2', which recommends using either the Cockcroft-Gault (CG) formula or the 'Modification of diet in renal disease' (MDRD) study formula to determine the estimated glomerular filtration rate (eGFR) as an indicator of renal function, in a cohort of patients with type-2 diabetes. Inventory. The eGFR was calculated using the CG formula, the body-mass index (BMI-)corrected CG formula and the MDRD formula in 6224 patients with type-2 diabetes who entered the 'Zwolle outpatient diabetes project integrating available care' (ZODIAC) study in 2005. Using the CG and MDRD formulas, 31% and 63% of patients, respectively, had an eGFR of 30-59 ml/min (units for MDRD are ml/ min/1.73 m2) for which referral is advisable. In addition, 1% and 11%, respectively, had an eGFR <30 ml/min (reference: >90 ml/min), for which referral is necessary. Most patients ag...

In patients with end-stage renal disease (ESRD), cardiovascular complications are the main cause of death. Increased oxidative stress is one of the risk factors for enhanced atherosclerosis in this population. Literature data vary partially dependent on differences in methodology. The present study compares three different methods: plasma lipid peroxides, the newly developed measurement of circulating oxidized LDL (Ox-LDL) particles and the frequently used copper-induced LDL oxidation lag time. We assessed plasma lipid peroxides, circulating Ox-LDL and in vitro copper-induced LDL oxidation lag time in 47 non-diabetic patients with ESRD, at the start of renal replacement therapy, and compared these with 41 age- and sex-matched controls. In ESRD, total cholesterol (4.6 +/- 1.1 vs. 5.6 +/- 0.9 mmol/l; p < 0.001), LDL cholesterol (2.8 +/- 0.8 vs. 3.5 +/- 0.7 mmol/l; p < 0.001) and HDL cholesterol (1.0 +/- 0.3 vs. 1.4 +/- 0.4 mmol/l; p < 0.001) were lower compared to controls. Plasma lipid peroxides were higher (1.1 +/- 0.5 vs. 0.8 +/- 0.5 micromol/l; p = 0.003) in ESRD. No differences were observed in plasma Ox-LDL (63.1 +/- 62.0 vs. 55.3 +/- 48.0 mg/l). However, due to the lower plasma LDL cholesterol in ESRD, LDL oxidation level was increased in ESRD (7.1 +/- 0.1 vs. 4.2 +/- 0.3%; p = 0.03). LDL lag time was slightly longer (89 +/- 11 vs. 84 +/- 11 min; p = 0.04) in ESRD. There were no significant differences regarding the amount and rate of dienes produced. Elevated levels of lipid peroxides and higher LDL oxidation levels support the theory that ESRD is associated with increased oxidative stress, which may explain the accelerated atherosclerosis. The measured amount of oxidative stress is not reflected by in vitro oxidizability of LDL.

The assumption that sensitization to aeroallergens is rare in preschool children is based on population studies in which most subjects have little or no symptoms of atopic disease. We assessed the prevalence of atopic sensitization in children 0 to 4 yr of age presenting with symptoms of allergic disease by reviewing results of all specific immunoglobulin (IgE) tests performed in our hospital laboratory in children 4 yr of age or younger between 1985 and 2003. Tests were ordered by general practitioners or hospital-based pediatricians in children presenting with symptoms of allergic disease. Specific IgE tests to a panel of common food and inhalant allergens were performed in 2946 children; a specific IgE concentration >0.35 kU/l was considered positive. Overall, 505 (17%) tests were positive to aeroallergens: 346 (12%) for house dust mite, 257 (9%) for dog dander, 240 (8%) for cat dander, and 197 (7%) for grass pollen. Positive tests were more common in boys (19.2%) than in girls (14.2%, p < 0.01), irrespective of age. Although sensitization to food allergens was more common in 0-<3 yr olds, aeroallergen and food allergen showed comparable prevalence rates in 3-<5 yr olds. Sensitization to aeroallergens is common in preschool children with symptoms of allergic disease, and more common in boys than in girls. Screening tests for allergy in infants and toddlers should include inhalant allergens.

The clinical interpretation of children sensitized to numerous allergens is challenging. We examined differences between children sensitized to zero, one, or more allergens. This was a retrospective analysis of all specific IgE tests in children 0-18 yrs of age sent to our laboratory by general practitioners and hospital-based specialists for allergy testing between 1990 and 2003. Of all 9044 children tested, 5439 (60.1%) were not sensitized to any of the aeroallergens or food allergens tested. Three thousand six hundred and five children (39.9%) had one or more positive specific IgE tests, 1120 of which (31.1%) were monosensitized (73% to aeroallergens and 27% to food allergens), 1709 (47.4%) were sensitized to two to four allergens, and 776 (21.5%) to five or more allergens (polysensitization). Polysensitization was more common in children 4-11 yrs of age (24.8%) than in younger (18.7%) or older children (18.3%, p < 0.001), and more common in boys (9.8%) than in girls (7.3%, p < 0.001). Median total IgE values increased with increasing number of positive specific IgE tests (p < 0.001). House dust mite more commonly showed monosensitization (22.2%) than other aeroallergens (grass pollen 10.5%, tree pollen 3.6%, cat 2.9%, and dog 1.5%); cow's milk (27.6%) more commonly than other food allergens (hen's egg 9.7%, peanut 4.6%, wheat 0.8%, soy 0.7%). Between 55.7% (cow's milk) and 87.9% (soy) of children sensitized to food were cosensitized to aeroallergens, while only 25.4% (house dust mite) to 39.5% (dog) of children sensitized to aeroallergens were cosensitized to food. Polysensitization is common in children, in particular in boys. It is most common in school-aged children. The strong association with total serum IgE values and the striking cosensitization between biologically unrelated allergens suggest that polysensitization is the expression of a distinct clinical, more severe, atopic phenotype, and not of biologic cross-reactivity to similar allergens.

Disturbed lipoproteins and increased oxidative stress are two of the "non-traditional" cardiovascular risk factors in chronic renal failure. There are very few prospective data of the influence of dialysis on these two factors. In the present study we investigated the effects of the initiation of both hemo- and peritoneal dialysis therapy on lipoproteins and parameters of LDL oxidation. In this prospective cohort study, we assessed lipoproteins, plasma lipid peroxides and in vitro copper-induced LDL oxidation in 46 patients with end-stage renal disease prior to the start of dialysis and after 6 months of treatment with either hemodialysis (n=33) or peritoneal dialysis (n=13). After 6 months of treatment with hemodialysis there was an increase in total cholesterol (4.6+/-1.1 vs. 5.0+/-1.3 mmol/l; p<0.05) and triglycerides (2.0+/-0.9 vs. 2.8+/-1.6 mmol/l; p<0.03). In the peritoneal dialysis group the lipoproteins did not change. Regarding lipid peroxides and in vitro copper-induced LDL oxidation, also no changes were observed after 6 months of treatment in both groups. Dyslipidemia aggravates after 6 months of hemodialysis but not after 6 months of peritoneal dialysis. During this period, no net effects on oxidative stress were demonstrated.