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Biomedical subjects

M Winkler

Publications and source records attributed to M Winkler.

At least 109 records · Page 6Linked to original sources
PubMed1998

Catalytic activities, protein- and mRNA-expression of cytochrome P450 isoenzymes in intestinal cell lines.

1. Certain chemicals and drugs in addition to metabolically activated carcinogens are substrates for intestinal cytochrome P450s (CYPs) and a number of cell lines are available which could be used in metabolism studies. These include the rat duodenal cell line IEC 6, rat ileal IEC 18, foetal human HuTu 80, foetal human small intestinal FHS 74, human duodenal HCT 8 and human colon CaCo-2 cells, but they lack thorough biochemical characterization. 2. The aim of the present study was therefore to investigate the mRNA and protein expression of CYP1A1, CYP1A2, CYP2C9/10, CYP2E1 and CYP3A. In addition, the metabolism of the immunosuppressant drug tacrolimus and of the procarcinogen 7,12-dimethyl-benz[a]anthracene (DMBA) was studied to obtain information on the functional activity on these cell lines. 3. Of all the cell lines tested only CaCo-2 cells expressed CYP1A1 at the protein and mRNA level, but the CYP2E1 and CYP3A protein was also detected in CaCo-2 and FHS 74 cells. It is of considerable interest that none of the other cell lines expressed CYP1A1, CYP1A2, CYP2C9/10 or CYP3A4 at the protein and mRNA level. 4. When the metabolism of DMBA (a model carcinogen) was studied, CaCo-2 cells produced the following metabolites: 7,12-dihydroxymethylbenz[a]anthracene, 7,12-dimethylbenz-[a]anthracene-di-hydrodiol, 7-methyl-12-hydroxymethylbenz[a]anthracene, 7-hydroxy-methyl-12-benz[a]anthracene and possibly the dihydrated product of the latter two derivatives. 5. CaCo-2 cells also catalysed the metabolism of the immunosuppressant drug tacrolimus resulting in the formation of 13-O-demethyl-tacrolimus bisdemethyl-hydroxy-tacrolimus and demethyl-dihydroxy-tacrolimus. Neither the foetal human small intestinal FHS 74 cell line nor any of the other cell lines were able to catalyse the biotransformation of tacrolimus. 6. In conclusion, only CaCo-2 cells were able to produce metabolites similar to those observed in in vivo metabolism studies, whereas all other cell lines were metabolically incompetent. Therefore, this cell line may be used in studies of intestinal biotransformation.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1998

Desflurane maintains intraocular pressure at an equivalent level to isoflurane and propofol during unstressed non-ophthalmic surgery.

We have investigated the effects of desflurane compared with isoflurane and propofol on intraocular pressure (IOP) in 48 ASA I-II patients undergoing elective non-ophthalmic surgery. Anaesthesia was induced with thiopental 3-5 mg kg-1, fentanyl 2-4 micrograms kg-1 and vecuronium 0.1 mg kg-1. Patients were allocated randomly to receive propofol (n = 16) 4-8 mg kg-1 h-1, isoflurane (n = 16) or desflurane (n = 16) for maintenance of anaesthesia. Fentanyl was added if necessary. The lungs were ventilated with 70% nitrous oxide in oxygen. Arterial pressure, electrocardiography, heart rate and end-tidal carbon dioxide were measured throughout anaesthesia. IOP was measured before surgery, during maintenance and after emergence from anaesthesia with applanation tonometry by an ophthalmologist blinded to the anaesthetic technique. There was a significant decrease in IOP after induction of anaesthesia which did not differ between groups. Desflurane maintained IOP at an equivalent level to isoflurane and propofol.

Adolescent↗
PubMed1998

Armored RNA technology for production of ribonuclease-resistant viral RNA controls and standards.

The widespread use of sensitive assays for the detection of viral and cellular RNA sequences has created a need for stable, well-characterized controls and standards. We describe the development of a versatile, novel system for creating RNase-resistant RNA. "Armored RNA" is a complex of MS2 bacteriophage coat protein and RNA produced in Escherichia coli by the induction of an expression plasmid that encodes the coat protein and an RNA standard sequence. The RNA sequences are completely protected from RNase digestion within the bacteriophage-like complexes. As a prototype, a 172-base consensus sequence from a portion of the human immunodeficiency virus type 1 (HIV-1) gag gene was synthesized and cloned into the packaging vector used to produce the bacteriophage-like particles. After production and purification, the resulting HIV-1 Armored RNA particles were shown to be resistant to degradation in human plasma and produced reproducible results in the Amplicor HIV-1 Monitor assay for 180 days when stored at -20 degreesC or for 60 days at 4 degreesC. Additionally, Armored RNA preparations are homogeneous and noninfectious.

HIV-1↗
PubMed1998

Cervical dilatation: induction by antigestagens via adhesion molecules. An in vitro examination in endothelial cell cultures.

Cervical ripening at term resembles an inflammatory reaction with invasion of activated granulocytes into the cervical stroma. Our objective was to investigate the influence of the antigestagen onapristone alone and in combination with other substances associated with cervical ripening on the expression of the inflammation-associated adhesion molecules ELAM-1, ICAM-1 and VCAM-1. Human endothelial cell cultures were stimulated with onapristone (200 ng/ml), TNF-alpha (100 U/ml), IL-8 (20 ng/ml) and PGE2 (3 ng/ml) separately and in combination (n = 6). The expression of adhesion molecules was determined qualitatively and quantitatively by immunofluorescence staining and flow cytometry with statistical evaluation by Kolmogorov-Smirnov analysis. Onapristone upregulated slightly the expression of ELAM-1 (19%). The costimulation of onapristone and TNF-alpha provoked an additive expression of VCAM-1 (64%) beyond the effect of TNF-alpha alone, while the costimulation of onapristone and PGE2 as well as the combination with IL-8 did not result in an additional stimulatory effect. All results were statistically significant (p < 0.001). This result supports our hypothesis that onapristone may lead to an increased adhesion of granulocytes to the capillary endothelium thereby initiating cervical ripening.

Cell Adhesion Molecules↗
PubMed1998

The importance of extracellular matrix in the induction of preterm delivery.

Evidence suggests that in the lower uterine segment at term an increased production of tumor necrosis factor (TNF) alpha and interleukin (IL)-1 beta induces an increase in the expression of adhesion molecules by the endothelium. The expression of endothelial-leukocyte adhesion molecule-1 and vascular cell adhesion molecule-1 were found to increase from a cervical dilatation of 2-3 cm and reaches a maximum at cervical dilatation of more than 6 cm. Increased adhesiveness of the endothelium leads to extravasation of neutrophils into the cervical stroma. The chemotaxis and degranulation of these cells is triggered by a rise in the concentration of IL-8 found in the lower uterine segment when cervical dilatation progresses. A significant increase in stroma invasion by neutrophils with progressive cervical dilatation was observed. This finding coincides with a rise in the granulocytic matrix metalloproteinases (MMP)-8 and MMP-9 concentrations up to complete cervical dilatation. We found similar patterns of cytokine concentrations in the lower uterine segment of patients with preterm delivery: at 2-3 cm dilatation the concentrations of IL-1 beta, IL-6- und IL-8 were significantly higher than at less than 2 cm. Concomitantly, we also found an increase in MMP-8, MMP-9 and tissue inhibitor of metalloproteinases-1 concentrations from less than 2 cm to 2-3 cm cervical dilatation. These findings suggest that the changes in the lower uterine segment during preterm parturition seem to be similar to those at term and both resemble an inflammatory process.

Cervix Uteri↗
PubMed1998

Effects of half the tidal volume during cardiopulmonary resuscitation on acid-base balance and haemodynamics in pigs.

The effects of half the tidal volume during cardiopulmonary resuscitation (CPR) on haemodynamics, acid-base balance, and oxygenation were studied in anaesthetized pigs. The animals were ventilated with a mean tidal volume of 12.5 +/- 0.5 ml/kg at a rate of 14/minute resulting in a mean arterial pCO2 of 40 +/- 3 mmHg. They were randomly assigned to two groups: in one group ventilation was performed with half the pre-arrest tidal volume (50% group; n = 9), in the other group ventilation remained unchanged during CPR (100% group; n = 10). After 8 minutes of CPR attempts were made to restore spontaneous circulation with epinephrine and countershocks. There were no differences in advanced life support requirements, haemodynamic parameters, and resuscitability between the two groups. During CPR up to 5 minutes after restoration of spontaneous circulation (ROSC) dead space ventilation was significantly higher in the 50% group compared with the 100% group (p < 0.05). During CPR (at 3 and 7 minutes) arterial pO2 values were significantly lower (218 +/- 136 and 221 +/- 120 mmHg vs. 381 +/- 130 and 352 +/- 147 mmHg; FiO2 1.0) and hyperventilation was less pronounced (34.4 +/- 7.3 and 31.3 +/- 7.7 mmHg vs. 26.2 +/- 5.6 and 26.9 +/- 6.3 mmHg) in the 50% group. Our results suggest that half the tidal volume during CPR is likely to establish a more physiological acid-base balance and has no adverse effects on haemodynamics in intubated pigs ventilated with pure oxygen.

Acid-Base Equilibrium↗
PubMed1997

Rapid and automated determination of the beta2-agonist reproterol in human plasma by atmospheric pressure chemical ionisation high-performance liquid chromatography-tandem mass spectrometry.

An on-line HPLC assay with tandem mass spectrometric detection for the fast and sensitive determination of reproterol in human plasma was developed, utilising a methylated structural analogue as the internal standard. Automated solid-phase extraction of diluted plasma samples, based on 250 microl plasma aliquots, at pH 6.5, allowed a reliable quantification of reproterol down to 400 pg/ml. Injection of 100 microl of plasma extracts onto a 30 mm x 4.6 mm reversed-phase guard column provided retention times ranging from 20 to 30 s for reproterol and the internal standard. The standard curves were linear from 0.2 to 200 ng/ml using weighted linear regression analysis (1/y2). The inter-assay and intra-assay accuracies were -0.9% and +3.2%, exhibiting a precision (C.V.) of +/- 11% and +/- 9.3%, respectively. Up to 100 unknowns may be analysed each 24 h per analyst.

Administration, Inhalation↗
PubMed1997

Humanization of an anti-vascular endothelial growth factor monoclonal antibody for the therapy of solid tumors and other disorders.

Vascular endothelial growth factor (VEGF) is a major mediator of angiogenesis associated with tumors and other pathological conditions, including proliferative diabetic retinopathy and age-related macular degeneration. The murine anti-human VEGF monoclonal antibody (muMAb VEGF) A.4.6.1 has been shown to potently suppress angiogenesis and growth in a variety of human tumor cells lines transplanted in nude mice and also to inhibit neovascularization in a primate model of ischemic retinal disease. In this report, we describe the humanization of muMAb VEGF A.4.6.1. by site-directed mutagenesis of a human framework. Not only the residues involved in the six complementarity-determining regions but also several framework residues were changed from human to murine. Humanized anti-VEGF F(ab) and IgG1 variants bind VEGF with affinity very similar to that of the original murine antibody. Furthermore, recombinant humanized MAb VEGF inhibits VEGF-induced proliferation of endothelial cells in vitro and tumor growth in vivo with potency and efficacy very similar to those of muMAb VEGF A.4.6.1. Therefore, recombinant humanized MAb VEGF is suitable to test the hypothesis that inhibition of VEGF-induced angiogenesis is a valid strategy for the treatment of solid tumors and other disorders in humans.

Adrenal Cortex↗
PubMed1997

[Results of surgical therapy of stomach carcinoma in the elderly].

In our hospital 35% of the patients undergoing resection for gastric carcinoma are over 70 years old. This group was divided into three age groups; clinico-pathological features, operative morbidity and mortality as well as long-term prognosis were compared with findings taken from a control group of subjects under 70 years old. No differences were noted in the distribution of the UICC stages and the incidence of early gastric cancer. The number of subtotal gastric resections was significantly higher in older patients. During the past 10 years the perioperative morbidity and mortality rates have decreased markedly and only minor differences in comparison with the control group have been recorded. However, the 5-year survival rate was significantly lower in the age group above 80 years old (13.7%) than in the control group (48.7%). Univariate analysis of prognostic factors showed the UICC stage, lymph node metastases, resection line involvement and the patient's age to be of significance. In contrast, in the multivariate analysis age was not of prognostic value.

Aged↗
PubMed1997

Surgery for early gastric cancer: a European one-center experience.

Between 1972 and 1995 a total of 251 patients with early gastric cancer underwent resection in our department of surgery. At the time of the operation 10.8% of the patients were proved to have lymph node involvement, and two already had distant metastases. A subtotal gastric resection was performed in 59.8% of cases (n = 150), a total gastrectomy in 33.8% (n = 85), and either a proximal or an atypical resection in 6.4% (n = 16). Since 1985 subtotal distal resection and total gastrectomy were accompanied by a systematic lymphadenectomy of compartments I and II. The overall postoperative morbidity was 18.3%, and the hospital mortality, 4.9%; it was only 1.6% within the last decade. Concerning these short-term results there were no statistically significant differences between the different surgical procedures. The cumulative overall 5-year-survival rate was 82.6%. There was no statistically significant influence of either the different surgical procedures or the histologic types according to the Japanese classification of early gastric cancer.

Adult↗
PubMed1997

Parturition: steroids, prostaglandin E2, and expression of adhesion molecules by endothelial cells.

OBJECTIVE: To determine whether 17 beta-estradiol, progesterone, and prostaglandin (PG) E2, alone or in combination with cytokines, influence the adhesiveness of vascular endothelium and thus play a role in the first stage of leukocyte infiltration of the uterine cervix during parturition. METHODS: Cultured umbilical vein endothelial cells obtained from 11 women after vaginal delivery at term were incubated with 17 beta-estradiol, progesterone, PGE2, tumor necrosis factor alpha (TNF-alpha), and interleukin-8, (IL-8), alone and in combination. The expression of endothelial leukocyte adhesion molecule-1, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 was investigated by immunofluorescence and flow cytometry. The Kolmogorov-Smirnov test was used for statistical analysis. RESULTS: We found that 17 beta-estradiol augmented the TNF-alpha-induced expression of endothelial leukocyte adhesion molecule-1, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 by 107, 9, and 39%, respectively. Alone, 17 beta-estradiol induced the expression of only intercellular adhesion molecule-1 (24%), as did PGE2 (13%). Neither progesterone nor IL-8 induced expression of any of these adhesion molecules. CONCLUSIONS: Unlike progesterone, 17 beta-estradiol and PGE2 stimulate the expression of adhesion molecules in vitro and may, therefore, promote adhesion of granulocytes to capillary endothelium.

Cells, Cultured↗
PubMed1997

Screening of European coffee final products for occurrence of ochratoxin A (OTA).

Samples (633) of final coffee products were drawn from the markets of different European countries relative to the market share of each product type and brand. These samples were analysed in a cooperative action with nine different laboratories. With low limits of detection (mean detection limit approximately 0.5 ng/g) no OTA was found in over half of the samples (334 negatives). In the remaining samples occurrence of OTA at a rather low level was seen. Only four samples (all instants) exceeded a level of 10 ng/g, whereas for both instants, and roast and grounds (R & G), over three-quarters of the samples were in the range from nondetectable to 1 ng/g. The overall mean for all R & Gs was 0.8 ng/g and for all instant 1.3 ng/g (for samples in which no OTA was detected, half of the detection limit was included in this calculation). In the brewing methods frequently used in Europe the OTA is essentially fully extracted. Consumption of four cups of coffee per day (approximately 24 g R & G or approximately 8 g instant coffee) contributes on average 19 or 10 ng/day respectively. Four cups/day is above the per caput consumption level in most European contries. Compared with the Provisional Tolerable Weekly Intake (PTWI) recently set by the Joint FAO/WHO Expert Committee on Food Additives at 100 ng/kg bodyweight/week, consumption of 28 cups/week contributes up to 2% to the PTWI.

Carcinogens↗
PubMed1997

Comparison of the western blot assay with the neutralizing-antibody and enzyme-linked immunosorbent assays for measuring antibody to verocytotoxin 1.

A Western blot (immunoblot) assay (WBA) was developed to detect immunoglobulin G (IgG) antibodies against Escherichia coli Verocytotoxin 1 (VT1) by using a chemiluminescence detection system. The assay was compared with a VT1-neutralizing-antibody (VT1-NAb) assay and an anti-VT1 IgG enzyme-linked immunosorbent assay (ELISA). When four human serum samples that were known to be positive by VT1-NAb assay and ELISA were titrated to the endpoint by the three assays, the WBA gave endpoint titers that were up to 8-fold higher than those by ELISA and up to 256-fold higher than those by the VT1-NAb assay. Of 32 serum samples that were known to be positive by VT1-NAb assay and ELISA, 31 (97%) were positive by WBA; the one sample with a discrepant result gave borderline results by the VT1-NAb assay and ELISA. Of 52 serum samples that were known to be negative by the VT1-NAb assay and ELISA, 50 (96%) were negative and 2 (4%) were positive by WBA. Of 44 serum samples that gave discrepant results by the VT1-NAb assay and ELISA, neither of the latter correlated with the results of WBA. In an investigation of 19 pairs of acute- and convalescent-phase serum samples from patients with hemolytic-uremic syndrome, 10 pairs that were positive by the VT1-NAb assay were also WBA positive, while 9 pairs that were NAb negative were also WBA negative. The WBA is inherently more specific and sensitive than either the NAb assay or the ELISA and may be used as a "gold standard" to detect IgG antibodies to VT1. Like the NAb assay and the ELISA for detecting antibodies to VT1, the WBA has little to offer in the diagnostic setting but is expected to play an important role in seroepidemiological studies.

Animals↗