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Biomedical subjects

M Winkler

Publications and source records attributed to M Winkler.

At least 19 recordsLinked to original sources
PubMed1996

Evidence for the presence of a large keratan sulphate proteoglycan in the human uterine cervix.

Profound changes occur in the uterine cervix during pregnancy. In particular, the extracellular matrix of the connective tissue is remodelled extensively. To elucidate the mechanisms involved in this process, we have analysed the proteoglycan pattern in the human cervix from pregnant and non-pregnant women. Proteoglycans of the cervix tissue specimen were extracted with 4 M guanidine hydrochloride and precipitated with 80% ethanol. Purification of proteoglycans was performed by several chromatographic steps. Characterization of proteoglycans was done by SDS/PAGE before and after digestion with glycosaminoglycan-specific enzymes. Proteoglycans were detected by combined Alcian Blue/silver staining or, after blotting of biotin-labelled proteoglycans on to poly(vinylidene difluoride) membrane, with peroxidase-conjugated avidin or by the use of keratan sulphate- or decorin-specific monoclonal antibodies. In contrast with previous reports, where only chondroitin/dermatan sulphate proteoglycans have been found in the uterine cervix, we have shown in the present study the existence of a large keratan sulphate proteoglycan with an M(r) > 220,000 in cervix samples from non-pregnant and pregnant women. This proteoglycan showed a strong reaction with the keratan sulphate-specific monoclonal antibody 5D4 and could be degraded by keratanases. The size of the core protein of this keratan sulphate proteoglycan was estimated to be about M(r) 220,000.

Cervix Uteri↗
PubMed1996

A new oral formulation of cyclosporine for early oral immunosuppressive therapy in liver transplant recipients.

CsA-ME is a new oral microemulsion formulation of CsA. Studies in stable liver grafted patients with cholestasis and subsequent poor absorption of the conventional cyclosporine formulation showed a substantial increase in CsA absorption after conversion to CsA-ME. To investigate its use in patients during the early course after liver transplantation we recruited 50 liver transplant recipients in two centers. During the first study phase A CsA-ME was administered to 20 patients in incremental doses after a short initial course of intravenous cyclosporine. During the second study phase B (30 patients) CsA-ME was administered from the time of transplantation. One year actual patient and graft survival of patients included in phase A and B of the trial was between 90% and 93.3%; 50% and 60% of the patients enrolled in phase A and phase B of the trial were free from rejection at month 3, respectively. Chronic rejection was diagnosed in one patient. No increase in the incidence of CsA related side effects was observed. The optimum CsA-ME starting dose was found to be 10 mg/kgbw/day for patients without external biliary diversion and 15 mg/kgbw/day for patients with a T tube in situ. Using these starting doses, 26 consecutive patients with external bile diversion via T tube were treated with CsA-ME from the day of transplantation. Intravenous CsA was necessary only in three patients. When CsA-ME absorption in patients with stable liver function was compared with that in patients with early liver dysfunction, no difference in the pharmacokinetic profiles was observed between the groups. Our results indicate that CsA-ME therapy is effective and well tolerated in liver graft recipients, even in patients with external biliary diversion during the early posttransplant phase. Thus, CsA-ME is a useful alternative to intravenous CsA treatment in these patients.

Administration, Oral↗
PubMed1996

Long-acting growth hormones produced by conjugation with polyethylene glycol.

Derivatives of human growth hormone (hGH) of increasing size were produced by reaction with the N-hydroxysuccinimide ester of polyethylene glycol-5000 (PEG5000), a 5-kDa reagent that selectively conjugates to primary amines. By adjusting the reaction conditions and purification procedure, it was possible to isolate hGH derivatives containing up to seven PEG moieties that altered the Stokes radius and thereby the effective molecular masses of the unmodified hormone from 22 to 300 kDa. Fortunately, the most reactive amines were ones that did not lie in either of the two sites important for receptor binding. Nonetheless, increasing the level of PEG modification linearly reduced the affinity of hGH for its receptor and increased the EC50 in a cell-based assay up to 1500-fold. Most of the reduction in affinity was the result of slowing the association rate for the receptor. The clearance rate of hGH in rats was inversely proportional to effective molecular weight and closely fit a filtration model. We have tested the potency of these analogs by injecting them daily or every 6 days into hypophysectomized rats and determining the effects on body and organ growth. The efficacy of these analogs was optimal for hGH conjugated with 5 eq of PEG5000, and the potency was increased by about 10-fold compared with unmodified hGH. Such PEG-hGH derivatives show promise as long-acting alternatives to daily injections of hGH. More generally these studies show that improving hormone clearance properties, even at the expense of reducing receptor binding affinity, can lead to dramatic increases in hormone efficacy.

Animals↗
PubMed1996

Combined effects of NO inhalation and intravenous PGF2 alpha on pulmonary circulation and gas exchange in an ovine ARDS model.

OBJECTIVES: Inhalation of nitric oxide (NO) selectively dilates pulmonary vessels in well-ventilated regions. Prostaglandin F2 alpha (PGF2 alpha) is a vasoconstrictor and is reported to enhance hypoxic pulmonary vasoconstriction. The objective of this study was to examine whether the combination of intravenous PGF2 alpha and inhaled NO in ARDS lungs has a beneficial effect on oxygenation. DESIGN: We investigated the effect of intravenous PGF2 alpha infusion (0.05-10.0 micrograms/kg per min) with and without NO inhalation (60 ppm) on the hemodynamics and gas exchange in an ovine ARDS model, examining the pulmonary artery pressure versus the flow plot by varying cardiac output. MEASUREMENTS AND RESULTS: After lung lavage, NO inhalation reduced the mean pulmonary arterial pressure (MPAP) by decreasing the zero-flow pressure intercept from 10.6 +/- 3.8 (mean +/- SD) to 8.5 +/- 3.8 mmHg (p < 0.05) with no significant change in slope. NO inhalation improved PaO2 from 56 +/- 12 to 84 +/- 38 mmHg (p < 0.005) and reduced pulmonary shunt from 65 +/- 5 to 53 +/- 8% (Qs/Qt) (p < 0.001). The dose-dependent effects of PGF2 alpha infusion were: (1) increased MPAP attributed to an increased slope in pulmonary artery pressure-flow plot; (2) decreased cardiac index; (3) decreased Qs/Qt with unchanged PaO2. The dose-dependent decrease in Qs/Qt after PGF2 alpha infusion was attributed to the decreased cardiac output. CONCLUSIONS: It is suggested that inhalation of NO reduced the critical vascular pressure near alveoli without affecting upstream vessels, while infused PGF2 alpha constricted the larger upstream pulmonary artery vessels without appreciably affecting the critical pressure. Inhalation of NO into well-ventilated lung areas shifted perfusion to well-oxygenated areas, and there was no supplemental shift in blood flow by adding an infusion of PGF2 alpha.

Administration, Inhalation↗
PubMed1996

[Does lung maturation therapy with 16-methylene-prednisolone modify maternal infection parameters in threatened premature labor?].

UNLABELLED: Silent intrauterine infection is a frequent cause of preterm labour. Maternal C-reactive protein (CRP) and leukocyte count are important predictors of such infections. Treatment with corticosteroids is known to elevate leukocyte count and in this manner can possibly interfere with its accuracy as a predictor of infection. Therefore, we investigated the impact of antenatal administration of 16-methylene-prednisolone (Decortilen solubile) on the maternal serum level of CRP and leukocyte count. Furthermore, we determined the haptoglobin, the platelet count as well as the percentage of stab cells and lymphocytes. PATIENTS AND METHODS: 20 patients with preterm labour between 25 + 6 and 34 + 2 weeks of gestation were enrolled in a prospective study. Premature rupture of the membranes, uterine bleeding, infection and treatment with antibiotics were criteria for exclusion. Three doses (60 mg) of Decortilen solubile were given intravenously 24 h apart. Blood samples were obtained before, twice a day (8.00 a.m. and p.m.) during treatment and until the day 4 after termination of corticosteroid treatment. For statistical analysis the Wilcoxon rank sum test was used. RESULTS: Before corticosteroid treatment the medians (range) were: CRP: 5.2 ( < 5.0-28.0) mg/l, haptoglobin: 1.4 (0.7-2.0) g/l, leukocytes: 10.5 (5.2-16.0) G/l, platelets: 246 (128-424) G/l, stabs: 9.5 (3.0-14.0)% and lymphocytes: 28.5 (16.0-50.0)%. During the after termination of corticosteroid administration no significant changes in the CRP and haptoglobin levels were seen. The leukocyte count was unchanged during treatment and decreased on day 1-2 after termination of treatment. The platelet count remained unchanged during corticosteroid treatment and increased significantly thereafter. The stab cell percentage increased slightly from day 1-2 to day 3-4 after termination of treatment. The lymphocyte percentage increased during treatment and decreased significantly from day 1-2 to day 3-4 after treatment. CONCLUSION: Decortilen solubile treatment is not associated with an increase in maternal CRP-level and leukocyte count. We emphasise that the accuracy especially of the CRP for early prediction of silent infection in preterm labour does not seem to be impaired by this corticosteroid in a dosage usually administered for prevention of respiratory distress syndrome.

C-Reactive Protein↗
PubMed1996

Mastitis in dairy cattle caused by Corynebacterium pseudotuberculosis and the feasibility of transmission by houseflies. I.

Morbidity due to Corynebacterium pseudotuberculosis infection occurred in 29 dairy herds. The disease appeared basically in three clinical forms: cutaneous, mastitic, and visceral. The appearance of the disease showed a marked seasonality: in 23 herds it occurred during the spring and summer months (dry season) (March-October). The mastitic form occurred in only 10 herds and the causative bacterium was isolated from 33 cows (5.8%). All the strains of C. pseudotuberculosis isolated from the milk samples were found not to be nitrate reducers. The bacterium was excreted in the milk of six cows from herd B during a period of 11 months. In the mastitic cows, a decrease in milk production and considerable increases in the somatic cell count were noted. C. pseudotuberculosis was isolated from houseflies collected over a cow lesion. Laboratory-reared houseflies were successfully infected with C. pseudotuberculosis-contaminated milk, broth and sugar cubes. Flies infected with the bacterium from contaminated milk excreted the bacterium in their droppings for up to 4 h and from their saliva for up to 3 h post infection. The bacterium survived on the external organs of houseflies for no longer than 10 min post infection, after the flies had been dipped in contaminated broth.

Animals↗
PubMed1996

Evaluation of the Pro-Trac tacrolimus monoclonal whole-blood enzyme-linked immunosorbent assay for monitoring of tacrolimus levels in patients after kidney, heart, and liver transplantation.

In a prospective study, we evaluated a novel enzyme-linked immunosorbent assay (ELISA) (Pro-Trac) for determining tacrolimus (FK 506) concentrations in whole blood. Results obtained by the ELISA were compared with those obtained either by microparticle enzyme immunoassay (MEIA) or by high-performance liquid chromatography/mass spectrometry (HPLC-MS). The lower limit of quantitation of the ELISA was 0.5 microgram/L. The within-series coefficient of variation (CV) was < 11%. For spiked blood samples containing different concentrations of tacrolimus, interassay CV was 23.6% at 2.5 micrograms/L; however, at 15 and 60 micrograms/L, interassay CV was 44.9 and 50.8%, respectively. In crossover studies including blood samples from patients after liver, heart, or kidney transplantation, ELISA results correlated with those of the HPLC-MS (r = 0.73) as well as with those generated by MEIA (r = 0.82). The ELISA and MEIA showed 52.3 and 56.2% cross-reactivity with 15-O-demethyltacrolimus, respectively, but only 5.0 and 5.4% cross-reactivity with 13-O-demethyltacrolimus. We conclude that if assay precision in the upper range is improved, the Pro-Trac ELISA might be a valuable alternative to the MEIA for therapeutic drug monitoring of tacrolimus.

Chromatography, High Pressure Liquid↗
PubMed1996

The anti-inflammatory drugs phenylbutazone and dipyrone in the treatment of field cases of bovine mastitis.

The efficacy of phenylbutazone vs. dipyrone for the treatment of acute clinical mastitis were compared in a clinical trial. All cows were treated with 20 g sulfadiazine and 4 g trimethoprim i.m. upon diagnosis and half dosage once daily thereafter. In addition, the NSAIDs treated cows received once daily either 4 g phenylbutazone or 20 g dipyrone i.m. for the duration of the antimicrobial therapy. In all treatment groups the major causative organisms were coliforms. Recovery rates for the controls, the phenylbutazone and dipyrone treatment groups were 81.8%, 89.4% and 86.6%, respectively. Recovery was evaluated by the logistic regression analysis, the odds ratios (OR) and their 95% confidence interval (CI) of treatment success for phenylbutazone and dipyrone treatments relative to the control treatment were calculated. Odds ratio of recovery was high for phenylbutazone (OR = 2.42; CI = 0.98-5.96; P = 0.054) as well as for dipyrone (OR = 1.71; CI = 0.98-3.00; P = 0.060), demonstrating a strong trend towards improved recovery in NSAID groups. The odds of treatment failure for the phenylbutazone group relative to the dipyrone group was 0.71 with 95% CI of 0.28-1.78. Clearly no significant difference could be demonstrated between phenylbutazone and dipyrone in this field trial.

Animals↗
PubMed1996

Neutralizing antibodies to Escherichia coli Vero cytotoxin 1 and antibodies to O157 lipopolysaccharide in healthy farm family members and urban residents.

An enzyme-linked immunosorbent assay (ELISA) to detect antibodies to Escherichia coli O157 lipopolysaccharide (LPS) was developed with sera from 63 children with confirmed recent E. coli O157 infection and from 256 age-stratified urban controls. The median ELISA values for control and case sera were 0.05 (interquartile range, 0 to 0.20; mean +/- standard deviation [SD], 0.15 +/- 0.22) and 1.41 (interquartile range, 1.11 to 1.59; mean +/- SD, 1.41 +/- 0.53), respectively (P < 0.001). With a breakpoint of 0.59 (mean ELISA value of the control sera + 2 SDs), the assay had a sensitivity, specificity, and positive and negative predictive values of 95, 94, 80, and 98%, respectively, for recent E. coli O157 infection. The O157 LPS assay and Vero cytotoxin (VT) 1-neutralizing-antibody (NAb) assay were used to compare the relative frequencies of O157 LPS antibodies and VT1-NAbs in an age-stratified urban population from Toronto, Ontario, Canada, and in 216 healthy family members from dairy farm in southern Ontario. The frequency of O157 LPS antibodies was about threefold higher in dairy farm residents (12.5%) than in urban residents (4.7%) (P < 0.01). Similarly, the frequency of VT1-NAbs was about sixfold higher in dairy farm residents (42.0%) than in urban residents (7.7%) (P < 0.001). These findings are consistent with a greater level of exposure of dairy farm residents to VT-producing E. coli (VTEC) strains. The high rate of seropositivity to VT1 in farm residents probably reflects the booster effect of repeated VTEC exposures and argues against a sustained generalized immunosuppressive effect of VT1. Seroepidemiological studies may help in assessing the level of exposure of different populations to VTEC strains.

Adolescent↗
PubMed1996

A specific subform of the human cytomegalovirus transactivator protein pUL69 is contained within the tegument of virus particles.

The polypeptide encoded by the open reading frame UL69 of human cytomegalovirus (HCMV), which is homologous to the immediate-early regulator ICP27 of herpes simplex virus, has recently been identified as a transactivator protein that exerts a broad stimulatory effect on gene expression (M. Winkler, S. A. Rice, and T. Stamminger, J. Virol. 68:3943-3954, 1994). Here, we provide evidence that pUL69 is a phosphorylated tegument protein of HCMV. This finding could be demonstrated by Western blot (immunoblot) analyses with purified virions and a specific antiserum against pUL69. These experiments revealed that one phosphorylated subform of the three pUL69 polypeptides that are synthesized in infected fibroblast cells is contained within the HCMV virion. After the treatment of purified virions with detergents, pUL69 could not be detected within the membrane fraction, suggesting that it is either a capsid or a tegument protein. Its presence within dense bodies, however, shows that pUL69 is a constituent of the viral tegument.

Cytomegalovirus↗
PubMed1996

[Serum concentrations of the MCA tumor marker during pregnancy].

MCA (Mucin like carcinoma associated antigen) is used as a marker for the follow-up control of breast cancer patients, and high levels of MCA have been observed during the course of pregnancy. The aim of our study was to determine the value of MCA as a tumor marker in the follow-up of young breast cancer patients with subsequent pregnancy. We determined the MCA concentration in the sera of 201 pregnant women in the course of gravidity in order to find the degree and time of increase MCA shows during normal pregnancy. 121 serum samples of healthy, non-pregnant women were analysed for MCA in comparison. Up to the 19th week of gestation, the concentration of MCA did not show any differences in comparison to non-pregnant healthy women (n = 121, median: 7 U/ml: 25% percentile: 7 U/ml: 75% percentile: 10 U/ml (control collective). Median: 6 U/ml: 25% percentile: 5 U/ml: 75% percentile: 10 U/ml (gestational week 5-19)). After the 23rd week of gestation, the MCA concentration was elevated. After the 35th week of gestation, all women had MCA levels which were significantly higher than in healthy, non-pregnant women. Median: 100 U/ml: 25% percentile: 71 U/ml: 75% percentile: 147 U/ml (gestational week 36-42). Therefore the tumor marker MCA can be used in the followup of breast cancer only until the 19th week of gestation. After the 23rd gestational week very high MCA concentrations are observed in some cases and can not be distinguished from MCA augmentation due to advanced breast cancer.

Antigens, Neoplasm↗
PubMed1996

Tacrolimus (FK506) metabolite patterns in blood from liver and kidney transplant patients.

The metabolite patterns of tacrolimus in blood were evaluated in 41 kidney and liver graft recipients. Trough concentrations of tacrolimus and its metabolites were measured by HPLC-mass spectrometry and microparticle enzyme immunoassay in parallel. A statistically significant correlation between results of both assays was observed for kidney and liver transplant patients (r = 0.77, P <0.001 and r = 0.71, P <0.001, respectively). The main metabolites in blood were demethyl, demethylhydroxy, didemethyl, didemethylhydroxy, and hydroxy tacrolimus. These metabolites added up to 42% (range 0-145%) of the tacrolimus concentration in liver transplant patients and to 44.8% (range 16-152%) in kidney transplant patients. During episodes of impaired liver function, concentrations of tacrolimus and its metabolites were increased compared with normal liver function, indicating accumulation of metabolites, in particular second-generation metabolites such as didemethyl and didemethylhydroxy tacrolimus. Stepwise regression analysis including tacrolimus, its metabolites, and liver function parameters suggested a model including serum activities of gamma-glutamyltransferase, alkaline phosphatase, and alanine aminotransferase as predictors for increased concentrations of demethyl tacrolimus, didemethyl tacrolimus, and the parent drug.

Adolescent↗
PubMed1995

Feed contamination with Candida krusei as a probable source of mycotic mastitis in dairy cows.

Over 3 months, yeasts were isolated in pure culture from milk samples obtained from 8 lactating cows with acute mastitis and from 1 cow with subacute mastitis. Eight of the isolates were identified as Candida krusei; 1 isolate was not submitted for identification. The affected cows were assigned to separate milking groups and had not been treated by intramammary administration of antibiotic before the outbreak. Remission of disease without treatment was observed, followed by shedding of the yeast in milk for 2 to 5 weeks. Median somatic cell counts in the affected cows before, during, and 2 months after the onset of clinical signs were 93,000; 1,793,000; and 135,000 cells/ml, respectively. Wheat silage was found to be the probable source of the infecting microorganism, whereas inadequate milking hygiene resulted in its persistence in the herd. Following replacement of the silage and improvement of the milking hygiene, the outbreak ceased. Candida krusei thus may cause mastitis in cattle not only following intramammary antibiotic treatment, but also in conditions of heavy environmental contamination, in conjunction with inadequate milking hygiene.

Animal Feed↗