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Biomedical subjects

W Köhler

Publications and source records attributed to W Köhler.

At least 55 records · Page 3Linked to original sources
PubMed1996

Conventional vs accelerated fractionation in head and neck cancer.

From October 1990 to March 1994, 90 patients entered a prospectively randomised trial in head and neck cancer. All patients had verified squamous cell carcinoma and were referred for primary radiation therapy. Tumours originated in the oral cavity in 25, oropharynx in 37, larynx in 15 and hypopharynx in 13 cases. Patients' stages were predominantely T3 and T4 (71/90) and had lymph node metastases (60/90). Seventy-nine male patients and 11 female patients, with a median age of 57 years (range 37-76 years) were treated. Patients were randomised to one of three treatment options: conventional fractionation (CF) consisting of 70 Gy in 35 fractions over 7 weeks or continuous hyperfractionated accelerated radiation therapy (Vienna-CHART) or Vienna-CHART with administration of a single dose of mitomycin C on day 5 of treatment (V-CHART + MMC). By the accelerated regimen a total dose of 55.3 Gy was given in 33 fractions within 17 consecutive days. Acute mucositis was the main toxicity recorded in those patients treated by accelerated fractionation, although the overall duration of mucosal reaction did not differ in the three treatment groups. There was no influence on local toxicity if MMC was added to radiation therapy or not. Those patients treated with additional MMC experienced a grade III/IV haematological toxicity in 4/28 cases. Complete remission (CR) was recorded in 48% following CF, 79% after Vienna-CHART (P < 0.05) and 71% after Vienna-CHART + MMC. The overall local failure rates were 73%, 59% and 42% (P = NS) for patients treated by CF, Vienna-CHART and Vienna-CHART + MMC respectively.

Adult↗
PubMed1995

[Cephalosporin treatment of maxillary sinusitis].

A new oral prodrug cephalosporin, cefcanel daloxate, was compared with cefaclor, another oral cephalosporin, in the treatment of acute purulent maxillary sinusitis. Three hundred fifty-four patients with purulent sinus secretion were recruited and randomized at thirteen European ENT clinics for the double blind study. The patients were treated with either cefcanel 300 mg tablets twice daily or cefaclor 250 mg capsules three times daily for ten days. Apart from the sinus puncture and secretion aspiration, the existence of sinus pain and purulent nasal secretion were inclusion criteria. Between days 12 and 15, reaspiration of the sinus was performed and signs and symptoms noted. The latter were checked again at the final visit between days 25 and 30. Samples of blood and urine were obtained for safety evaluation at inclusion, during and after treatment. The bacteriological culture of the purulent sinus secretion in 58.4% of patients revealed Haemophilus influenzae (37.9%) as the most prevalent pathogen followed by Streptococcus pneumonia (19.7%), while Streptococcus pyogenes, Branhamella catarrhalis, and Staphylococcus aureus amounted each to 6-7% only. The overall clinical outcome was defined as cure/improvement in 83.3% of the patients in the cefcanel group and 89.3% in the cefaclor group, respectively. No statistically significant differences with regard to efficacy were found between the two treatment groups. Adverse events, mostly related to gastrointestinal reactions, were reported by 36 patients (15.7%) in the cefcanel group and by 16 (13.4%) in the cefaclor group. Both cefcanel 300 mg twice daily and cefaclor 250 mg three times daily were highly effective with only minimal adverse events in the treatment of acute purulent maxillary sinusitis.

Acute Disease↗
PubMed1995

Isolation and characterization of a mitogen characteristic of group A streptococci (Streptococcus pyogenes).

It has been supposed for many years that group A streptococci may elaborate more than the three well known erythrogenic toxins A, B or C (ETA, ETB, ETC). The analysis of the culture supernatant of streptococcal strain 27297 carrying neither genes for ETA nor ETC revealed mitogenic activity at pH 7.3 in isoelectric focusing. This mitogen of strain 27297 was purified by hydrophobic adsorption to Phenyl-Sepharose following FPLC chromatography on a Mono S column resulting in two proteins with mitogenic activity called AX and BX, respectively. Both differed in only one aminoterminal residue. The mitogenic activity of BX lacking one aminoterminal arginine was found to be about 100 times higher than that of AX. The aminoterminus of BX does not correspond to a predictable cleavage site for signal peptidase. We assume that BX was produced after translation by cleavage of the mature protein or the AX molecule with streptococcal proteinase (ETB) or an arginylaminopeptidase which is detectable on whole cells. The purified proteins BX and AX showed molecular weights of about 27 kDa in SDS electrophoresis and isoelectric points of 8.3 (AX) and 7.3 (BX) in isoelectric focusing, respectively. Both proteins were produced by practically all group A strains tested but not by groups B, C, G or H streptococci. Therefore, AX or BX seem to be proteins characteristic of group A streptococci.

Amino Acid Sequence↗
PubMed1994

Kinetics of growth and product formation in cultures from streptococci of groups A and C.

During growth of streptococci of Lancefield groups A and C in a culture medium containing glucose, yeast extract and peptone, two main growth phases occur: growth phase I and growth phase II (diauxic growth). They are separated by a short stationary phase (1st stationary phase). The diauxic growth is caused by transient limitations as well as the availability of new sources of the amino acids L-serine and L-arginine. Growth phase I consists of an exponential and a nearly linear part. These growth kinetics are reflected by the kinetics of gas metabolism as well as by product formation. Hyaluronic acid is formed during the nearly linear phase whereas the enzyme alkaline phosphatase, is exclusively excreted in the 1st stationary phase. Also carbon dioxide and L-lactate are mainly produced in a growth phase-dependent mode. In the late stationary phase (2nd stationary phase) more oxygen is consumed whereas the demand for oxygen in the 1st stationary phase is nearly zero.

Amino Acids↗
PubMed1994

Production and partial characterization of monoclonal antibodies against erythrogenic toxins type A and C from Streptococcus pyogenes.

Hybridoma cell lines producing monoclonal antibodies against streptococcal erythrogenic toxins type A and C were established from fusions of immunized BALB/c mice splenocytes with P3X63-Ag8.653 myeloma cells. Six MAbs recognize ETA and 11 MAbs bind to ETC. Two MAbs (designated ETA-2 and ETC-10) were produced in ascitic fluid and further characterized. ETA-2 (IgG2a) binds to ETA with an affinity constant of 1.8 x 10(10) M-1 and ETC-10 (IgG1) binds to ETC with an affinity constant of 3.5 x 10(9) M-1. The specificities of the MAbs were evaluated by ELISA and immunoblotting. Both MAbs ETA-2 and ETC-10 are useful in developing specific double-sandwich ELISAs, in which the MAbs were used as solid-phase capture antibodies for the quantitative determinations of ETA and ETC.

Animals↗
PubMed1994

Streptococcal erythrogenic toxin type C is not a phosphorylated protein. Description of two different purification procedures and investigation of its phosphorylation state.

Erythrogenic toxin type C (ETC) from different streptococcal group A strains was successively purified by absorption on phenylsepharose, acidic dialysis of the eluate at 40% saturated ammonium sulphate solution, CM-Sepharose chromatography, finally by immunoaffinity chromatography on monoclonal antibodies. Second, after growing of bacteria in the presence of [32P]orthophosphate to phosphorylate ETC, the ETC was purified with phenylsepharose following immunoaffinity chromatography. The occurrence of phosphoamino acids in the purified ETC was investigated by an immunoassay. No phosphoamino acids could be detected in the ETC molecule. Also after radiolabelling with 32P it was not possible to demonstrate a radioactive signal. The treatment with alkaline phosphatase has no influence on the mitogenicity or position of ETC in isoelectric focusing. The results obtained led to the conclusion that in contrast to the literature, ETC is not a phosphorylated protein.

Amino Acid Sequence↗
PubMed1994

Effects of bright light on circadian patterns of cyclic adenosine monophosphate, melatonin and cortisol in healthy subjects.

Bright light is known as a strong zeitgeber on human circadian rhythms and influences several endocrine and neuroendocrine functions. In the present study we examined the influence of a 3-h bright light stimulus, given at different times during the day (morning or evening), on circadian patterns of cyclic adenosine monophosphate (cAMP), melatonin and cortisol. Two groups of synchronized healthy volunteers (lights on: 05.00-23.00 h) were exposed to bright light (2500 lux) for 3 h over 6 days either in the morning (05.00-08.00 h) or in the evening (18.00-21.00 h). The results showed a significant phase advance in the circadian rhythms of melatonin and cortisol when bright light was given in the morning but not when given in the evening. Rhythm in plasma cAMP basically was not affected by either light treatment.

Adult↗
PubMed1994

Pitch is influenced by spontaneous otoacoustic emissions.

Binaural frequency discrimination was studied in musically trained normal subjects. The frequency chosen was that of a spontaneous oto-acoustic emission (SOAE) in one ear. The other ear was free from SOAEs in the test frequency range. Subjective frequency sensation was found to drift away from the frequency of the peak of the emission: Frequencies lower than that of the emission were perceived as being lower than they actually were; frequencies higher than that of the emission were felt to be higher than they actually were (test range, emission frequency +/- 30 Hz).

Adult↗
PubMed1994

[Identification of group C Streptococcus with Helix pomatia lectin in use for blood grouping].

The lectin anti-AHP (Helix lectin) obtained from the albumen gland of an edible snail, Helix pomatia, was used for the identification of group C streptococci. All 1,045 strains of group C streptococci (S. equisimilis, S. zooepidemicus, and S. dysgalactiae) were agglutinated with the Helix lectin, but none of the 12,264 strains of group A, 1,346 strains of group G, 330 strains of group B, and 121 strains of other streptococcal groups was not. A few strains of S. equisimilis which belongs to group C have on the cell surface specific T antigen of group A streptococci, but it did not affect the agglutination with Helix lectin. Some strains of S. milleri carrying or not carrying group G specific carbohydrate and S. sanguis with group H antigen were agglutinated with the Helix lectin (Anti-AHP). This suggests that terminal N-acetyl-D-galactsamine of the cell-wall carbohydrate of these bacteria reacts with the Helix lectin (Anti-AHP). Therefore, the Helix lectin is a very useful tool for identification of group C streptococci.

Agglutination Tests↗
PubMed1993

Stimulation of human T cells by streptococcal "superantigen" erythrogenic toxins (scarlet fever toxins).

The pyrogenic (erythrogenic) exotoxins A and C (SPEA and SPEC) of Streptococcus pyogenes belong to the family of mitogenic toxins of which the staphylococcal enterotoxins are the prototypes. The erythrogenic toxin B (SPEB) is a proteinase precursor. All SPE have been reported to be superantigens. Here we have analyzed the human T cell response to these toxins. We used highly purified preparations of SPEA, SPEB, and SPEC from different S. pyogenes strains. These toxins were apparently homogenous in SDS-PAGE, IEF, and HPLC. In addition, recombinant SPEA and SPEC were produced in Escherichia coli. In cultures of PBMC, all three toxins expanded preferentially a fraction of T cells. Using mAb against V beta 2, -5, -6, -8, and -12, we investigated the phenotype of the stimulated cells. Natural SPEA, SPEB, and SPEC strongly stimulated V beta 8+ T cells, whereas recombinant SPEA and SPEC did not. Both natural and recombinant SPEA stimulated V beta 12+ cells and both natural and recombinant SPEC stimulated V beta 2+ cells. In accordance with these findings, a human V beta 8+ line responded to all three toxins derived from S. pyogenes but not to the recombinant proteins. An antiserum against natural SPEC neutralized specifically the V beta 2-stimulating activity of SPEC and the V beta 8-stimulating activity of all three toxins, but had no effect on the response to other superantigens. This shows that trace amounts of a potent novel V beta 8-stimulating activity not identical to SPEA and SPEC are responsible for the stimulation of V beta 8+ T cells by natural SPEA and SPEC reported previously. In a preliminary screening of S. pyogenes strains from patients, we found that this novel superantigen appears to be more widely distributed than SPEA and SPEC. Furthermore, we present evidence that also the superantigenic properties of SPEB are due to contaminations with this V beta 8 stimulator. The response to SPEB usually required 1000 times higher concentrations than to SPEA or SPEC. Antisera to SPEC but not to SPEB inhibited the response of PBMC and V beta 8+ Jurkat cells to SPEB. Furthermore, more stringent purification of SPEB yielded SPEB preparations devoid of mitogenic activity. These results indicate that the mitogenicity that is commonly attributed to SPEB is due to minute contaminations of the V beta 8 stimulator. These results raise two important caveats for the work with these highly potent T cell mitogens.(ABSTRACT TRUNCATED AT 400 WORDS)

Antigens, Bacterial↗
PubMed1993

Erythrogenic toxin type A (ETA): epidemiological analysis of gene distribution and protein formation in clinical Streptococcus pyogenes strains causing scarlet fever and the streptococcal toxic shock-like syndrome (TSLS).

Erythrogenic toxin type A (ETA) is assumed to play a causative role in both scarlet fever and the streptococcal toxic shock-like syndrome (TSLS). For a molecular epidemiological analysis of the gene of erythrogenic toxin type A (speA) we used altogether 497 clinical isolates of Streptococcus pyogenes belonging to three groups: a) isolates from patients with scarlet fever, b) isolates from cases with TSLS, c) isolates from patients with other streptococcal infections (like otitis media, tonsillitis, impetigo) (general group). We found that less than 50% of the scarlet fever-associated strains possessed the speA gene as compared to 25% of the general group. Only 5 to 30% of these strains secreted the toxin under experimental conditions in very low quantities. Among strains isolated from TSLS, 67% appeared to contain the speA gene. The amount of ETA secreted into the medium was also extremely low. Southern hybridization patterns proved to be the same with an speA-specific probe in all three groups of streptococcal isolates (HaeIII, HindIII). Increased occurrence of the speA gene among scarlet fever and TSLS-associated strains does not seem to be sufficient to support the hypothesis that ETA may have a causative role in both diseases since a considerable number of strains in these groups did not possess the speA gene.

Bacterial Proteins↗
PubMed1993

Studies on binding of glycosaminoglycans to Streptococcus pyogenes by using 125I-heparan sulphate as a probe.

Binding of 125I-heparan sulphate to the cell surface of Streptococcus pyogenes is mediated by proteins, that could be released from the streptococcal cell wall by using alkaline buffer. SDS-electrophoresis revealed two bands with molecular weights of 63 and 58 kDa. Binding of the 125I-labelled heparan sulphate probe to streptococci seems to be due to charge interactions, as the same probe was displaced by unlabelled heparan sulphate, other negatively charged molecules such as heparin, dextran sulphate, dermatan sulphate or by high ionic strength. The interaction was also strongly influenced by pH. The binding constant at pH 7.2 was estimated to be 9.8 x 10(6) mol/l, suggesting a moderate affinity. The presence of collagen of different types enhanced binding of 125I-labelled heparan sulphate to streptococci, whereas fibronectin and vitronectin had an inhibitory effect. The cooperation between heparan sulphate and collagen could be important for the adhesion of streptococci to connective tissue.

Bacterial Outer Membrane Proteins↗
PubMed1993

Multiple binding of type 3 streptococcal M protein to human fibrinogen, albumin and fibronectin.

M proteins are major virulence factors of group A streptococci which enable the bacteria to resist phagocytic attack. Their binding capacity for different plasma proteins seems to be one reason for the antiphagocytic activity of M protein. In the present study we demonstrate that M3 protein, isolated from the streptococcal culture supernatant of strain 4/55, and the recombinant form (rM3), purified from an E. coli lysate after cloning in phage lambda-EMBL3, show a multiple binding to fibrinogen, albumin and fibronectin in Western blot and dot binding assays. Binding of M3 protein to the multifunctional extracellular matrix and plasma protein fibronectin may not only influence phagocytosis but may also contribute to the adherence of these bacteria to endothelial and epithelial cells.

Amino Acid Sequence↗
PubMed1993

Susceptibility of chicken embryos to group A streptococci: correlation with fibrinogen binding.

One problem in investigating group A streptococcal infections and virulence is the lack of appropriate in vivo models. In this study we introduce the chicken embryo model for determining virulence of Streptococcus pyogenes. We found that M protein positive strains, if administered intravenously, were highly virulent for 12-day-old chicken embryos. The LD50 of the strains tested could be correlated directly with the amount of cell wall exposed M protein, which has been determined by the capacity of streptococci to bind fibrinogen and by the ability of streptococci to survive in fresh normal human blood. The number of colony forming units (cfu) of M+ strains necessary to kill 50% of embryonated eggs was significantly lower (< 10(2) cfu) than for M-variants (> 10(4) cfu). Albumin and/or IgG binding streptococcal cells, which can also take place in proteins of the M protein family which do not bind to fibrinogen, did not show that clear correlation to the virulence in chicken embryos that did fibrinogen binding. Application of anti-streptococcal M protein antisera from chicken and rabbit reduced the lethality of the chicken embryos. In contrast, no correlation was found between lethality of chicken embryos and the in vitro production of erythrogenic toxins by the administered strains. Thus the results indicate that the presence of M-protein with its fibrinogen binding activity on the streptococcal cell surface is necessary for virulence of group A streptococci in the chicken embryo model.

Animals↗
PubMed1993

Streptococcal and staphylococcal superantigens (ETA, SEB): presentation by human epidermal cells and induction of autologous T cell proliferation in vitro.

Streptococcal and staphylococcal toxins are responsible for skin-related clinical conditions, e.g. scarlet fever and toxic shock syndrome. Skin involvement may result from a hypersensitivity reaction to these toxins; however, their precise mode of action has still to be elucidated. The aim of the present study was to investigate the capacity of human epidermal cells to present streptococcal erythrogenic toxin A (ETA) or staphylococcal enterotoxin B (SEB) to autologous T-lymphocytes in vitro. We found a significant T-lymphocyte proliferation response to minute amounts of ETA (p < 0.01) and SEB (p < 0.02) when co-cultured with freshly isolated autologous human epidermal cells. We suggest that human skin may serve not only as an entry for microbial toxin-producing strains but also as an important target for streptococcal and staphylococcal toxin-binding and subsequent T cell proliferation.

Adult↗