Wir entwickelten ein DNA- Mikroarray zur kostengünstigen und schnellen Geno- und Pathotypisierung von Pseudomonas aeruginosa. Mit diesem Hilfsmittel ist es möglich, Einsicht in die Populationsstruktur, Verbreitung und Evolution von P.aeruginosa zu erhalten. Durch Verwendung von pathogenitätsassoziierten Genen und SNPs von hoch konservierten P.aeruginosa Genen kann für die Typisierung eine Zuverlässigkeit von >99,8% erreicht werden. Zusätzlich wird untersucht, welche der P.aeruginosa- typischen Geninseln im jeweiligen Isolat vertreten sind. Der kommerziell erhältliche Chip wurde zur Erzeugung von Typisierungsdaten entwickelt, die in einem standardisierten, binären Format gelagert oder mit anderen Zentren ausgetauscht werden können. Durch diese universelle Form ist es möglich, die Populationsstruktur und Verbreitung von P.aeruginosa sowohl lokal als auch global zu überwachen. In einer ersten Studie untersuchten wir mehr als 200 europäische P.aeruginosa- Isolate von CF- Patienten, Intensivstationen und verschiedenen Umweltproben. Zu unserer Überraschung stellten nur 15 klonale Linien mehr als 50% aller Isolate, wobei die bereits sequenzierten Stämme PA14 und PAO zwei der häufigsten Klone repräsentieren. Die häufigsten Klone wurden aus verschiedensten Quellen isoliert, ohne dabei ein spezifisches Habitat zu bevorzugen. Ko-kolonisierende P.aeruginosa waren in den meisten Fällen unverwandt, allerdings fanden wir zu jedem dieser Stämme verwandte klonale Varianten, die oft aus anderen Ländern oder sogar Kontinenten stammten. Das Auftreten der Geninseln dagegen kann sogar in einem Klon innerhalb eines spezifischen Habitats variieren. Dies zeigt, dass die evolutionäre, molekulare Uhr in P.aeruginosa mit deutlich unterschiedlichen Geschwindigkeiten für das akzessorische und das Kern- Genom läuft.

There are significant bioperiodicities for hormonal, neural, cellular, and humoral factors as well as for mediators. A combination of these findings is an explanation for the increased hyperreactivity in patients with allergic diseases during the night. In the morning hours between 2 and 6 a.m., the histamine concentration shows a peak, adrenaline and cyclic AMP have their minimum, while cortisol secretion is just ascending. Circadian variations are also seen with respect to the density of beta-receptors. Thromboxane A2 shows a peak during the night, PGE2 is depressed, a finding also in favour of bronchial constriction. Total plasma protein IgA, IgM, IgG and IgE show a distinct bioperiodicity with a minimum during the night, cellular elements like T11, T4 and B-lymphocytes, and Leu8 have a maximum. The nocturnal symptom exacerbation must be given the fullest attention in choosing the time of administration of the appropriate medication.

CD2-associated molecules were identified by means of in vitro kinase assays of CD2 immunoprecipitates obtained from nontransformed human T lymphocytes lysed in the detergent brij 58. Under these conditions CD2 was found to be associated with the protein tyrosine kinases p56lck and p59fyn as well as two low molecular weight phosphoproteins. The latter were identified as the zeta and epsilon chains, the major signaling components of the CD/7 TcR complex. Importantly, induction of T cell unresponsiveness towards CD2-mediated stimuli by means of CD3/T cell receptor (TcR) modulation results in uncoupling of zeta and epsilon from the CD2 molecule, while its associations with p56lck and p59fyn remain unaffected. Moreover, despite the incapacity of T lymphocytes to undergo DNA synthesis in the CD3/TcR-modulated state, CD2 triggering still results in tyrosine phosphorylation of some unknown protein substrates. Thus, the same zeta and epsilon chains which are components of a functional TcR complex appear to also couple to the CD2 molecular complex. Moreover, dissociation of TcR and CD2 complexes in intact cells seems to block CD2-mediated T cell growth but does not result in complete abolishment of the signal transducing capacity of the CD2 receptor.

In human T lymphocytes a functional complex is formed between the protein tyrosine phosphatase CD45, the protein tyrosine kinase p56lck and a phospho‐protein, pp32, a possible common substrate. Here we demonstrate that the previously described pp32 protein is composed of two distinct molecules (pp29 and pp32) in both resting human T lymphocytes and continuously proliferating T lymphoma lines. Importantly, T lymphocyte activation employing CD2 monoclonal antibodies (mAb), CD3 mAb or phorbol 12, 13 dibutyrate results in loss of pp29 and pp32 from the CD45/p56lck molecular complex and concomitant association of two distinct phosphoproteins with different molecular weights (pp30 and pp31). These events appear to be unrelated to clonal T cell growth but rather depend on receptor‐mediated differentiation signals. Reprecipitation experiments employing an antiserum directed at a consensus sequence of GTP‐binding proteins suggest that all four pp29‐pp32 molecules might represent proteins with GTP‐binding properties. Biochemical analysis of pp29‐pp32 employing V8‐protease digestion indicates that they differ in low‐molecular weight fragments of 8, 5, 4.5, 4 and 3 kDa, respectively.

The long-term stability of morning serum theophylline concentrations (STC) and peak expiratory flow (PEF) was investigated for at least one year in 21 asthmatic patients who received individualised theophylline doses in addition to a baseline medication (beta 2-agonists, corticosteroids, anticholinergics by MDI). All patients had previously participated in a randomised crossover study with the new sustained-release theophylline formulation Euphylong, so that 24-hour steady-state STC profiles were available. During the 15 (13-18) months' follow-up, the theophylline dose was increased by about 6% on average; the largest increase of 33% (from 750 mg to 1000 mg) occurred in 4 patients. The morning STCs between 8 and 9 a.m. were intra-individually averaged over the longest interval with a constant dose; median and range were 11.6 (9.7-15.8) mg/l. The coefficient of variation was less than 10% in 8 patients and between 11 and 20% in 10 patients. The corresponding peak-flow values were 474 (239-622) l/min with a coefficient of variation of up to 10% in 16 patients and between 11 and 20% in 3 patients. The long-term study over more than one year confirmed the reproducibility of the Euphylong plateau profile previously found in controlled short-term pharmacokinetic studies and clinical trials.

In this randomised crossover study in 26 outpatients with bronchial asthma the efficacy of a new once-daily theophylline formulation given in addition to a baseline medication was investigated; moreover, under steady state conditions, the effect of three evening intake times (6, 8 and 10 p.m.) on 24-hour pharmacokinetics and peak-expiratory flow profiles was evaluated. The theophylline dose had been individually titrated. The pharmacodynamic results show a marked improvement in 24-hour lung function data after adding theophylline to a drug therapy including inhalative anti-cholinergics in 50% of the treated outpatients. No significant differences between the pharmacokinetic characteristics and the 24-hour averages (mesors) of peak expiratory flow at the three different intake times 6, 8 and 10 p.m. were found; however, intake at 10 p.m. resulted in the highest nocturnal excess of serum theophylline concentrations and the highest peak expiratory flow during the early morning hours between 2 and 6 a.m.

In this randomized crossover study of 26 outpatients with bronchial asthma the efficacy of a new once-daily theophylline formulation given in addition to a baseline medication was investigated; moreover, under steady state conditions, the effect of three evening intake times (6, 8 and 10 p.m.) on 24 h pharmacokinetics and peak-expiratory flow profiles was evaluated. The theophylline dose had been individually titrated. The pharmacodynamic results show a marked improvement of 24 h peak expiratory flow values after adding theophylline to a drug therapy including inhalative beta 2-agonists and corticosteroids in nearly all and inhalative anticholinergics in 50% of the treated outpatients. No significant differences between the pharmacokinetic characteristics and the 24 hr averages (mesors) of peak expiratory flow at the three different intake times 6, 8 and 10 p.m. were found; however, intake at 10 p.m. resulted in the highest nocturnal excess of serum theophylline concentrations and the highest peak expiratory flow during the early morning hours between 2 and 6 a.m.

In the majority of patients one of the primary features of bronchial asthma is the occurrence of nocturnal symptoms. There are significant bioperiodicities for hormonal, neural, cellular and humoral factors and for mediators, which are all in favour of reducing bronchial patency during the night. In the morning hours between 2 and 6 a.m. the histamine concentration shows a peak, the adrenaline and cyclic AMP have their minimum, while the cortisol secretion with its delayed onset of action is already ascending. The kallikrein kinin system is activated by histamine. Circadian variations have also been found for the receptor density of beta-receptors, cAMP, adenylcyclase and phosphodiesterase. While circadian rhythms are not known for the platelet activating factor (PAF) at the present time, the known nocturnal peak of thromboxane A2 may influence PAF-liberation as well. The bronchodilating metabolite of the cyclooxygenase pathway, PGE2, was found to have depressed levels during the night. Total plasma, total protein as well as IgA, IgM, IgG and IgE have a minimum during the night, cellular elements like T11-, T4-, B-lymphocytes and Leu8 have a maximum. Slow release theophylline is today the most important drug for nocturnal asthma. Theophylline is known to interfere with histamine release from mast cells, mediator release of arachidonic acid metabolites, the cyclic AMP concentration and suppressor cell activity. Important work remains to be done to clarify the therapeutic and immunological role of theophylline in nocturnal asthma and to identify the subgroups of patients having the greatest benefit of theophylline treatment.