A molecular model of type I allergy: identification and characterization of a nonanaphylactic anti-human IgE antibody fragment that blocks the IgE-FcepsilonRI interaction and reacts with receptor-bound IgE.

BACKGROUND: The IgE-mediated activation of effector cells and antigen-presenting cells through the high-affinity receptor for IgE (FcepsilonRI) represents a key pathomechanism in type I allergy and many forms of asthma. OBJECTIVE: We sought to establish an in vitro molecular model for the interaction of human FcepsilonRI, IgE, and the corresponding allergen and to identify monoclonal anti-human IgE antibodies with a therapeutic profile different from previously established anti-IgE antibodies. METHODS: Human FcepsilonRI alpha chain, a human monoclonal allergen-specific IgE antibody (chimeric Bip 1), and the corresponding allergen, the major birch pollen allergen Bet v 1, were produced as recombinant proteins and analyzed by means of circular dichroism and native overlays, respectively. Using this molecular model, as well as negative stain immunoelectron microscopic analysis, and in vitro cultivated human basophils, we characterized mouse anti-human IgE antibodies. RESULTS: We established a molecular model for the interaction of human IgE with FcepsilonRI. Using this molecular model, we identified a nonanaphylactic anti-human IgE antibody fragment (Fab12), which blocked the IgE-FcepsilonRI interaction and reacted with effector cell-bound IgE. CONCLUSION: Fab12 represents a candidate molecule for therapy of atopy and asthma because it can be used for the depletion of circulating IgE antibodies, as well as for the depletion of IgE-bearing cells.     

Effects of urapidil on catecholamine turnover and release in the central nervous system of the rat

The actions of urapidil, prazosin, idazoxan, and haloperidol on the turnover of noradrenaline in the hypothalamus and dopamine in the nucleus accumbens of the rat were investigated using changes in the ratios of 3-methoxy-4-hydroxyphenyl-glycol/noradrenaline (MHPG/NA) and 3,4-dihydroxyphenylacetic acid/dopamine (DOPAC/DA), respectively, as measures for drug-induced effects. Urapidil (2.5-30 mg kg-1 i.v.) increased the ratios of MHPG/NA and DOPAC/DA. Its effects on NA turnover were maximal at 60 min (160% of control at 30 mg kg-1), and on DA turnover at 30 min (138% of control at 30 mg kg-1). Prazosin (0.5-2.5 mg kg-1 i.v.) had no effect, but the high dose of 5 mg kg-1 i.v. significantly increased the ratio of MHPG/NA in the hypothalamus. Idazoxan (2-50 mg kg-1 i.v.) and haloperidol (0.02-0.5 mg kg-1 i.v.) selectively enhanced turnover of NA and DA, respectively. In experiments on field-stimulated overflow of tritium from slices of hypothalamus and nucleus accumbens labelled with [3H]NA or [3H]DA, respectively, urapidil (1 mumol L-1) facilitated the evoked responses in both regions. Prazosin (0.1 mumol L-1) had no effect in either of the two areas. Idazoxan (0.1 mumol L-1) increased stimulated overflow of [3H]NA from the hypothalamus but not of [3H]DA from the nucleus accumbens. Conversely, haloperidol (0.1 mumol L-1) greatly enhanced evoked overflow of [3H]DA but not of [3H]NA. From the present results it is concluded that urapidil has an antagonistic effect at central alpha 2-adrenoceptors and also a weak antagonistic action at central dopamine D2-receptors.     

Thiolated Polymers: Development and Evaluation of Transdermal Delivery Systems for Progesterone

Purpose. To evaluate the possible use of polycarbophil-cysteine (PCP-Cys) as polymeric matrix for transdermal progesterone application. Methods. Thiolated polycarbophil was synthesised by the covalent attachment of cysteine to the basis polymer. The adhesive properties of PCP-Cys in comparison to polyvinylpyrrolidone/hydroxypropyl- methylcellulose (PVP/HPMC) and polyvinylpyrrolidone/polyvinyl- alcohol (PVP/PVA) were investigated by testing the total work of adhesion (TWA) on porcine skin. Release studies in Franz diffusion cells and standard in vitro permeation experiments with porcine skin were performed analysing the progesterone content by high-per- formance liquid chromatography.Results. Films based on PCP-Cys displayed very high cohesive properties due to the formation of interchain disulfide bonds. The TWA of the thiolated polymer on porcine skin was significantly (P <0.05) the highest. In addition progesterone permeation was also the highest from PCP-Cys compared with PVP/HPMC and PVP/PVA within 24 hours. Conclusion. PCP-Cys—a partly thiolated polymer—might be a novel polymer matrix for transdermal progesterone delivery with excellent adhesiveness on porcine skin.     

On the transfer of the Fusarium toxins deoxynivalenol (DON) and zearalenone (ZON) from sows to their fetuses during days 35-70 of gestation

Eleven pregnant sows with a body weight between 153 and 197 kg were fed a control diet (CON, 0.15 mg DON and 0.0035 mg ZON/kg diet) or a diet containing 15% of Fusarium toxin contaminated triticale (MYCO, 4.42 mg DON and 0.048 mg ZON/kg diet) in the period of day 35 and 70 of gestation. The indirect effect of feed intake was separated from the direct effects of the Fusarium toxins by the restricted feeding regimen where all sows were fed the same amount of feed (2000 g/d) over the whole study. At the end of experiment, fetuses were delivered by Caesarian section and samples of serum, bile, urine, liver, kidney and spleen of euthanatized sows and fetuses were taken to analyze the concentrations of DON, ZON and their metabolites. Feeding the Fusarium toxin contaminated diet to pregnant sows caused neither adverse effects on performance, organ weights and maintenance of pregnancy of sows nor on fetus weight and length. Furthermore, no teratogenic or embryolethal effects could be observed in the MYCO group. Hematological and clinical-chemical parameters of sows and fetuses were not affected by feeding, with the exception of significantly lower GLDH (glutamate dehydrogenase) serum activities in MYCO sows. The carry over of DON and ZON from the diet to the sow or fetus tissues was calculated by the diet ratio (sum of concentrations of all metabolites in the physiological specimen divided by the dietary toxin concentration), while the fetus ratio was evaluated by the sum of concentrations of all metabolites in the physiological specimen of the fetus divided by that of the sows. DON and deepoxy-DON were found in urine, bile, serum, liver, kidney and spleen of sows of the MYCO group, but not in the bile of fetuses (spleen not analyzed). ZON and its metabolite alpha-zearalenol (alpha-ZOL) were detected in urine and bile of sows, while all specimens of fetuses as well as serum and liver of sows were negative for ZON metabolites. The maximum diet ratios for urine and bile in sows of the MYCO group were 0.84 and 0.05 for DON metabolites and 1.2 and 3.8 for ZON metabolites, underscoring the differences in metabolism and excretion of both toxins. The maximum diet ratio of DON and deepoxy-DON into liver, kidney and spleen of MYCO sows were 0.003, 0.007 and 0.003, respectively. The maximum fetus ratio of DON and deepoxy-DON into urine, bile, serum, liver and kidney of fetuses were 0.006, 0, 0.5, 0.88, and 0.33, while the maximum placental ratio (sum of toxin concentrations in the physiological specimen of the fetus divided by the toxin serum concentration of the sow) were 0.64, 0, 0.50, 0.70 and 0.52, respectively. Therefore, it can be concluded that the developing fetus is exposed to DON between the gestation days 35 and 70 when the sows are fed a Fusarium toxin contaminated diet. ZON concentration in the MYCO diet was too low to get reliable results for fetus or placental ratios.     

Isolation and structure determination of Pachybasium cerebrosides which potentiate the antifungal activity of aculeacin

A set of four cerebrosides was isolated from a Pachybasium species and purified by preparative reversed-phase HPLC. All four products displayed activity in a natural product screen aimed at detecting novel cell wall-active antifungal agents based on synergy with the known glucan synthetase inhibitor, aculeacin. Based on degradation studies, fast atom bombardment mass spectrometry and 13C and high field 1H NMR techniques, the structure of the major cerebroside was determined to be (4E,8E)-N-D-2'-hydroxy-(E)-3'- hexadecenoyl-1-O-beta-D-glucopyranosyl-9-methyl-4,8-sphingadiene. The other components were found to be the corresponding 2'-hydroxypalmitic acid analog with one less double bond and an analogous pair containing 2'-hydroxystearic acid with and without the 3' double bond.     

Real-time breath-hold triggering of myocardial perfusion imaging with a novel cadmium-zinc-telluride detector gamma camera

Purpose  

The aim of this study was to assess the ability of real-time breath-hold-triggered myocardial perfusion imaging (MPI) using a novel cadmium-zinc-telluride (CZT) gamma camera to discriminate artefacts from true perfusion defects.     

Influence of 6-ketocholestanol on skin permeation of 5-aminolevulinic acid and evaluation of chemical stability

The ability of 6-ketocholestanol to increase the skin permeation of the prodrug aminolevulinic acid (5-ALA) was investigated. 6-Ketocholestanol was incorporated together with 5-ALA in four different formulations. Preparations used were a liquid solution/suspension of 5-ALA in buffer, 5-ALA in phospholipid liposomal formulations with and without gelating agent, and finally, a complex cream formulation also including phospholipids. Standard diffusion experiments of 5-ALA using Franz-type diffusion cells and porcine skin were performed. Drug stability was monitored by analyzing the 5-ALA content in the different formulations over time and viewing the preparation for microbial contamination. The analysis of 5-ALA as a nonfluorescent probe was performed after chemical reaction, leading to a fluorescent derivative. The 5-ALA permeation through porcine skin was increased threefold by 6-ketocholestanol in the cream formulation. The chemical stability of 5-ALA in the tested formulations was in the range of about 33 to 72% after an observation period of 28 days. After that time point microbial stability was no longer evident for formulations 2 and 3. Formulation 1 could be observed until day 34, and only formulation 4 showed a microbial stability over the whole observation period of 42 days.     

Cardiac fusion imaging with low-dose computed tomography using prospective electrocardiogram gating

In a 66-year-old patient with prolonged episodes of chest pain, catheter angiography revealed total occlusions of the right coronary artery and the distal circumflex artery (CX) as well as 2 sequential significant stenoses in the proximal CX and one significant stenosis in the proximal left anterior descending artery (LAD). To identify the culprit lesions with their respective territory, noninvasive assessment of viability was performed by F-18 FDG positron emission tomography (PET) and fused with a low-dose computed tomography coronary angiography using prospective electrocardiogram gating. Fused PET/computed tomography coronary angiography images demonstrated a large scar in the inferior myocardium, corresponding to the total occlusion in the right coronary artery, viable myocardium in the territory of the CX, and infarcted scar tissue with partially preserved viability in the anterior myocardium, corresponding to the presumably recanalized lesion in the LAD. The patient was scheduled for revascularization of the lesions in the LAD and the CX to reverse dysfunctional but viable myocardial segments.     

Identification of medication non-adherence factors in adolescent transplant patients: the patient's viewpoint

Studies report a clear association between medication non-adherence and an unfavorable transplant outcome. The adolescent population, in particular, has difficulty adhering to post-transplant medication regimens. The purpose of this study is to identify, categorize and understand the opinions of adolescent transplant patients regarding why they may not take their medications as prescribed. From January to August 2005, nine adolescent kidney transplant patients at an urban medical center were surveyed and asked to rank-order 33 statements regarding their opinions on why adolescents may not take their medications as prescribed. Q-methodology, a powerful tool in subjective study, was used to identify and categorize the viewpoints of adolescents on this subject. Three factors emerged and were labeled to reflect their distinct viewpoints: (1) Medication Issues (e.g. taste, size, frequency, schedule), (2) Troubled Adolescent (e.g. poor home life, depression, overwhelming situation), and (3) Deliberate Non-Adherer (e.g. attention-seeker, infallible attitude). By understanding these different viewpoints and the factors that contribute to them, it may be easier to identify which management approach to non-adherence works best in specific subgroups of patients.     

Quantification of myocardial blood flow with <Superscript>82</Superscript>Rb positron emission tomography: clinical validation with <Superscript>15</Superscript>O-water

Purpose

Quantification of myocardial blood flow (MBF) with generator-produced ⁸²Rb is an attractive alternative for centres without an on-site cyclotron. Our aim was to validate ⁸²Rb-measured MBF in relation to that measured using ¹⁵O-water, as a tracer 100% of which can be extracted from the circulation even at high flow rates, in healthy control subject and patients with mild coronary artery disease (CAD).

Methods

MBF was measured at rest and during adenosine-induced hyperaemia with ⁸²Rb and ¹⁵O-water PET in 33 participants (22 control subjects, aged 30?±?13 years; 11 CAD patients without transmural infarction, aged 60?±?13 years). A one-tissue compartment ⁸²Rb model with ventricular spillover correction was used. The ⁸²Rb flow-dependent extraction rate was derived from ¹⁵O-water measurements in a subset of 11 control subjects. Myocardial flow reserve (MFR) was defined as the hyperaemic/rest MBF. Pearson’s correlation r, Bland-Altman 95% limits of agreement (LoA), and Lin’s concordance correlation ρ _c (measuring both precision and accuracy) were used.

Results

Over the entire MBF range (0.66–4.7 ml/min/g), concordance was excellent for MBF (r?=?0.90, [⁸²Rb–¹⁵O-water] mean difference?±?SD?=?0.04?±?0.66 ml/min/g, LoA?=??1.26 to 1.33 ml/min/g, ρ _c?=?0.88) and MFR (range 1.79–5.81, r?=?0.83, mean difference?=?0.14?±?0.58, LoA?=??0.99 to 1.28, ρ _c?=?0.82). Hyperaemic MBF was reduced in CAD patients compared with the subset of 11 control subjects (2.53?±?0.74 vs. 3.62?±?0.68 ml/min/g, p?=?0.002, for ¹⁵O-water; 2.53?±?1.01 vs. 3.82?±?1.21 ml/min/g, p?=?0.013, for ⁸²Rb) and this was paralleled by a lower MFR (2.65?±?0.62 vs. 3.79?±?0.98, p?=?0.004, for ¹⁵O-water; 2.85?±?0.91 vs. 3.88?±?0.91, p?=?0.012, for ⁸²Rb). Myocardial perfusion was homogeneous in 1,114 of 1,122 segments (99.3%) and there were no differences in MBF among the coronary artery territories (p?>?0.31).

Conclusion

Quantification of MBF with ⁸²Rb with a newly derived correction for the nonlinear extraction function was validated against MBF measured using ¹⁵O-water in control subjects and patients with mild CAD, where it was found to be accurate at high flow rates. ⁸²Rb-derived MBF estimates seem robust for clinical research, advancing a step further towards its implementation in clinical routine.