Effect of Short-Term Fasting on Systemic Cytochrome P450-Mediated Drug Metabolism in Healthy Subjects: A Randomized, Controlled, Crossover Study Using a Cocktail Approach

Background and Objective: Short-term fasting can alter drug exposure but it is unknown whether this is an effect of altered oral bioavailability and/or systemic clearance. Therefore, the aim of our study was to assess the effect of short-term fasting on oral bioavailability and systemic clearance of different drugs. Methods: In a randomized, controlled, crossover trial, 12 healthy subjects received a single administration of a cytochrome P450 (CYP) probe cocktail, consisting of caffeine (CYP1A2), metoprolol (CYP2D6), midazo

Prospective comparative study of spiral computer tomography and magnetic resonance imaging for detection of hepatocellular carcinoma

BACKGROUND: Hepatocellular carcinoma (HCC) is often detected at a relatively late stage when tumour size prohibits curative surgery. Screening to detect HCC at an early stage is performed for patients at risk. AIM: The aim of this study was to compare prospectively the diagnostic accuracy and classification for management of the two state of the art secondline imaging techniques: triphasic spiral computer tomography (CT) and super paramagnetic iron oxide (SPIO) enhanced magnetic resonance imaging (MRI). PATIENTS: Sixty one patients were evaluated between January 1996 and January 1998. Patients underwent CT and MRI within a mean interval of 6.75 days. METHODS: CT and MRI were evaluated blindly for the presence and number of lesions, characterisation of these lesions, and classification for management. For comparison of the data on characterisation, the CT and MRI findings were compared with histopathological studies of the surgical specimens and/or follow up imaging. Data of patients not lost to follow up were available to January 2001. RESULTS: SPIO enhanced MRI detected more lesions and overall smaller lesions than triphasic spiral CT (number of lesions 189 v 124; median diameter 1.0 v 1.8 cm; Spearman rank's correlation coefficient 0.63, p<0.001). There was no significant difference in accuracy between CT and MRI for lesion characterisation. The agreement in classification for management was very good (weighted kappa 0.91, 95% CI 0.83-0.99). CONCLUSION: SPIO enhanced MRI detects more and smaller lesions, but both techniques are comparable in terms of classification for management. SPIO enhanced MRI may be preferred as there is no exposure to ionising radiation

Production of Slit2 LRR domains in mammalian cells for structural studies and the structure of human Slit2 domain 3

Slit2 and Roundabout 1 (Robo1) provide a key ligand-receptor interaction for the navigation of commissural neurons during the development of the central nervous system. Slit2 is a large multidomain protein containing an unusual domain organization of four tandem leucine-rich repeat (LRR) domains at its N-terminus. These domains are well known to mediate protein-protein interactions; indeed, the Robo1-binding region has been mapped to the concave face of the second LRR domain. It has also been shown that the fourth LRR domain may mediate Slit dimerization and that both the first and second domains can bind heparin. Thus, while roles have been ascribed for three of the LRR domains, there is still no known role for the third domain. Each of the four LRR domains from human Slit2 have now been successfully expressed in milligram quantities using expression in mammalian cells. Here, the crystallization of the second and third LRR domains and the structure of the third LRR domain are presented. This is the first structure of an LRR domain from human Slit2, which has an extra repeat compared with the Drosophila homologue. It is proposed that a highly conserved patch of surface residues on the concave face may mediate any protein-protein interactions involving this LRR domain, a result that will be useful in guiding further studies on Slit2

Cloning, expression, crystallization and preliminary X-ray analysis of the first two Ig domains from human Roundabout 1 (Robo1).

Activation of Roundabout 1 (Robo1) by Slit proteins results in axon repulsion from the midline. Robo1 is a large transmembrane receptor expressed on the axon growth cone and the minimal Robo1-binding region required for Slit activation has been mapped to the N-terminal Ig1-2 domains. The cDNA encoding the first two Ig domains of Robo1 has been cloned and the protein has been expressed in HEK293 EBNA-1 mammalian cells. Here, the purification and crystallization conditions of this Robo1 construct are reported. The crystals are orthorhombic, space group P21212, with unit-cell parameters a = 38.8, b = 69.4, c = 103.3 Å and one molecule in the asymmetric unit. X-ray diffraction data have been collected to 2.8 Å resolution on beamline ID29 at the ESRF

Characterization of four lactose monophosphates by application of 31P-, 13C-, and 1H-N.M.R. spectroscopy

By a combination of ion-exchange chromatography and h.p.l.c., two fractions (A and B) have been obtained from pharmaceutical-grade lactose, each containing a mixture of lactose monophosphates. 31P-N.m.r. and 13C-n.m.r. spectroscopic analysis indicated A to contain the 6- and 6-phosphates and B to contain the 3- and 4-phosphates. Application of 2D-13C-1H COSY and 2D-1H-1H COSY afforded 1H-n.m.r. assignments for all protons in all four compounds. The observed 31P-1H and 31C-1H couplings are interpreted in terms of preferred orientations of the phosphate group in each compound

Crystal structures of TAFI elucidate the inactivation mechanism of activated TAFI: a novel mechanism for enzyme autoregulation

Thrombin-activatable fibrinolysis inhibitor (TAFI) is a pro-metallocarboxypeptidase that can be proteolytically activated (TAFIa). TAFIa is unique among carboxypeptidases in that it spontaneously inactivates with a short half-life, a property that is crucial for its role in controlling blood clot lysis.We studied the intrinsic instability of TAFIa by solving crystal structures of TAFI, a TAFI inhibitor (GEMSA) complex and a quadruple TAFI mutant (70-fold more stable active enzyme). The crystal structures show that TAFIa stability is directly related to the dynamics of a 55-residue segment (residues 296-350) that includes residues of the active site wall. Dynamics of this flap are markedly reduced by the inhibitor GEMSA, a known stabilizer of TAFIa, and stabilizing mutations. Our data provide the structural basis for a model of TAFI auto-regulation: in zymogen TAFI the dynamic flap is stabilized by interactions with the activation peptide. Release of the activation peptide increases dynamic flap mobility and in time this leads to conformational changes that disrupt the catalytic site and expose a cryptic thrombincleavage site present at Arg302. This represents a novel mechanism of enzyme control that enables TAFI to regulate its activity in plasma in the absence of specific inhibitors. (Blood. 2008;112: 2803-2809)

Translation and validation of the Revised Dental Beliefs Survey (DBS-R) in China

Patient perceptions of behaviours and attitudes of dentists are associated with dental fear and poor dental attendance in Western countries. However, there is a paucity of research exploring patient perceptions of the dentist in China. One reason for this may be the lack of a valid and reliable scale in Chinese (Standard Mandarin) to measure this. This study aimed to translate the Revised Dental Beliefs Survey (DBS-R) into Chinese and then explore the reliability and validity of this measure (both the short and longer versions) in a Chinese population. We translated the DBS-R using the forwards-backwards method and pilot tested it on a small sample of adults in China. Following this, 480 Chinese adults completed the newly translated scale, as well as well as a standardised dental anxiety questionnaire (the Modified Dental Anxiety Scale Chinese version) to test convergent validity. 109 participants completed the DBS-R again 2 weeks later for test-retest reliability. Both versions of the Chinese DBS-R were internally consistent and demonstrated convergent validity; test-retest reliability was also good. Both versions of the scale performed similarly, but for now we would suggest the 28-item version may be superior as items relating to the technical competence of the dentist appear important to Chinese adults

Structural insights into the Slit-Robo complex

Slits are large multidomain leucine-rich repeat (LRR)-containing proteins that provide crucial guidance cues in neuronal and vascular development. More recently, Slits have been implicated in heart morphogenesis, angiogenesis, and tumor metastasis. Slits are ligands for the Robo (Roundabout) receptors, which belong to the Ig superfamily of transmembrane signaling molecules. The Slit-Robo interaction is mediated by the second LRR domain of Slit and the two N-terminal Ig domains of Robo, but the molecular details of this interaction and how it induces signaling remain unclear. Here we describe the crystal structures of the second LRR domain of human Slit2 (Slit2 D2), the first two Ig domains of its receptor Robo1 (Ig1-2), and the minimal complex between these proteins (Slit2 D2-Robo1 Ig1). Slit2 D2 binds with its concave surface to the side of Ig1 with electrostatic and hydrophobic contact regions mediated by residues that are conserved in other family members. Surface plasmon resonance experiments and a mutational analysis of the interface confirm that Ig1 is the primary domain for binding Slit2. These structures provide molecular insight into Slit-Robo complex formation and will be important for the development of novel cancer therapeutics

Aortic valve calcification and mild tricuspid regurgitation but no clinical heart disease after 8 years of dopamine agonist therapy for prolactinoma

Objective: Treatment with ergot-derived dopamine agonists, pergolide, and cabergoline has been associated with an increased frequency of valvular heart disease in Parkinson's disease. The aim of the present study was to assess the prevalence of valvular heart disease in patients treated with dopamine agonists for prolactinomas. Design: This was a cross-sectional study. Patients: We performed two-dimensional and Doppler echocardiography in 78 consecutive patients with prolactinoma ( mean age 47 +/- 1.4 yr, 26% male, 31% macroprolactinoma) treated with dopamine agonists for at least 1 yr ( mean 8 +/- 0.6 yr) and 78 control subjects. Patients were classified according to treatment: patients treated with cabergoline ( group 1: n = 47) and patients not treated with cabergoline ( group 2: n = 31). Results: Clinically relevant valvular heart disease was present in 12% of patients (nine of 78) vs. 17% of controls ( 13 of 78) ( P = 0.141) and 17% ( eight of 47) of patients treated with cabergoline vs. 3% ( one of 31) of patients not treated with cabergoline ( P = 0.062). Mild tricuspid regurgitation was present in 41% of patients vs. 26% of controls ( P = 0.042), and aortic valve calcification was present in 40% of patients, compared with 18% of controls ( P = 0.003). There was no relation between the cumulative dose of cabergoline and the presence of mild, moderate, or severe valve regurgitation. Conclusion: Several years of dopamine agonist treatment in patients with prolactinomas is associated with increased prevalence of aortic valve calcification and mild tricuspid regurgitation but not with clinically relevant valvular heart disease. Therefore, additional studies on the adverse cardiac effects of dopaminergic drugs in prolactinoma are warranted, especially in patients with much longer use of these drugs