Characterization of novel StAR (steroidogenic acute regulatory protein) mutations causing non-classic lipoid adrenal hyperplasia

Context Steroidogenic acute regulatory protein (StAR) is crucial for transport of cholesterol to mitochondria where biosynthesis of steroids is initiated. Loss of StAR function causes lipoid congenital adrenal hyperplasia (LCAH). Objective StAR gene mutations causing partial loss of function manifest atypical and may be mistaken as familial glucocorticoid deficiency. Only a few mutations have been reported. Design To report clinical, biochemical, genetic, protein structure and functional data on two novel StAR mutations, and to compare them with published literature. Setting Collaboration between the University Children's Hospital Bern, Switzerland, and the CIBERER, Hospital Vall d'Hebron, Autonomous University, Barcelona, Spain. Patients Two subjects of a non-consanguineous Caucasian family were studied. The 46,XX phenotypic normal female was diagnosed with adrenal insufficiency at the age of 10 months, had normal pubertal development and still has no signs of hypergonodatropic hypogonadism at 32 years of age. Her 46,XY brother was born with normal male external genitalia and was diagnosed with adrenal insufficiency at 14 months. Puberty was normal and no signs of hypergonadotropic hypogonadism are present at 29 years of age. Results StAR gene analysis revealed two novel compound heterozygote mutations T44HfsX3 and G221S. T44HfsX3 is a loss-of-function StAR mutation. G221S retains partial activity (~30%) and is therefore responsible for a milder, non-classic phenotype. G221S is located in the cholesterol binding pocket and seems to alter binding/release of cholesterol. Conclusions StAR mutations located in the cholesterol binding pocket (V187M, R188C, R192C, G221D/S) seem to cause non-classic lipoid CAH. Accuracy of genotype-phenotype prediction by in vitro testing may vary with the assays employed.

Vibronic Spectra of Jet-Cooled 2-Aminopurine center dot H2O Clusters Studied by UV Resonant Two-Photon Ionization Spectroscopy and Quantum Chemical Calculations

For understanding the major- and minor-groove hydration patterns of DNAs and RNAs, it is important to understand the local solvation of individual nucleobases at the molecular level. We have investigated the 2-aminopurine center dot H2O. monohydrate by two-color resonant two-photon ionization and UV/UV hole-burning spectroscopies, which reveal two isomers, denoted A and B. The electronic spectral shift delta nu of the S-1 <- S-0 transition relative to bare 9H-2-aminopurine (9H-2AP) is small for isomer A (-70 cm(-1)), while that of isomer B is much larger (delta nu = 889 cm(-1)). B3LYP geometry optimizations with the TZVP basis set predict four cluster isomers, of which three are doubly H-bonded, with H2O acting as an acceptor to a N-H or -NH2 group and as a donor to either of the pyrimidine N sites. The "sugar-edge" isomer A is calculated to be the most stable form with binding energy D-e = 56.4 kJ/mol. Isomers B and C are H-bonded between the -NH2 group and pyrimidine moieties and are 2.5 and 6.9 kJ/mol less stable, respectively. Time-dependent (TD) B3LYP/TZVP calculations predict the adiabatic energies of the lowest (1)pi pi* states of A and B in excellent agreement with the observed 0(0)(0) bands; also, the relative intensities of the A and B origin bands agree well with the calculated S-0 state relative energies. This allows unequivocal identification of the isomers. The R2PI spectra of 9H-2AP and of isomer A exhibit intense low-frequency out-of-plane overtone and combination bands, which is interpreted as a coupling of the optically excited (1)pi pi* state to the lower-lying (1)n pi* dark state. In contrast, these overtone and combination bands are much weaker for isomer B, implying that the (1)pi pi* state of B is planar and decoupled from the (1)n pi* state. These observations agree with the calculations, which predict the (1)n pi* above the (1)pi pi* state for isomer B but below the (1)pi pi* for both 9H-2AP and isomer A.

Analysis of lorazepam and its 30-glucuronide in human urine by capillary electrophoresis: evidence for the formation of two distinct diastereoisomeric glucuronides

Lorazepam (LOR) is a 3-hydroxy-1,4-benzodiazepine that is chiral and undergoes enantiomerization at room temperature. In humans, about 75% of the administered dose of LOR is excreted in the urine as its 30-glucuronide. CE-MS with negative ESI was used to confirm the presence of LOR-30-glucuronide in urines that stemmed from a healthy individual who ingested 1 or 2 mg LOR, whereas free LOR could be detected in extracts prepared from enzymatically hydrolyzed urines. As the 30-glucuronidation reaction occurs at the chiral center of the molecule, two diastereoisomers can theoretically be formed, molecules that can no longer interconvert. The stereoselective formation of LOR glucuronides in humans and in vitro was investigated. MEKC analysis of extracts of the nonhydrolyzed urines suggested the presence of the two different LOR glucuronides in the urine. The formation of the same two diastereoisomers was also observed in vitro employing incubations of LOR with human liver microsomes in the presence of uridine 5'-diphospho-glucuronic acid as coenzyme. The absence of other coenzymes excluded the formation of phase I or other phase II metabolites of LOR. Both results revealed a stereoselectivity, one diastereoisomer being formed in a higher amount than the other. After enzymatic hydrolysis using beta-glucuronidase, these peaks could not be detected any more. Instead, LOR was monitored. Analysis of the extracts prepared from enzymatically hydrolyzed urines by MEKC in the presence of 2-hydroxypropyl-beta-CD revealed the enantiomerization process of LOR (observation of two peaks of equal magnitude connected with a plateau zone). The data presented provide for the first time the evidence of the stereoselectivity of the LOR glucuronidation in humans.

Subcolumnar Dendritic and Axonal Organization of Spiny Stellate and Star Pyramid Neurons within a Barrel in Rat Somatosensory Cortex

Excitatory neurons at the level of cortical layer 4 in the rodent somatosensory barrel field often display a strong eccentricity in comparison with layer 4 neurons in other cortical regions. In rat, dendritic symmetry of the 2 main excitatory neuronal classes, spiny stellate and star pyramid neurons (SSNs and SPNs), was quantified by an asymmetry index, the dendrite-free angle. We carefully measured shrinkage and analyzed its influence on morphological parameters. SSNs had mostly eccentric morphology, whereas SPNs were nearly radially symmetric. Most asymmetric neurons were located near the barrel border. The axonal projections, analyzed at the level of layer 4, were mostly restricted to a single barrel except for those of 3 interbarrel projection neurons. Comparing voxel representations of dendrites and axon collaterals of the same neuron revealed a close overlap of dendritic and axonal fields, more pronounced in SSNs versus SPNs and considerably stronger in spiny L4 neurons versus extragranular pyramidal cells. These observations suggest that within a barrel dendrites and axons of individual excitatory cells are organized in subcolumns that may confer receptive field properties such as directional selectivity to higher layers, whereas the interbarrel projections challenge our view of barrels as completely independent processors of thalamic input.

Erythropoietin or Darbepoetin for patients with cancer--meta-analysis based on individual patient data

BACKGROUND: Erythropoiesis-stimulating agents (ESAs) reduce anemia in cancer patients and may improve quality of life, but there are concerns that ESAs might increase mortality. OBJECTIVES: Our objectives were to examine the effect of ESAs and identify factors that modify the effects of ESAs on overall survival, progression free survival, thromboembolic and cardiovascular events as well as need for transfusions and other important safety and efficacy outcomes in cancer patients. SEARCH STRATEGY: We searched the Cochrane Library, Medline, Embase and conference proceedings for eligible trials. Manufacturers of ESAs were contacted to identify additional trials. SELECTION CRITERIA: We included randomized controlled trials comparing epoetin or darbepoetin plus red blood cell transfusions (as necessary) versus red blood cell transfusions (as necessary) alone, to prevent or treat anemia in adult or pediatric cancer patients with or without concurrent antineoplastic therapy. DATA COLLECTION AND ANALYSIS: We performed a meta-analysis of randomized controlled trials comparing epoetin alpha, epoetin beta or darbepoetin alpha plus red blood cell transfusions versus transfusion alone, for prophylaxis or therapy of anemia while or after receiving anti-cancer treatment. Patient-level data were obtained and analyzed by independent statisticians at two academic departments, using fixed-effects and random-effects meta-analysis. Analyses were according to the intention-to-treat principle. Primary endpoints were on study mortality and overall survival during the longest available follow-up, regardless of anticancer treatment, and in patients receiving chemotherapy. Tests for interactions were used to identify differences in effects of ESAs on mortality across pre-specified subgroups. The present review reports only the results for the primary endpoint. MAIN RESULTS: A total of 13933 cancer patients from 53 trials were analyzed, 1530 patients died on-study and 4993 overall. ESAs increased on study mortality (combined hazard ratio [cHR] 1.17; 95% CI 1.06-1.30) and worsened overall survival (cHR 1.06; 95% CI 1.00-1.12), with little heterogeneity between trials (I(2) 0%, p=0.87 and I(2) 7.1%, p=0.33, respectively). Thirty-eight trials enrolled 10441 patients receiving chemotherapy. The cHR for on study mortality was 1.10 (95% CI 0.98-1.24) and 1.04; 95% CI 0.97-1.11) for overall survival. There was little evidence for a difference between trials of patients receiving different cancer treatments (P for interaction=0.42). AUTHORS' CONCLUSIONS: ESA treatment in cancer patients increased on study mortality and worsened overall survival. For patients undergoing chemotherapy the increase was less pronounced, but an adverse effect could not be excluded.

The importance of partnership factors and individual factors associated with absent or inconsistent condom use in heterosexuals: a cross-sectional study.

OBJECTIVES Decisions to use condoms are made within partnerships. We examined the associations between inconsistent or no condom use and individual and partnership characteristics. We also examined the relative importance of individual versus partnership factors. METHODS Cross-sectional study of heterosexual individuals enrolled from the sexually transmitted infections (STI) outpatient clinic in Amsterdam, the Netherlands, from May to August 2010. Participants completed a questionnaire about sexual behaviour with the last four partners in the preceding year. Participant and partnership factors associated with inconsistent or no condom use in steady and casual partnerships were identified. RESULTS 2144 individuals were included, reporting 6401 partnerships; 54.7% were female, the median age was 25 (IQR 22-30) years and 79.9% were Dutch. Inconsistent or no condom use occurred in 13.9% of 2387 steady partnerships and in 33.5% of 4014 casual partnerships. There was statistical evidence of associations between inconsistent condom use in steady partnerships and ethnic concordance, longer duration, higher number of sex acts, practising anal sex, and sex-related drug use. In casual partnerships, associations were found with having an older partner, ethnic concordance, longer duration, higher number of sex acts, anal sex, sex-related drug use, ongoing partnerships and concurrency. In multivariable models, partnership factors explained 50.9% of the variance in steady partnerships and 70.1% in casual partnerships compared with 10.5% and 15.4% respectively for individual factors. CONCLUSIONS Among heterosexual STI clinic attendees in Amsterdam, partnership factors are more important factors related with inconsistent condom use than characteristics of the individual.

0 and Metacognition: Relations and individual differences in 2nd graders

Introduction. Prospective Memory (PM), defined as the ability to remember to perform intended activities at some point in the future (Kliegel & Jäger, 2007), is crucial to succeed in everyday life. PM seems to increase over the childhood years (Zimmermann & Meier, 2006), but yet little is known about PM competences in children in general, but also about factors that influence its development. Currently, a number of studies has focused on factors that might influence PM performance, with EF being potentially influencing mechanisms (Ford, Driscoll, Shum & Macaulay, 2012; Mahy & Moses, 2011). Also metacognitive processes (MC: monitoring and control) are assumed to be involved while learning or optimizing one’s performance (Krebs & Roebers, 2010; 2012; Roebers, Schmid, & Roderer, 2009). Yet, the empirical relation between PM, EF and MC remains rather unclear. We intend to examine relations and explain individual differences in PM performance. Method. An empirical cross-sectional study on 120 2nd graders will be presented. Participants completed six EF tasks (a Stroop, two Updating Tasks, two Shifting Tasks, a Flanker Task), a computerised event-based PM Task and a MC spelling task. Children were tested individually in two sessions of 30 minutes each. Each of the three EF components defined by Miyake, Friedman, Emerson, Witzki & Howerter (2002) was represented by two variables. PM performance was represented by PM accuracy. Metacognitive processes (control, monitoring) were represented separately. Results. Preliminary analyses (SEM) indicate a substantial association between EF (updating, inhibition) and PM. Further, MC seems to be significantly related only to EF. We will explore whether metacognitive monitoring is related to PM monitoring (Roebers, 2002; Mantylä, 2007). As to EF and MC, we expect the two domains to be empirically well distinguishable and nevertheless substantially interrelated. Discussion. The results are discussed on a broader and interindividual level.

Long-term outcomes of biodegradable versus durable polymer drug-eluting stents in patients with acute ST-segment elevation myocardial infarction: a pooled analysis of individual patient data from three randomised trials

Aims: Arterial plaque rupture and thrombus characterise ST-elevation myocardial infarction (STEMI) and may aggravate delayed arterial healing following durable polymer drug-eluting stent (DP-DES) implantation. Biodegradable polymer (BP) may improve biocompatibility. We compared long-term outcomes in STEMI patients receiving BP-DES vs. durable polymer sirolimus-eluting stents (DP-SES). Methods and results: We pooled individual patient-level data from three randomised clinical trials (ISAR-TEST-3, ISAR-TEST-4 and LEADERS) comparing outcomes from BP-DES with DP-SES at four years. The primary endpoint (MACE) comprised cardiac death, MI, or target lesion revascularisation (TLR). Secondary endpoints were TLR, cardiac death or MI, and definite or probable stent thrombosis. Of 497 patients with STEMI, 291 received BP-DES and 206 DP-SES. At four years, MACE was significantly reduced following treatment with BP-DES (hazard ratio [HR] 0.59, 95% CI: 0.39-0.90; p=0.01) driven by reduced TLR (HR 0.54, 95% CI: 0.30-0.98; p=0.04). Trends towards reduction were seen for cardiac death or MI (HR 0.63, 95% CI: 0.37-1.05; p=0.07) and definite or probable stent thrombosis (3.6% vs. 7.1%; HR 0.49, 95% CI: 0.22-1.11; p=0.09). Conclusions: In STEMI, BP-DES demonstrated superior clinical outcomes to DP-SES at four years. Trends towards reduced cardiac death or myocardial infarction and reduced stent thrombosis require corroboration in specifically powered trials.

Individual and spousal education, mortality and life expectancy in Switzerland: a national cohort study.

BACKGROUND Household measures of socioeconomic position may better account for the shared nature of material resources, lifestyle, and social position of cohabiting persons, but household measures of education are rarely used. We aimed to evaluate the association of combined educational attainment of married couples on mortality and life expectancy in Switzerland. METHODS The study included 3 496 163 ever-married persons aged ≥30 years. The 2000 census was linked to mortality records through 2008. Mortality by combined educational attainment was assessed by gender-age-specific HRs, with 95% CIs from adjusted models, life expectancy was derived using abridged life tables. RESULTS Having a less educated partner was associated with increased mortality. For example, the HR comparing men aged 50-64 years with tertiary education married to women with tertiary education to men with compulsory education married to women with compulsory education was 2.05 (1.92-2.18). The estimated remaining life expectancy in tertiary educated men aged 30 years married to women with tertiary education was 4.6 years longer than in men with compulsory education married to women with compulsory education. The gradient based on individual education was less steep: the HR comparing men aged 50-64 years with tertiary education with men with compulsory education was 1.74 (1.67-1.81). CONCLUSIONS Using individual educational attainment of married persons is common in epidemiological research, but may underestimate the combined effect of education on mortality and life expectancy. These findings are relevant to epidemiologic studies examining socio-demographic characteristics or aiming to adjust results for these characteristics.

Evidence for mental ability related individual differences in the attentional blink obtained by an analysis of the P300 component

Attentional blink (AB) refers to impaired identification of a target (T2) when this target follows a preceding target (T1) after about 150-450 ms within a stream of rapidly presented stimuli. Previous research on a possible relation between AB and mental ability (MA) turned out to be highly ambiguous. The present study investigated MA-related individual differences in consolidation of T2 in working memory during the AB as indicated by the P300 component of the event-related potential. Thirty high (HA) and 30 low MA (LA) female participants performed an AB task while their brain activity was recorded. The AB did not differ between the two groups. HA individuals exhibited a larger P300 amplitude and longer P300 latencies during the AB suggesting higher mental effort. This higher mental effort, however, did not result in better performance presumably because of more competition between target and distractor stimuli in HA than LA individuals.

Individual Video Training iVT - Development of a National Infrastructure for Video-Supported Learning

Introduction: Video‐Supported Learning is particularly effective when it comes to skills and behaviors. Video registration of patient‐physician interviews, class room instruction or practical skills allow it to learners themselves, their peers, and their tutors to assess the quality of the learner's performance, to give specific feedback, and to make suggestions for improvement. Methods: In Switzerland, four pedagogical universities and two medical faculties joined to initiate the development of a national infrastructure for Video Supported Learning. The goal was to have a system that is simple to use, has most steps automated, provides the videos over the Internet, and has a sophisticated access control. Together with SWITCH, the national IT‐Support‐Organisation for Swiss Universities, the program iVT (Individual Video Training) was developed by integrating two preexisting technologies. The first technology is SWITCHcast, a podcast system. With SWITCHcast, videos are automatically uploaded to a server as soon as the registration is over. There the videos are processed and converted to different formats. The second technology is the national Single Logon System AAI (Authentification and Authorization Infrastructure) that enables iVT to link each video with the corresponding learner. The learner starts the registration with his Single Logon. Thus, the video can unambiguously be assigned. Via his institution's Learning Management System (LMS), the learner can access his video and give access to his video to peers and tutors. Results: iVT is now used at all involved institutions. The system works flawlessly. In Bern, we use iVT for the communications skills training in the forth and sixth year. Since students meet with patient actors alone, iVT is also used to certify attendance. Students are encouraged to watch the videos of the interview and the feedback of the patient actor. The offer to discuss a video with a tutor was not used by the students. Discussion: We plan to expand the use of iVT by making peer assessment compulsory. To support this, annotation capabilities are currently added to iVT. We also want to use iVT in training of practical skills, again for self as well as for peer assessment.  At present, we use iVT for quality control of patient actor's performance.

Controlled assembly and single electron charging of monolayer protected Au 144 clusters: an electrochemistry and scanning tunneling spectroscopy study

Single gold particles may serve as room temperature single electron memory units because of their size dependent electronic level spacing. Here, we present a proof-of-concept study by electrochemically controlled scanning probe experiments performed on tailor-made Au particles of narrow dispersity. In particular, the charge transport characteristics through chemically synthesized hexane-1-thiol and 4-pyridylbenzene-1-thiol mixed monolayer protected Au144 clusters (MPCs) by differential pulse voltammetry (DPV) and electrochemical scanning tunneling spectroscopy (EC-STS) are reported. The pyridyl groups exposed by the Au-MPCs enable their immobilization on Pt(111) substrates. By varying the humidity during their deposition, samples coated by stacks of compact monolayers of Au-MPCs or decorated with individual, laterally separated Au-MPCs are obtained. DPV experiments with stacked monolayers of Au144-MPCs and EC-STS experiments with laterally separated individual Au144-MPCs are performed both in aqueous and ionic liquid electrolytes. Lower capacitance values were observed for individual clusters compared to ensemble clusters. This trend remains the same irrespective of the composition of the electrolyte surrounding the Au144-MPC. However, the resolution of the energy level spacing of the single clusters is strongly affected by the proximity of neighboring particles.

High-Spin Molecules: Synthesis, X-ray Characterization, and Magnetic Behavior of Two New Cyano-Bridged Ni II 9 Mo V 6 and Ni II 9 W V 6 Clusters with a S = 12 Ground State

The preparations, X-ray structures, and magnetic characterizations are presented for two new pentadecanuclear cluster compounds:  [NiII{NiII(MeOH)3}8(μ-CN)30{MV(CN)3}6]·xMeOH·yH2O (MV = MoV (1) with x = 17, y = 1; MV = WV (2) with x = 15, y = 0). Both compounds crystallize in the monoclinic space group C2/c, with cell dimensions of a = 28.4957(18) Å, b = 19.2583(10) Å, c = 32.4279(17) Å, β = 113.155(6)°, and Z = 4 for 1 and a = 28.5278(16) Å, b = 19.2008(18) Å, c = 32.4072(17) Å, β = 113.727(6)°, and Z = 4 for 2. The structures of 1 and 2 consist of neutral cluster complexes comprising 15 metal ions, 9 NiII and 6 MV, all linked by μ-cyano ligands. Magnetic susceptibilities and magnetization measurements of compounds 1 and 2 in the crystalline and dissolved state indicate that these clusters have a S = 12 ground state, originating from intracluster ferromagnetic exchange interactions between the μ-cyano-bridged metal ions of the type NiII−NC−MV. Indeed, these data show clearly that the cluster molecules stay intact in solution. Ac magnetic susceptibility measurements reveal that the cluster compounds exhibit magnetic susceptibility relaxation phenomena at low temperatures since, with nonzero dc fields, χ‘ ‘M has a nonzero value that is frequency dependent. However, there appears no out-of-phase (χ‘ ‘M) signal in zero dc field down to 1.8 K, which excludes the expected signature for a single molecule magnet. This finding is confirmed with the small uniaxial magnetic anisotropy value for D of 0.015 cm-1, deduced from the high-field, high-frequency EPR measurement, which distinctly reveals a positive sign in D. Obviously, the overall magnetic anisotropy of the compounds is too low, and this may be a consequence of a small single ion magnetic anisotropy combined with the highly symmetric arrangement of the metal ions in the cluster molecule.

Tracking individual membrane proteins and their biochemistry: The power of direct observation

The advent of single molecule fluorescence microscopy has allowed experimental molecular biophysics and biochemistry to transcend traditional ensemble measurements, where the behavior of individual proteins could not be precisely sampled. The recent explosion in popularity of new super-resolution and super-localization techniques coupled with technical advances in optical designs and fast highly sensitive cameras with single photon sensitivity and millisecond time resolution have made it possible to track key motions, reactions, and interactions of individual proteins with high temporal resolution and spatial resolution well beyond the diffraction limit. Within the purview of membrane proteins and ligand gated ion channels (LGICs), these outstanding advances in single molecule microscopy allow for the direct observation of discrete biochemical states and their fluctuation dynamics. Such observations are fundamentally important for understanding molecular-level mechanisms governing these systems. Examples reviewed here include the effects of allostery on the stoichiometry of ligand binding in the presence of fluorescent ligands; the observation of subdomain partitioning of membrane proteins due to microenvironment effects; and the use of single particle tracking experiments to elucidate characteristics of membrane protein diffusion and the direct measurement of thermodynamic properties, which govern the free energy landscape of protein dimerization. The review of such characteristic topics represents a snapshot of efforts to push the boundaries of fluorescence microscopy of membrane proteins to the absolute limit.