Do we truly see what we think we see? The role of cognitive bias in pathological interpretation.

In the histomorphological grading of prostate carcinoma, pathologists have regularly assigned comparable scores for the architectural Gleason and the now-obsolete nuclear World Health Organization (WHO) grading systems. Although both systems demonstrate good correspondence between grade and survival, they are based on fundamentally different biological criteria. We tested the hypothesis that this apparent concurrence between the two grading systems originates from an interpretation bias in the minds of diagnostic pathologists, rather than reflecting a biological reality. Three pathologists graded 178 prostatectomy specimens, assigning Gleason and WHO scores on glass slides and on digital images of nuclei isolated out of their architectural context. The results were analysed with respect to interdependencies among the grading systems, to tumour recurrence (PSA relapse > 0.1 ng/ml at 48 months) and robust nuclear morphometry, as assessed by computer-assisted image analysis. WHO and Gleason grades were strongly correlated (r = 0.82) and demonstrated identical prognostic power. However, WHO grades correlated poorly with nuclear morphology (r = 0.19). Grading of nuclei isolated out of their architectural context significantly improved accuracy for nuclear morphology (r = 0.55), but the prognostic power was virtually lost. In conclusion, the architectural organization of a tumour, which the pathologist cannot avoid noticing during initial slide viewing at low magnification, unwittingly influences the subsequent nuclear grade assignment. In our study, the prognostic power of the WHO grading system was dependent on visual assessment of tumour growth pattern. We demonstrate for the first time the influence a cognitive bias can have in the generation of an error in diagnostic pathology and highlight a considerable problem in histopathological tumour grading.

The effects of CYP2D6 and CYP3A activities on the pharmacokinetics of immediate release oxycodone

Background and purpose: There is high interindividual variability in the activity of drug-metabolizing enzymes catalysing the oxidation of oxycodone [cytochrome P450 (CYP) 2D6 and 3A], due to genetic polymorphisms and/or drug-drug interactions. The effects of CYP2D6 and/or CYP3A activity modulation on the pharmacokinetics of oxycodone remains poorly explored. Experimental approach: A randomized crossover double-blind placebo-controlled study was performed with 10 healthy volunteers genotyped for CYP2D6 [six extensive (EM), two deficient (PM/IM) and two ultrarapid metabolizers (UM)]. The volunteers randomly received on five different occasions: oxycodone 0.2 mg·kg−1 and placebo; oxycodone and quinidine (CYP2D6 inhibitor); oxycodone and ketoconazole (CYP3A inhibitor); oxycodone and quinidine+ketoconazole; placebo. Blood samples for plasma concentrations of oxycodone and metabolites (oxymorphone, noroxycodone and noroxymorphone) were collected for 24 h after dosing. Phenotyping for CYP2D6 (with dextromethorphan) and CYP3A (with midazolam) were assessed at each session. Key results: CYP2D6 activity was correlated with oxymorphone and noroxymorphone AUCs and Cmax (−0.71 < Spearman correlation coefficient ρs < −0.92). Oxymorphone Cmax was 62% and 75% lower in PM than EM and UM. Noroxymorphone Cmax reduction was even more pronounced (90%). In UM, oxymorphone and noroxymorphone concentrations increased whereas noroxycodone exposure was halved. Blocking CYP2D6 (with quinidine) reduced oxymorphone and noroxymorphone Cmax by 40% and 80%, and increased noroxycodone AUC∞ by 70%. Blocking CYP3A4 (with ketoconazole) tripled oxymorphone AUC∞ and reduced noroxycodone and noroxymorphone AUCs by 80%. Shunting to CYP2D6 pathway was observed after CYP3A4 inhibition. Conclusions and implications: Drug-drug interactions via CYP2D6 and CYP3A affected oxycodone pharmacokinetics and its magnitude depended on CYP2D6 genotype.

Deer as a potential wildlife reservoir for Parachlamydia species.

Wildlife populations represent an important reservoir for emerging pathogens and trans-boundary livestock diseases. However, detailed information relating to the occurrence of endemic pathogens such as those of the order Chlamydiales in such populations is lacking. During the hunting season of 2008, 863 samples (including blood, conjunctival swabs, internal organs and faeces) were collected in the Eastern Swiss Alps from 99 free-living red deer (Cervus elaphus) and 64 free-living roe deer (Capreolus capreolus) and tested using ELISA, PCR and immunohistochemistry for members of the family Chlamydiaceae and the genus Parachlamydia. Parachlamydia spp. were detected in the conjunctival swabs, faeces and internal organs of both species of deer (2.4% positive, with a further 29.5% inconclusive). The very low occurrence of Chlamydiaceae (2.5%) was in line with serological data (0.7% seroprevalence for Chlamydia abortus). Further investigations are required to elucidate the zoonotic potential, pathogenicity, and distribution of Parachlamydia spp. in wild ruminants.

Energy expenditure during walking and running in obese and nonobese prepubertal children.

We measured body composition and energy expenditure during walking and running on a treadmill in 40 prepubertal children: 23 obese children (9.3 +/- 1.1 years of age; 46 +/- 10 kg (mean +/- SD)) and 17 nonobese matched control children (9.2 +/- 0.6 years of age; 30 +/- 5 kg). Energy expenditure was assessed by indirect calorimetry with a standard open-circuit method. At the same speed of exercise, the energy expenditure was significantly (p < 0.01) greater in obese than in control children, in both boys and girls. Expressed per kilogram of body weight or per kilogram of fat-free mass, the energy expenditure was comparable in the two groups. Obese children had a significantly (p < 0.01) larger pulmonary ventilatory response to exercise than did control children. Heart rate was comparable in boys and girls combined but significantly higher (p < 0.05) in obese subjects, if boys and girls were analyzed separately. These data indicate that walking and running are energetically more expensive for obese children than for children of normal body weight. The knowledge of these energy costs could be useful in devising a physical activity program to be used in the treatment of obese children.

Correlation between vasoconstrictor roles and mRNA expression of alpha1-adrenoceptor subtypes in blood vessels of genetically engineered mice.

We examined the contribution of each alpha(1)-adrenoceptor (AR) subtype in noradrenaline (NAd)-evoked contraction in the thoracic aortas and mesenteric arteries of mice. Compared with the concentration-response curves (CRCs) for NAd in the thoracic aortas of wild-type (WT) mice, the CRCs of mutant mice showed a significantly lower sensitivity. The pD(2) value in rank order is as follows: WT mice (8.21) > alpha(1B)-adrenoceptor knockout (alpha(1B)-KO) (7.77) > alpha(1D)-AR knockout (alpha(1D)-KO) (6.44) > alpha(1B)- and alpha(1D)-AR double knockout (alpha(1BD)-KO) (5.15). In the mesenteric artery, CRCs for NAd did not differ significantly between either WT (6.52) and alpha(1B)-KO mice (7.12) or alpha(1D)-KO (6.19) and alpha(1BD)-KO (6.29) mice. However, the CRC maximum responses to NAd in alpha(1D)- and alpha(1BD)-KO mice were significantly lower than those in WT and alpha(1B)-KO mice. Except in the thoracic aortas of alpha(1BD)-KO mice, the competitive antagonist prazosin inhibited the contraction response to NAd with high affinity. However, prazosin produced shallow Schild slopes in the vessels of mice lacking the alpha(1D)-AR gene. In the thoracic aorta, pA(2) values in WT mice for KMD-3213 and BMY7378 were 8.25 and 8.46, respectively, and in alpha(1B)-KO mice they were 8.49 and 9.13, respectively. In the mesenteric artery, pA(2) values in WT mice for KMD-3213 and BMY7378 were 8.34 and 7.47, respectively, and in alpha(1B)-KO mice they were 8.11 and 7.82, respectively. These pharmacological findings were in fairly good agreement with findings from comparison of CRCs, with the exception of the mesenteric arteries of WT and alpha(1B)-KO mice, which showed low affinities to BMY7378. We performed a quantitative analysis of the mRNA expression of each alpha(1)-AR subtype in these vessels in order to examine the correlation between mRNA expression level and the predominance of each alpha(1)-AR subtype in mediating vascular contraction. The rank order of each alpha(1)-AR subtype in terms of its vasoconstrictor role was in fairly good agreement with the level of expression of mRNA of each subtype, that is, alpha(1D)-AR > alpha(1B)-AR > alpha(1A)-AR in the thoracic aorta and alpha(1D)-AR > alpha(1A)-AR > alpha(1B)-AR in the mesenteric artery. No dramatic compensatory change of alpha(1)-AR subtype in mutant mice was observed in pharmacological or quantitative mRNA expression analysis.

The transcriptome of the arbuscular mycorrhizal fungus Glomus intraradices (DAOM 197198) reveals functional tradeoffs in an obligate symbiont.

? The arbuscular mycorrhizal symbiosis is arguably the most ecologically important eukaryotic symbiosis, yet it is poorly understood at the molecular level. To provide novel insights into the molecular basis of symbiosis-associated traits, we report the first genome-wide analysis of the transcriptome from Glomus intraradices DAOM 197198. ? We generated a set of 25,906 nonredundant virtual transcripts (NRVTs) transcribed in germinated spores, extraradical mycelium and symbiotic roots using Sanger and 454 sequencing. NRVTs were used to construct an oligoarray for investigating gene expression. ? We identified transcripts coding for the meiotic recombination machinery, as well as meiosis-specific proteins, suggesting that the lack of a known sexual cycle in G. intraradices is not a result of major deletions of genes essential for sexual reproduction and meiosis. Induced expression of genes encoding membrane transporters and small secreted proteins in intraradical mycelium, together with the lack of expression of hydrolytic enzymes acting on plant cell wall polysaccharides, are all features of G. intraradices that are shared with ectomycorrhizal symbionts and obligate biotrophic pathogens. ? Our results illuminate the genetic basis of symbiosis-related traits of the most ancient lineage of plant biotrophs, advancing future research on these agriculturally and ecologically important symbionts.

In vivo localisation of radiolabelled monoclonal antibody in human gliomas.

F(ab')2-fragments of the anti-melanoma monoclonal antibody MeI-14 were labelled with 123I for external scanning and with 125I for tissue measurement of radioactivity and injected intravenously into patients scheduled for surgical resection of a glioma. The paired-label study was performed by injecting simultaneously 131I-labelled control (F(ab')2-fragments. The patients were scanned by computerised tomoscintigraphy. After surgery, the activities of 125I and 131I were counted in tumour and normal tissues. The results indicate that there was a low but definite uptake of the antibody in the tumour due to its specificity. The external detection was difficult because of accumulation of antibody fragments in the skull.

Measured and predicted resting metabolic rate in obese and nonobese adolescents.

OBJECTIVES: The validity of equations for the calculation of resting metabolic rate (RMR) were studied and new predictive equations were developed. STUDY DESIGN: The RMR was measured in a sample of 371 10- to 16-year-old prepubertal and postpubertal children. The study group included 193 male (116 nonobese and 77 obese) and 178 female (119 nonobese and 59 obese) subjects; for each group the RMRs predicted from five equations recommended for this age group were compared. The RMR was assessed by indirect calorimetry with a ventilated hood system for 45 minutes after an overnight fast. Body composition was estimated from skin-fold measurements. RESULTS: The mean +/- SD RMR was found to be 5600 +/- 972 kJ/24 hr and 7223 +/- 1220 kJ/24 hr in nonobese and obese boys, and 5112 +/- 632 kJ/24 hr and 6665 +/- 1106 kJ/24 hr in nonobese and obese girls, respectively. All five equations applicable to 10- to 16-year-old children overestimated RMR by 7.5% to 18.1% (p < 0.001 for each equation). Stepwise regression analysis, with independent variables such as age, weight, height, and gender, allowed development of new predictive equations for the calculation of RMR in 10- to 16-year-old boys (RMR = 50.9 Weight (kg) + 25.3 Height (cm) -50.3 Age (yr) + 26.9; R2 = 0.884, p < 0.0001) and girls (RMR = 51.2 Weight (kg) + 24.5 Height (cm) - 207.5 Age (yr) + 1629.8; R2 = 0.824, p < 0.0001). These predictive equations were tested in a second, independent cohort of children (80 male and 61 female subject) and were found to give a reliable estimate of RMR in 10- to 16-year-old obese and nonobese adolescents. CONCLUSIONS: The currently used predictive equations overestimate RMR in 10- to 16-year-old children. The use of the newly developed equations is recommended.

Peak and persistent excess of genetic diversity following an abrupt migration increase.

Genetic diversity is essential for population survival and adaptation to changing environments. Demographic processes (e.g., bottleneck and expansion) and spatial structure (e.g., migration, number, and size of populations) are known to shape the patterns of the genetic diversity of populations. However, the impact of temporal changes in migration on genetic diversity has seldom been considered, although such events might be the norm. Indeed, during the millions of years of a species' lifetime, repeated isolation and reconnection of populations occur. Geological and climatic events alternately isolate and reconnect habitats. We analytically document the dynamics of genetic diversity after an abrupt change in migration given the mutation rate and the number and sizes of the populations. We demonstrate that during transient dynamics, genetic diversity can reach unexpectedly high values that can be maintained over thousands of generations. We discuss the consequences of such processes for the evolution of species based on standing genetic variation and how they can affect the reconstruction of a population's demographic and evolutionary history from genetic data. Our results also provide guidelines for the use of genetic data for the conservation of natural populations.

Mutations inducing divergent shifts of constitutive activity reveal different modes of binding among catecholamine analogues to the beta(2)-adrenergic receptor.

1. We compared the changes in binding energy generated by two mutations that shift in divergent directions the constitutive activity of the human beta(2) adrenergic receptor (beta(2)AR). 2. A constitutively activating mutant (CAM) and the double alanine replacement (AA mutant) of catechol-binding serines (S204A, S207A) in helix 5 were stably expressed in CHO cell lines, and used to measure the binding affinities of more than 40 adrenergic ligands. Moreover, the efficacy of the same group of compounds was determined as intrinsic activity for maximal adenylyl cyclase stimulation in wild-type beta(2)AR. 3. Although the two mutations had opposite effects on ligand affinity, the extents of change were in both cases largely correlated with the degree of ligand efficacy. This was particularly evident if the extra loss of binding energy due to hydrogen bond deletion in the AA mutant was taken into account. Thus the data demonstrate that there is an overall linkage between the configuration of the binding pocket and the intrinsic equilibrium between active and inactive receptor forms. 4. We also found that AA mutation-induced affinity changes for catecholamine congeners gradually lacking ethanolamine substituents were linearly correlated to the loss of affinity that such modifications of the ligand cause for wild-type receptor. This indicates that the strength of bonds between catechol ring and helix 5 is critically dependent on the rest of interactions of the beta-ethanolamine tail with other residues of the beta(2)-AR binding pocket.

Diagnosis and treatment of resistant hypertension.

Hypertension resistant to lifestyle interventions and antihypertensive medications is a common problem encountered by physicians in everyday practice. It is most often defined as a blood pressure remaining ≥ 140/90 mmHg despite the regular intake of at least three drugs lowering blood pressure by different mechanisms, one of them being a diuretic. It now appears justified to include, unless contraindicated or not tolerated, a blocker of the renin-angiotensin system and a calcium channel blocker in this drug regimen, not only to gain antihypertensive efficacy, but also to prevent or regress target organ damage and delay the development of cardiorenal complications. A non-negligible fraction of treatment-resistant hypertension have normal "out of office" blood pressures. Ambulatory blood pressure monitoring and/or home blood pressure recording should therefore be routinely performed to identify patients with true resistant hypertension, i.e. patients who are more likely to benefit from treatment intensification.

Opinions on tobacco control policies in Lausanne, Switzerland, 2003-2006.

OBJECTIVE: To assess the determinants of opinions regarding tobacco control policies in the Swiss general population. METHODS: Cross-sectional study conducted between 2003 and 2006 on a random sample of adult residents of Lausanne, Switzerland, aged 35-75 years (2601 women and 2398 men). Nine questions on smoking policies were applied. RESULTS: Ninety-five percent of responders supported policies that would help smokers to quit, 92% no selling of tobacco to subjects aged less than 16 years, 87% a smoking ban in public places and 86% a national campaign against smoking. A further 77% supported a total ban on tobacco advertising, 74% the reimbursement of nicotine replacement therapies and 70% an increase in the price of cigarettes. A lower support was found for two non-evidence-based interventions total ban of tobacco sales (35%) and promotion of light cigarettes (22%). Never smokers, women, physically active subjects, teetotallers and subjects with lower educational level were more likely to favour stronger measures while no differences were found between age groups. Reimbursement of nicotine replacement therapies was favoured more by current smokers and inactive subjects. CONCLUSION: The vast majority of responders supported the recommended tobacco control policies. Opinions regarding specific interventions vary according to the policy and subjects' characteristics.