The Wilcoxon–Mann–Whitney test under scrutiny

The Wilcoxon–Mann–Whitney (WMW) test is often used to compare the means or medians of two independent, possibly nonnormal distributions. For this problem, the true significance level of the large sample approximate version of the WMW test is known to be sensitive to differences in the shapes of the distributions. Based on a wide ranging simulation study, our paper shows that the problem of lack of robustness of this test is more serious than is thought to be the case. In particular, small differences in variances and moderate degrees of skewness can produce large deviations from the nominal type I error rate. This is further exacerbated when the two distributions have different degrees of skewness. Other rank‐based methods like the Fligner–Policello (FP) test and the Brunner–Munzel (BM) test perform similarly, although the BM test is generally better. By considering the WMW test as a two‐sample T test on ranks, we explain the results by noting some undesirable properties of the rank transformation. In practice, the ranked samples should be examined and found to sufficiently satisfy reasonable symmetry and variance homogeneity before the test results are interpreted. Copyright © 2009 John Wiley & Sons, Ltd.     

A simulation study provided sample size guidance for differential item functioning (DIF) studies using short scales

ObjectiveDifferential item functioning (DIF) analyses are increasingly used to evaluate health-related quality of life (HRQoL) instruments, which often include relatively short subscales. Computer simulations were used to explore how various factors including scale length affect analysis of DIF by ordinal logistic regression.Study Design and settingSimulated data, representative of HRQoL scales with four-category items, were generated. The power and type I error rates of the DIF method were then investigated when, respectively, DIF was deliberately introduced and when no DIF was added. The sample size, scale length, floor effects (FEs) and significance level were varied.ResultsWhen there was no DIF, type I error rates were close to 5%. Detecting moderate uniform DIF in a two-item scale required a sample size of 300 per group for adequate (>80%) power. For longer scales, a sample size of 200 was adequate. Considerably larger sample sizes were required to detect nonuniform DIF, when there were extreme FEs or when a reduced type I error rate was required.ConclusionThe impact of the number of items in the scale was relatively small. Ordinal logistic regression successfully detects DIF for HRQoL instruments with short scales. Sample size guidelines are provided.     

Somatostatin effects on the proteome of the LNCaP cell-line

Some clinical results indicate that somatostatin (sms) might be useful in the treatment of advanced prostate cancer (HRPC). Because of its transient in vivo half-life only more stable derivatives of sms are of interest in this context. Recent studies have shown that natural sms can be conjugated to a carbohydrate (smsdx) with preservation of sms-like effects on the prostatic tumor cell proteome. The present study identifies some of the affected proteins in an effort to elucidate pathways and proteins that might be of importance for the potential usefulness of sms treatment in HRPC. After incubating the LNCaP cell-line with sms14/smsdx, comparative proteomics was used for analysing and identifying affected proteins. Protein expression patterns were analysed with two-dimensional polyacrylamide gel electrophoresis and mass spectrometry. Catalytic mitochondrial and mitochondrial-associated proteins were significantly affected (fold change approximately 2 or higher) and they were in general up-regulated. Apoptosis-related proteins were both up-regulated (VDAC1, VDAC2) and down-regulated (PRDX2, TCTP). The fold change was >2 for PRDX2 and <2 for the others. There was a strong agreement between sms and smsdx on the up- and down-regulation of proteins. Sms/smsdx triggered up-regulation of catalytic mitochondrial proteins and seemed to affect apoptosis-related proteins. This could indicate important pathways on which smsdx might be able to curb the progression of HRPC. The results encourage a pending clinical phase II study.     

Elevated ACE activity is not associated with asthma, COPD, and COPD co-morbidity

The angiotensin-converting enzyme (ACE) gene is a potential candidate gene for risk of asthma, COPD, and COPD co-morbidity. In 9034 Danish adults, we determined whether individuals homozygous or heterozygous for the ACE D allele are at greater risk of asthma, COPD, or COPD co-morbidity compared with ACE II homozygous individuals. In the general population, serum ACE activity increased with the number of D alleles (Kruskal-Wallis ANOVA: II vs. ID, p < 0.001; ID vs. DD, p < 0.001); however, this did not translate into altered risk of asthma or COPD. In the general population, the odds ratio (95% confidence interval) for asthma was 1.2 (0.9–1.4) for ID individuals and 1.2 (0.9–1.5) for DD individuals compared with II individuals. In the general population, the odds ratio for COPD was 0.9 (0.8–1.1) for ID individuals and 1.0 (0.8–1.2) for DD individuals compared with II individuals. Among patients with COPD, the odds ratio for ischemic heart disease was 1.1 (0.8–1.6) for ID individuals and 1.2 (0.8–1.7) for DD individuals compared with II individuals; corresponding odds ratios for hypertension were 1.1 (0.7–1.5) and 0.8 (0.5–1.2), and for low physical activity 0.9 (0.5–1.4) and 0.7 (0.4–1.2). The results were similar upon adjustment for sex, age, smoking status, body mass index, total cholesterol, and ACE inhibitor/angiotensin II type 1 receptor blocker use. These data suggest that lifelong genetically elevated ACE activity is not a major risk factor for asthma or COPD, or for ischemic heart disease, hypertension, and low physical activity in COPD patients.     

Neurogenesis in the adult spinal cord in an experimental model of multiple sclerosis

Multiple sclerosis is an inflammatory disease of the central nervous system characterized by inflammation, demyelination, axonal degeneration and accumulation of neurological disability. Previously, we demonstrated that stem cells constitute a possible endogenous source for remyelination. We now addressed the question of whether neurogenesis can occur in neuroinflammatory lesions. We demonstrated that, in experimental autoimmune encephalomyelitis, induced in rats 1,1'-dioctadecyl-6,6'-di(4sulphopentyl)-3,3,3',3'tetramethylindocarbocyanin(DiI)-labelled ependymal cells not only proliferated but descendants migrated to the area of neuroinflammation and differentiated into cells expressing the neuronal markers beta-III-tubulin and NeuN. Furthermore, these cells were immunoreactive for bromodeoxyuridine and PCNA, markers for cells undergoing cell proliferation. Using the whole-cell patch-clamp technique on freshly isolated 1, DiI-labelled cells from spinal cord lesions we demonstrated the ability of these cells to fire overshooting action potentials similar to those of immature neurones. We thus provide the first evidence for the initiation of neurogenesis in neuroinflammatory lesions in the adult spinal cord.     

Disparate cholinergic currents in rat principal trigeminal sensory nucleus neurons mediated by M1 and M2 receptors: a possible mechanism for selective gating of afferent sensory neurotransmission

Neurons situated in the principal sensory trigeminal nucleus (PSTN) convey orofacial sensory inputs to thalamic relay regions and higher brain centres, and the excitability of these ascending tract cells is modulated across sleep/wakefulness states and during pain conditions. Moreover, acetylcholine release changes profoundly across sleep/wakefulness states and ascending sensory neurotransmission is altered by cholinergic agonists. An intriguing possibility is, therefore, that cholinergic mechanisms mediate such state-dependent modulation of PSTN tract neurons. We tested the hypotheses that cholinergic agonists can modulate PSTN cell excitability and that such effects are mediated by muscarinic receptor subtypes, using patch-clamp methods in rat and mouse. In all examined cells, carbachol elicited an electrophysiological response that was independent of action potential generation as it persisted in the presence of tetrodotoxin. Responses were of three types: depolarization, hyperpolarization or a biphasic response consisting of hyperpolarization followed by depolarization. In voltage-clamp mode, carbachol evoked corresponding inward, outward or biphasic currents. Moreover, immunostaining for the vesicle-associated choline transporter showed cholinergic innervation of the PSTN. Using muscarinic receptor antagonists, we found that carbachol-elicited PSTN neuron hyperpolarization was mediated by M2 receptors and depolarization, in large part, by M1 receptors. These data suggest that acetylcholine acting on M1 and M2 receptors may contribute to selective excitability enhancement or depression in individual, rostrally projecting sensory neurons. Such selective gating effects via cholinergic input may play a functional role in modulation of ascending sensory transmission, including across behavioral states typified by distinct cholinergic tone, e.g. sleep/wakefulness arousal levels or neuropathic pain conditions.     

Assessing burden of illness following acute deep vein thrombosis: data quality, reliability and validity of the Norwegian version of VEINES-QOL/Sym, a disease-specific questionnaire

The data quality, reliability and validity of the Norwegian version of VEINES-QOL/Sym were assessed in 74 patients with deep vein thrombosis (DVT). This patient-reported questionnaire produces two scale scores of venous disease-specific quality of life and venous symptoms. Items had low levels of missing data. Item-total correlations ranged from 0.41 to 0.78 with the exception of 0.29 for the symptom item 'night cramps'. Internal consistency was supported by Cronbach's alpha of 0.88 and 0.94 for VEINES-Sym and VEINES-QOL, respectively. Test–retest reliability assessed for 40 patients gave intraclass correlation coefficients of 0.83 and 0.88 for VEINES-Sym and VEINES-QOL, respectively. Assessment of correlation between the two scales and other clinical measures supports the construct validity of the scales. The results indicate acceptable internal consistency, test–retest reliability and validity of the Norwegian version of the VEINES-QOL/Sym questionnaire in patients with DVT. The results follow those of previous studies, and support the use of VEINES-QOL/Sym in the evaluation of patient outcomes and burden of illness in clinical studies of venous thrombosis.     

Cervid herpesvirus 2 experimentally reactivated in reindeer can produce generalized viremia and abortion

Cervid herpesvirus 2 (CvHV2) has never been isolated from reindeer in Norway, but serological data and investigations by PCR indicate that the virus is endemic in the country, with horizontal and vertical transmission, systemic spread, and latency in the trigeminal ganglion. In this study two seropositive reindeer, one of which was pregnant, were administered dexamethasone, to reactivate CvHV2 latent infection. One control animal received sterile water. All animals including the control reactivated, as shown by amplification of CvHV2 DNA from nasal swabs. The pregnant animal showed lesions in the lip mucosa 10 days after the first dexamethasone injection and CvHV2 was visualized by electron microscopy and isolated from those lesions, as well as from nasal and vaginal swabs. On day 13 she aborted and CvHV2 was isolated from both the aborted calf and the mother. CvHV2 was isolated from the other animal administered dexamethasone. Despite amplification of viral DNA in the control animal, it was never possible to isolate the virus. Molecular characterization of the new isolates confirmed these to be CvHV2, and similar to the previous known strain Salla82.Present results represent the first isolation of CvHV2 in Norway and reconfirm that this virus can cause systemic infections in reindeer even after reactivation episodes, and infect the fetus in utero despite a prompt immune response. While it is not possible to atribute the abortion to CvHV2 alone, present data together with previous reports of vertical transmission of CvHV2 and neonatal death, point to an abortogenic potential, which should be further investigated.