Release of extracellular particles by recombinant vaccinia virus over-expressing the major envelope protein p37K.

During the replication cycle of vaccinia virus, four different forms of viral particles are produced. The two extracellular enveloped forms, cell-associated enveloped virus and extracellular enveloped virus, are responsible for cell-to-cell transmission and long-range spread of infection both in vivo and in vitro. Despite the biological importance of the enveloped forms, the mechanism of envelopment and the components involved in this process have been analysed only recently. Therefore the individual steps and the rate-limiting factors of the envelopment process are still unknown. The protein p37K, an unglycosylated but acylated envelope protein of molecular mass 37 kDa, has been shown to be essential for envelopment. However, this study shows that over-expression of p37K by vaccinia virus recombinants reduces rather than increases the yield of infectious enveloped virus which is mainly due to the enveloped virions exhibiting a strongly diminished specific infectivity.

Leishmania major-specific B cells are necessary for Th2 cell development and susceptibility to L. major LV39 in BALB/c mice.

B lymphocytes are considered to play a minimal role in host defense against Leishmania major. In this study, the contribution of B cells to susceptibility to infection with different strains of L. major was investigated in BALB/c mice lacking mature B cells due to the disruption of the IgM transmembrane domain (microMT). Whereas BALB/c microMT remained susceptible to infection with L. major IR173 and IR75, they were partially resistant to infection with L. major LV39. Adoptive transfer of naive B cells into BALB/c microMT mice before infection restored susceptibility to infection with L. major LV39, demonstrating a role for B cells in susceptibility to infection with this parasite. In contrast, adoptive transfer of B cells that express an IgM/IgD specific for hen egg lysozyme (HEL), an irrelevant Ag, did not restore disease progression in BALB/c microMT mice infected with L. major LV39. This finding was likely due to the inability of HEL Tg B cells to internalize and present Leishmania Ags to specific T cells. Furthermore, specific Ig did not contribute to disease progression as assessed by transfer of immune serum in BALB/c microMT mice. These data suggest that direct Ag presentation by specific B cells and not Ig effector functions is involved in susceptibility of BALB/c mice to infection with L. major LV39.

Interpretation of array comparative genome hybridization data: a major challenge.

The advent and application of high-resolution array-based comparative genome hybridization (array CGH) has led to the detection of large numbers of copy number variants (CNVs) in patients with developmental delay and/or multiple congenital anomalies as well as in healthy individuals. The notion that CNVs are also abundantly present in the normal population challenges the interpretation of the clinical significance of detected CNVs in patients. In this review we will illustrate a general clinical workflow based on our own experience that can be used in routine diagnostics for the interpretation of CNVs.

Management of refractory status epilepticus in adults: still more questions than answers.

Refractory status epilepticus (RSE) is defined as status epilepticus that continues despite treatment with benzodiazepines and one antiepileptic drug. RSE should be treated promptly to prevent morbidity and mortality; however, scarce evidence is available to support the choice of specific treatments. Major independent outcome predictors are age (not modifiable) and cause (which should be actively targeted). Recent recommendations for adults suggest that the aggressiveness of treatment for RSE should be tailored to the clinical situation. To minimise intensive care unit-related complications, focal RSE without impairment of consciousness might initially be approached conservatively; conversely, early induction of pharmacological coma is advisable in generalised convulsive forms of the disorder. At this stage, midazolam, propofol, or barbiturates are the most commonly used drugs. Several other treatments, such as additional anaesthetics, other antiepileptic or immunomodulatory compounds, or non-pharmacological approaches (eg, electroconvulsive treatment or hypothermia), have been used in protracted RSE. Treatment lasting weeks or months can sometimes result in a good outcome, as in selected patients after encephalitis or autoimmune disorders. Well designed prospective studies of RSE are urgently needed.

Redundant Notch1 and Notch2 signaling is necessary for IFNγ secretion by T helper 1 cells during infection with Leishmania major.

The protective immune response to intracellular parasites involves in most cases the differentiation of IFNγ-secreting CD4(+) T helper (Th) 1 cells. Notch receptors regulate cell differentiation during development but their implication in the polarization of peripheral CD4(+) T helper 1 cells is not well understood. Of the four Notch receptors, only Notch1 (N1) and Notch2 (N2) are expressed on activated CD4(+) T cells. To investigate the role of Notch in Th1 cell differentiation following parasite infection, mice with T cell-specific gene ablation of N1, N2 or both (N1N2(ΔCD4Cre)) were infected with the protozoan parasite Leishmania major. N1N2(ΔCD4Cre) mice, on the C57BL/6 L. major-resistant genetic background, developed unhealing lesions and uncontrolled parasitemia. Susceptibility correlated with impaired secretion of IFNγ by draining lymph node CD4(+) T cells and increased secretion of the IL-5 and IL-13 Th2 cytokines. Mice with single inactivation of N1 or N2 in their T cells were resistant to infection and developed a protective Th1 immune response, showing that CD4(+) T cell expression of N1 or N2 is redundant in driving Th1 differentiation. Furthermore, we show that Notch signaling is required for the secretion of IFNγ by Th1 cells. This effect is independent of CSL/RBP-Jκ, the major effector of Notch receptors, since L. major-infected mice with a RBP-Jκ deletion in their T cells were able to develop IFNγ-secreting Th1 cells, kill parasites and heal their lesions. Collectively, we demonstrate here a crucial role for RBP-Jκ-independent Notch signaling in the differentiation of a functional Th1 immune response following L. major infection.

p.L1612P, a novel voltage-gated sodium channel Nav1.7 mutation inducing a cold sensitive paroxysmal extreme pain disorder.

BACKGROUND: Mutations in the SCN9A gene cause chronic pain and pain insensitivity syndromes. We aimed to study clinical, genetic, and electrophysiological features of paroxysmal extreme pain disorder (PEPD) caused by a novel SCN9A mutation. METHODS: Description of a 4-generation family suffering from PEPD with clinical, genetic and electrophysiological studies including patch clamp experiments assessing response to drug and temperature. RESULTS: The family was clinically comparable to those reported previously with the exception of a favorable effect of cold exposure and a lack of drug efficacy including with carbamazepine, a proposed treatment for PEPD. A novel p.L1612P mutation in the Nav1.7 voltage-gated sodium channel was found in the four affected family members tested. Electrophysiologically the mutation substantially depolarized the steady-state inactivation curve (V1/2 from -61.8 ± 4.5 mV to -30.9 ± 2.2 mV, n = 4 and 7, P < 0.001), significantly increased ramp current (from 1.8% to 3.4%, n = 10 and 12) and shortened recovery from inactivation (from 7.2 ± 5.6 ms to 2.2 ± 1.5 ms, n = 11 and 10). However, there was no persistent current. Cold exposure reduced peak current and prolonged recovery from inactivation in wild-type and mutated channels. Amitriptyline only slightly corrected the steady-state inactivation shift of the mutated channel, which is consistent with the lack of clinical benefit. CONCLUSIONS: The novel p.L1612P Nav1.7 mutation expands the PEPD spectrum with a unique combination of clinical symptoms and electrophysiological properties. Symptoms are partially responsive to temperature but not to drug therapy. In vitro trials of sodium channel blockers or temperature dependence might help predict treatment efficacy in PEPD.

The "help" question doesn't help when screening for major depression: external validation of the three-question screening test for primary care patients managed for physical complaints.

BACKGROUND: Major depression, although frequent in primary care, is commonly hidden behind multiple physical complaints that are often the first and only reason for patient consultation. Major depression can be screened by two validated questions that are easier to use in primary care than the full DSM-IV criteria. A third question, called the "help" question, improves the specificity without apparently decreasing the sensitivity of this screening procedure. We validated the abbreviated screening procedure for major depression with and without the "help" question in primary care patients managed for a physical complaint. METHODS: This diagnostic accuracy study used data from a cohort study called SODA (for SOmatisation Depression Anxiety ) conducted by 24 general practitioners (GPs) in western Switzerland that included patients over 18 years of age with at least one physical complaint at index consultation. Major depression was identified with the full Patient Health Questionnaire. GPs were asked to screen patients for major depression with the three screening questions one year after inclusion. RESULTS: Out of 937 patients with at least one physical complaint, 751 were eligible one year after index consultation. Major depression was diagnosed in 69/724 (9.5%) patients. The sensitivity and specificity of the two-question method alone were 91.3% (95% confidence interval 81.4-96.4%) and 65.0% (95% confidence interval 61.2-68.6%), respectively. Adding the "help" question decreased the sensitivity (59.4% ; 95% confidence interval 47.0-70.9%) but improved the specificity (88.2% ; 95% confidence interval 85.4-90.5%) of the three-question method. CONCLUSIONS: The use of two screening questions for major depression was associated with high sensitivity and low specificity in primary care patients presenting a physical complaint. Adding the "help" question improved the specificity but clearly decreased the sensitivity; when using the "help" question; four out of ten patients with depression will be missed, compared to only one out of ten with the two-question method. Therefore, the "help" question is not useful as a screening question, but may help discussing management strategies.

Anti-leishmania effector functions of CD4+ Th1 cells and early events instructing Th2 cell development and susceptibility to Leishmania major in BALB/c mice.

Resistance and susceptibility to infection with the intracellular parasite, Leishmania major, are mediated by parasite-specific CD4+ Th1 and Th2 cells, respectively. It is well established that the protective effect of parasite-specific CD4+ Th1 cells is largely dependent upon the IFN-gamma produced. However, recent results indicate that the effect of Th1 cells on resolution of lesions induced by L. major in genetically resistant mice also requires a functional Fas-FasL pathway of cytotoxicity. In contrast to resistant mice, susceptible BALB/c mice develop aberrant Th2 responses following infection with L. major and consequently suffer progressive disease. These outcomes clearly depends upon the production of interleukin 4 (IL-4) early after infection. We have shown that a burst of IL-4 mRNA, peaking in draining lymph nodes of BALB/c mice 16 hrs after infection, occurs within CD4+ T cells that express V beta 4-V alpha 8 T cell receptors. In contrast to control and V beta 6-deficient mice, V beta 4-deficient BALB/c mice were resistant to infection, demonstrating the role of these cells in Th2 development. The early IL-4 response was absent in these mice, and Th1 responses occurred following infection. The LACK antigen of L. major induced comparable IL-4 production in V beta 4-V alpha 8 CD4+ T cells. Thus, the IL-4 required for Th2 development and susceptibility to L. major is produced by a restricted population of V beta 4-V alpha 8 CD4+ T cells after cognate interaction with a single antigen from this complex parasite. The IL-4 produced rapidly by these CD4+ T cells induces within 48 hours a state of unresponsiveness to IL-12 among parasite-specific CD4+ T cell precursors by downregulating the IL-12 receptor beta 2 chain expression.

Quality of Web-based information on obsessive compulsive disorder.

BACKGROUND: The Internet is increasingly used as a source of information for mental health issues. The burden of obsessive compulsive disorder (OCD) may lead persons with diagnosed or undiagnosed OCD, and their relatives, to search for good quality information on the Web. This study aimed to evaluate the quality of Web-based information on English-language sites dealing with OCD and to compare the quality of websites found through a general and a medically specialized search engine. METHODS: Keywords related to OCD were entered into Google and OmniMedicalSearch. Websites were assessed on the basis of accountability, interactivity, readability, and content quality. The "Health on the Net" (HON) quality label and the Brief DISCERN scale score were used as possible content quality indicators. Of the 235 links identified, 53 websites were analyzed. RESULTS: The content quality of the OCD websites examined was relatively good. The use of a specialized search engine did not offer an advantage in finding websites with better content quality. A score ≥16 on the Brief DISCERN scale is associated with better content quality. CONCLUSION: This study shows the acceptability of the content quality of OCD websites. There is no advantage in searching for information with a specialized search engine rather than a general one. Practical implications: The Internet offers a number of high quality OCD websites. It remains critical, however, to have a provider-patient talk about the information found on the Web.

Major influencing factors of indoor radon concentrations in Switzerland.

PURPOSE: In Switzerland, nationwide large-scale radon surveys have been conducted since the early 1980s to establish the distribution of indoor radon concentrations (IRC). The aim of this work was to study the factors influencing IRC in Switzerland using univariate analyses that take into account biases caused by spatial irregularities of sampling. METHODS: About 212,000 IRC measurements carried out in more than 136,000 dwellings were available for this study. A probability map to assess risk of exceeding an IRC of 300 Bq/m(3) was produced using basic geostatistical techniques. Univariate analyses of IRC for different variables, namely the type of radon detector, various building characteristics such as foundation type, year of construction and building type, as well as the altitude, the average outdoor temperature during measurement and the lithology, were performed comparing 95% confidence intervals among classes of each variable. Furthermore, a map showing the spatial aggregation of the number of measurements was generated for each class of variable in order to assess biases due to spatially irregular sampling. RESULTS: IRC measurements carried out with electret detectors were 35% higher than measurements performed with track detectors. Regarding building characteristics, the IRC of apartments are significantly lower than individual houses. Furthermore, buildings with concrete foundations have the lowest IRC. A significant decrease in IRC was found in buildings constructed after 1900 and again after 1970. Moreover, IRC decreases at higher outdoor temperatures. There is also a tendency to have higher IRC with altitude. Regarding lithology, carbonate rock in the Jura Mountains produces significantly higher IRC, almost by a factor of 2, than carbonate rock in the Alps. Sedimentary rock and sediment produce the lowest IRC while carbonate rock from the Jura Mountains and igneous rock produce the highest IRC. Potential biases due to spatially unbalanced sampling of measurements were identified for several influencing factors. CONCLUSIONS: Significant associations were found between IRC and all variables under study. However, we showed that the spatial distribution of samples strongly affected the relevance of those associations. Therefore, future methods to estimate local radon hazards should take the multidimensionality of the process of IRC into account.

Clinical outcome after trimipramine in patients with delusional depression - a pilot study

The treatment of delusional depression is a major challenge in psychopharmacology. Hypothalamic-pituitary-adrenocortical (HPA) overdrive may contribute, via increased dopaminergic activity, to the pathophysiology of the disorder. Trimipramine appears to be an interesting potential candidate, since it is an atypical antidepressant that is known to inhibit HPA activity. In a four-week open trial we investigated its effects in 15 inpatients with delusional depression. The dosage was increased within 7 days up to 300 - 400 mg/d and was then maintained for three weeks. Psychometric assessments and safety monitoring were conducted weekly. Assessment of the HPA activity was achieved by a combined dexamethasone suppression/corticotropin-releasing hormone stimulation (Dex/CRH) test before and after four weeks of treatment. Therapeutic response was defined as a decrease in the HAMD-score of at least 50 %. Eight out of 13 completers were rated as responders. Therapeutic response was associated with L, D-trimipramine concentrations higher than 160 ng/ml. Intent-to-treat analysis showed significant improvement in psychometric variables. Despite the high dosage, the substance was generally well tolerated, with the exception of one patient who suffered from a hypotensive reaction. Mean +/- SD concentration of L-trimipramine and D-trimipramine were 138 +/- 61 ng/ml and 119 +/- 50 ng/ml at a final dose of 346 +/- 50 mg/d. The ACTH and cortisol area under the curve in the Dex/CRH tests decreased significantly, reflecting a decrease of activity in the HPA system. We suggest that the clinical use of high-dose trimipramine in delusional depression seems to be a promising treatment strategy.

IL-4 rapidly produced by V beta 4 V alpha 8 CD4+ T cells instructs Th2 development and susceptibility to Leishmania major in BALB/c mice.

BALB/c mice develop aberrant T helper 2 (Th2) responses and suffer progressive disease after infection with Leishmania major. These outcomes depend on the production of interleukin-4 (IL-4) early after infection. Here we demonstrate that the burst of IL-4 mRNA, peaking in draining lymph nodes of BALB/c mice 16 hr after infection, occurs within CD4+ T cells that express V beta 4 V alpha 8 T cell receptors. In contrast to control and V beta 6-deficient BALB/c mice, V beta 4-deficient BALB/c mice were resistant to infection, demonstrating the role of these cells in Th2 development. The early IL-4 response was absent in these mice, and T helper 1 responses occurred following infection. Recombinant LACK antigen from L. major induced comparable IL-4 production in V beta 4 V alpha 8 CD4+ cells. Thus, the IL-4 required for Th2 development and susceptibility to L. major is produced by a restricted population of V beta 4 V alpha 8 CD4+ T cells after cognate interaction with a single antigen from this complex organism.