Use of Large-volume Cone-Beam Computed Tomography in Identification and Localization of Horizontal Root Fracture in the Presence and Absence of Intracanal Metallic Post

Introduction: The objective of the study was to evaluate the ability of large-volume cone-beam computed tomography (CBCT) to detect horizontal root fracture and to test the influence of a metallic post. Methods: Through the examination of 40 teeth by large-volume CBCT (20-cm height and 15-cm diameter cylinder) at 0.2-mm voxel resolution, 2 observers analyzed the samples for the presence and localization of horizontal root fracture. Results: The values of accuracy in the groups that had no metallic post ranged from 33%-68%, whereas for the samples with the metallic post, values showed a wide variation (38%-83%). Intraobserver agreement showed no statistically significant difference between the groups with/without metallic post; both ranged from very weak to weak (kappa, 0.09-0.369). Conclusions: The low accuracy and low intraobserver and interobserver agreement reflect the difficulty in performing an adequate diagnosis of horizontal root fractures through a large-volume CBCT by using a small voxel reconstruction. (J Endod 2012;38:856-859)

Lipopolysaccharide-Induced Sickness Behaviour Evaluated in Different Models of Anxiety and Innate Fear in Rats

The fact that there is a complex and bidirectional communication between the immune and nervous systems has been well demonstrated. Lipopolysaccharide (LPS), a component of gram-negative bacteria, is widely used to systematically stimulate the immune system and generate profound physiological and behavioural changes, also known as sickness behaviour (e.g. anhedonia, lethargy, loss of appetite, anxiety, sleepiness). Different ethological tools have been used to analyse the behavioural modifications induced by LPS; however, many researchers analysed only individual tests, a single LPS dose or a unique ethological parameter, thus leading to disagreements regarding the data. In the present study, we investigated the effects of different doses of LPS (10, 50, 200 and 500 mu g/kg, i.p.) in young male Wistar rats (weighing 180200 g; 89 weeks old) on the ethological and spatiotemporal parameters of the elevated plus maze, light-dark box, elevated T maze, open-field tests and emission of ultrasound vocalizations. There was a dose-dependent increase in anxiety-like behaviours caused by LPS, forming an inverted U curve peaked at LPS 200 mu g/kg dose. However, these anxiety-like behaviours were detected only by complementary ethological analysis (stretching, grooming, immobility responses and alarm calls), and these reactions seem to be a very sensitive tool in assessing the first signs of sickness behaviour. In summary, the present work clearly showed that there are resting and alertness reactions induced by opposite neuroimmune mechanisms (neuroimmune bias) that could lead to anxiety behaviours, suggesting that misunderstanding data could occur when only few ethological variables or single doses of LPS are analysed. Finally, it is hypothesized that this bias is an evolutionary tool that increases animals security while the body recovers from a systemic infection.

A General Latent Class Model for Performance Evaluation of Diagnostic Tests in the Absence of a Gold Standard: An Application to Chagas Disease

We propose a new general Bayesian latent class model for evaluation of the performance of multiple diagnostic tests in situations in which no gold standard test exists based on a computationally intensive approach. The modeling represents an interesting and suitable alternative to models with complex structures that involve the general case of several conditionally independent diagnostic tests, covariates, and strata with different disease prevalences. The technique of stratifying the population according to different disease prevalence rates does not add further marked complexity to the modeling, but it makes the model more flexible and interpretable. To illustrate the general model proposed, we evaluate the performance of six diagnostic screening tests for Chagas disease considering some epidemiological variables. Serology at the time of donation (negative, positive, inconclusive) was considered as a factor of stratification in the model. The general model with stratification of the population performed better in comparison with its concurrents without stratification. The group formed by the testing laboratory Biomanguinhos FIOCRUZ-kit (c-ELISA and rec-ELISA) is the best option in the confirmation process by presenting false-negative rate of 0.0002% from the serial scheme. We are 100% sure that the donor is healthy when these two tests have negative results and he is chagasic when they have positive results.

ABSENCE OF STABILIZATION POINT OF THE SPECIES ACCUMULATION CURVE IN TROPICAL FORESTS

The definition of the sample size is a major problem in studies of phytosociology. The species accumulation curve is used to define the sampling sufficiency, but this method presents some limitations such as the absence of a stabilization point that can be objectively determined and the arbitrariness of the order of sampling units in the curve. A solution to this problem is the use of randomization procedures, e. g. permutation, for obtaining a mean species accumulation curve and empiric confidence intervals. However, the randomization process emphasizes the asymptotical character of the curve. Moreover, the inexistence of an inflection point in the curve makes it impossible to define objectively the point of optimum sample size.

Absence of Functional LIN28B Mutations in a Large Cohort of Patients with Idiopathic Central Precocious Puberty

Aim: To investigate LIN28B gene variants in children with idiopathic central precocious puberty (CPP). Patients and Methods: We studied 178 Brazilian children with CPP (171 girls, 16.8% familial cases). A large multiethnic group (1,599 subjects; Multiethnic Cohort, MEC) was used as control. DNA analysis and biochemical in vitro studies were performed. Results: A heterozygous LIN28B variant, p. H199R, was identified in a girl who developed CPP at 5.2 years. This variant was absent in 310 Brazilian control individuals, but it was found in the same allele frequency in women from the MEC cohort, independent of the age of menarche. Functional studies revealed that when ectopically expressed in cells, the mutant protein was capable of binding pre-let-7 microRNA and inhibiting let-7 expression to the same extent as wild-type Lin28B protein. Other rare LIN28B variants (p.P173P, c.198+32_33delCT, g.9575731A>C and c.-11C>T) were identified in CPP patients and controls. Therefore, no functional mutation was identified. Conclusion: In vitro studies revealed that the rare LIN28B p.H199R variant identified in a girl with CPP does not affect the Lin28B function in the regulation of let-7 expression. Although LIN28B SNPs were associated with normal pubertal timing, rare variations in this gene do not seem to be commonly involved in the molecular pathogenesis of CPP. Copyright (C) 2012 S. Karger AG, Basel

Leishmania Metacyclogenesis Is Promoted in the Absence of Purines

Leishmania parasites, the causative agent of leishmaniasis, are transmitted through the bite of an infected sand fly. Leishmania parasites present two basic forms known as promastigote and amastigote which, respectively, parasitizes the vector and the mammalian hosts. Infection of the vertebrate host is dependent on the development, in the vector, of metacyclic promastigotes, however, little is known about the factors that trigger metacyclogenesis in Leishmania parasites. It has been generally stated that "stressful conditions" will lead to development of metacyclic forms, and with the exception of a few studies no detailed analysis of the molecular nature of the stress factor has been performed. Here we show that presence/absence of nucleosides, especially adenosine, controls metacyclogenesis both in vitro and in vivo. We found that addition of an adenosine-receptor antagonist to in vitro cultures of Leishmania amazonensis significantly increases metacyclogenesis, an effect that can be reversed by the presence of specific purine nucleosides or nucleobases. Furthermore, our results show that proliferation and metacyclogenesis are independently regulated and that addition of adenosine to culture medium is sufficient to recover proliferative characteristics for purified metacyclic promastigotes. More importantly, we show that metacyclogenesis was inhibited in sand flies infected with Leishmania infantum chagasi that were fed a mixture of sucrose and adenosine. Our results fill a gap in the life cycle of Leishmania parasites by demonstrating how metacyclogenesis, a key point in the propagation of the parasite to the mammalian host, can be controlled by the presence of specific purines.

AURICULOTHERAPY EFFECTIVENESS IN THE REDUCTION OF ANXIETY IN NURSING STUDENTS

The objective of this single-blinded randomized controlled trial was to assess anxiety levels in nursing school students of the Beneficencia Portuguesa Hospital (Sao Paulo) and the effectiveness of auriculotherapy in the reduction of these levels. The Trait-Anxiety Inventory State was applied at the beginning of the study, after 8 and 12 sessions, and at follow-up (15 days). The sample was comprised of 71 students divided into 3 groups: control without intervention (25), auriculotherapy (24), and placebo (22). The analysis of variance (ANOVA) showed statistically significant differences post hoc between the control and auriculotherapy groups at 2nd (p=0.000), 3rd (p=0.012) and 4th assessments (p=0.005), and between placebo and control groups at 2nd assessment (p=0.003). Auriculotherapy with Shenmen and Brain Stem points was more effective (20.97%) than sham points (13.74%) for reduction of anxiety levels in Nursing students, but studies with more representative samples are recommended.

Absence of inactivating mutations and deletions in the DMRT1 and FGF9 genes in a large cohort of 46,XY patients with gonadal dysgenesis

Despite advances in our understanding of the mechanisms involved in sex determination and differentiation, the specific roles of many genes in these processes are not completely understood in humans. Both DMRT1 and FGF9 are among this group of genes. Dmrt1 controls germ cell differentiation, proliferation, migration and pluripotency and Sertoli cell proliferation and differentiation. Fgf9 has been considered a critical factor in early testicular development and germ cell survival in mice. We screened for the presence of DMRT1 and FGF9 mutations in 33 patients with 46,XY gonadal dysgenesis. No deletions in either DMRT1 or FGF9 were identified using the MLPA technique. Eight allelic variants of DMRT1 were identified, and in silico analysis suggested that the novel c.968-15insTTCTCTCT variant and the c.774G>C (rs146975077) variant could have potentially deleterious effects on the DMRT1 protein. Nine previously described FGF9 allelic variants and six different alleles of the 3' UTR microsatellite were identified. However, none of these DMRT1 or FGF9 variants was associated with increased 46,XY gonadal dysgenesis. In conclusion, our study suggests that neither DMRT1 nor FGF9 abnormalities are frequently involved in dysgenetic male gonad development in patients with non-syndromic 46,XY disorder of sex development. (C) 2012 Published by Elsevier Masson SAS.

Absence of effects of contralateral group I muscle afferents on presynaptic inhibition of Ia terminals in humans and cats

Mezzarane RA, Kohn AF, Couto-Roldan E, Martinez L, Flores A, Manjarrez E. Absence of effects of contralateral group I muscle afferents on presynaptic inhibition of Ia terminals in humans and cats. J Neurophysiol 108: 1176-1185, 2012. First published June 6, 2012; doi:10.1152/jn.00831.2011.-Crossed effects from group I afferents on reflex excitability and their mechanisms of action are not yet well understood. The current view is that the influence is weak and takes place indirectly via oligosynaptic pathways. We examined possible contralateral effects from group I afferents on presynaptic inhibition of Ia terminals in humans and cats. In resting and seated human subjects the soleus (SO) H-reflex was conditioned by an electrical stimulus to the ipsilateral common peroneal nerve (CPN) to assess the level of presynaptic inhibition (PSI_control). A brief conditioning vibratory stimulus was applied to the triceps surae tendon at the contralateral side (to activate preferentially Ia muscle afferents). The amplitude of the resulting H-reflex response (PSI_conditioned) was compared to the H-reflex under PSI_control, i.e., without the vibration. The interstimulus interval between the brief vibratory stimulus and the electrical shock to the CPN was -60 to 60 ms. The H-reflex conditioned by both stimuli did not differ from that conditioned exclusively by the ipsilateral CPN stimulation. In anesthetized cats, bilateral monosynaptic reflexes (MSRs) in the left and right L 7 ventral roots were recorded simultaneously. Conditioning stimulation applied to the contralateral group I posterior biceps and semitendinosus (PBSt) afferents at different time intervals (0-120 ms) did not have an effect on the ipsilateral gastrocnemius/soleus (GS) MSR. An additional experimental paradigm in the cat using contralateral tendon vibration, similar to that conducted in humans, was also performed. No significant differences between GS-MSRs conditioned by ipsilateral PBSt stimulus alone and those conditioned by both ipsilateral PBSt stimulus and contralateral tendon vibration were detected. The present results strongly suggest an absence of effects from contralateral group I fibers on the presynaptic mechanism of MSR modulation in relaxed humans and anesthetized cats.

Genomic analysis of ERVWE2 locus in patients with multiple sclerosis: absence of genetic association but potential role of human endogenous retrovirus type W elements in molecular mimicry with myelin antigen

Human endogenous retroviruses (HERVs) arise from ancient infections of the host germline cells by exogenous retroviruses, constituting 8% of the human genome. Elevated level of envelope transcripts from HERVs-W has been detected in CSF, plasma and brain tissues from patients with Multiple Sclerosis (MS), most of them from Xq22.3, 15q21.3, and 6q21 chromosomes. However, since the locus Xq22.3 (ERVWE2) lack the 5' LTR promoter and the putative protein should be truncated due to a stop codon, we investigated the ERVWE2 genomic loci from 84 individuals, including MS patients with active HERV-W expression detected in PBMC. In addition, an automated search for promoter sequences in 20 kb nearby region of ERVWE2 reference sequence was performed. Several putative binding sites for cellular cofactors and enhancers were found, suggesting that transcription may occur via alternative promoters. However, ERVWE2 DNA sequencing of MS and healthy individuals revealed that all of them harbor a stop codon at site 39, undermining the expression of a full-length protein. Finally, since plaque formation in central nervous system (CNS) of MS patients is attributed to immunological mechanisms triggered by autoimmune attack against myelin, we also investigated the level of similarity between envelope protein and myelin oligodendrocyte glycoprotein (MOG). Comparison of the MOG to the envelope identified five retroviral regions similar to the Ig-like domain of MOG. Interestingly, one of them includes T and B cell epitopes, capable to induce T effector functions and circulating Abs in rats. In sum, although no DNA substitutions that would link ERVWE2 to the MS pathogeny was found, the similarity between the envelope protein to MOG extends the idea that ERVEW2 may be involved on the immunopathogenesis of MS, maybe facilitating the MOG recognizing by the immune system. Although awaiting experimental evidences, the data presented here may expand the scope of the endogenous retroviruses involvement on MS pathogenesis

Prevalence of anxiety, depression and quality of life in HTLV-1 infected patients

The HAM/TSP caused by HTLV-1 infection usually affects patients to disabling states, and sometimes can lead them to paraplegia presenting symptoms of depression and anxiety, impacting on quality of life. Objective: The purpose of this study was to evaluate the frequency of depression and anxiety and its impact on quality of life in HTLV-1-infected TSP/HAM patients. Material and Methods: This was a cross-sectional study including 67 asymptomatic (control group) and 63 with TSP/HAM subjects. The instruments used were a demographic questionnaire, scales for anxiety and depression diagnosis (BDI and BAI), questionnaire for the assessment of Quality of Life of the World Health Organization (WHOQOL-Brief) and neurological scale to measure the disability level (Osame’s Disability Status Scale). All patients had HTLV-I diagnosis by serological and molecular approaches, monitored at Instituto de Infectologia Emílio Ribas from May 2008 to July 2009. Data were analyzed statistically by frequencies, the Mann-Whitney test and the Spearman correlation test. Data among groups were analyzed and correlated with functional and severity aspects. Results: The results showed that patients with HAM/TSP compared to asymptomatic carriers had higher rates of depression (p < 0.001) and anxiety (p < 0.001), and impairment on quality of life in the areas of: dissatisfaction with health (p < 0.001), physical (p < 0.001) and the environment (p = 0.003). The main factors that correlated with levels of depression and anxiety and the domains of the WHOQOL-brief were: education, family income and social class. Conclusion: A well conducted evaluation and counseling may help in treatment, for a better quality of life of these patients.

Absence of placeholders abolishes exogenous attention effect in a peripheral cueing procedure

Most studies of exogenous visuospatial attention use placeholders indicating the regions where the stimuli appear on the screen. Preliminary results from our laboratory provided evidence that the attentional effect is more frequently observed when placeholders are used in these experimental procedures. Four experiments were carried out. Experiment 1 aimed at confirming the finding that the attentional effect of a spatially non-informative cue (S1) observed in the presence of placeholders disappears in their absence. The results confirmed this finding. Experiments 2, 3, and 4 examined several possible processes that could explain this finding. Experiment 2 investigated if the contribution of a faster disengagement of attention from the cued location or a stronger forward masking could explain the absence of attentional effect when no placeholders were used. Experiment 3 investigated if increased difficulty in discrimination of the target (S2) from S1 would favor the appearance of the attentional effect in the absence of placeholders. Experiment 4 investigated if an insufficient focusing of attention towards the cued location could explain the absence of attentional effect when no placeholders were used. The results of the three experiments indicated that placeholders act by reducing the discriminability of the S2. This would presumably lead to the adoption of an attentional set that favors the mobilization of attention by the S1

Absence of somatostatin SST(2) receptor internalization in vivo after intravenous SOM230 application in the AR42J animal tumor model

Among clinically relevant somatostatin functions, agonist-induced somatostatin receptor subtype 2 (sst(2)) internalization is a potent mechanism for tumor targeting with sst(2) affine radioligands such as octreotide. Since, as opposed to octreotide, the second generation multi-somatostatin analog SOM230 (pasireotide) exhibits strong functional selectivity, it appeared of interest to evaluate its ability to affect sst(2) internalization in vivo. Rats bearing AR42J tumors endogenously expressing somatostatin sst(2) receptors were injected intravenously with SOM230 or with the [Tyr(3), Thr(8)]-octreotide (TATE) analog; they were euthanized at various time points; tumors and pancreas were analyzed by immunohistochemistry for the cellular localization of somatostatin sst(2) receptors. SOM230-induced sst(2) internalization was also evaluated in vitro by immunofluorescence microscopy in AR42J cells. At difference to the efficient in vivo sst(2) internalization triggered by intravenous [Tyr(3), Thr(8)]-octreotide, intravenous SOM230 did not elicit sst(2) internalization: immunohistochemically stained sst(2) in AR42J tumor cells and pancreatic cells were detectable at the cell surface at 2.5min, 10min, 1h, 6h, or 24h after SOM230 injection while sst(2) were found intracellularly after [Tyr(3), Thr(8)]-octreotide injection. The inability of stimulating sst(2) internalization by SOM230 was confirmed in vitro in AR42J cells by immunofluorescence microscopy. Furthermore, SOM230 was unable to antagonize agonist-induced sst(2) internalization, neither in vivo, nor in vitro. Therefore, SOM230 does not induce sst(2) internalization in vivo or in vitro in AR42J cells and pancreas, at difference to octreotide derivatives with comparable sst(2) binding affinities. These characteristics may point towards different tumor targeting but also to different desensitization properties of clinically applied SOM230.