Exocytosis-related genes and response to methylphenidate treatment in adults with ADHD.

Experimental studies have demonstrated that methylphenidate (MPH) modulates the synaptic vesicle trafficking and synaptotagmin-1 (SytI) mRNA levels. SytI is a regulatory protein of the SNARE complex, a neurotransmitter exocytosis mediator. Despite this evidence, most SNARE complex-related genes have never been evaluated in attention-deficit/hyperactivity disorder (ADHD) pharmacogenetics. This study evaluates, for we believe the first time, polymorphisms on the SNARE complex-related genes STX1A (rs2228607), VAMP2 (26bp Ins/Del) and SYT1 (rs1880867 and rs2251214) on the response to immediate-release methylphenidate (IR-MPH) in a naturalistic sample of adults with ADHD. The sample comprised 433 subjects, of which 272 (62.8%) have completed the short-term IR-MPH treatment (at least 30 days). The main outcome measure was the categorical variable of short-term response to IR-MPH based on the Swanson, Nolan and Pelham Rating Scale version 4 (SNAP-IV), and on the clinical global impression-improvement scale. Additional analyses evaluated the percentage of SNAP-IV symptom reduction for each dimension as well as short- and long- (7 years) term treatment persistence. SYT1-rs2251214 was associated with the categorical short-term response to IR-MPH (P=0.006, PFDR=0.028), and with the percentage of inattention and oppositional defiant disorder symptoms reduction (P=0.007, PFDR=0.028 and P=0.017, PFDR=0.048, respectively). SYT1-rs2251214 was also associated with short-term treatment persistence (P=0.018, PFDR=0.048), and with months of treatment (P=0.002, PFDR=0.016) in the long-term protocol. Our findings suggest that SYT1-rs2251214 presents a broad influence in IR-MPH response variability in adults with ADHD, being involved with both symptom response and treatment persistence. If such findings are replicated, SytI could represent a key element in MPH pharmacodynamics in adults with ADHD.

Impact of chlorpyrifos on human villous trophoblasts and chorionic villi.

Placental barrier regulates maternal-fetal interchange protecting the baby from damage caused by substances found in the uterine environment or circulating in the vascular system. Organophosphate (OP) pesticides are a paramount group of environmental pollutants used in intensive agriculture for protection against diseases and pests. While many studies have reported an increased risk of pregnancy alterations in pregnant women exposed to OPs, few have analyzed the effects caused by these pesticides in the placenta. Herein, we evaluated the effects of chlorpyrifos (CPF), one of the most widely used OP insecticides, on human placenta using in vitro and ex vivo exposure models. Villous cytotrophoblast cells isolated from normal human term placentas maintained their cell viability, differentiated into syncytiotrophoblast-like structures, and increased the expression of ß-hCG, ABCG2, and P-gp in the presence of CPF at concentrations of 10 to 100µM. The same doses of CPF induced marked changes in chorionic villi samples. Indeed, CPF exposure increased stroma cell apoptosis, altered villi matrix composition, basement membrane thickness, and trophoblastic layer integrity. Histomorphological and ultrastructural alterations are compatible with those found in placentas where maternal-placenta injury is chronic and able to impair the placental barrier function and nutrient transport from mother to the fetus. Our study shows that placental ex vivo exposure to CPF produces tissue alterations and suggest that human placenta is a potential target of CPF toxicity. In addition, it highlights the importance of using different models to assess the effects of a toxic on human placenta.

Use of PCR-DHPLC with fluorescence detection for the characterization of the bacterial diversity during cassava (Manihot esculenta Crantz) fermentation.

Denaturing high-performance liquid chromatography (DHPLC) has been described as a suitable method to study DNA polymorphisms. Here, cassava (Manihot esculenta Crantz) fermentation liquor was examined using DHPLC analysis to characterize the bacterial diversity during the fermentation process. GC-clamped amplicons corresponding to a variable region of the bacterial community 16S rDNA were synthesized using polymerase chain reaction (PCR) and then resolved on a base-composition basis using preparative DHPLC. Eluate fractions were collected at random and used as a source of whole community DNA that could be used to determine the bacterial diversity. As a first approach, GC-clamps were removed from the eluted DNA fragments using PCR to avoid the possible bias these clamps could cause during the construction of clone libraries. As a second approach, a clone library of each eluate sample was constructed, preserving the GC-clamps of the DNA fragments. The first approach generated 132 bacterial rDNA sequences with an average size of 200 bp, 45% of which had similarity to unculturable or non-classified bacteria. The second approach produced 194 sequences identified as Proteobacteria (48%), uncultured or non-classified environmental bacteria (40%) and Firmicutes (12%). We detected a remarkably greater bacterial diversity using the first approach than the second approach. The DHPLC-PCR method allowed for the fast and non-laborious detection of a vast bacterial diversity that was associated with cassava fermentation, and we conclude that it is a promising alternative for the characterization of the overall microbial diversity in complex samples.

Parental stress and dental caries experience in adolescents: analysis of data from a birth cohort study in Pelotas, Southern Brazil.

PURPOSE: Analyze the association between parental stress and dental caries experience in adolescents in southern Brazil using data from the Pelotas 2004 Birth Cohort. METHODS: Interviews and oral health examinations for the determination of the main exposure and outcome of the study were performed in the homes of the adolescents. The outcome was dental caries experience in the permanent dentition analyzed using the Decayed, Missing and Filling Surfaces (DMFS) index. The main exposure was parental stress measured using the Parenting Stress Index-Short Form administered to the parents of the adolescents. Demographic/socioeconomic characteristics, oral health characteristics and oral health-related quality of life were considered potential confounding factors. Negative binomial regressions estimated mean ratios (MR) and 95% confidence intervals (CI). RESULTS: Nine hundred ninety-six adolescents were evaluated at 12 and 13 years of age. The prevalence of dental caries experience in the adolescents was 36.9% (95% CI: 33.8-40.0) and 15.1% (95% CI: 12.8-17.3) of the parents had parental stress. After adjusting for confounding factors, parental stress was associated with a higher mean number of decayed, missing and filling surfaces in the adolescents (MR = 1.10; 95% CI: 1.01-1.26; p = 0.045). CONCLUSION: Adolescents of parents with parental stress have more dental caries experience compared to those whose parents do not have parental stress.

Low oxygen tension induces Krüppel-Like Factor 6 expression in trophoblast cells.

The transcription factor Krüppel-Like Factor 6 (KLF6) has important roles in cell differentiation, angiogenesis, apoptosis, and proliferation. Furthermore, there is evidence that KLF6 is required for proper placental development. While oxygen is a critical mediator of trophoblast differentiation and function, the involvement of oxygen in the regulation of KLF6 expression remains unexplored. In the present study we examined the expression of KLF6 in placental tissue from uncomplicated and preeclamptic pregnancies, the latter often characterized by an inadequately perfused placenta. We also determined the effect of hypoxia and the involvement of Hypoxia-Inducible Factor 1α (HIF-1α) on the expression of KLF6 in cultured trophoblast cells and placental tissues. Results revealed that villous, interstitial and endovascular extravillous cytotrophoblasts from placentas from normal and preeclamptic pregnancies express KLF6. In addition, KLF6 immunoreactivity was higher in the placental bed of preeclamptic pregnancies than in those of uncomplicated pregnancies. We demonstrated that hypoxia induced an early and transient increase in KLF6 protein levels in HTR8/SVneo extravillous cytotrophoblast cells and in placental explants. Reoxygenation returned KLF6 protein to basal levels. Moreover, hypoxia-induced up-regulation of KLF6 expression was dependent on HIF-1α as revealed by siRNA knockdown in HTR8/SVneo cells. These results indicate that KLF6 may mediate some of the effects of hypoxia in placental development. The regulation of KLF6 protein levels by oxygen has significant implications for understanding its putative role in diseases affected by tissue hypoxia.

Vascular endothelial growth factor (VEGF) and VEGF-receptor expression in placenta of hyperglycemic pregnant women.

Hyperglycemia occurs in a variety of conditions such as overt diabetes, gestational diabetes and mild hyperglycemia, all of which are generally defined based on the oral glucose tolerance test and glucose profiles. Whereas diabetes has received considerable attention in recent decades, few studies have examined the mechanisms of mild hyperglycemia and its associated disturbances. Mild gestational hyperglycemia is associated with macrosomia and a high risk of perinatal mortality. Morphologically, the placenta of these women is characterized by an increase in the number of terminal villi and capillaries, presumably as part of a compensatory mechanism to maintain homeostasis at the maternal-fetal interface. In this study, we analised the expression of VEGF and its receptors VEGFR-1 (Flt-1) and VEGFR-2 (KDR) in placentas from mildly hyperglycemic women. This expression was compared with that of normoglycemic women and women with gestational and overt diabetes. Immunohistochemistry revealed strong staining for VEGF and VEGFR-2 in vascular and trophoblastic cells of mildly hyperglycemic women, whereas the staining for VEGFR-1 was discrete and limited to the trophoblast. The pattern of VEGF and VEGF-receptor reactivity in placentas from women with overt diabetes was similar to that of normoglycemic women. In women with gestational diabetes, strong staining for VEGFR-1 was observed in vascular and trophoblastic cells whereas VEGF and VEGFR-2 were detected only in the trophoblast. The expression of these proteins was confirmed by western blotting, which revealed the presence of an additional band of 75 kDa. In the decidual compartment, only extravillous trophoblast reacted with all antibodies. Morphological analysis revealed collagen deposition around large arteries in all groups with altered glycemia. These findings indicate a placental response to altered glycemia that could have important consequences for the fetus. The change in the placental VEGF/VEGFR expression ratio in mild hyperglycemia may favor angiogenesis in placental tissue and could explain the hypercapillarization of villi seen in this gestational disturbance.

Genomics and proteomics approaches to the study of cancer-stroma interactions.

BACKGROUND: The development and progression of cancer depend on its genetic characteristics as well as on the interactions with its microenvironment. Understanding these interactions may contribute to diagnostic and prognostic evaluations and to the development of new cancer therapies. Aiming to investigate potential mechanisms by which the tumor microenvironment might contribute to a cancer phenotype, we evaluated soluble paracrine factors produced by stromal and neoplastic cells which may influence proliferation and gene and protein expression. METHODS: The study was carried out on the epithelial cancer cell line (Hep-2) and fibroblasts isolated from a primary oral cancer. We combined a conditioned-medium technique with subtraction hybridization approach, quantitative PCR and proteomics, in order to evaluate gene and protein expression influenced by soluble paracrine factors produced by stromal and neoplastic cells. RESULTS: We observed that conditioned medium from fibroblast cultures (FCM) inhibited proliferation and induced apoptosis in Hep-2 cells. In neoplastic cells, 41 genes and 5 proteins exhibited changes in expression levels in response to FCM and, in fibroblasts, 17 genes and 2 proteins showed down-regulation in response to conditioned medium from Hep-2 cells (HCM). Nine genes were selected and the expression results of 6 down-regulated genes (ARID4A, CALR, GNB2L1, RNF10, SQSTM1, USP9X) were validated by real time PCR. CONCLUSIONS: A significant and common denominator in the results was the potential induction of signaling changes associated with immune or inflammatory response in the absence of a specific protein.

Análise eletromiográfica e força do grupo muscular extensor do punho durante isquemia induzida / Electromyographic analysis and strength of the wrist extensor muscle group during induced ischemia

OBJETIVO: Avaliar o efeito da isquemia induzida sobre os parâmetros do sinal eletromiográfico e a força do grupo muscular extensor do punho (GMEP) em mulheres saudáveis. MÉTODOS: Participaram 13 voluntárias, destras, sedentárias, com idade de 23,38±2,32 anos e índice de massa corporal (IMC) de 20,68±1,87kg/m². Para determinar a força do GMEP, foram realizadas 3 contrações isométricas voluntárias máximas (CIVM), utilizando-se uma célula de carga por 15 segundos, com intervalos de 2 minutos entre cada contração, sendo todo procedimento repetido por 3 dias não consecutivos. A isquemia foi realizada por 5 minutos, utilizando um esfigmomanômetro posicionado no braço dominante e inflado até a ausência do fluxo sanguíneo, confirmada pelo ultrassom Doppler. Para coleta do sinal eletromiográfico do GMEP, utilizou-se o equipamento EMG1000 (Lynx®) com eletrodo de superfície diferencial (Lynx®). Foram coletadas 3 CIVM por 15 segundos, com intervalo de 30 segundos entre elas, nas situações de pré-isquemia; isquemia; pós-isquemia imediata (pós-1) e pós-isquemia tardia (pós-2 - após 10 minutos do início da isquemia). Para análise dos parâmetros do sinal eletromiográfico, root mean square (RMS), e frequência mediana do espectro de potência do sinal foi utilizado o software MATLAB 6.5.1. Para análise estatística, foram utilizados os testes de Friedman e ANOVA two-way. RESULTADOS: A isquemia promoveu redução significativa (p<0,05) da força do GMEP. Entretanto, não provocou alterações significativas nos parâmetros eletromiográficos RMS (p=0,05) e frequência mediana do espectro de potência do sinal (p=0,09). CONCLUSÃO: A isquemia induzida promoveu fadiga do GMEP quando relacionada à produção da força muscular. Porém, não provocou fadiga eletromiográfica do grupo muscular avaliado.

OBJECTIVE: To analyze the effect of induced ischemia on the parameters of electromyographic signals and the strength of the wrist extensor muscle group (WEMG) in healthy women. METHODS: Thirteen right-handed sedentary subjects aged 23.38±2.32 years old, with body mass index (BMI) of 20.68±1.87kg/m², took part. To determine WEMG strength, three maximal voluntary isometric contractions (MVIC) were performed using a load cell for 15 seconds, with 2 minutes intervals between contractions. The entire procedure was repeated on three nonconsecutive days. Ischemia was induced for 5 minutes using a sphygmomanometer placed on the dominant arm and inflated until blood flow was absent, as confirmed by Doppler ultrasound. The EMG1000 module (Lynx®) was used with differential surface electrodes (Lynx®) to record the electromyographic signal of the WEMG. Three MVIC were recorded for 15 seconds, with 30 seconds intervals between them, under the following conditions: pre-ischemia, ischemia, immediate post-ischemia (post-1) and later post-ischemia (post-2: 10 minutes after the onset of ischemia). The MATLAB 6.5.1 software was used to analyze the parameters for the electromyographic signal, the root mean square (RMS) and the median frequency of the signal power spectrum. For statistical analysis, two-way ANOVA and the Friedman test were used. RESULTS: Ischemia caused a significant reduction (p<0.05) in WEMG strength. However, there were no significant changes in the RMS electromyographic parameters (p=0.05) or the median frequency of the signal power spectrum (p=0.09). CONCLUSION: Induced ischemia caused WEMG fatigue in relation to muscle strength production. However, it did not cause electromyographic fatigue in the evaluated muscle group.

Fosseta colobomatosa da papila associada com descolamento seroso central: apresentaçäo de um caso tratado com êxito pela fotocoagulaçäo com Xenônio / Colobomatous fossette of the papila associated with central serous detachment: report of a case treated with xenon photocoagulation

Faz-se uma breve revisäo nas características desta afecçäo, apresenta-se um caso, seu tratamento pela fotocoagulaçäo com xenônio e a evoluçäo após 6 anos de seguimento. A técnica adotada foi o bloqueio paramacular temporal com dupla fileira, carga III, diafragma 1,5

Amaurosis congénita de Leber y el electroretinograma / Congenital amaurosis of Leber and the electroretinogram

Los autores presentan 29 casos de Amaurosis congénita de Leber, mostrando la dificultad diagnóstica por fundoscopía, debido a la variedad de formas clínicas. Hacen una revisión bibliográfica y muestran la importancia de esta entidad patológica como causa de ceguera congénita y resaltan el electroretinograma como método propedéutico y diagnóstico fundamental

Neurofibromatose de Von Recklinghausen: a propósito de dois casos

Os autores apresentam dois casos clínicos de neurofibromatose, sendo um deles na forma frusta e outro na forma clássica. Fazem, também, uma revisäo detalhada das características desta doença

Síndrome de Sturge Weber / Sturge-Weber syndrome

Faz-se uma revisäo bibliográfica do assunto, descrevem-se algumas características desta síndrome e seu tratamento. Apresentam-se 2 casos e a intervençäo feita