Development of an in vitro system to study the developmental stages of Toxoplasma gondii using a genetically modified strain expressing markers for tachyzoites and bradyzoites.

Toxoplasma gondii, the agent of toxoplasmosis, is an intracellular parasite that can infect a wide range of vertebrate hosts. Toxoplasmosis causes severe damage to immunocompromised hosts and its treatment is mainly based on the combination of pyrimethamine and sulfadiazine, which causes relevant side effects primarily observed in AIDS patients, including bone marrow suppression and hematological toxicity (pyrimethamine) and/or hypersensitivity and allergic skin reactions (sulfadiazine). Thus, it is important to investigate new compounds against T. gondii, particularly those that may act on bradyzoites, which are present in cysts during the chronic disease phase. We propose an in vitro model to simultaneously study new candidate compounds against the two main causative stages of Toxoplasma infection in humans, using the EGS-DC strain that was modified from a type I/III strain (EGS), isolated from a case of human congenital toxoplasmosis in Brazil and engineered to express markers for both stages of development. One feature of this strain is that it presents tachyzoite and bradyzoite in the same culture system and in the same host cell under normal culture conditions. Additionally, this strain presents stage-specific fluorescent protein expression, allowing for easy identification of both stages, thus making this strain useful in different studies. HFF cells were infected and after 4 and 7 days post infection the cells were treated with 10 µM of pyrimethamine or atovaquone, for 48 or 72 h. We used high-throughput screening to quantify the extent of parasite infection. Despite a reduction in tachyzoite infection caused by both treatments, the atovaquone treatment reduced the bradyzoite infection while the pyrimethamine one increased it. Ultrastructural analysis showed that after treatment with both drugs, parasites displayed altered mitochondria. Fluorescence microscopy of cells labeled with MitoTracker CMXRos showed that the cysts present inside the cells lost their mitochondrial membrane potential. Our results indicate that this experimental model is adequate to simultaneously analyze new active compounds against tachyzoite and bradyzoite forms.

Escala de Gravedad de Síntomas Revisada (EGS-R) del trastorno de estrés postraumático según el DSM-5: propiedades psicométricas / Posttraumatic stress disorder symptom Severity Scale-Revised (EGS-R) according to DSM-5 criteria: psychometric properties

La escala de gravedad de síntomas Revisada del trastorno de estrés postraumático (TEPT) es una entrevista estructurada que consta de 21 ítems, basada en los criterios diagnósticos del DSM-5, y sirve para evaluar la gravedad de los síntomas de este cuadro clínico. En este trabajo se describen las propiedades psicométricas de este instrumento. la muestra contó con 526 víctimas de agresiones sexuales o de violencia familiar y con una muestra normativa de 193 participantes. El instrumento global mostró una alta consistencia interna (α = .91), así como una buena validez discriminante (g = 1.27) y convergente (rbp = .78 con el diagnóstico). Los resultados del análisis factorial confirmatorio apoyan los cuatro núcleos de síntomas del DSM-5. Un punto de corte de 20, con una eficacia diagnóstica del 82.48%, es apropiada para discriminar a las víctimas con un TEPT. Esta escala resulta útil para planificar el tratamiento y las investigaciones clínicas.

the posttraumatic stress Disorder (ptsD) symptom severity scale-revised is a 21-item structured interview based in DSM-5 criteria and intended to assess the severity of the symptoms of this mental disorder. This paper describes the psychometric properties of this instrument for assessing PTSD. The sample consisted of 526 patients who had been victims of sexual aggression or family violence and 193 people from the general population. The global instrument showed high internal consistency (α = .91), as well as good discriminant (g = 1.27) and concurrent validity (rpb = .78 with diagnosis of PTSD). The results of confirmatory factor analysis are presented and give support to the DSM-5 four symptom clusters. a cut-off point of 20, with a diagnostic efficacy of 82.48%, is appropriate to discriminate the victims with PTSD. The interview appears to be a sound instrument and should prove useful for treatment planning and research.

Ezh1 and Ezh2 differentially regulate PSD-95 gene transcription in developing hippocampal neurons.

Polycomb Repressive Complex 2 (PRC2) mediates transcriptional silencing by catalyzing histone H3 lysine 27 trimethylation (H3K27me3), but its role in the maturation of postmitotic mammalian neurons remains largely unknown. We report that the PRC2 paralogs Ezh1 and Ezh2 are differentially expressed during hippocampal development. We show that depletion of Ezh2 leads to increased expression of PSD-95, a critical plasticity gene, and that reduced PSD-95 gene transcription is correlated with enrichment of Ezh2 at the PSD-95 gene promoter; however, the H3K27me3 epigenetic mark is not present at the PSD-95 gene promoter, likely due to the antagonizing effects of the H3S28P and H3K27Ac marks and the activity of the H3K27 demethylases JMJD3 and UTX. In contrast, increased PSD-95 gene transcription is accompanied by the presence of Ezh1 and elongation-engaged RNA Polymerase II complexes at the PSD-95 gene promoter, while knock-down of Ezh1 reduces PSD-95 transcription. These results indicate that Ezh1 and Ezh2 have antagonistic roles in regulating PSD-95 transcription.