4-Quinolinylhydrazone analogues kill Leishmania (Leishmania) amazonensis by inducing apoptosis and mitochondria-dependent pathway cell death.

Despite efforts, available alternatives for the treatment of leishmaniasis are still scarce. In this work we tested a class of 15 quinolinylhydrazone analogues and presented data that support the use of the most active compound in cutaneous leishmaniasis caused by Leishmania amazonensis. In general, the compounds showed activity at low concentrations for both parasitic forms (5.33-37.04 µM to promastigotes, and 14.31-61.98 µM to amastigotes). In addition, the best compound (MHZ15) is highly selective for the parasite. Biochemical studies indicate that the treatment of promastigotes with MHZ15 leads the loss of mitochondrial potential and increase in ROS levels as the primary effects, which triggers accumulation of lipid droplets, loss of plasma membrane integrity and apoptosis hallmarks, including DNA fragmentation and phosphatidylserine exposure. These effects were similar in the intracellular form of the parasite. However, in this parasitic form there is no change in plasma membrane integrity in the observed treatment time, which can be attributed to metabolic differences and the resilience of the amastigote. Also, ultrastructural changes such as vacuolization suggesting autophagy were observed. The in vivo effectiveness of MHZ15 in the experimental model of cutaneous leishmaniasis was carried out in mice of the BALB/c strain infected with L. amazonensis. The treatment by intralesional route showed that MHZ15 acted with great efficiency with significantly reduction in the parasite load in the injured paws and draining lymph nodes, without clinical signs of distress or compromise of animal welfare. In vivo toxicity was also evaluated and null alterations in the levels of hepatic enzymes aspartate aminotransferase, and alanine aminotransferase was observed. The data presented herein demonstrates that MHZ15 exhibits a range of favorable characteristics conducive to the development of an antileishmanial agent.

Açai (Euterpe oleracea Mart.) supplementation promotes histological and ultrastructural changes in rats' alveolar bone.

The açai juice contains high concentrations of phenolic compounds, including cyanidin-3-glucoside and others flavonoids. The aim of this study was to evaluate the impact of açai supplementation on healthy mandibular alveolar bone in male albino rats of the Wistar strain. 24 rats were divided into 3 groups, in which one group received a daily dose of saline solution and the other two groups were treated with daily doses of clarified açai juice for 14 or 28 days. After the experiment, hemimandibles were collected and analyzed using Scanning Electron Microscopy (SEM), histological assessments, and micro-CT. Results showed changes in the integrity of the alveolar bone as seen in SEM, increased osteocyte density and higher collagen matrix area in the açai group compared to the control group as seen in histological analysis, and increased bone volume, trabecular thickness and number, and cortical bone as seen in micro-CT analysis. The space between bone trabeculae showed no difference among the groups. These results suggest that açai supplementation may have a structural change effect on alveolar bone, but further research is needed to confirm these findings in humans and to determine the exact mechanisms behind these effects.

T cell maturation is significantly affected by SARS-CoV-2 infection.

Coronavirus disease 2019 (COVID-19) is a respiratory tract infection caused by the new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). An adequate T cell response is essential not only for fighting disease but also for the creation of immune memory. Thus, the present study aims to evaluate the T cells of patients with moderate, severe and critical COVID-19 not only at the time of illness but also 2 months after diagnosis to observe whether changes in this compartment persist. In this study, 166 COVID-19 patients were stratified into moderate/severe and critical disease categories. The maturation and activation of T cells were evaluated through flow cytometry. In addition, Treg cells were analysed. Until 15 days after diagnosis, patients presented a reduction in absolute and relative T lymphocyte counts. After 2 months, in moderate/severe patients, the counts returned to a similar level as that of the control group. In convalescent patients who had a critical illness, absolute T lymphocyte values increased considerably. Patients with active disease did not show differentiation of T cells. Nonetheless, after 2 months, patients with critical COVID-19 showed a significant increase in CD4+ EMRA (CD45RA+ effector memory) T lymphocytes. Furthermore, COVID-19 patients showed delayed T cell activation and reduced CD8+ suppressor T cells even 2 months after diagnosis. A reduction in CD4+ Treg cells was also observed, and their numbers returned to a similar level as that of healthy controls in convalescent patients. The results demonstrate that COVID-19 patients have a delayed activation and differentiation of T cells. In addition, these patients have a great reduction of T cells with a suppressor phenotype.

Influence of Apical Diameter on Filling Material Extrusion during Retreatment - A Micro-CT and CBCT evaluation.

AIM: To investigate whether foraminal widening performed at primary treatment has an effect on the amount of apically extruded obturator material during retreatment and to evaluate the sensitivity of cone beam computed tomography (CBCT) in detecting extruded obturator material. METHODS: Forty palatal roots of maxillary molars were selected based on micro-CT and divided into two groups (n=20): with foraminal widening (WE) and without foraminal widening (NE). To standardize the apical foramen, all specimens were instrumented to the foramen using the Protaper Next system, up to instrument X3. The WE group was instrumented to the foramen up to instrument X5, and the NE group was instrumented 1 mm lower. The canals were obturated 1 mm below the apical foramen with gutta-percha and AH Plus and stored for 7 days at 37 °C and 95% humidity. Roots were fixed in microtubes filled with 1.5% agar gel. The obturation material was removed with Reciproc R50. Scans of the teeth and agar were performed using micro-CT and CBCT. Comparison between groups and between methods was performed using Mann-Withney test (p ≤0.05). RESULTS: No statistical difference was found when comparing the extruded material between groups using micro-CT (p = 0.589) or CBCT (p = 0.953). CBCT measured a greater volume of extruded material than micro- CT (p = 0.0004). CONCLUSION: Foraminal widening had no effect on the extrusion of filling material during retreatment. The CBCT favored the evaluation of apically extruded filling material.

Influence of Apical Diameter on Filling Material Extrusion during Retreatment - A Micro-CT and CBCT evaluation

Abstract Aim: To investigate whether foraminal widening performed at primary treatment has an effect on the amount of apically extruded obturator material during retreatment and to evaluate the sensitivity of cone beam computed tomography (CBCT) in detecting extruded obturator material. Methods: Forty palatal roots of maxillary molars were selected based on micro-CT and divided into two groups (n=20): with foraminal widening (WE) and without foraminal widening (NE). To standardize the apical foramen, all specimens were instrumented to the foramen using the Protaper Next system, up to instrument X3. The WE group was instrumented to the foramen up to instrument X5, and the NE group was instrumented 1 mm lower. The canals were obturated 1 mm below the apical foramen with gutta-percha and AH Plus and stored for 7 days at 37 °C and 95% humidity. Roots were fixed in microtubes filled with 1.5% agar gel. The obturation material was removed with Reciproc R50. Scans of the teeth and agar were performed using micro-CT and CBCT. Comparison between groups and between methods was performed using Mann-Withney test (p ≤0.05). Results: No statistical difference was found when comparing the extruded material between groups using micro-CT (p = 0.589) or CBCT (p = 0.953). CBCT measured a greater volume of extruded material than micro- CT (p = 0.0004). Conclusion: Foraminal widening had no effect on the extrusion of filling material during retreatment. The CBCT favored the evaluation of apically extruded filling material.

Resumo Objetivo: Investigar se o alargamento do foramen realizado durante o tratamento primário tem um efeito na quantidade de material obturador apicalmente extruído durante oretratamento e verificar a sensibilidade da tomografia computorizada de feixe cônico (TCFC) na detecção de material obturador extruído. Métodos: Quarenta raízes palatinas de molares superiores foram selecionadas de acordo com a microtomografia computorizada, e foram divididas em dois grupos (n=20): com alargamento do foraminal (CA) e sem alargamento do foraminal (SA). Para padronizar o forame apical, todas as amostras foram instrumentadas com o sistema ProTaper Next até ao instrumento X3 até o forame. O grupo CA foi instrumentado até ao instrumento X5 até o forame, e o grupo SA foi instrumentado 1 mm aquém. Os canais foram obturados 1 mm abaixo do forame apical com gutta-percha e AH Plus e armazenados durante 7 dias a 37 °C e 95% de umidade. As raízes foram fixadas em microtubos preenchidos com gel de ágar a 1,5%. O material obturador foi removido com Reciproc R50. Os escaneamentos dos dentes e do ágar foram realizados com micro-CT e CBCT. A comparação entre grupos e entre métodos foi realizada utilizando o teste Mann-Withney (p ≤0.05). Resultados: Não foi encontrada diferença estatística ao comparar o material extruído entre grupos utilizando o Micro-CT (p = 0,589) ou TCFC (p = 0,953). Foi medido um volume maior de material extruído com a TCFC do que com a Micro- CT (p = 0,0004). Conclusão: O alargamento foraminal não teve qualquer efeito na extrusão do material obturador durante o retratamento. A TCFC favoreceu a avaliação do material de obturador apicalmente extruído.

Subtilisin of Leishmania amazonensis as Potential Druggable Target: Subcellular Localization, In Vitro Leishmanicidal Activity and Molecular Docking of PF-429242, a Subtilisin Inhibitor.

Subtilisin proteases, found in all organisms, are enzymes important in the post-translational steps of protein processing. In Leishmania major and L. donovani, this enzyme has been described as essential to their survival; however, few compounds that target subtilisin have been investigated for their potential as an antileishmanial drug. In this study, we first show, by electron microscopy and flow cytometry, that subtilisin has broad localization throughout the cytoplasm and membrane of the parasite in the promastigote form with foci in the flagellar pocket. Through in silico analysis, the similarity between subtilisin of different Leishmania species and that of humans were determined, and based on molecular docking, we evaluated the interaction capacity of a serine protease inhibitor against both life cycle forms of Leishmania. The selected inhibitor, known as PF-429242, has already been used against the dengue virus, arenaviruses, and the hepatitis C virus. Moreover, it proved to have antilipogenic activity in a mouse model and caused hypolipidemia in human cells in vitro. Here, PF-429242 significantly inhibited the growth of L. amazonensis promastigotes of four different strains (IC50 values = 3.07 ± 0.20; 0.83 ± 0.12; 2.02 ± 0.27 and 5.83 ± 1.2 µM against LTB0016, PH8, Josefa and LV78 strains) whilst having low toxicity in the host macrophages (CC50 = 170.30 µM). We detected by flow cytometry that there is a greater expression of subtilisin in the amastigote form; however, PF-429242 had a low effect against this intracellular form with an IC50 of >100 µM for intracellular amastigotes, as well as against axenic amastigotes (94.12 ± 2.8 µM for the LV78 strain). In conclusion, even though PF-429242 does not affect the intracellular forms, this drug will serve as a tool to explore pharmacological and potentially leishmanicidal targets.

The relationship between peripheral immune response and disease severity in SARS-CoV-2-infected subjects: A cross-sectional study.

Coronavirus disease 2019 (COVID-19) is a respiratory infection caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and marked by an intense inflammatory response and immune dysregulation in the most severe cases. In order to better clarify the relationship between peripheral immune system changes and the severity of COVID-19, this study aimed to evaluate the frequencies and absolute numbers of peripheral subsets of neutrophils, monocytes, and dendritic cells (DCs), in addition to quantifying the levels of inflammatory mediators. One hundred fifty-seven COVID-19 patients were stratified into mild, moderate, severe, and critical disease categories. The cellular components and circulating cytokines were assessed by flow cytometry. Nitric oxide (NOx) and myeloperoxidase (MPO) levels were measured by colourimetric tests. COVID-19 patients presented neutrophilia, with signs of emergency myelopoiesis. Alterations in the monocytic component were observed in patients with moderate to critical illness, with an increase in classical monocytes and a reduction in nonclassical monocytes, in addition to a reduction in the expression of HLA-DR in all subtypes of monocytes, indicating immunosuppression. DCs, especially plasmacytoid DCs, also showed a large reduction in moderate to critical patients. COVID-19 patients showed an increase in MPO, interleukin (IL)-12, IL-6, IL-10, and IL-8, accompanied by a reduction in IL-17A and NOx. IL-10 levels ≥14 pg/ml were strongly related to the worst outcome, with a sensitivity of 78·3% and a specificity of 79·1%. The results of this study indicate the presence of systemic effects induced by COVID-19, which appear to be related to the pathophysiology of the disease, highlighting the potential of IL-10 as a possible prognostic biomarker for COVID-19.

Influence of conservative endodontic access cavities on instrumentation of oval-shaped straight root canals.

AIM: To evaluate the influence of the design of endodontic access cavities on the percentage of unprepared areas of canal walls and flexural fatigue of instruments activated by reciprocating movement in oval-shaped straight root canals of extracted teeth. METHODOLOGY: Forty-two mandibular incisors with oval canals were scanned by a microcomputed tomography (micro-CT) device for homogeneous selection and distribution of the samples. Then, the teeth were divided into two groups (n = 21) according to the design of access cavity being tested: ultraconservative endodontic access cavity (UltraAC) and traditional access cavity. The canals were accessed with the aid of a surgical microscope, instrumented with the WaveOne Gold Medium system and irrigated with 2.5% NaOCl and 17% EDTA. The unprepared areas of the canal wall were analysed by overlaying images before and after instrumentation and expressed as percentages. micro-CT data were analysed using t-test, Mann-Whitney and Wilcoxon tests. The endodontic instruments used during instrumentation were subjected to static flexural fatigue testing using an artificial stainless steel canal with a 60° angle of curvature and a radius of 5 mm, located 5 mm from the tip of the instrument. The instruments were activated until fracture occurred, and the time in seconds for the fracture was recorded using a digital timer. The number of cycles to fracture was calculated and analysed statistically. For flexural fatigue data, an anova test complemented by a Tukey range test was used. The significance level of 5% was used for all analyses. RESULTS: There was no significant difference between the groups related to unprepared areas by the instrument during canal preparation (p > .05). The difference in flexural fatigue resistance between the groups was not significant. CONCLUSION: The use of UltraAC did not interfere with the canal instrumentation of extracted mandibular incisors with straight and oval canals. There was no difference in the flexural fatigue resistance of the instruments in relation to access cavity design.

Impact of irrigation protocols with some chelators and mechanical agitation on intratubular decontamination.

This study evaluated main canal and intratubular decontamination using different irrigation solutions followed by adjunctive agitation steps for infected root canals. Sixty-eight lower incisors were contaminated with Enterococcus faecalis and allocated to groups according to canal treatment (n = 10): G1, NaOCl followed by ethylenediaminetetraacetic acid (EDTA); G2, a mixture of NaOCl with hydroxyethylidene bisphosphonate (HEBP); and G3, NaOCl followed by EDTA-T (EDTA with sodium lauryl ether sulfate). All three groups of teeth were agitated with passive ultrasonic irrigation (PUI) using saline solution, whereby G4, G5, and G6 were prepared as above, and agitation was performed using an XP-Endo Finisher instrument. Microbiological samples were collected from the root canals with paper points at three times: before and after chemomechanical preparation and after agitation. The colony-forming units (CFU)/mL count was determined, and bacterial intratubular viability was analyzed via confocal laser scanning microscopy using Live/Dead staining. Statistical analysis was performed using a Kruskal-Wallis test followed by Dunn tests. A Friedman test was applied for colony-counting data (α = 0.05). CFU/mL counting indicated equally effective decontamination in the experimental groups (p > 0.05). According to microscopy images, the use of irrigation solutions followed by agitation with the XP-Endo Finisher yielded better results. Moreover, NaOCl+EDTA-T followed by XP-Endo Finisher resulted in significantly lower viability than in the PUI-activated groups (p < 0.05). The cervical and medium thirds of the specimens presented similar results. Overall, NaOCl+EDTA-T exhibited the best intratubular antibacterial activity, mainly for canals that were subsequently agitated using XP-Endo Finisher.

Molhabilidade de um cimento endodôntico à base de resina epóxi na dentina tratada com diferentes protocolos de quelação / Wettability of an epoxy resin-based root canal sealer on dentin treated with different chelating protocols

Objetivo: Avaliar a molhabilidade do cimento AH Plus em contato com a dentina radicular após diferentes protocolos quelantes envolvendo ácido etidrônico (HEBP) e o EDTA. Métodos: Cinquenta e seis fatias de dentina radicular foram utilizadas e irrigadas com 5.25% de hipoclorito de sódio (NaOCl) ou com uma mistura de 5.25%NaOCl/18%HEBP para simular a irrigação durante o preparo químico-mecânico. As amostras irrigadas com NaOCl foram divididas em 5 grupos, de acordo com o agente quelante: G1- água destilada (AD); G2-17%EDTA; G3-17%EDTA+2.5%NaOCl; G4-18%HEBP; e G5-18%HEBP+2.5%NaOCl. As amostras irrigadas com a mistura de NaOCl/HEBP foram dividias em 2 grupos: G6-AD; G7-NaOCl/HEBP+2.5%- NaOCl. Todos os protocolos receberam irrigação de AD entre as substâncias irrigadoras e como lavagem final. O goniômetro Rame-Hart foi utilizado para mesurar o ângulo de contato entre a superfície da dentina radicular e o cimento. O teste estatístico Kruskal-Wallis e Dunn foram aplicados (p<0.05). Resultados: Os grupos onde a smear layer foi removida apresentaram menor ângulo de contato (p<0.05), com exceção do G7. O G6 apresentou o menor ângulo de contato do AH Plus, entretanto, a irrigação final com NaOCl (G7) aumentou significativamente o ângulo de contato. G2 e G4 apresentaram comportamento similar e a irrigação final com NaOCL (G3 e G4) não modificou a molhabilidade da dentina. Conclusão: O tratamento da dentina radicular com a mistura NaOCl/HEBP, quando usada como irrigante, conferiu boa molhabilidade desta superfície ao cimento AH Plus(AU).

Evaluate the wettability of AH Plus in contact with root dentin after different chelating protocols involving etidronic acid (HEBP) and EDTA. Material and Methods: Fifty six human polished root dentin slices were used. They were irrigated with 5.25% sodium hypochlorite (NaOCl) or a mixture of 5.25%NaOCl/18%HEBP to simulate irrigation during chemomechanical preparation. The specimens irrigated with NaOCl were divided into 5 groups regarding chelating agents: G1-destiled water (DW); G2- 17%EDTA; G3-17%EDTA+2.5%NaOCl; G4-18%HEBP; and G5-18%HEBP+2.5%NaOCl. The specimens irrigated with the mixture NaOCl/HEBP were divided into 2 groups: G6-DW; G7-NaOCl/HEBP+2.5%NaOCl. All protocols received irrigation with DW between irrigants and as final rinse. Rame-Hart goniometer was used to measure the contact angle between the dentin surfaces and the sealer. Kruskal-Wallis and Dunn tests were applied (p<0.05). Results: Groups in which the smear layer was removed showed a lower contact angle (p<0.05), except for G7. The G6 showed the lowest contact angle of AH Plus, but the NaOCl final irrigation (G7) increased the angle. G2 and G4 have similar behaviour and final irrigation with NaOCl (G3 and G5) did not change wettability when these chelators were used. Conclusions: The mixture NaOCl/HEBP showed good effect on the wettability of sealer on to the root canal dentine, when used as main irrigant (AU).

Effects of curcumin-mediated antimicrobial photodynamic therapy associated to different chelators against Enterococcus faecalis biofilms.

BACKGROUND: The aim of this study was to evaluate curcumin-mediated antimicrobial photodynamic therapy (aPDT) action combined or not with ethylenediaminetetraacetic acid (EDTA) and hydroxyethylidene bisphosphonate (HEBP) on Enterococcus faecalis biofilms. METHODS: Enterococcus faecalis biofilms were grown on dentin bovine discs in brain heart infusion (BHI) medium with 1% glucose, in aerobic conditions at 37°C for 7 days. Then, they were randomly distributed to one of experimental conditions, as follows: control, 75 J.cm-2 LED, 600 µmol.L-1 curcumin, 17% EDTA, 18% HEBP, 600 µmol.L-1 curcumin plus 75 J.cm-2 LED, 600 µmol.L-1 curcumin plus 17% EDTA, 600 µmol.L-1 curcumin plus 18% HEBP, 600 µmol.L-1 curcumin plus 17% EDTA and 75 J.cm-2 LED or 600 µmol.L-1 curcumin plus 18% HEBP and 75 J.cm-2 LED. The viability of microorganisms and the vitality of biofilms were determined by colony forming unit counts and confocal scanning laser microscopy (CSLM), respectively. Statistical analysis was conducted by Kruskal Wallis and Dunn's post-hoc tests (α = 0.05). RESULTS: The results showed that all combinations of aPDT with chelators significantly reduced the viability of microbial cells and the vitality of biofilms in comparison to control, even when considering deeper layers of biofilms. CONCLUSION: The combination of curcumin with EDTA and HEBP similarly improved the effect of aPDT on E. faecalis biofilms.

The Impact of the Implementation of the Picture Exchange Communication System - PECS on Understanding Instructions in Children with Autism Spectrum Disorders. / O impacto da implementação do Picture Exchange Communication System - PECS na compreensão de instruções em crianças com Transtorno do Espectro do Autismo.

PURPOSE: The purpose of this study was to analyze the impact of the implementation of the Picture Exchange Communication System (PECS) on the comprehension of instructions by children with Autism Spectrum Disorder (ASD). METHODS: This is a longitudinal study (N° 0809/2018). The sample consisted of 20 children with nonverbal ASDs, 15 boys and 5 girls, aged 6 to 12 years, evaluated and diagnosed by a multidisciplinary team according to the DSM-5. For assessment of the comprehension of instructions, we used eight visual instructions and eight oral instructions, which were applied at two points in the PECS Implementation Program: early phases II and IV. The program consisted of 24 individual speech therapy sessions with the presence of a family member and followed the six phases originally proposed by the PECS Training Manual. RESULTS: There was an expressive increase in the comprehension of all instructions, in the comparison between the two moments of the study; and this increase was statistically significant in six of the oral instructions (p=0.001) and five of the visual ones (p=0.002). CONCLUSION: It was possible to observe the positive impact of the use of PECS in the comprehension of both visual and oral instructions, showing that this system not only provides an augmentative or alternative communication tool for the children to express themselves but also promotes significant improvement in the understanding of contextual information.

OBJETIVO: O objetivo deste estudo foi analisar o impacto da implementação do Picture Exchange Communication System ­ PECS na compreensão de instruções de crianças com Transtorno do Espectro do Autismo (TEA). MÉTODO: Trata-se de estudo longitudinal. A amostra foi constituída por 20 crianças com TEA não verbais, sendo 15 meninos e 5 meninas, na faixa etária de 6 a 12 anos, avaliadas e diagnosticadas por equipe multidisciplinar, segundo os critérios do DSM-5. Para avaliação da compreensão de instruções foram aplicadas 8 instruções visuais e 8 instruções orais, em dois momentos do Programa de Implementação do PECS: no início das fases II e IV. O programa foi composto por 24 sessões de terapia fonoaudiológica individual com a presença do familiar e obedeceu às seis fases propostas originalmente pelo Manual de Treinamento do PECS. RESULTADOS: Houve aumento expressivo na compreensão de todas as instruções, na comparação entre os dois momentos do estudo; sendo que em seis das instruções orais (p=0,001) e cinco das instruções visuais (p=0,002), esse aumento foi estatisticamente significante. CONCLUSÃO: Foi possível observar o impacto positivo do uso do PECS na compreensão de instruções tanto visuais quanto orais, mostrando que esse sistema não apenas fornece uma ferramenta de comunicação aumentativa ou alternativa para a criança se expressar, mas também promove melhora significativa na compreensão das informações contextuais.

PF-429242, a Subtilisin Inhibitor, Is Effective in vitro Against Leishmania infantum.

PF-429242 is an inhibitor of subtilisin, an important protease found in Leishmania. However, studies regarding the effect of PF-429242 on Leishmania are scarce. In this work we evaluated the antileishmanial effect of PF-429242 against Leishmania infantum and the mechanism involved in the death of the parasite. PF-429242 had low toxicity against mammalian cells (peritoneal macrophages) (CC50 = 189.07 µM) and presented activity against L. infantum promastigotes (IC50 = 2.78 µM) and intracellular amastigotes (IC50 = 14.07 µM), indicating selectivity toward the parasite. Transmission electron microscopy (TEM), as well as staining of L. infantum promastigotes with MitoTracker® Red, rhodamine 123 and MitoSOX, revealed that the mitochondria was a potential target of PF-429242. In addition, PF-429242 caused an accumulation of neutral lipids in promastigotes, which was demonstrated by Nile Red staining and TEM, and induced oxidative stress (H2DCFDA staining). Furthermore the formation of autophagic vacuoles in L. infantum promastigotes was observed by MDC staining and TEM. However, the killing induced by PF-429242 in L. infantum promastigotes appeared to be unrelated to apoptosis and/or necrosis as there was no phosphatidylserine externalization, DNA fragmentation or alterations in the permeability of the parasite plasma membrane, as assessed by annexin V-FITC, TUNEL and propidium iodide staining, respectively. The morphological and ultrastructural evaluation of the promastigotes by optical microscopy, scanning electron microscopy (SEM) and TEM, revealed the presence of parasites with flagellar defects. TEM analysis of the intracellular amastigotes indicated that mitochondrial damage and autophagy could also be involved in the death of these forms after treatment with PF-429242. In addition, PF-429242 treatment stimulated NO production from infected macrophage, but only at a high concentration (100 µM), as well as an increase of TNF levels after treatment with 10 µM of PF-429242. The compound did not stimulate ROS or IL-10 production. Together, these data highlight the antileishmanial potential of PF-429242, inducing several cellular alterations in the parasite, such as mitochondrial damage, neutral lipids accumulation, oxidative stress and autophagy which culminate in the death of L. infantum, as well as modulating host cellular responses that favor the development of an immune response against the parasite.

Influência do acesso minimamente invasivo na fratura de dentes tratados endodonticamente - uma revisão de literatura / Influence of minimally invasive access on fracture of endodonti-cally treated teeth- a literature review

O acesso à cavidade pulpar é a etapa do tratamento endodôntico que tem como objetivo expor a embocadura dos canais radiculares. Por muito tempo, o formato ideal da cavidade era aquele que proporcionasse a criação de uma trajetória reta ao canal, com a remoção completa do teto da câmara pulpar. Porém, nas últimas décadas, foi investigado um desgaste excessivo de dentina que possibilita a redução da resistência do dente. Então, propuseram novo formato de cavidade de acesso, que permite a preservação máxima possível das estruturas de suporte, objetivando aumentar a resistência de dentes tratados endodonticamente. Apesar das vantagens, supostamente atribuídas aos acessos minimamente invasivos, esse formato tem sido questionado por dificultar a visibilidade da entrada dos canais, localização, e possibilidade de deixar áreas intocadas nas paredes dos canais. Diante disso, este trabalho realizou um levantamento bibliográfico a fim de verificar se há consistência científica quanto à interferência do acesso coronário na resistência dentária. Concluiu-se que o acesso minimamente invasivo não apresentou diferença quanto ao aumento da resistência à fratura de dentes tratados endodonticamente quando comparado ao acesso tradicional, entretanto, o tema ainda é relevante e os benefícios dessa técnica devem ser investigados clinicamente a longo prazo.

Access to the pulp cavity is the stage of endodontic treatment that aims to expose the mouth of the root canals. For a long time, creating a straight path to the canal, with the complete removal of the pulp chamber roof, was the ideal cavity format. However, in recent decades, excessive dentin wear, which makes it possible to reduce tooth strength, has been investi-gated. So, a new access cavity format, which allows the maximum possible preservation of the support structures, was proposed to increase the resistance of endodontically treated teeth. Despite the advantages supposedly attributed to minimally invasive accesses, this format has been questioned for hindering the visibility of the entrance to the channels, the location, and the possibility of leaving untouched areas on the walls of the channels. Therefore, this work carried out a bibliographical survey to verify the scientific consistency regarding the interference of coronary access in dental resistance. The minimally inva-sive access showed no difference regarding increased fracture resistance of endodontically treated teeth compared to the traditional access. However, the topic is still relevant, and its benefits, in the long term, should be clinically investigated.

O impacto da implementação do Picture Exchange Communication System - PECS na compreensão de instruções em crianças com Transtorno do Espectro do Autismo / The Impact of the Implementation of the Picture Exchange Communication System - PECS on Understanding Instructions in Children with Autism Spectrum Disorders

RESUMO Objetivo O objetivo deste estudo foi analisar o impacto da implementação do Picture Exchange Communication System - PECS na compreensão de instruções de crianças com Transtorno do Espectro do Autismo (TEA). Método Trata-se de estudo longitudinal. A amostra foi constituída por 20 crianças com TEA não verbais, sendo 15 meninos e 5 meninas, na faixa etária de 6 a 12 anos, avaliadas e diagnosticadas por equipe multidisciplinar, segundo os critérios do DSM-5. Para avaliação da compreensão de instruções foram aplicadas 8 instruções visuais e 8 instruções orais, em dois momentos do Programa de Implementação do PECS: no início das fases II e IV. O programa foi composto por 24 sessões de terapia fonoaudiológica individual com a presença do familiar e obedeceu às seis fases propostas originalmente pelo Manual de Treinamento do PECS. Resultados Houve aumento expressivo na compreensão de todas as instruções, na comparação entre os dois momentos do estudo; sendo que em seis das instruções orais (p=0,001) e cinco das instruções visuais (p=0,002), esse aumento foi estatisticamente significante. Conclusão Foi possível observar o impacto positivo do uso do PECS na compreensão de instruções tanto visuais quanto orais, mostrando que esse sistema não apenas fornece uma ferramenta de comunicação aumentativa ou alternativa para a criança se expressar, mas também promove melhora significativa na compreensão das informações contextuais.

ABSTRACT Purpose The purpose of this study was to analyze the impact of the implementation of the Picture Exchange Communication System (PECS) on the comprehension of instructions by children with Autism Spectrum Disorder (ASD). Methods This is a longitudinal study (N° 0809/2018). The sample consisted of 20 children with nonverbal ASDs, 15 boys and 5 girls, aged 6 to 12 years, evaluated and diagnosed by a multidisciplinary team according to the DSM-5. For assessment of the comprehension of instructions, we used eight visual instructions and eight oral instructions, which were applied at two points in the PECS Implementation Program: early phases II and IV. The program consisted of 24 individual speech therapy sessions with the presence of a family member and followed the six phases originally proposed by the PECS Training Manual. Results There was an expressive increase in the comprehension of all instructions, in the comparison between the two moments of the study; and this increase was statistically significant in six of the oral instructions (p=0.001) and five of the visual ones (p=0.002). Conclusion It was possible to observe the positive impact of the use of PECS in the comprehension of both visual and oral instructions, showing that this system not only provides an augmentative or alternative communication tool for the children to express themselves but also promotes significant improvement in the understanding of contextual information.

Impact of irrigation protocols with some chelators and mechanical agitation on intratubular decontamination

Abstract: This study evaluated main canal and intratubular decontamination using different irrigation solutions followed by adjunctive agitation steps for infected root canals. Sixty-eight lower incisors were contaminated with Enterococcus faecalis and allocated to groups according to canal treatment (n = 10): G1, NaOCl followed by ethylenediaminetetraacetic acid (EDTA); G2, a mixture of NaOCl with hydroxyethylidene bisphosphonate (HEBP); and G3, NaOCl followed by EDTA-T (EDTA with sodium lauryl ether sulfate). All three groups of teeth were agitated with passive ultrasonic irrigation (PUI) using saline solution, whereby G4, G5, and G6 were prepared as above, and agitation was performed using an XP-Endo Finisher instrument. Microbiological samples were collected from the root canals with paper points at three times: before and after chemomechanical preparation and after agitation. The colony-forming units (CFU)/mL count was determined, and bacterial intratubular viability was analyzed via confocal laser scanning microscopy using Live/Dead staining. Statistical analysis was performed using a Kruskal-Wallis test followed by Dunn tests. A Friedman test was applied for colony-counting data (α = 0.05). CFU/mL counting indicated equally effective decontamination in the experimental groups (p > 0.05). According to microscopy images, the use of irrigation solutions followed by agitation with the XP-Endo Finisher yielded better results. Moreover, NaOCl+EDTA-T followed by XP-Endo Finisher resulted in significantly lower viability than in the PUI-activated groups (p < 0.05). The cervical and medium thirds of the specimens presented similar results. Overall, NaOCl+EDTA-T exhibited the best intratubular antibacterial activity, mainly for canals that were subsequently agitated using XP-Endo Finisher.

Cytotoxicity assessment of beta-lapachone in endothelial cells

The selective activity of an antineoplastic drug is related to its ability to promote cytotoxic action on tumor cells and preserve the integrity of non-neoplastic cells. Beta-lapachone is extracted from the sawdust of Ipe wood, a thick bark tree from the Ipe wood found in the Brazilian Cerrado biome. This study aimed to evaluate the cytotoxic action of beta-lapachone in an endothelial cell line. The EA.hy926 cells were seeded in two groups, G1 and G2, cultured and exposed to beta-lapachone at concentrations of 0.0, 0.01, 0.03, 0.1, 0.3, 1 and 3 & 956;M for 24 hours. G1 remained under normal cultivation conditions and G2 was subjected to oxidative stress through an ischemia and reperfusion assay, in a deoxygenated sealed chamber. The cytotoxicity assay was performed using the tetrazolium reduction method. In G1, the cytotoxicity ranged from 0.0 to 10.0%; and in G2 between 0.0 and 6.3%. No statistically significant difference was observed between the obtained values. Moreover, we found no cytotoxic action of beta-lapachone on endothelial cells, and the results point out that the drug might have preserved the cell’s integrity against oxidative stress under the conditions of this experiment. This promising result suggests the possibility of beta-lapachone as a chemotherapy drug with selective activity.

Vitamin D increases killing of intracellular Leishmania amazonensis in vitro independently of macrophage oxidative mechanisms.

Vitamin D has been reported to activate macrophage microbicidal mechanisms by inducing the production of antimicrobial peptides and nitric oxide (NO), but conversely has been shown to contribute to a greater susceptibility to Leishmania amazonensis infection in mice. Thus, this study aimed to evaluate the role of vitamin D during intracellular infection with L. amazonensis by examining its effect on macrophage oxidative mechanisms and parasite survival in vitro. Vitamins D2 and D3 significantly inhibited promastigote and amastigote growth in vitro. Vitamin D3 was not able to induce NO and reactive oxygen species (ROS) production in uninfected macrophages or macrophages infected with L. amazonensis. In addition, vitamin D3 in combination with interferon (IFN)-γ did not enhance amastigote killing and in fact, significantly reduced NO and ROS production when compared with the effect of IFN-γ alone. In this study, we demonstrated that vitamin D directly reduces parasite growth in infected macrophages (approximately 50-60% at 50 µm) but this effect is independent of the activation of macrophage oxidative mechanisms. These findings will contribute to a better understanding of the role of vitamin D in cutaneous leishmaniasis.

Safety of hospitalized older adult patients: an analysis of the risk of falls. / Segurança do paciente idoso hospitalizado: uma análise do risco de quedas.

OBJECTIVE: To identify the risk of falls of older adults in a hospital in the Trairi region in Rio Grande do Norte, Brazil, and to describe the relationship between risk of falls and the sociodemographic characteristics of the participants. METHOD: A descriptive, transversal, and quantitative study conducted in a regional hospital, respecting the ethical principles in force. The Morse Scale was adopted for data collection, which took place in July and August 2018, and adapted with sociodemographic questions. A descriptive and inferential statistical analysis was performed. RESULTS: There were 46 participants in the study, most of whom were women with low education, and the most frequent reasons for hospitalization were surgical treatment and lung disease. More than half had a high risk of falling (54.35%), followed by moderate (32.61%) and low (13.04%). There was an association between high risk of falls, having lung disease as a reason for hospitalization and diabetes as a comorbidity. The high risk of falls was less frequent among older adult patients hospitalized for surgical treatment. CONCLUSION: The high risk of falls was found in more than half of the older adults, which varied according to the reason for hospitalization and comorbidities, being more frequent in those hospitalized for lung disease and in those who had Diabetes.

Magnitude and Dynamics of the T-Cell Response to SARS-CoV-2 Infection at Both Individual and Population Levels

T cells are involved in the early identification and clearance of viral infections and also support the development of antibodies by B cells. This central role for T cells makes them a desirable target for assessing the immune response to SARS-CoV-2 infection. Here, we combined two high-throughput immune profiling methods to create a quantitative picture of the T-cell response to SARS-CoV-2. First, at the individual level, we deeply characterized 3 acutely infected and 58 recovered COVID-19 subjects by experimentally mapping their CD8 T-cell response through antigen stimulation to 545 Human Leukocyte Antigen (HLA) class I presented viral peptides (class II data in a forthcoming study). Then, at the population level, we performed T-cell repertoire sequencing on 1,815 samples (from 1,521 COVID-19 subjects) as well as 3,500 controls to identify shared "public" T-cell receptors (TCRs) associated with SARS-CoV-2 infection from both CD8 and CD4 T cells. Collectively, our data reveal that CD8 T-cell responses are often driven by a few immunodominant, HLA-restricted epitopes. As expected, the T-cell response to SARS-CoV-2 peaks about one to two weeks after infection and is detectable for at least several months after recovery. As an application of these data, we trained a classifier to diagnose SARSCoV-2 infection based solely on TCR sequencing from blood samples, and observed, at 99.8% specificity, high early sensitivity soon after diagnosis (Day 3-7 = 85.1% [95% CI = 79.9-89.7]; Day 8-14 = 94.8% [90.7-98.4]) as well as lasting sensitivity after recovery (Day 29+/convalescent = 95.4% [92.1-98.3]). These results demonstrate an approach to reliably assess the adaptive immune response both soon after viral antigenic exposure (before antibodies are typically detectable) as well as at later time points. This blood-based molecular approach to characterizing the cellular immune response has applications in clinical diagnostics as well as in vaccine development and monitoring.