Glucose metabolism sustains heme-induced Trypanosoma cruzi epimastigote growth in vitro.

Chagas disease is caused by the protozoan parasite, Trypanosoma cruzi. This parasite alternates between an insect vector and a mammalian host. T. cruzi epimastigotes reside in the insect vector and coexist with the blood components of the vertebrate host. The metabolic profile of T. cruzi has been extensively studied; however, changes in its metabolism in response to signaling molecules present in the vector are poorly understood. Heme acts as a physiological oxidant that triggers intense epimastigote proliferation and upregulates the expression of genes related to glycolysis and aerobic fermentation in vitro. Here, heme-cultured epimastigotes increased D-glucose consumption. In fact, heme-cultured parasites secreted more succinate (the end product of the so-called succinic fermentation) followed by glucose intake. Increased succinate levels reduced the extracellular pH, leading to acidification of the supernatant. However, the acidification and proliferation stimulated by heme was impaired when glycolysis was inhibited. Otherwise, when glucose amount is enhanced in supernatant, heme-cultured parasites increased its growth whereas the glucose depletion caused a delay in proliferation. Heme supplementation increased epimastigote electron transport system-related O2 consumption rates, while glucose addition reduced both the electron transport system-related O2 consumption rates and spare respiratory capacity, indicating a Crabtree-like effect. These results show that glycolysis predominated in heme-cultured epimastigotes over oxidative phosphorylation for energy supply when glucose is present to sustain its high proliferation in vitro. Furthermore, it provided an insight into the parasite biology in the vector environment that supply glucose and the digestion of blood generates free heme that can lead to the growth of T. cruzi epimastigotes.

Hypoxia Effects on Trypanosoma cruzi Epimastigotes Proliferation, Differentiation, and Energy Metabolism.

Trypanosoma cruzi, the causative agent of Chagas disease, faces changes in redox status and nutritional availability during its life cycle. However, the influence of oxygen fluctuation upon the biology of T. cruzi is unclear. The present work investigated the response of T. cruzi epimastigotes to hypoxia. The parasites showed an adaptation to the hypoxic condition, presenting an increase in proliferation and a reduction in metacyclogenesis. Additionally, parasites cultured in hypoxia produced more reactive oxygen species (ROS) compared to parasites cultured in normoxia. The analyses of the mitochondrial physiology demonstrated that hypoxic condition induced a decrease in both oxidative phosphorylation and mitochondrial membrane potential (ΔΨm) in epimastigotes. In spite of that, ATP levels of parasites cultivated in hypoxia increased. The hypoxic condition also increased the expression of the hexokinase and NADH fumarate reductase genes and reduced NAD(P)H, suggesting that this increase in ATP levels of hypoxia-challenged parasites was a consequence of increased glycolysis and fermentation pathways. Taken together, our results suggest that decreased oxygen levels trigger a shift in the bioenergetic metabolism of T. cruzi epimastigotes, favoring ROS production and fermentation to sustain ATP production, allowing the parasite to survive and proliferate in the insect vector.

Does outpatient dental treatment have impact on the quality of life of children with early childhood caries?

AIM: To assess the impact of outpatient dental treatment on oral health-related quality of life (OHRQoL) of children and their families. MATERIALS AND METHODS: This prospective observational study was conducted with children with untreated dental caries, recruited from a public university/the pediatric dentistry clinic of a public university in the northeastern Brazil. Sociodemographic data were collected, a clinical examination was carried out before and after dental treatment, and the Early Childhood Oral Health Impact Scale (ECOHIS) questionnaire was applied to those responsible for the children, before and after dental treatment. Descriptive and bivariate statistics were used, and the Wilcoxon and Student's t tests were applied (p < 0.05). RESULTS: The study included 64 children, of both sexes, aged between 3 and 5 years old. The majority had a dmft index higher than or equal to 6 (60.9%), with a mean value of 7.11 (± 4.11). Sociodemographic conditions such as sex, age, and socioeconomic status had no impact on their OHRQoL (p > 0.05). The mean total ECOHIS questionnaire scores and those of its domains decreased after completion of the treatments (p < 0.05), except for scores in the self-image and distress domains of the parents (p > 0.05). This study reinforces the relevance of adopting oral health policies aimed at the prevention and treatment of ECC. CONCLUSION: Outpatient dental treatment had a positive impact on the OHRQoL of children and their families. CLINICAL RELEVANCE: The possibility of evaluating the services provided by the pediatric dentistry clinic.

LSTR Antibiotic Paste Versus Zinc Oxide and Eugenol Pulpectomy for the Treatment of Primary Molars with Pulp Necrosis: A Randomized Controlled Trial.

Purpose: The purpose of this study was to compare the effectiveness of lesion sterilization and tissue repair (LSTR) antibiotic paste comprised of chloramphenicol, tetracycline, and zinc oxide and eugenol (CTZ) versus zinc oxide eugenol (ZOE) pulpectomy in the treatment of primary molars with pulp necrosis. Methods: A total of 70 three- to eight-year-old subjects with 88 primary mandibular molars with pulp necrosis were included. The teeth were randomized to the CTZ group or ZOE group. The time taken to perform both techniques was recorded. The parents of the children and the dentist who performed clinical evaluations were blind to the group assignment, although the radiographic evaluator could see the difference in treatments. Clinical and radiographic assessments were performed at three, six, nine, and 12 months. Results: At the 12-month evaluation, the clinical success was 86.4 percent for CTZ and 90.9 percent for ZOE (P=0.50), the radiographic success was 75.0 percent for CTZ and 72.7 percent for ZOE (P=0.81), and the overall success was 70.5 percent for CTZ and 72.7 percent for ZOE (P=0.81). The mean time taken to perform was 61.4 (±20.5 standard deviation) minutes for CTZ and 145.1 (±53.2) minutes for ZOE (P<0.001). Conclusions: At 12 months, both techniques presented no significant difference in success rates for nonvital pulp therapy in primary molars with necrosis. The lesion sterilization and tissue repair procedure time using chloramphenicol, tetracycline, zinc oxide, and eugenol was significantly shorter than for a zinc oxide eugenol pulpectomy.

[Equity and access to mental health care in three Northeastern states]. / Equidade e acesso aos cuidados em saúde mental em três estados nordestinos.

The scope of this study is to discuss equity and access to mental health care by means of analysis of the distribution of primary care teams (Community Health Agents; Family Health Teams; the Expanded Family Health Nucleus) and the coverage of the Psychosocial Care Network in three Northeastern states (Ceará, Piauí and Rio Grande do Norte) and their respective health regions. It is a descriptive study, supported by exploratory quantitative analysis. For this purpose, the database of the Department of Basic Care/DATASUS was used to collect secondary data in relation to the historical trajectory of training of Primary Care teams. Regarding the points of care of RAPS, the database made available by the National Mental Health Coordination of the Ministry of Health was consulted. The conclusion of the investigation was that prior to 2018 there was a major drive in the country into the interior and expansion of coverage in relation to primary and psychosocial care, impacting the expansion of equity in territories of lesser economic and social development. However, it was also observed that Psychosocial Care Network services are more prevalent in the interior while the other related services are more developed in capitals and large municipalities.

Objetiva-se discutir equidade e acesso em saúde mental por meio da análise da distribuição das equipes da atenção primária (Agentes comunitários de Saúde; Equipes de saúde da Família; Núcleo Ampliado de Saúde da Família) e da cobertura da Rede de Atenção Psicossocial (RAPS) em três estados nordestinos (Ceará, Piauí e Rio Grande do Norte) nas suas respectivas regiões de saúde. Trata-se de um estudo descritivo, apoiado por análise quantitativa exploratória. Para tanto, recorremos à base de dados do Departamento de Atenção Básica/DATASUS para a coleta dos dados secundários em relação às séries históricas de habilitação das equipes da Atenção Primária. Quanto aos pontos de atenção da RAPS, utilizamos a base de dados disponibilizada pela Coordenação Nacional de Saúde Mental do Ministério da Saúde. A investigação concluiu que até 2018 observou-se no país um importante movimento de interiorização e expansão da cobertura em relação à atenção primária e psicossocial, impactando na ampliação da equidade nos territórios de menor desenvolvimento econômico e social. Contudo, verificou-se que os serviços de APS estão mais interiorizados, enquanto os demais dispositivos da RAPS estão mais desenvolvidos nas capitais e grandes municípios.

The Potent Trypanocidal Effect of LQB303, a Novel Redox-Active Phenyl-Tert-Butyl-Nitrone Derivate That Causes Mitochondrial Collapse in Trypanosoma cruzi.

Chagas disease, which is caused by Trypanosoma cruzi, establishes lifelong infections in humans and other mammals that lead to severe cardiac and gastrointestinal complications despite the competent immune response of the hosts. Furthermore, it is a neglected disease that affects 8 million people worldwide. The scenario is even more frustrating since the main chemotherapy is based on benznidazole, a drug that presents severe side effects and low efficacy in the chronic phase of the disease. Thus, the search for new therapeutic alternatives is urgent. In the present study, we investigated the activity of a novel phenyl-tert-butyl-nitrone (PBN) derivate, LQB303, against T. cruzi. LQB303 presented trypanocidal effect against intracellular [IC50/48 h = 2.6 µM] and extracellular amastigotes [IC50/24 h = 3.3 µM] in vitro, leading to parasite lysis; however, it does not present any toxicity to host cells. Despite emerging evidence that mitochondrial metabolism is essential for amastigotes to grow inside mammalian cells, the mechanism of redox-active molecules that target T. cruzi mitochondrion is still poorly explored. Therefore, we investigated if LQB303 trypanocidal activity was related to the impairment of the mitochondrial function of amastigotes. The investigation showed there was a significant decrease compared to the baseline oxygen consumption rate (OCR) of LQB303-treated extracellular amastigotes of T. cruzi, as well as reduction of "proton leak" (the depletion of proton motive force by the inhibition of F1Fo ATP synthase) and "ETS" (maximal oxygen consumption after uncoupling) oxygen consumption rates. Interestingly, the residual respiration ("ROX") enhanced about three times in LQB303-treated amastigotes. The spare respiratory capacity ratio (SRC: cell ability to meet new energy demands) and the ATP-linked OCR were also impaired by LQB303 treatment, correlating the trypanocidal activity of LQB303 with the impairment of mitochondrial redox metabolism of amastigotes. Flow cytometric analysis demonstrated a significant reduction of the ΔΨm of treated amastigotes. LQB303 had no significant influence on the OCR of treated mammalian cells, evidencing its specificity against T. cruzi mitochondrial metabolism. Our results suggest a promising trypanocidal activity of LQB303, associated with parasite bioenergetic inefficiency, with no influence on the host energy metabolism, a fact that may point to an attractive alternative therapy for Chagas disease.

Equidade e acesso aos cuidados em saúde mental em três estados nordestinos / Equity and access to mental health care in three Northeastern states

Resumo Objetiva-se discutir equidade e acesso em saúde mental por meio da análise da distribuição das equipes da atenção primária (Agentes comunitários de Saúde; Equipes de saúde da Família; Núcleo Ampliado de Saúde da Família) e da cobertura da Rede de Atenção Psicossocial (RAPS) em três estados nordestinos (Ceará, Piauí e Rio Grande do Norte) nas suas respectivas regiões de saúde. Trata-se de um estudo descritivo, apoiado por análise quantitativa exploratória. Para tanto, recorremos à base de dados do Departamento de Atenção Básica/DATASUS para a coleta dos dados secundários em relação às séries históricas de habilitação das equipes da Atenção Primária. Quanto aos pontos de atenção da RAPS, utilizamos a base de dados disponibilizada pela Coordenação Nacional de Saúde Mental do Ministério da Saúde. A investigação concluiu que até 2018 observou-se no país um importante movimento de interiorização e expansão da cobertura em relação à atenção primária e psicossocial, impactando na ampliação da equidade nos territórios de menor desenvolvimento econômico e social. Contudo, verificou-se que os serviços de APS estão mais interiorizados, enquanto os demais dispositivos da RAPS estão mais desenvolvidos nas capitais e grandes municípios.

Abstract The scope of this study is to discuss equity and access to mental health care by means of analysis of the distribution of primary care teams (Community Health Agents; Family Health Teams; the Expanded Family Health Nucleus) and the coverage of the Psychosocial Care Network in three Northeastern states (Ceará, Piauí and Rio Grande do Norte) and their respective health regions. It is a descriptive study, supported by exploratory quantitative analysis. For this purpose, the database of the Department of Basic Care/DATASUS was used to collect secondary data in relation to the historical trajectory of training of Primary Care teams. Regarding the points of care of RAPS, the database made available by the National Mental Health Coordination of the Ministry of Health was consulted. The conclusion of the investigation was that prior to 2018 there was a major drive in the country into the interior and expansion of coverage in relation to primary and psychosocial care, impacting the expansion of equity in territories of lesser economic and social development. However, it was also observed that Psychosocial Care Network services are more prevalent in the interior while the other related services are more developed in capitals and large municipalities.

Adolescent internet addiction - role of parental control and adolescent behaviours.

INTRODUCTION: Excessive Internet use can negatively affect academic performance, family relationships and emotional development among the youth. Such issues have been identified as Internet addiction (IA). We aimed to determine the prevalence of IA among Portuguese adolescents and assess how parental control can relate to IA. METHODS: An observational cross-sectional study was performed at public schools within a Portuguese region, using Young's Internet Addiction Test survey. General sociodemographic and emotional well-being data were obtained. A descriptive and bivariate analysis was done among Internet-addicted and average users, followed by a logistic regression analysis. Adjusted odds ratios (aORs) were computed with two-sided P values < .05 for statistical significance. RESULTS: A total of 1916 eligible responses were obtained. Mean age was 15 ± 1.8 years, with a slight predominance of female (53.3%) participants. In our sample, 16.5% were deemed Internet addicted and less likely to have any parental control over Internet use (aOR 0.74, P <.05). Moreover, 28% of the Internet-addicted users were less likely to have control over time spent online (aOR 0.72, P < .05), and close to half were unlikely to have online content restrictions (aOR 0.56, P < .01). CONCLUSIONS: Our findings reported a significant rate of Internet-addicted youth. IA was negatively related to parental control. Whenever any kind of parental control over Internet use was reported, IA was less likely to occur. Healthcare professionals should be aware of the risks of IA in adolescents to improve its prevention and intervention.

New perspectives for hydrogen peroxide in the amastigogenesis of Trypanosoma cruzi in vitro.

Trypanosoma cruzi has a complex life cycle involving four life stages: the replicative epimastigotes and metacyclic trypomastigotes in the invertebrate host digestive tract, and intracellular amastigotes and bloodstream trypomastigotes in the mammalian host. Trypomastigotes can invade any nucleated cell, including macrophages, which produce ROS that enhance intracellular infection. However, how ROS modulate T. cruzi infection in the mammalian cell remains unclear. Therefore, the present work aimed to investigate the role of ROS during the stimulation of amastigogenesis in vitro. Our results showed that H2O2 improves the differentiation process in vitro and that it was impaired by Peg-Catalase. However, the antioxidants GSH and NAC had no influence on induced amastigogenesis, which suggests the specificity of H2O2 to increase intracellular differentiation. Amastigogenesis physiologically occurs in low pH, thus we investigated whether parasites are able to produce ROS during amastigogenesis. Interestingly, after 60 min of differentiation induction in vitro, we observed an increase in H2O2 production, which was inhibited by the mitochondrial-targeted antioxidant, mitoTEMPO and Cyclosporine A (a mitochondrial permeability transition pore -mPTP- inhibitor), suggesting mitochondrion as a H2O2 source. Indeed, quantitative real time (qPCR) showed an increase of the mitochondrial superoxide dismutase (FeSODA) gene expression after 60 min of induced amastigogenesis, reinforcing the hypothesis of mitochondrial ROS induction during intracellular differentiation of T. cruzi. The reduction of cellular respiration and the decreased ΔΨm observed during amastigogenesis can explain the increased mitochondrial ROS through mPTP opening. In conclusion, our results suggest that H2O2 is involved in the amastigogenesis of T. cruzi.

Oleic acid and hydroxytyrosol present in olive oil promote ROS and inflammatory response in normal cultures of murine dermal fibroblasts through the NF-κB and NRF2 pathways.

Few studies have evaluated the effects of olive oil on normal tissues like skin and its components. Hence, we investigated whether olive oil could increase the production of ROS and oxidative damage in murine dermal fibroblast cultures in a short-term exposition. In addition, we evaluated the role of oleic acid and hydroxytyrosol, which are the two most important components of olive oil, in the associated mechanisms of action, and the metabolism of long-chain fatty acids from olive oil. To study this, neonatal murine dermal fibroblasts (NMDF) were incubated with olive oil, oleic acid, or hydroxytyrosol for 24 or 72 h. The NMDF incubated with olive oil or oleic acid showed an increase in the production of ROS after 24 h, lipid peroxidation, and protein carbonylation after 72 h, as well as increased expression of nuclear factor-kappa B (NF-κB) p65 and cyclooxygenase-2 (COX-2) after 72 h. However, NMDF treated with olive oil or hydroxytyrosol demonstrated an increase in the expression of nuclear factor-erythroid2-related factor 2 (NRF2) and heme oxygenase-1 (HO-1) after 72 h. In addition, NMDF treated with olive oil also showed an increase in the protein expression of diacylglycerol acyltransferase1 (DGAT1), which promotes triacylglycerol synthesis, and in the levels of triacylglycerols. The microscopic analysis showed Nile red-positive lipid droplets inside olive oil-treated NMDF after 72 h. Moreover, gas chromatography-mass spectrometry demonstrated high levels of oleic acid in the olive oil-treated NMDF after 72 h. In conclusion, oleic acid present in the olive oil promotes the production of ROS and oxidative damage in murine dermal fibroblasts, which leads to NF-κB p65 and COX-2 expression, while hydroxytyrosol promotes NRF2 and HO-1 expression. In addition, NMDF area capable of absorbing long-chain fatty acids derived from olive oil, which promotes the synthesis and the accumulation of triacylglycerols into cytoplasm of NMDF through DGAT1 activation.

Gene expression profiling of Trypanosoma cruzi in the presence of heme points to glycosomal metabolic adaptation of epimastigotes inside the vector.

Chagas disease, also known as American trypanosomiasis, is a potentially life-threatening illness caused by the protozoan parasite, Trypanosoma cruzi, and is transmitted by triatomine insects during its blood meal. Proliferative epimastigotes forms thrive inside the insects in the presence of heme (iron protoporphyrin IX), an abundant product of blood digestion, however little is known about the metabolic outcome of this signaling molecule in the parasite. Trypanosomatids exhibit unusual gene transcription employing a polycistronic transcription mechanism through trans-splicing that regulates its life cycle. Using the Deep Seq transcriptome sequencing we characterized the heme induced transcriptome of epimastigotes and determined that most of the upregulated genes were related to glucose metabolism inside the glycosomes. These results were supported by the upregulation of glycosomal isoforms of PEPCK and fumarate reductase of heme-treated parasites, implying that the fermentation process was favored. Moreover, the downregulation of mitochondrial gene enzymes in the presence of heme also supported the hypothesis that heme shifts the parasite glycosomal glucose metabolism towards aerobic fermentation. These results are examples of the environmental metabolic plasticity inside the vector supporting ATP production, promoting epimastigotes proliferation and survival.

Author Correction: Clinical Manifestations and Ophthalmic Outcomes of Ocular Syphilis at a Time of Re-Emergence of the Systemic Infection.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has not been fixed in the paper.

Clinical Manifestations and Ophthalmic Outcomes of Ocular Syphilis at a Time of Re-Emergence of the Systemic Infection.

Recent reports from different world regions suggest ocular syphilis is re-emerging, in parallel with an increasing incidence of the systemic infection globally. We conducted a large observational study of 127 persons consecutively treated for ocular syphilis at public medical centers in Brazil over a 2.5-year period ending July 2015. Of 104 individuals serologically tested for human immunodeficiency virus (HIV), 34.6% were positive. Ophthalmological evaluations included measurement of Snellen visual acuity and intraocular pressure, and assessment of inflammation by slit lamp examination and dilated posterior eye examination. Involvements in 214 eyes were anterior (6.1%), intermediate (8.4%), posterior (76.2%) and pan- (8.4%) uveitis, and scleritis (0.9%). Multiple anterior and posterior eye complications were observed, including cataract in the anterior eye (incidence rate, 0.18/eye-year) and epiretinal membrane in the posterior eye (incidence rate, 0.09/eye-year); incidence rates of reduction in best-corrected visual acuity to ≤20/50 and ≤20/200 were 0.10 and 0.06/eye-year, respectively. Rates of complications and visual acuity loss did not differ significantly between HIV- positive and negative individuals. In an era of re-emergence, syphilis has ocular complications that may compromise vision, despite treatment with appropriate anti-microbial drugs.

Uso tópico do inteferon alfa-2b para neoplasia escamosa da superficie ocular em um caso de paciente com xeroderma pigmentoso / Topical Interferon alfa-2b for ocular surface squamous neoplasia in one case of patient with xeroderma pigmentosum

Resumo Relato de um caso clínico de Xeroderma Pigmentoso com carcinoma espinocelular de conjuntiva bilateral que apresentou regressão importante das dimensões tumorais com o uso de Interferon alfa-2b tópico. Relato de caso: Paciente feminina com Xeroderma Pigmentoso em estágio avançado, com ausência de pele sadia, tendo sido submetida a cerca de 60 exéreses de lesões de pele malignas. A paciente compareceu com tumoração conjuntival em ambos os olhos, correspondendo a carcinoma espinocelular de conjuntiva e neoplasia intraepitelial de conjuntiva em olho esquerdo. Devido as dificuldades cirúrgicas, alta taxa de recidiva e elevada probabilidade de formação de simbléfaro foi-se iniciado terapêutica com Interferon alfa-2beta 1.000.000 unidades tópico, obtendo-se bons resultados com importante regressão do tamanho da lesão e resolução dos sintomas. Conclusão: O uso tópico de interferon alfa-2beta em neoplasia escamosa de conjuntiva, mostrou-se uma boa opção terapêutica em situações de elevado risco cirúrgico e de complicações pós operatórias.

Abstract Report of a case of xeroderma pigmentosum with squamous cell carcinoma of bilateral conjunctiva that showed a significant regression in tumor size with the use of interferon alfa-2b topic. Case report: Female patient with Xeroderma pigmentosum in an advanced stage, with no healthy skin, having been subjected to about 60 excisions of malignant skin lesions. The patient appeared with conjunctival tumors in both eyes, corresponding to squamous cell carcinoma of the conjunctiva. Due to surgical difficulties, high relapse rate and high probability of symblepharon formation, therapy was started with interferon alpha 2beta 1,000,000 topic units, obtaining good results with a significant decrease in lesion size and resolution of symptoms. Conclusion: Topical use of alpha-interferon in 2beta squamous neoplasia of the conjunctiva proved to be a good therapeutic option for high surgical risk and situations of postoperative complications.

Heme modulates Trypanosoma cruzi bioenergetics inducing mitochondrial ROS production.

Trypanosoma cruzi is the causative agent of Chagas disease and has a single mitochondrion, an organelle responsible for ATP production and the main site for the formation of reactive oxygen species (ROS). T. cruzi is an obligate intracellular parasite with a complex life cycle that alternates between vertebrate and invertebrate hosts, therefore the development of survival strategies and morphogenetic adaptations to deal with the various environments is mandatory. Over the years our group has been studying the vector-parasite interactions using heme as a physiological oxidant molecule that triggered epimastigote proliferation however, the source of ROS induced by heme remained unknown. In the present study we demonstrate the involvement of heme in the parasite mitochondrial metabolism, decreasing oxygen consumption leading to increased mitochondrial ROS and membrane potential. First, we incubated epimastigotes with carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone (FCCP), an uncoupler of oxidative phosphorylation, which led to decreased ROS formation and parasite proliferation, even in the presence of heme, correlating mitochondrial ROS and T. cruzi survival. This hypothesis was confirmed after the mitochondria-targeted antioxidant ((2-(2,2,6,6 Tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl) triphenylphosphonium chloride (MitoTEMPO) decreased both heme-induced ROS and epimastigote proliferation. Furthermore, heme increased the percentage of tetramethylrhodamine methyl ester (TMRM) positive parasites tremendously-indicating the hyperpolarization and increase of potential of the mitochondrial membrane (ΔΨm). Assessing the mitochondrial functional metabolism, we observed that in comparison to untreated parasites, heme-treated epimastigotes decreased their oxygen consumption, and increased the complex II-III activity. These changes allowed the electron flow into the electron transport system, even though the complex IV (cytochrome c oxidase) activity decreased significantly, showing that heme-induced mitochondrial ROS appears to be a consequence of the enhanced mitochondrial physiological modulation. Finally, the parasites that were submitted to high concentrations of heme presented no alterations in the ultrastructure. Consequently, our results suggest that heme released by the insect vector after the blood meal, modify epimastigote mitochondrial physiology to increase ROS as a metabolic mechanism to maintain epimastigote survival and proliferation.

Mutagenic and Cytotoxicity LQB 123 Profile: Safety and Tripanocidal Effect of a Phenyl-t-Butylnitrone Derivative.

The therapeutic options for Chagas disease are limited and its treatment presents a number of drawbacks including toxicity, drug resistance, and insufficient effectiveness against the chronic stage of the disease. Therefore, new therapeutical options are mandatory. In the present work, we evaluated the effect of a phenyl-tert-butylnitrone (PBN) derivate, LQB 123, against Trypanosoma cruzi forms. LQB 123 presented a trypanocidal effect against bloodstream trypomastigotes (IC50 = 259.4 ± 6.1 µM) and intracellular amastigotes infecting peritoneal macrophages (IC50 = 188.2 ± 47.5 µM), with no harmful effects upon the mammalian cells (CC50 values greater than 4 mM), resulting in a high selectivity index (CC50/IC50 > 20). Additionally, metacyclic trypomastigotes submitted to LQB 123 presented an IC50 of about 191.8 ± 10.5 µM and epimastigotes forms incubated with different concentrations of LQB 123 presented an inhibition of parasite growth with an IC50 of 255.1 ± 3.6 µM. Finally, we investigated the mutagenic potential of the nitrone by the Salmonella/microsome assay and observed no induction of mutagenicity even in concentrations as high as 33000 µM. Taken together, these results present a nonmutagenic compound, with trypanocidal activity against all relevant forms of T. cruzi, offering new insights into CD treatment suggesting additional in vivo tests.

Purification and physico-chemical study of serum albumins of two neotropical fish species from the São Francisco River Bassin, Brazil / Purificação e estudo físico-químico de albuminas de duas espécies de peixes neotropicais da Bacia do Rio São Francisco, Brasil

This paper presents results from a study of albumin from pacu (Piaractus mesopotamicus, Holmberg 1887) and the catfish pintado (Pseudoplatystoma corruscans, Spix & Agassiz, 1829), two neotropical fish species inhabitants of Brazilian rivers, comparing their molecular mass and discussing their secondary structures based on spectropolarimetric (circular dychroism) measurements. Genetic controlled specimens were obtained from two fish hatcheries, located in Mococa (pacu) and in São João da Boa Vista (pintado), both in São Paulo State, Brazil. After a period of adaptation in holding tanks, fish blood samples were taken by punctioning their abdominal aorta. Purified albumin was obtained by gel filtration. SDS-PAGE electrophoresis was performed for the molecular mass estimation. Circular Dichroism spectra were registered for albumins of the two fish species over the range of 190-250 nm (far-UV), which shown two negative bands at 217 and 208 nm, a positive peak at 196 nm and a crossover at 200 nm. This profile is compatible with proteins that content predominantly alpha-helix structure.

Este artigo apresenta os resultados de um estudo sobre as albuminas de pacu (Piaractus mesopotamicus, Holmberg 1887) e pintado (Pseudoplatystoma corruscans, Spix & Agassiz, 1829), duas espécies neotropicais de peixes nativas do Brasil, determinando as suas massas moleculares e discutindo suas estruturas secundárias, com base em medidas de espectropolarimetria (dicroísmo circular). Espécimes controlados geneticamente foram obtidos de duas diferentes pisciculturas, uma localizada na cidade de Mococa (pacu) e a outra, na cidade de São João da Boa Vista (pintado), ambas no Estado de São Paulo, Brasil. Após um período de adaptação em tanques apropriados, amostras de sangue foram coletadas por punção da aorta abdominal dos peixes. Albumina pura foi obtida por gel filtração dessas amostras e as massas moleculares foram determinadas a partir dos dados da eletroforese SDS-PAGE. Espectros de dicroísmo circular das albuminas dos peixes foram registrados na região de 190-250 nm (far-UV), os quais mostraram duas bandas negativas, a 217 e 208 nm, um pico positivo a 196 nm e um crossover a 200 nm; perfil este compatível com proteínas que contem predominantemente estrutura alfa-hélice.

Punctate Inner Choroidopathy.

Purpose. To report a case of bilateral punctate inner choroidopathy (PIC). Case Report. A 26-year-old Caucasian woman presented with bilateral blurred vision with one year of evolution. There was no relevant systemic disease or family history. Best-corrected visual acuity in the right eye was 20/30 and in the left eye was 20/20; there was no clinically significant refractive error. Fundoscopy evidenced multiple, small, round, yellow-white lesions limited to the posterior pole of both eyes, with greater macular involvement in the RE. There were no signs of inflammation in the anterior chamber or vitreous cavity. Fluorescein angiography revealed the presence of multiple hyperfluorescent lesions more evident in the later stages of the angiogram in both eyes. On indocyanine green angiography, these lesions appeared hypofluorescent in both early and late phases. Optical coherence tomography showed the presence of focal elevations of the retinal pigment epithelium with underlying hyporeflective space, bilaterally. Laboratory and imaging evaluation for evidence of autoimmune and infectious diseases were negative. Conclusion. The PIC is a relatively uncommon condition. In this report, an attempt has been made to describe a classic clinic presentation of this disease in a young and female patient.

Proliferation and differentiation of Trypanosoma cruzi inside its vector have a new trigger: redox status.

Trypanosoma cruzi proliferate and differentiate inside different compartments of triatomines gut that is the first environment encountered by T. cruzi. Due to its complex life cycle, the parasite is constantly exposed to reactive oxygen species (ROS). We tested the influence of the pro-oxidant molecules H2O2 and the superoxide generator, Paraquat, as well as, metabolism products of the vector, with distinct redox status, in the proliferation and metacyclogenesis. These molecules are heme, hemozoin and urate. We also tested the antioxidants NAC and GSH. Heme induced the proliferation of epimastigotes and impaired the metacyclogenesis. ß-hematin, did not affect epimastigote proliferation but decreased parasite differentiation. Conversely, we show that urate, GSH and NAC dramatically impaired epimastigote proliferation and during metacyclogenesis, NAC and urate induced a significant increment of trypomastigotes and decreased the percentage of epimastigotes. We also quantified the parasite loads in the anterior and posterior midguts and in the rectum of the vector by qPCR. The treatment with the antioxidants increased the parasite loads in all midgut sections analyzed. In vivo, the group of vectors fed with reduced molecules showed an increment of trypomastigotes and decreased epimastigotes when analyzed by differential counting. Heme stimulated proliferation by increasing the cell number in the S and G2/M phases, whereas NAC arrested epimastigotes in G1 phase. NAC greatly increased the percentage of trypomastigotes. Taken together, these data show a shift in the triatomine gut microenvironment caused by the redox status may also influence T. cruzi biology inside the vector. In this scenario, oxidants act to turn on epimastigote proliferation while antioxidants seem to switch the cycle towards metacyclogenesis. This is a new insight that defines a key role for redox metabolism in governing the parasitic life cycle.

The effect of thiamine deficiency on inflammation, oxidative stress and cellular migration in an experimental model of sepsis.

BACKGROUND: Sepsis is a prevalent condition in critically ill patients and may be associated with thiamine deficiency (TD). The aim of this study was to evaluate the effect of TD on inflammation, oxidative stress and cellular recruitment in a sepsis model. METHODS: The experimental sepsis model, cecal ligation and puncture (CLP), was utilized on mice in comparison with a sham procedure. The following four groups were compared against each other: SHAM with AIN93G complete chow, SHAM with thiamine deficient (TD) chow, CLP with AIN93G complete chow, and CLP with TD chow. Thiamine pyrophosphate (TPP) blood concentrations were determined, and blood and peritoneal fluid were evaluated for differences in TNF-alpha, IL-1, IL-6, KC and MCP-1/CCL2 levels. In addition, the levels of 4-HNE adducts in liver proteins were evaluated by Western Blot. RESULTS: The mean TPP blood concentration from the mice fed with the complete chow was 303.3 ± 42.6 nmol/L, and TD occurred within 10 days. TNF-α and MCP-1 concentrations in the peritoneal fluid were significantly greater in the CLP with TD chow group when compared with the other groups. The blood IL-1ß level, however, was lower in the CLP with TD chow group. Liver 4-HNE levels were highest in the TD chow groups. Blood mononuclear cell numbers, as well as peritoneal total leukocyte, mononuclear cell and neutrophil numbers were greater in the CLP with TD chow group. Peritoneal bacterial colony forming units (CFU) were significantly lower in the CLP with TD chow group. CONCLUSION: TD was associated with greater bacterial clearance, oxidative stress and inflammatory response changes.