Search

Objective: Almost two thirds of the world's aneurysmal subarachnoid hemorrhage (aSAH) are in low- and middle-income countries. Herein, we aimed to evaluate the impact of complications on the outcome of aSAH in a middle-income country., Methods: Baseline data (age, sex, World Federation of Neurosurgical Society, time ictus-treatment, treatment modality) and medical and neurologic complications from a cohort in Brazil (2016-2019) were evaluated: delayed cerebral ischemia; hydrocephalus; meningitis; seizures; intracranial hypertension; infections (pneumonia, bloodstream, urinary tract infection infection of undetermined source); sodium disturbances; acute kidney injury; and cardiac and pulmonary complications. The primary outcome was the modified Rankin scale (mRS) at hospital discharge. Univariate and multivariate models were employed., Results: From 212 patients (71.7% female, age 52.7 ± 12.8), 92% developed at least 1 complication (any infection-43.9%, hydrocephalus-34.4%, intracranial hypertension-33%, infection of undetermined source-20.8%, hypernatremia-20.8%, hyponatremia-19.8%, delayed cerebral ischemia-related infarction-18.7%, pneumonia-18.4%, acute kidney injury-16.5%, and seizures-11.8%). In unadjusted analysis, all but hyponatremia and urinary tract infection were associated with mRS 3-6 at discharge; however, complications explained only 12% of the variation in functional outcome (mRS). Most patients were treated by clipping (66.5%), and 15.6% (33 patients) did not receive a definitive treatment. The median time ictus-admission and ictus-treatment were 5 and 9 days, respectively., Conclusions: While medical and neurologic complications are a recognized opportunity to improve aSAH care, low- and middle-income countries comprise 70% of the world population and still encounter difficulties concerning early definitive aneurysm treatment, rebleeding, and human and material resources., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Objectives: Since oral conditions negatively affect oral health-related quality of life (OHRQoL), this study evaluated which oral clinical condition, signs, and symptoms are associated with the impact on OHRQoL, its domains, and specific daily life activities among adults., Materials and Methods: Data from a probabilistic sample of adults (35-44 years old) was used. The Oral Impacts on Daily Performance (OIDP) was used to evaluate OHRQoL, its domains (physical, psychological, and social), and nine daily life activities. Signs and symptoms of oral diseases (dental caries, periodontal disease, need for dental prothesis, and tooth loss) and oral self-perception were considered. Descriptive, bivariate, and multi-level analyses were conducted., Results: A total of 5,834 adults were included, of which 52.9% had some negative impact of oral conditions on OHRQoL. Difficulty in eating was the most affected daily life activity. For multiple models, dental caries lesions (cavities), filled teeth with caries, gingival bleeding, periodontal pocket, dental pain, need for upper or lower dental prosthesis, and oral health self-perception were associated (p < 0.05) with overall OHRQoL or at least one of its domains. The impact on daily life activities of each individual was associated with at least one oral condition. Dental caries lesions (cavity) and dental pain were associated (p < 0.05) with the impact on most daily life activities evaluated., Conclusions: Different signs and symptoms of prevalent oral diseases are associated with the impact on specific daily life activities among adults, which may compromise the OHRQoL., Clinical Relevance: Knowledge of how signs, symptoms, and oral conditions affect OHRQoL and daily life activities can provide essential information for clinicians to establish proper disease management and preventive strategies focusing on improving patients' lives., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The use of saliva as a protein source prior to microbiological and biological assays requires previous processing. However, the effect of these processing methods on the proteomic profile of saliva has not been tested. Stimulated human saliva was collected from eight healthy volunteers. Non-processed saliva was compared with 0.22 μm filtered, 0.45 μm filtered, and pasteurized saliva, by liquid chromatography-mass spectrometry. Data are available via ProteomeXchange with identifier PXD039248. The effect of processed saliva on microbial adhesion was tested using bacterial and fungus species and in biological cell behavior using HaCaT immortalized human keratinocytes. Two hundred and seventy-eight proteins were identified in non-processed saliva, of which 54 proteins (≈19%) were exclusive. Saliva processing reduced identified proteins to 222 (≈80%) for the 0.22 μm group, 219 (≈79%) for the 0.45 μm group, and 201 (≈72%) for the pasteurized saliva, compared to non-processed saliva. The proteomic profile showed similar molecular functions and biological processes. The different saliva processing methods did not alter microbial adhesion (ANOVA, p > 0.05). Interestingly, pasteurized saliva reduced keratinocyte cell viability. Saliva processing methods tested reduced the proteomic profile diversity of saliva but maintained similar molecular functions and biological processes, not interfering with microbial adhesion and cell viability, except for pasteurization, which reduced cell viability.

Objective: The present study aimed to establish an optimized protocol for biofilm removal from titanium (Ti) surfaces using citric acid (CA) solutions., Background: Biofilm accumulation is the main factor to trigger peri-implant infections and to increase the risk of treatment failures. Although CA has been suggested as the anti-infective agent with highest potential for biofilm removal on Ti, there is no consensus that CA could improve the anti-infective treatment and its effect., Methods: Physical and chemical alterations, electrochemical behavior, cytotoxicity, and antimicrobial effect of CA on Ti discs were evaluated using four concentrations (1, 10, 20, and 40%) and two application methods (immersion and rubbing). Negative control using 0.9% NaCl was used in all experiments. To evaluate whether different application times can have similar response, polymicrobial biofilm (microcosm model) was formed on Ti and treated with CA for 1, 2, 4, and 8 min. An in situ study was conducted to verify whether the established protocol is equally effective in biofilms formed on machined and sandblasted, large-grit, and acid-etched (SLA) Ti surfaces., Results: CA 40% induced significantly higher surface alterations observed by confocal images and profilometry. In general, rubbing protocol decreased the surface roughness and increased the wettability (p < 0.05), exhibiting better surface cleaning by biofilm removal. CA 10% presented no indirect cytotoxicity and, when applied by rubbing for 8 min, presented proper in vitro antibacterial action and potential corrosion inhibition. When CA 10% was rubbed on Ti surfaces for 4 min, it displayed optimum cleaning ability as 8 min, working equally to remove in situ biofilm on machined and SLA surfaces., Conclusions: The application of CA 10% by rubbing for at least 4 min demonstrated to be a promising protocol to eliminate biofilms formed in smooth and rougher surfaces, which could improve implant-related infection therapies., (© 2021 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Objective: To evaluate the effect of titanium (Ti) particles and ions on oral biofilm growth and composition., Background: Particles and ions of Ti released from dental implants can trigger unfavorable biological responses in human cells. However, their effect on oral biofilms composition has not been tested., Methods: In this blind in situ study, volunteers wore a palatal appliance containing Ti disks for 7 days to allow biofilm formation. Disks were then collected and biofilms were treated, in vitro, with Ti particles (0.75% and 1%), ions (10 and 20 ppm), or a combination of both (1% particles + 20 ppm ions). Biofilms exposed only to medium was used as control group. After 24 hours, biofilms were collected and analyzed by checkerboard DNA-DNA hybridization. Direct effects of Ti particles and ions on biofilm/cellular morphology were evaluated by transmission electron microscopy (TEM)., Results: Ti particles affected biofilm composition, increasing population of four bacterial species (P < .05), while Ti ions showed higher levels of putative pathogens from the orange complex with reduction in species from the yellow complex (P < .05), compared with control. The combination of particles + ions increased green complex and reduced yellow complex proportions (P < .05). TEM showed clusters of particles agglomerated in extracellular environment, while Ti ions were precipitated in both extracellular and intracellular sites., Conclusions: Ti products, especially Ti ions, have the potential to change the microbiological composition of biofilms formed on Ti surfaces. Therefore, the presence of Ti products around dental implants may contribute to microbial dysbiosis and peri-implantitis., (© 2019 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Titanium-zirconium (Ti-Zr) alloy has been widely used as a biomaterial for implant devices, and it is commonly treated by sandblasting followed by acid etching (SLA) to improve biological responses. Although protein adsorption is the first biological response, the effect of this SLA treatment on the proteomic profile of proteins adsorbed from saliva and blood plasma has not been tested. In this study, the proteomic profile was evaluated by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Streptococcus sanguinis was used to test whether the protein layer affects bacterial adhesion. SLA treatment affected the proteomic profile, showing exclusive proteins adsorbed from saliva (14) and plasma (3). However, both groups exhibited close patterns of intensity for common proteins, molecular functions and biological processes mediated by proteins. Interestingly, Ti-Zr SLA showed higher bacterial adhesion (∼1.9 fold over) for the surface coated with plasma proteins. Therefore, SLA treatment of Ti-Zr alloy changed the proteomic profile, which may affect bacterial adhesion.

Trypanosoma cruzi is a pathogenic protozoan that still has an impact on public health, despite the decrease in the number of infection cases along the years. T. cruzi possesses an heteroxenic life cycle in which it differentiates in at least four forms. Among the differentiation processes, metacyclogenesis has been exploited in different views by researchers. An intriguing question that rises is how metacyclogenesis is triggered and controlled by cell signaling and which are the differentially expressed proteins and posttranslational modifications involved in this process. An important cell signaling pathway is the protein phosphorylation, and it is reinforced in T. cruzi in which the gene expression control occurs almost exclusively posttranscriptionally. Additionally, the number of protein kinases in T. cruzi is relatively high compared to other organisms. A way to approach these questions is evaluating the cells through phosphoproteomics and proteomics. In this chapter, we will describe the steps from the cell protein extraction, digestion and fractionation, phosphopeptide enrichment, to LC-MS/MS analysis as well as a brief overview on peptide identification. In addition, a published method for in vitro metacyclogenesis will be detailed.

Objective: The main goal of this study was to examine the influence of hydroxyapatite (HAp) macroaggreate concentrations on thermal and mechanical properties of radioactive bone cement and to study the relation of glass transition Tg with its mechanical properties., Methods: The bone cement as (1-x)PMMA-xHAp binary system was prepared in six [x] distinct concentration parameters of 0.0 up to 0.5. The HAp was synthesized using a solgel procedure following calcination by thermal treatment. The composite was prepared in cold based (non-radioactive) mixing polymethyl methacrylate (PMMA) and HAp. Differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and mechanical compressive strength (CS) were used to measure the thermal and mechanical properties., Results: The DSC and TGA thermal profiles in function to concentration parameter [x] were presented. The CS lies in a range of 3.71-7.37 MPa and the glass transition temperature Tg = 126.27 °C. There was a direct relationship between the PMMA-HAp thermoplastic properties with mechanical and thermal properties in function of HAp concentrations., Conclusion: The specific PMMA-HAp composite, with a concentration ratio of 1:1 and HAp thermal treatment at the Tg, provides a material with a compression strength of 7.37 MPa and a suitable amount of porous similar to a trabecular bone, possible to apply in bone cement implants, regardless of whether they are radioactive or not.

Phosphorylation is an important event in cell signaling that is modulated by kinases and phosphatases. In Trypanosoma cruzi, the etiological agent of Chagas disease, approximately 2% of the protein-coding genes encode for protein kinases. This parasite has a heteroxenic life cycle with four different development stages. In the midgut of invertebrate vector, epimastigotes differentiate into metacyclic trypomastigotes in a process known as metacyclogenesis. This process can be reproduced in vitro by submitting parasites to nutritional stress (NS). Aiming to contribute to the elucidation of mechanisms that trigger metacyclogenesis, we applied super-SILAC (super-stable isotope labeling by amino acids in cell culture) and LC-MS/MS to analyze different points during NS. This analysis resulted in the identification of 4205 protein groups and 3643 phosphopeptides with the location of 4846 phosphorylation sites. Several phosphosites were considered modulated along NS and are present in proteins associated with various functions, such as fatty acid synthesis and the regulation of protein expression, reinforcing the importance of phosphorylation and signaling events to the parasite. These modulated sites may be triggers of metacyclogenesis.

Background: Citric acid (CA) has been suggested as an effective antimicrobial agent against biofilms formed on titanium. However, the antimicrobial effect of CA in biofilms formed in the oral environment and its effects on the physical properties of titanium surface remain unknown. Therefore, this study evaluated the antimicrobial effect of CA on in situ biofilm, whether this treatment favors the bacteria recolonization and its effect on the electrochemical properties of titanium., Methods: In the in situ test, volunteers wore a palatal appliance containing titanium discs. After 7 days, the discs (N = 21) were exposed in vitro to the following treatments: immersion in 0.9% sodium chloride (control); 40% CA immersion; and 40% CA rubbing. Afterwards, these discs (N = 18) were exposed in vitro to new bacterial adhesion with Streptococcus sanguinis. New discs (N = 18) without biofilm were exposed to the treatments and subjected to electrochemical tests and surface characterization. Data were analyzed by one-way ANOVA followed by Tukey's HSD test., Results: The CA groups showed a significant reduction (≈ 5-log reduction) in the biofilm formed in situ compared with the control group (p < 0.05), but no difference was found between CA application methods (p = 0.680). The acid treatment did not favor the recolonization of bacteria (p = 0.629). CA treatment did not influence the polarization resistance and capacitance of the oxide film, but statistically enhanced the electrochemical stability of titanium., Conclusion: Citric acid appears to be an effective clinical alternative for treatment of the main etiologic factor in dental implant failure, biofilm formation, enhancing electrochemical behavior of titanium., (© 2018 American Academy of Periodontology.)

The scope of this study was to evaluate dissatisfaction with dental services among dentate and edentulous elderly Brazilians and its association with contextual and individual variables. The data from the National Oral Health Survey conducted in 2010 and contextual variables related to cities were used. Descriptive, bivariate and multilevel analysis was conducted among 1,989 elderly individuals. Of these, 11.2% of dentate and 22.1% of edentulous elderly people were dissatisfied. Among the dentate individuals, dissatisfaction was associated with contextual variables (location of municipality, human development index, GINI Index, the presence of Dental Specialty Centers and the coverage of oral health in public service) and persons related to health care and oral health conditions. For the edentulous individuals, no associated contextual variables were identified. Only the motive for the use of the service and satisfaction with teeth and mouth were associated with dissatisfaction with dental services. A considerable prevalence of dissatisfaction with the services was identified, being higher among edentulous elderly Brazilians. Among dentate elderly Brazilians, an association with contextual variables was detected.

Knowledge about fluoride delivery to oral fluids from foods cooked with fluoridated water and salt is scarce, and no study has evaluated fluoride concentrations in saliva or biofilm during meal consumption. In this randomized double-blind crossover study, 12 volunteers ingested meals (rice, beans, meat, and legumes) prepared with nonfluoridated water and salt (control group), fluoridated water (0.70 mg F/L; water group), and fluoridated salt (183.7 mg F/kg; salt group). Whole saliva was collected before meal ingestion, during mastication, and up to 2 h after meal ingestion. Dental biofilm was collected before and immediately after meal ingestion. Fluoride concentrations in saliva and dental biofilm were determined by an ion-specific electrode. The mean (±standard deviation; n = 4) fluoride concentrations in meals prepared for the control, water, and salt groups were 0.039 ± 0.01, 0.43 ± 0.04, and 1.71 ± 0.32 μg F/g, respectively. The three groups had significantly different fluoride concentrations in saliva collected during mastication (p < 0.0001) and after meal ingestion (p < 0.04; salt > water > control). The fluoride concentration in saliva returned to baseline 30 min after meal ingestion in the water group but remained high for up to 2 h in the salt group (p = 0.002). The fluoride concentration in biofilm fluid differed only between the salt and control groups (p = 0.008). The mastication of foods cooked with fluoridated water and salt increases fluoride concentrations in oral fluids and may contribute to the local effect of these community-based fluoride interventions on caries control., (© 2018 S. Karger AG, Basel.)

This study aimed to test the dose-response effect of chlorhexidine on multispecies biofilms formed on commercially pure titanium (cpTi) and titanium-zirconium (TiZr) alloy. Biofilms were formed on cpTi and TiZr discs and treated two times per day with five different chlorhexidine concentrations (0.12, 0.20, 0.50, 1, 2%). The biofilms were collected for microbiological, biochemical and microscopic analyses. The significance of differences among groups was evaluated by linear regression, ANOVA, Bonferroni and Tukey tests. The mean number of colony-forming units decreased as the chlorhexidine concentration increased for both cpTi and TiZr (p < 0.05). The maximum effect was observed with the 0.5% concentration. Confocal microscopy images suggested an increase in the number of dead bacterial cells with increased chlorhexidine concentration. The biofilm pH increased after chlorhexidine exposure (p < 0.05). Chlorhexidine showed an antimicrobial dose-response effect in controlling biofilm on cpTi and TiZr. 0.5% chlorhexidine can be used to achieve the maximum antimicrobial effect on both materials.

Data about total fluoride intake in children living in a tropical semi-arid climate city is scarce, thus we conducted this study. Fifty-eight children aged two to five years, living in a Brazilian tropical city with optimally fluoridated water were selected. Dietary samples were collected using the duplicate diet method on two non-consecutive days in the children's home toothpaste was determined by subtracting the amount of fluoride recovered after brushing from the amount placed on the toothbrush. The mean total dose (SD) of fluoride intake was 0.043(0.016) mg F·kg-1·d-1, with the major (60.6%) contribution from water. The factors associated with the ingestion of fluoride from toothpaste were fluoride concentration of the toothpaste (p = 0.03) and the use of kids toothpaste (p = 0.02). The findings suggest that children have a low fluoride intake, measured by at-home meals and use of fluoride toothpaste; drinking water is the main source of fluoride ingestion.

Protein phosphorylation and dephosphorylation events regulate many cellular processes. The identification of all phosphorylation sites and their association to a respective protein kinase or phosphatase is a challenging and crucial step to have a deeper understanding of the effects of signaling networks on cells. Pathogenic trypanosomatids have a large number of protein kinases and phosphatases in comparison to other organisms, which reinforces the relevance of the phosphorylation process in these early eukaryotes, nevertheless little is known about protein phosphorylation in these protozoa. In this context, the role of a MAP kinase-like kinase (MAPKLK1), observed to be essential to proliferation of procyclic Trypanosoma brucei, was studied. After silencing MAPKLK1 expression by RNAi, the cells were evaluated by SILAC MS-based proteomics and RNA-Seq. We identified 1756 phosphorylation sites of which 384 were not previously described in T. brucei. Despite being essential, few modulations were observed at the phosphorylation patterns and gene expression levels of MAPKLK1 knockdown. These indirect targets and potential substrates of MAPKLK1 are related to key cellular processes enriched to mRNA processing and stability control., Significance: The field of cell signaling is a promising topic of study for trypanosomatids, since little is known about this topic and the gene expression regulation occurs at post-transcriptional level. In this sense, the present work increases the knowledge on protein phosphorylation process in Trypanosoma brucei. We depleted one MAP kinase (MAPKLK1) of T. brucei and evaluated the effects on the cell. We showed that MAPKLK1 is essential to the cell, while few modulations on phosphoproteome, proteome and transcriptome are observed with its depletion. Although in low number, the changes in phosphoproteome were significant, presenting possible substrate candidates of MAPKLK1 and indirect targets related to mRNA processing and stability control, metabolic pathways, among others. This result provides insights in the phosphorylation network of T. brucei, a model organism that impacts human and animal health., (Copyright © 2016. Published by Elsevier B.V.)

Background: Homeless men are highly vulnerable to acquisition of the hepatitis C virus (HCV) compared to the general population. In Brazil, a country of continental dimensions, the extent of HCV infection in this population remains unknown. The objective of this study is to investigate the epidemiological profile of exposure to HCV in homeless men in Central Brazil., Methods: A Cross-sectional study was conducted in 481 men aged over 18 years attending therapeutic communities specialized in the recovery and reintegration of homeless people. Participants were tested for anti-HCV markers using rapid tests. Poisson regression analysis was used to verify the risk factors associated with exposure to HCV., Results: The prevalence of HCV exposure was 2.5% (95.0% CI: 1.4 to 4.3%) and was associated with age, absence of family life, injection drug use, number of sexual partners, and history of sexually transmitted infections (STI). Participants reported multiple risk behaviors, such as alcohol (78.9%), cocaine (37.1%) and/or crack use (53.1%), and inconsistent condom use (82.6%). Injection drug use was reported by 8.7% of participants., Conclusions: The prevalence of HCV infection among homeless men was relatively high. Several risk behaviors were commonly reported, which shows the high vulnerability of this population. These findings emphasize the need for the development of specific strategies to reduce the risk of HCV among homeless men.

Our objective was to evaluate whether there is an association with the different levels of dental fluorosis and the presence of dental trauma amongst school children. A transversal study was conducted amongst school children from the age of 12. Dental examinations were conducted by 24 well trained and fully qualified dental surgeons. Data was collected from 36 randomly selected public schools amongst 89 schools in a municipality. The criteria used to diagnose dental fluorosis was based on the Dean's fluorosis Index and for diagnosing dental trauma we looked for clinical signs of crown fractures and dental avulsions. Multiple descriptive analysis, which was bivariate, was carried out. Amongst the 2,755 school children that took part in the study 1,089 (39.6%) were diagnosed with dental fluorosis and 106 (3.8%) had one tooth or more with dental trauma. We noted a high prevalence of dental fluorosis, independent of the level of severity, amongst individuals with one tooth or more who had dental trauma. This association was even more evident where there were severely high levels of fluorosis. We also noted that the presence of fluorosis was greater amongst those that actively paid more attention to discoloration on their teeth and who received treatment from a dental professional at their schools. Nevertheless dental fluorosis was associated with the presence of dental trauma, independent of its severity.

Purpose: The purpose of this study was to evaluate the prevalence of dental caries, toothbrushing habits, fluoride intake, and factors associated with the use of toothpaste in Brazilian children according to socioeconomic status (SES)., Methods: A cross-sectional observational study was conducted with 148 three- to four-year-olds of both low and high socioeconomic status. Data collection consisted of administering a questionnaire to guardians, brushing to determine fluoride intake, and examining the children's teeth to evaluate dental caries., Results: In this study, 42 percent of children from high SES and 2.7 percent of children from low SES used toothpaste without fluoride or with a concentration below 1,000 ppm F. Fluoride intake was associated with SES, frequency of brushing, the amount of toothpaste used, and the concentration of fluoride present in toothpastes (P<0.05). Low SES children had a higher fluoride intake during brushing (0.045 mg F/kg body weight/day), compared to high SES children (0.023 mg F/kg body weight/day). Unlike low SES children (68.9 percent), all high SES children were caries free., Conclusions: A lower socioeconomic status was associated with a higher prevalence of dental caries and a higher intake of fluoride from toothpaste.

Objective: In the present study, the authors investigated the in vitro behavior of radio-resistant breast adenocarcinoma (MDA-MB-231) cells line and radiosensitive peripheral blood mononuclear cells (PBMC), as a function of different radiation doses, dose rates and postirradiation time kinetics, with a view to the interest of clinical radiotherapy., Materials and Methods: The cells were irradiated with Co-60, at 2 and 10 Gy and two different exposure rates, 339.56 cGy.min(-1) and the other corresponding to one fourth of the standard dose rates, present over a 10-year period of cobalt therapy. Post-irradiation sampling was performed at pre-established kinetics of 24, 48 and 72 hours. The optical density response in viability assay was evaluated and a morphological analysis was performed., Results: Radiosensitive PBMC showed decrease in viability at 2 Gy, and a more significant decrease at 10 Gy for both dose rates. MDAMB- 231 cells presented viability decrease only at higher dose and dose rate. The results showed MDA-MB-231 clone expansion at low dose rate after 48-72 hours post-radiation., Conclusion: Low dose rate shows a possible potential clinical impact involving decrease in management of radio-resistant and radiosensitive tumor cell lines in cobalt therapy for breast cancer.

The main sources of fluoride intake by children are fluoridated water and toothpaste. Little has been studied regarding fluoride intake from these sources in regions with tropical climates and high temperatures throughout the year. This study aimed to determine the amount of fluoride ingested from diet and tooth brushing by children who live in a city with a tropical climate. Sixty-seven children from Teresina, Piauí, Brazil, took part in this study. The city's water supply was optimally fluoridated. The duplicate-diet method was used to determine the fluoride intake from diet. The intake of fluoride from dentifrice was determined by subtracting the amount of fluoride placed on the toothbrush and that recovered after brushing. The concentration of fluoride was measured using an ion-specific electrode and is expressed as milligrams/kilogram of body weight/day. The mean (±SD) total amount was 0.071 ± 0.036 mg F/kg body weight/day, and the relative contributions of diet and toothpaste were 0.025 ± 0.010 and 0.046 ± 0.035, respectively. The factors associated with fluoride intake from toothpaste were: use of children's toothpaste (p = 0.003), use of large amounts of toothpaste (p < 0.001), and a high frequency of tooth brushing (p = 0.003). Sixty-four percent of children had an intake of less than 0.07 mg F/kg body weight/day, which is considered the upper limit for an aesthetically tolerable fluorosis risk. The results suggest that the amount of fluoride ingested by most children who live in a Brazilian city with a tropical climate is considered safe in terms of the risk of dental fluorosis., (© 2015 S. Karger AG, Basel.)

Background: Trypanosoma cruzi, the etiologic agent of Chagas disease, alternates between distinct morphological and functional forms during its life cycle. Axenic multiplication and differentiation processes of this protozoan parasite can be reproduced in vitro, enabling the isolation and study of the different evolutionary forms. Although there are several publications attempting the cultivation of T. cruzi under chemically defined conditions, in our experience none of the published media are capable of maintaining T. cruzi in continuous growth., Results: In this work we modified a known chemically defined medium for Trypanosoma brucei growth. The resulting LM14 and LM14B defined media enabled cultivation of five different strains of T. cruzi for more than forty passages until now. The parasite's biological characteristics such as morphology and differentiation to metacyclic trypomastigotes were maintained when defined media is used., Conclusions: The establishment of a defined medium for T. cruzi cultivation is an important tool for basic biological research allowing several different approaches, providing new perspectives for further studies related to cell biology of this parasite.

Heteroxenic and monoxenic trypanosomatids were screened for the presence of actin using a mouse polyclonal antibody produced against the entire sequence of the Trypanosoma cruzi actin gene, encoding a 41.9 kDa protein. Western blot analysis showed that this antibody reacted with a polypeptide of approximately 42 kDa in the whole-cell lysates of parasites targeting mammals (T. cruzi, Trypanosoma brucei and Leishmania major), insects (Angomonas deanei, Crithidia fasciculata, Herpetomonas samuelpessoai and Strigomonas culicis) and plants (Phytomonas serpens). A single polypeptide of approximately 42 kDa was detected in the whole-cell lysates of T. cruzi cultured epimastigotes, metacyclic trypomastigotes and amastigotes at similar protein expression levels. Confocal microscopy showed that actin was expressed throughout the cytoplasm of all the tested trypanosomatids. These data demonstrate that actin expression is widespread in trypanosomatids.

Trypanosoma cruzi is a rare example of an eukaryote that has genes for two threonine proteases: HslVU complex and 20S proteasome. HslVU is an ATP-dependent protease consisting of two multimeric components: the HslU ATPase and the HslV peptidase. In this study, we expressed and obtained specific antibodies to HslU and HslV recombinant proteins and demonstrated the interaction between HslU/HslV by coimmunoprecipitation. To evaluate the intracellular distribution of HslV in T. cruzi we used an immunofluorescence assay and ultrastructural localization by transmission electron microscopy. Both techniques demonstrated that HslV was localized in the kinetoplast of epimastigotes. We also analyzed the HslV/20S proteasome co-expression in Y, Berenice 62 (Be-62) and Berenice 78 (Be-78) T. cruzi strains. Our results showed that HslV and 20S proteasome are differently expressed in these strains. To investigate whether a proteasome inhibitor could modulate HslV and proteasome expressions, epimastigotes from T. cruzi were grown in the presence of PSI, a classical proteasome inhibitor. This result showed that while the level of expression of HslV/20S proteasome is not affected in Be-78 strain, in Y and Be-62 strains the presence of PSI induced a significantly increase in Hslv/20S proteasome expression. Together, these results suggest the coexistence of the protease HslVU and 20S proteasome in T. cruzi, reinforcing the hypothesis that non-lysosomal degradation pathways have an important role in T. cruzi biology., (Copyright © 2011 Elsevier Inc. All rights reserved.)

In this study, we evaluated the bradykinin potentiating activity and ACE inhibitory activity of several Ang-(1-7)-related peptides: Ang-(2-7), Ang-(3-7), Ang-(4-7), Ang-(1-6), Ang-(1-5) and the selective antagonist of Ang-(1-7): D-[Ala7]Ang-(1-7) (A-779). In vivo experiments were performed in freely moving Wistar rats. ACE activity was evaluated by a fluorometric assay in rat plasma using Hip-His-Leu as a substrate. Intravenous injections of Ang-(1-7) (2.2 nmol) transformed the effect of a single dose of bradykinin (1 nmol) into the effect produced by a double dose. A similar bradykinin potentiating activity was demonstrated for Ang-(2-7) and Ang-(3-7). On the other hand, Ang-(1-5), Ang-(1-6), Ang-(4-7) and A-779 did not change the hypotensive effect of bradykinin in doses ranging from 8 up to 25 nmols. The hypotensive effect of bradykinin was increased by intravenous infusion (0.3 ng/min) of Ang-(1-7) > Ang-(2-7) > Ang-(3-7). Conversely, Ang-(1-5), Ang-(1-6), Ang-(4-7) or A-779 did not change the hypotensive effect of bradykinin. ACE inhibition with Ang-(1-7) related peptides occurred in the order: Ang-(2-7) > or = Ang-(3-7) > Ang-(1-7) [>>] Ang-(1-5) > Ang-(4-7) > or = Ang-(1-6) > or = A-779. A-779 in concentrations up to 10(-5) M did not change the ACE inhibitory activity of Ang-(1-7). These results suggest that Ang-(1-7), Ang-(2-7) and Ang-(3-7) can modulate bradykinin actions in vivo. More important, our data pointed out that alternative mechanisms besides interaction with ACE are required to explain the bradykinin potentiating activity of Ang-(1-7).

In this study we evaluated the effect of angiotensin-(1-7) on the hypotensive action of bradykinin (BK) in normotensive rats, renal hypertensive rats (RHR), and spontaneously hypertensive rats (SHR). In addition, we evaluated the effect of angiotensin-converting enzyme (ACE) inhibition with enalaprilat treatment (10 mg/kg I.V.) on the BK-potentiating activity of Ang-(1-7). Renal hypertension was produced by aorta coarctation between the origin of renal arteries. Ang-(1-7) (0.3 pmol/min) or saline (0.9% NaCl, 5 microL/min) was infused intravenously in conscious male Wistar rats, adult SHR, or RHR. Intravenous bolus injections of BK (0.1 to 1.6 nmol in RHR and SHR; 0.625 to 5 nmol in Wistar rats) were made before and within 30 and 60 minutes of Ang-(1-7) infusion. Ang-(1-7) infusion did not change mean arterial pressure (MAP) of Wistar rats (MAP=97+/-3 mm Hg), RHR (MAP=173+/-3 mm Hg), or SHR (MAP=177+/-5 mm Hg). In Wistar rats, Ang-(1-7) increased the BK hypotensive effect by 24+/-6% within 60 minutes of infusion. No significant changes were observed at 30 minutes of infusion. In additional groups of rats, Ang-(1-7) (5 pmol/min, n=5) was infused alone or combined with its selective antagonist D-Ala7-Ang-(1-7) (A-779) (5 pmol/min, n=6). The bradykinin-potentiating activity of Ang-(1-7) was completely abolished by A-779. In SHR and RHR, Ang-(1-7) significantly increased the hypotensive effect of BK by 59+/-8% and 57+/-9.8%, respectively, within 60 minutes of infusion. No significant changes were observed with saline infusion. In Wistar rats, enalaprilat treatment increased the BK-potentiating activity of Ang-(1-7) transforming the effect of 0.3 pmol/min into that observed with a rate 16-fold higher (5 pmol/min). On the other hand, in SHR enalaprilat did not change the Ang-(1-7) effect, while it abolished the BK potentiation in RHR. Our data show that the BK-potentiating activity of Ang-(1-7) is preserved and even augmented in hypertensive rats. The finding that the BK-potentiating activity of Ang-(1-7) could be demonstrated at a very low infusion rate suggests that this angiotensin can act as an endogenous modulator of the vascular actions of kinins. ACE inhibition can influence differently the BK-potentiating activity of Ang-(1-7) in normotensive and hypertensive rats.

Treatment with angiotensin-converting enzyme inhibitors increases the angiotensin-(1-7) [Ang-(1-7)] and bradykinin concentrations in plasma and tissue. In this study we evaluated the interaction between these peptides by determining the effect of Ang-(1-7) on the hypotensive action of bradykinin in conscious rats. Administration of Ang-(1-7) (5 nmol) did not change mean arterial pressure or heart rate. However, the hypotensive effect of bradykinin, produced by an intravenous or intra-arterial route, was potentiated by Ang-(1-7) in a dose-dependent manner. The Ang-(1-7) doses necessary to transform the effect of a single dose of bradykinin into that produced by a double dose (potentiating unit) were 2 nmol i.v. and 5 nmol IA. The Ang-(1-7) dose used did not change either the pressor effect of Ang II or the hypotensive effect of sodium nitroprusside. The bradykinin-potentiating Ang-(1-7) activity was significantly attenuated by pretreatment with indomethacin (5 mg/kg IM, n = 4). In an additional group the bradykinin-potentiating activity of Ang-(1-7) was evaluated 30 minutes after treatment with the angiotensin-converting enzyme inhibitor enalaprilat (10 mg/kg i.v., n = 9). Under this condition the bradykinin-potentiating activity of Ang-(1-7) was substantially increased, resulting in a potentiating unit of approximately 0.2 nmol IV. Pretreatment with indomethacin (5 mg/kg IM, n = 7) also attenuated the bradykinin-potentiating activity of Ang-(1-7) in enalaprilat-treated rats. These results show that Ang-(1-7) is a bradykinin-potentiating peptide in vivo. Furthermore, the data obtained with indomethacin suggest that prostaglandins participate in the mechanism of the bradykinin potentiation by Ang-(1-7). More importantly, these data suggest that the interaction between Ang-(1-7) and bradykinin can contribute to the pharmacological effects of angiotensin-converting enzyme inhibitors.