Exosomes consist of membrane vesicles that are secreted by several cell types, including tumors and have been found in biological fluids. Exosomes interact with other cells and may serve as vehicles ...for the transfer of protein and RNA among cells.
SKOV3 exosomes were labelled with carboxyfluorescein diacetate succinimidyl-ester and collected by ultracentrifugation. Uptake of these vesicles, under different conditions, by the same cells from where they originated was monitored by immunofluorescence microscopy and flow cytometry analysis. Lectin analysis was performed to investigate the glycosylation properties of proteins from exosomes and cellular extracts.
In this work, the ovarian carcinoma SKOV3 cell line has been shown to internalize exosomes from the same cells via several endocytic pathways that were strongly inhibited at 4°C, indicating their energy dependence. Partial colocalization with the endosome marker EEA1 and inhibition by chlorpromazine suggested the involvement of clathrin-dependent endocytosis. Furthermore, uptake inhibition in the presence of 5-ethyl-N-isopropyl amiloride, cytochalasin D and methyl-beta-cyclodextrin suggested the involvement of additional endocytic pathways. The uptake required proteins from the exosomes and from the cells since it was inhibited after proteinase K treatments. The exosomes were found to be enriched in specific mannose- and sialic acid-containing glycoproteins. Sialic acid removal caused a small but non-significant increase in uptake. Furthermore, the monosaccharides D-galactose, α-L-fucose, α-D-mannose, D-N-acetylglucosamine and the disaccharide β-lactose reduced exosomes uptake to a comparable extent as the control D-glucose.
In conclusion, exosomes are internalized by ovarian tumor cells via various endocytic pathways and proteins from exosomes and cells are required for uptake. On the other hand, exosomes are enriched in specific glycoproteins that may constitute exosome markers. This work contributes to the knowledge about the properties and dynamics of exosomes in cancer.
Protein inclusions rich in mutant Cu,Zn superoxide dismutase (SOD1) have been found in tissues from patients with familial amyotrophic lateral sclerosis (ALS). Here, the mouse motor neuron-like ...NSC-34 cell line transiently transfected with human SOD1
G93A fused to enhanced green fluorescent protein exhibited aggregates contrary to cells overexpressing wild-type human SOD1. The aggregates were immunoreactive for ubiquitin but not for the TAR DNA binding protein (TDP-43) that was found in the nucleus. These characteristics mimicked the pathology of mutant SOD1 associated familial ALS. Aggregate formation and mutant SOD1 detergent insolubility were significantly decreased in the presence of millimolar concentrations of trehalose possibly due to its capacity to induce autophagy or to its properties as chemical chaperone. Mutant SOD1, aggregated and non-aggregated, caused decreased levels of concomitantly expressed secretory (β-trace protein and erythropoietin) and plasma membrane (L1 cell adhesion molecule) glycoproteins, which were not due to their intracellular accumulation. These cells may be used to study mechanisms of pathogenesis associated with ALS and to test potential therapeutic compounds.
Little is known about the existence of drug-resistant Toxoplasma gondii strains and their possible impact on clinic outcomes. To expand our knowledge about the existence of natural variations on drug ...susceptibility of T. gondii strains in Brazil, we evaluated the in vitro and in vivo susceptibility to sulfadiazine (SDZ) and pyrimethamine (PYR) of three atypical strains (Wild2, Wild3, and Wild4) isolated from free-living wild birds. In vitro susceptibility assay showed that the three strains were equally susceptible to SDZ and PYR but variations in the susceptibility were observed to SDZ plus PYR treatment. Variations in the proliferation rates in vitro and spontaneous conversion to bradyzoites were also accessed for all strains. Wild2 showed a lower cystogenesis capacity compared to Wild3 and Wild4. The in vivo analysis showed that while Wild3 was highly susceptible to all SDZ and PYR doses, and their combination, Wild2 and Wild4 showed low susceptibility to the lower doses of SDZ or PYR. Interestingly, Wild2 presented low susceptibility to the higher doses of SDZ, PYR and their combination. Our results suggest that the variability in treatment response by T. gondii isolates could possibly be related not only to drug resistance but also to the strain cystogenesis capacity.
O item -mente, do lat. mens, mentis ‘espírito, alma’, participa da formação de advérbios na grande maioria das línguas românicas. No português setecentista escrito no Brasil, apresenta-se ligado ...graficamente ao substantivo anterior a ele sob três formas: com um espaço em branco, com um hífen ou sem espaço algum. Assim, testa-se a hipótese de que esse item estaria com sua clitização finalizada já no século XVIII, utilizando-se como corpus estatutos e compromissos de irmandades e termos de devassa do estado de Minas Gerais, Brasil. Para tal, é preciso utilizarse do arcabaouço teórico-metodológico não apenas da cliticização, mas também da gramaticalização. Conclui-se que, no português do século XVIII, o -mente estava no fim de seu processo de cliticização, mas que esse processo ainda estava em curso, devido ao número pequeno, mas ainda existente, de advérbios em que há separação gráfica entre o item, analisado com um afixo, e o substantivo ao qual está relacionado.
This study aimed to evaluate the effect of genetic composition and arrangement between female and male parents on tolerance to delayed drying of maize seeds, evaluating the physiological quality and ...enzyme expression. Ears were harvested close to the stage of physiological maturity (around 35% moisture) and the genotypes were identified as line 1 (L1), line 2 (L2), the hybrid (HB - ♀L1 and ♂L2), and the reciprocal hybrid (HR - ♀L2 and ♂L1). For assessment of physiological quality, CDR (4x6x2) was used, consisting of four genotypes, six times of delay before artificial drying (10, 18, 24, 28, 32, and 40 hours), and two drying delay temperatures (42 and 48 °C). DIC (4x3) was used for enzymatic expression, consisting of four genotypes and three delay times before artificial drying (10, 24 and 40 hours) at 48 °C. Analysis of variance F (p < 0.05), Tukey’s test (p < 0.05), and analysis of polynomial regressions were performed on the data. Lineage arrangement affects seed tolerance to drying delay. Therefore, susceptible lines should not be used as female parents. The seeds of the line most susceptible (L2) to delay in drying exhibit less expression of α-amylase (α-AM).
Resumo: Objetivou-se avaliar o efeito da composição genética e arranjo entre progenitores fêmea e macho sobre a tolerância ao atraso na secagem de sementes de milho, avaliando a qualidade fisiológica e a expressão enzimática. As espigas foram colhidas próximo ao ponto de maturidade fisiológica (cerca de 35% de umidade) e genótipos identificados como linhagem 1 (L1), linhagem 2 (L2), híbrido (HB - ♀L1 e ♂L2) e o híbrido recíproco (HR - ♀L2 e ♂L1). Para avaliação da qualidade fisiológica, foi utilizado DIC (4x6x2), constituído de quatro genótipos, seis tempos de atraso antes da secagem artificial (10, 18, 24, 28, 32 e 40 horas) e duas temperaturas no retardamento de secagem (42 e 48 °C). Para avaliar a expressão enzimática, utilizou-se DIC (4x3), consistindo de quatro genótipos e três tempos de atraso antes da secagem artificial (10, 24 e 40 horas) à 48 °C. Em posse dos dados, foram realizados a análise de variância F (p < 0,05), teste de Tukey (p < 0,05) e análise de regressão polinomial. O arranjo das linhagens afeta a tolerância das sementes ao retardamento na secagem. Linhagens suscetíveis não devem ser usadas como parentais femininos. As sementes da linhagem mais suscetível (L2) ao retardamento da secagem apresentam menor expressão de α-amilase (α-AM).
To assess the prevalence and factors associated with smoking in adolescents with cleft lip and/or palate.
This is a cross-sectional study, developed in a Brazilian public and tertiary hospital ...between November 2018 and August 2019. Adolescents aged between 12 and 19 years old, previously submitted to cheiloplasty and/or palatoplasty surgeries were included. Data collection was carried out through interviews, guided by questionnaires referring to sociodemographic issues, use and factors associated with smoking. For statistical analysis, Fisher's Exact Test, Chi-square, Student's t test and bivariate logistic regression were used, all with a significance level of 5% (p≤0.05).
102 adolescents participated. The prevalence of smoking was 20.6% (n=21). Having cleft lip and palate was associated with smoking (p=0.012). The bivariate analysis showed that not dating (p=0.001; OR=0.19) and owning a home (p=0.032; OR=0.33) were configured as protective factors for the act of being a smoker, while believing that smoking facilitates interaction with young people or with the group of friends (p=0.043; OR=2.95), having friends who use alcoholic beverages (p=0.002; OR=8.40), having friends who use drugs, except cigarettes (p=0.002; OR=5.33), use alcoholic beverages (p<0.001; OR=10.67) and sexual initiation (p<0.001; OR=7.00), were associated with being a smoker.
From the knowledge of the profile of adolescents most vulnerable to tobacco use, it is possible to plan and implement educational and preventive actions.
Seeds harvested on the ears have high moisture content. On that account, this study aimed at evaluating the loss of physiological quality of corn seeds harvested on the ears, as a function of ...different drying-delay times. Hybrid corn ears were harvested at 31% moisture and then had their drying postponed for 0, 12, 24 and 36 h, while subjected to temperatures of 30, 40, 50 and 60 °C. The physiological quality was evaluated after 0, 4, 8 and 12 months of storage. A completely randomized design was employed, in a 4 x 4 x 4 factorial scheme, with four replications. In addition, a study was performed in a seed-processing unit, reporting the average waiting time before drying and the temperatures of all loads of a corn hybrid received at the facility. The physiological quality was not affected by temperatures below 40 °C, considering 36 h of waiting before drying. At 50 ºC during the drying delay, the germination was impaired 36 h afterward, and the vigor was compromised after 24 h, with the damage effects intensifying as the storage advanced. At the temperature of 60 ºC, the deficits in germination and vigor occurred within the first hours of drying delay.
Resumo: Sementes colhidas em espiga possuem elevados teores de água. Assim, o objetivo neste trabalho foi avaliar a perda de qualidade fisiológica de sementes de milho colhidas em espiga, em função do retardamento de secagem. As espigas de milho híbrido foram colhidas com 31% de umidade e submetidas ao atraso da secagem por 0, 12, 24 e 36 horas, sob as temperaturas de 30, 40, 50 e 60 °C. As avaliações da qualidade fisiológica foram realizadas aos 0, 4, 8 e 12 meses de armazenamento. Utilizou-se o delineamento inteiramente casualizado, em um esquema fatorial 4 x 4 x 4, com quatro repetições. Adicionalmente, foi realizado um levantamento do tempo médio até a secagem e temperatura de todas as cargas de um híbrido recebidas em uma unidade de beneficiamento de sementes. A qualidade fisiológica não foi afetada pelas temperaturas abaixo de 40 ºC, considerando-se 36 horas de espera até a secagem. Com a temperatura no retardamento da secagem de 50 °C, a germinação foi prejudicada com 36 horas e o vigor com 24 horas de atraso, e os danos foram maiores com o avanço do armazenamento. Para a temperatura de 60 ºC, as perdas de qualidade ocorrem nas primeiras horas de retardamento de secagem.
Currently, the demand is not only for more productive corn hybrids, but also for those with high physiological seed quality. Seed quality is considered the sum of genetic, physical, physiological and ...sanitary attributes that directly interfere in plant vigor. The objective of this study was to evaluate the physiological parameters of the seeds of parents and maize inter varietal hybrids obtained in a reciprocal recurrent selection program through physiological and image analysis techniques and either to study the feasibility of simultaneously selecting for seed physiological quality and grain yield. Two experiments were performed, one in the field and the other in the laboratory. The evaluated traits in the field were days of female flowering, days of male flowering, insertion of the 1st ear, plant height and grain yield. For the physiological quality, the attributes of germination at four days, germination at seven days, vigor by the cold test, emergence speed index, and the ratio of the root length to shoot length were obtained using GroundEye®. Heterosis was measured for the agronomic and physiological traits. Through the contrast between the inter varietal crosse hybrids and reciprocals we measured the maternal effect. The magnitudes of heterosis allowed us to infer that as greater as the number of selection and recombination cycles, on average, greater heterosis for the traits grain yield and seed germination. The correlated response indicated that, high yield intervarietal hybrids have better seed physiological quality.
RESUMO: Atualmente a demanda não é somente para híbridos de milho mais produtivos, mas também que apresentem alta qualidade fisiológica de sementes. A qualidade de sementes é considerada o somatório dos atributos genéticos, físicos, fisiológicos e sanitários que interferem diretamente no vigor das plantas. Objetivou-se avaliar os parâmetros fisiológicos de sementes dos parentais e híbridos de milho obtidos, por meio de análises fisiológicas e técnicas de análise de imagens, bem como estudar a viabilidade da seleção simultânea para a qualidade fisiológica de sementes e produtividade de grãos. Foram realizados dois experimentos, um à campo e o outro em laboratório. Os parâmetros avaliados em campo foram dias para o florescimento feminino, dias para o florescimento masculino, inserção da 1ª espiga, altura de plantas e produtividade de grãos. Para a qualidade fisiológica foram obtidos os atributos de germinação aos quatro dias, germinação aos sete dias, vigor pelo teste de frio, índice de velocidade de emergência e a razão do comprimento da raiz pelo comprimento da parte aérea através do GroundEye®. Obteve-se a heterose para os caracteres agronômicos e fisiológicos. O desdobramento dos efeitos por meio do contraste entre híbridos e recíprocos permitiu testar o efeito materno. As magnitudes da heterose permitem inferir que quanto maior o número de ciclos seletivos e de recombinação, em média maior será a heterose para os caracteres produtividade de grãos e germinação das sementes. A resposta correlacionada indica que híbridos intervarietais mais produtivos apresentam melhor qualidade fisiológica de sementes.
Purpose
Myelomeningocele (MMC) is the most frequent form of spina bifida. Moreover, 90% of patients with MMC have hydrocephalus and require shunt placement. However, shunt failure management in such ...patients typically involves several shunt revisions, increasing the risk for morbidity and mortality. In this study, we report our experience with endoscopic third ventriculostomy (ETV) in MMC patients with shunts in a Brazilian institution.
Methods
Patients with MMC who presented with shunt failure and underwent ETV alone between January 1996 and December 2016 were included in this study. Patients who had undergone endoscopic choroid plexus cauterization (CPC) or had non-shunted MMC hydrocephalus were excluded. Various data related to the ETV procedure were collected for each patient, including features, operative characteristics, complications, and ETV success. The follow-up period for the patients was 6–12 months. ETV success scores (ETVSSs) were calculated retrospectively for each patient to compare the predicted success of the procedure with the actual success at 6 months after ETV.
Results
Forty-three MMC patients with shunt failure were included in this study. Of them, 16 (37.2%) had a previous central nervous system (CNS) infection and 4 (9.3%) had complications. Two neuroendoscopies were interrupted, one because of anatomic impossibility due to thickening of the interthalamic adhesion, obliterating the floor of the third ventricle, and another because of intraoperative hemorrhage. One patient developed hyponatremia during the postoperative period, whereas another developed reversible paresis of the oculomotor nerve. The ETVSS predicted a success rate of 49.6% compared with the actual ETV success rate of 53.48% (23 patients) at 6 months. At 12 months, the actual success rate was 41.9% (18 patients). All patients survived the procedure.
Conclusion
Our findings indicate that ETV is an effective procedure for managing MMC-related hydrocephalus after shunt failure. The expertise of the neurosurgeon in neuroendoscopy procedures is often the deciding factor when choosing a management option. Accordingly, less experienced neurosurgeons should be warned of the risks of this procedure.
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