Role of Caveolin-1 in microglial phenotype: impact on Glioblastoma

Glioblastoma multiform (GBM) is a lethal brain tumour composed by many distinct cell types that are closely connected and dependent on their surrounding environment. Microglia are the brain immune cells, which are highly abundant in GBM and create an immunosuppressive microenvironment that promotes tumour progression. Caveolin-1 (Cav1) is the most important protein of caveolae and it is involved in cell signalling activity. In the GBM, Cav1 promotes the tumour invasion and it is correlated with a poor prognosis. In immunes cells, its role is not well explored, however it can be involved in immune response. Our hypothesis was that Cav1 could have an impact in the response of human microglia to the environment, influencing tumour progression. To test our hypothesis, a human microglia cell line and an iPSC cell line were used to generate Cav1 knockout clones using CRISPR-Cas9 technology. The iPSC was used to generated human microglia cells. Primary human microglia expressed low levels of Cav1, which could be regulated upon activation. The viral immortalized human microglia cells expressed strong Cav1 protein levels, possibly correlated with the immortalization procedure with SV40 large T antigen. This infection in combination with the culture conditions might lead to a constitutive pro-inflammatory phenotype, impacting the ability of microglia to react to other stimulus and to do phagocytosis. A slightly modified protocol to generate microglia from iPSC allowed the differentiated cells to be polarized towards pro-inflammatory and anti-inflammatory phenotype and to perform phagocytosis. In microglia, Cav1 was involved in the regulation of the inflammatory response, cell migration, phagocytosis, and sensitivity to temozolomide. The microglia cell line did not impact the tumour behaviour, likely due to the profile presented by the cells. However, the deletion of Cav1 in microglia derived from iPSC promoted the tumour invasion

A human co‐culture cell model incorporating microglia supports glioblastoma growth and migration, and confers resistance to cytotoxics

Despite the importance of the tumor microenvironment in regulating tumor progression, few in vitro models have been developed to understand the effects of non‐neoplastic cells and extracellular matrix (ECM) on drug resistance in glioblastoma (GBM) cells. Using CellTrace‐labeled human GBM and microglial (MG) cells, we established a 2D co‐culture including various ratios of the two cell types. Viability, proliferation, migration, and drug response assays were carried out to assess the role of MG. A 3D model was then established using a hyaluronic acid‐gelatin hydrogel to culture a mixture of GBM and MG and evaluate drug resistance. A contact co‐culture of fluorescently labeled GBM and MG demonstrated that MG cells modestly promoted tumor cell proliferation (17%‐30% increase) and greater migration of GBM cells (>1.5‐fold increase). Notably, the presence of MG elicited drug resistance even when in a low ratio (10%‐20%) relative to co‐cultured tumor cells. The protective effect of MG on GBM was greater in the 3D model (>100% survival of GBM when challenged with cytotoxics). This new 3D human model demonstrated the influence of non‐neoplastic cells and matrix on chemoresistance of GBM cells to three agents with different mechanisms of action suggesting that such sophisticated in vitro approaches may facilitate improved preclinical testing

ACIDENTES MAIS FREQUENTES NA UTILIZAÇÃO DO HIPOCLORITO DE SÓDIO EM ENDODONTIA

Introdução: O tratamento endodôntico compreende a área da odontologia que visa tratar afecções da polpa dentária quando sua vitalidade está comprometida. Sendo assim, o mais importante é o que se retira do interior do canal. Partindo deste princípio, o irrigante auxiliar no preparo dos canais radiculares torna-se relevante nesta etapa, pois a solução promove uma circulação hidráulica promovendo a remoção das raspas de dentina, com o auxílio da aspiração. Alguns critérios são indispensáveis na escolha do irrigante, este deve apresentar: ação antimicrobiana, dissolução tecidual, inativação de endotoxinas e ser biocompatível. Esta solução apresenta concentrações de 0,5% até 5,25%, por este motivo é interessante que o profissional tenha conhecimento de que quanto maior sua concentração, maior seu poder de limpeza. Porém, o poder irritante também é maior, podendo provocar danos aos tecidos vivos apicais e periapicais. Reações alérgicas, obstrução das vias aéreas, e extrusão da solução nos tecidos adjacentes são alguns dos acidentes durante o tratamento. Estudos concluíram ser raro mas não incomuns, e embora alguns pacientes possam ser sensíveis a solução, este é um problema que pode ser abordado antes do tratamento, questionando se o paciente já apresentou sensibilidade quando em contato com água sanitária ou cloro de piscina. Objetivo: Realizar uma revisão de literatura acerca dos acidentes mais frequentes com hipoclorito de sódio em endodontia que possam comprometer o sucesso do tratamento. Metodologia: Para elaboração deste estudo, foi escolhido o método de revisão de literatura. Os materiais foram pesquisados e selecionados conforme o objetivo proposto, em bases de dados como Medline, PubMed e EBSCO, compreendendo o período de 2006 a 2016. Conclusão: Conclui-se que embora raros os acidentes com a utilização da solução não são incomuns. Portanto atenção minuciosa na hora da anamnese e os cuidados da manipulação tornam-se importantes aliados para assim evitá-los

Caveolin-1, a key mediator across multiple pathways in glioblastoma and an independent negative biomarker of patient survival

Glioblastoma (GB) remains an aggressive malignancy with an extremely poor prognosis. Discovering new candidate drug targets for GB remains an unmet medical need. Caveolin-1 (Cav-1) has been shown to act variously as both a tumour suppressor and tumour promoter in many cancers. The implications of Cav-1 expression in GB remains poorly understood. Using clinical and genomic databases we examined the relationship between tumour Cav-1 gene expression (including its spatial distribution) and clinical pathological parameters of the GB tumour and survival probability in a TCGA cohort (n=155) and CGGA cohort (n=220) of GB patients. High expression of Cav-1 represented a significant independent predictor of shortened survival (HR = 2.985, 5.1 vs 14.9 months) with a greater statistically significant impact in female patients and in the Proneural and Mesenchymal GB subtypes. High Cav-1 expression correlated with other factors associated with poor prognosis: IDH w/t status, high histological tumour grade and low KPS score. A total of 4879 differentially expressed genes (DEGs) in the GB tumour were found to correlate with Cav-1 expression (either positively or negatively). Pathway enrichment analysis highlighted an over-representation of these DEGs to certain biological pathways. Focusing on those that lie within a framework of epithelial to mesenchymal transition and tumour cell migration and invasion we identified 27 of these DEGs. We then examined the prognostic value of Cav-1 when used in combination with any of these 27 genes and identified a subset of combinations (with Cav-1) indicative of co-operative synergistic mechanisms of action. Overall, the work has confirmed Cav-1 can serve as an independent prognostic marker in GB, but also augment prognosis when used in combination with a panel of biomarkers or clinicopathologic parameters. Moreover, Cav-1 appears to be linked to many signalling entities within the GB tumour and as such this work begins to substantiate Cav-1 or its associated signalling partners as candidate target for GB new drug discovery

Search for bioindicators of pollution in the Guanabara Bay: integrations of ecologic patterns

Guanabara Bay, since its discovery, has largely changed with the human occupation causing large amounts of deposited sediment and waste, as well as domestic and industry sewage. Surface sediment was analysed for foraminifera and ostracoda distribution, diversity and dominance studies. These results were compared with TOC analyses aiming the determination of pollution bioindicators. In general, foraminifera dominant species were Ammonia tepida, Buliminella elegantissima and Quinqueloculina seminulum. The foraminifera assemblages presented distinct abundance and diversity values in different regions of the bay. The diversity was higher in the entrance (south) and in the central region than in the north region of the bay. The dominant species, that are characteristic of stressed environments, presented higher values of abundance in the north region. The TOC values increased from south to north regions, and were inversely proportional to foraminifera diversity. The very high TOC values in very polluted areas suggest sediment deposition in anoxic-dysoxic environment. The ostracoda Gen. Cyprideis was dominant and its occurrence increased from south to north region. Occurrence of Callistocythere sigmocostata, Xestoleberis sp., Aurila sp., and Paracypris sp. were restricted to the entrance and central area, indicating a preference for less restricted conditions, like marine conditions. Foraminifera and ostracoda characteristic responses to the environment conditions related high TOC values showed their importance as bioindicators of stressed environments caused by anthropogenic pollution, in the Guanabara Bay.A Baía de Guanabara, desde o seu descobrimento, tem sido bastante modificada pela ocupação humana, causando o acúmulo de grande quantidade de sedimento depositado e lixo, bem como lançamento de esgotos domésticos e industriais. Sedimentos superficiais foram analisados visando o estudo da distribuição, diversidade e dominância de foraminíferos e ostracodes. Estes estudos foram comparados com análises de COT objetivando a determinação de bioindicadores de poluição. Em geral as espécies dominantes de foraminíferos foram Ammonia tepida,Buliminella elegantissima and Quinqueloculina seminulum. As associações de foraminíferos apresentam distintos valores de abundância e diversidade em diferentes regiões da Baía. A diversidade foi mais elevada na entrada (sul) e na região central do que na região norte da Baía. As espécies dominantes, que são características de ambientes sob estresse, apresentaram valores altos de abundância na região norte. Os valores de COT aumentaram da região sul para o norte e foram inversamente proporcionais à diversidade de foraminíferos. O elevados valores de COT em áreas muito poluídas sugerem deposição sedimentar em ambiente anóxico-desóxico. O Gen. Cyprideis, de ostracode, foi dominante e sua ocorrência aumentou da região sul para o norte. A ocorrência de Callistocythere sigmocostata, Xestoleberis sp., Aurila sp. e Paracypris sp. foi estrita à entrada e região central, indicando uma preferência por condições menos restritas, como condições marinhas. As respostas características dos foraminíferos e ostracodes às condições ambientais relacionadas aos elevados valores de COT mostraram sua importância como bioindicadores de ambientes sob estresse causado por poluição antropogênica na Baía de Guanabara

Poly(ethylene glycol) based nanotubes for tuneable drug delivery to glioblastoma multiforme

Glioblastoma multiforme (GBM) is the most aggressive type of malignant brain tumour, which is associated with a poor two-year survival rate and a high rate of fatal recurrence near the original tumour. Focal/local drug delivery devices hold promise for improving therapeutic outcomes for GBM by increasing drug concentrations locally at the tumour site, or by facilitating the use of potent anti-cancer drugs that are poorly permeable across the blood brain barrier (BBB). For inoperable tumours, stereotactic delivery to the tumour necessitates the development of nanoscale/microscale injectable drug delivery devices. Herein we assess the ability of a novel class of polymer nanotube (based on poly(ethylene glycol) (PEG)) to load doxorubicin (a mainstay breast cancer therapeutic with poor BBB permeability) and release it slowly. The drug loading properties of the PEG nanotubes could be tuned by varying the degree of carboxylic acid functionalisation and hence the capacity of the nanotubes to electrostatically bind and load doxorubicin. 70% of the drug was released over the first seven days followed by sustained drug release for the remaining two weeks tested. Unloaded PEG nanotubes showed no toxicity to any of the cell types analysed, whereas doxorubicin loaded nanotubes decreased GBM cell viability (C6, U-87 and U-251) in a dose dependent manner in 2D in vitro culture. Finally, doxorubicin loaded PEG nanotubes significantly reduced the viability of in vitro 3D GBM models whilst unloaded nanotubes showed no cytotoxicity. Taken together, these findings show that polymer nanotubes could be used to deliver alternative anti-cancer drugs for local therapeutic strategies against brain cancers

Heparin-based, injectable microcarriers for controlled delivery of interleukin-13 to the brain

Interleukin-13 (IL-13) drives cells of myeloid origin towards a more anti-inflammatory phenotype, but delivery to the brain remains problematic. Herein, we show that heparin-based cryogel microcarriers load high amounts of IL-13, releasing it slowly. Intra-striatal injection of loaded microcarriers caused local up-regulation of ARG1 in myeloid cells for pro-regenerative immunomodulation in the brain