In vivo glioblastoma growth is reduced by apyrase activity in a rat glioma model

BACKGROUND: ATP is an important signalling molecule in the peripheral and central nervous system. Both glioma growth and tumor resection induces cell death, thus liberating nucleotides to the extracellular medium. Nucleotides are hydrolyzed very slowly by gliomas when compared with astrocytes and induce neuronal cell death and glioma proliferation. The objective of the present study was to test the involvement of extracellular ATP in glioblastoma growth in a rat glioma model. METHODS: To deplete the extracellular ATP, the enzyme apyrase was tested on the treatment of gliomas implanted in the rats CNS. One million glioma C6 cells in 3 microliters of DMEM/FCS were injected in the right striata of male Wistar rats, 250–270 g. After 20 days, the rats were decapitated and the brain sectioning and stained with hematoxylin and eosine. We performed immunohistochemical experiments with Ki67, CD31 and VEGF. Total RNA was isolated from cultured glioma C6 cells and the cDNA was analyzed by Real Time-PCR with primers for the NTPDase family. RESULTS: C6 glioma cells effectively have a low expression of all NTPDases investigated, in comparison with normal astrocytes. The implanted glioma co-injected with apyrase had a significant reduction in the tumor size (p < 0.05) when compared with the rats injected only with gliomas or with gliomas plus inactivated apyrase. According to the pathological analysis, the malignant gliomas induced by C6 injection and co-injected with apyrase presented a significant reduction in the mitotic index and other histological characteristics that indicate a less invasive/proliferative tumor. Reduction of proliferation induced by apyrase co-injection was confirmed by counting the percentage of Ki67 positive glioma cell nuclei. According to counts with CD31, vessel density and neoformation was higher in the C6 group 20 days after implantation. Confirming this observation, rats treated with apyrase presented less VEGF staining in comparison to the control group. CONCLUSION: These results indicate that the participation of extracellular ATP and the ecto-nucleotidases may be associated with the development of this type of brain tumor in an in vivo glioma model

Increased Level of Extracellular ATP at Tumor Sites: In Vivo Imaging with Plasma Membrane Luciferase

There is growing awareness that tumour cells build up a "self-advantageous" microenvironment that reduces effectiveness of anti-tumour immune response. While many different immunosuppressive mechanisms are likely to come into play, recent evidence suggests that extracellular adenosine acting at A2A receptors may have a major role in down-modulating the immune response as cancerous tissues contain elevated levels of adenosine and adenosine break-down products. While there is no doubt that all cells possess plasma membrane adenosine transporters that mediate adenosine uptake and may also allow its release, it is now clear that most of extracellularly-generated adenosine originates from the catabolism of extracellular ATP. METHODOLOGY/PRINCIPAL FINDINGS: Measurement of extracellular ATP is generally performed in cell supernatants by HPLC or soluble luciferin-luciferase assay, thus it generally turns out to be laborious and inaccurate. We have engineered a chimeric plasma membrane-targeted luciferase that allows in vivo real-time imaging of extracellular ATP. With this novel probe we have measured the ATP concentration within the tumour microenvironment of several experimentally-induced tumours. CONCLUSIONS/SIGNIFICANCE: Our results show that ATP in the tumour interstitium is in the hundreds micromolar range, while it is basically undetectable in healthy tissues. Here we show that a chimeric plasma membrane-targeted luciferase allows in vivo detection of high extracellular ATP concentration at tumour sites. On the contrary, tumour-free tissues show undetectable extracellular ATP levels. Extracellular ATP may be crucial for the tumour not only as a stimulus for growth but also as a source of an immunosuppressive agent such as adenosine. Our approach offers a new tool for the investigation of the biochemical composition of tumour milieu and for development of novel therapies based on the modulation of extracellular purine-based signalling

Evolução da prevalência de parasitoses intestinais em escolares em Caxias do Sul, RS Evolution of the prevalence of intestinal parasitosis among schoolchildren in Caxias do Sul, RS

Relatos da prevalência das parasitoses intestinais no Brasil são pontuais e têm sido descritos em diferentes populações, tornando difícil um diagnóstico abrangente. Visando estudar a variação em 35 anos da prevalência de enteroparasitoses em escolares de Caxias do Sul, RS, foram avaliados 9.787 exames parasitológicos de fezes realizados por centrífugo-sedimentação. Resultaram positivas 5.655 (58%) amostras sendo mais prevalente a infestação por Ascaris lumbricoides (47%), Trichuris trichiura (36%), Enterobius vermicularis (8%) e os protozoários: Giardia lamblia (24%) e Entamoeba coli (20%). A prevalência geral diminuiu de 89% para 37%, com um decréscimo médio de 1,4% ao ano. Houve redução na prevalência de Ascaris lumbricoides de 61 para 26% e de Trichuris trichiura de 38 a 18%. Para Giardia lamblia não houve alteração significativa. A prevalência de Entamoeba coli cresceu de 29 a 46%. Os decréscimos obtidos na prevalência dos helmintos são provavelmente devidos às melhorias da infra-estrutura e às ações formativas desenvolvidas nas escolas.<br>Reports on the prevalence of intestinal parasitosis in Brazil have been local in nature, with descriptions of different populations, which makes comprehensive diagnosis difficult. With the aim of studying the variation in the prevalence of intestinal parasitosis among schoolchildren in Caxias do Sul, Rio Grande do Sul, over a 35-year period, 9,787 parasitological stool tests that had been performed using centrifugal sedimentation were evaluated. There were positive results from 5,655 samples (58%), and the most prevalent infestations were of Ascaris lumbricoides (47%), Trichuris trichiura (36%), Enterobius vermicularis (8%) and the protozoa Giardia lamblia (24%) and Entamoeba coli (20%). The overall prevalence diminished from 89% to 37%, indicating an average decrease of 1.4% per year. Reductions in prevalence were observed for Ascaris lumbricoides (61 to 26%) and Trichuris trichiura (38 to 18%). No significant change was observed for Giardia lamblia. The prevalence of Entamoeba coli increased from 29 to 46%. The decreases in helminth prevalence were probably due to infrastructure improvements and educational actions undertaken in schools