Influence of Zika virus on the cytotoxicity, cell adhesion, apoptosis and inflammatory markers of glioblastoma cells.

Glioblastoma (GBM) is one of the most common types of brain tumor in adults. Despite the availability of treatments for this disease, GBM remains one of the most lethal and difficult types of tumors to treat, and thus, a majority of patients die within 2 years of diagnosis. Infection with Zika virus (ZIKV) inhibits cell proliferation and induces apoptosis, particularly in developing neuronal cells, and thus could potentially be considered an alternative for GBM treatment. In the present study, two GBM cell lines (U-138 and U-251) were infected with ZIKV at different multiplicities of infection (0.1, 0.01 and 0.001), and cell viability, migration, adhesion, induction of apoptosis, interleukin levels and CD14/CD73 cell surface marker expression were analyzed. The present study demonstrated that ZIKV infection promoted loss of cell viability and increased apoptosis in U-138 cells, as measured by MTT and triplex assay, respectively. Changes in cell migration, as determined by wound healing assay, were not observed; however, the GBM cell lines exhibited an increase in cell adhesion when compared with non-tumoral cells (Vero). The Luminex immunoassay showed a significant increase in the expression levels of IL-4 specifically in U-251 cells (MOI 0.001) following exposure to ZIKV. There was no significant change in the expression levels of IFN-γ upon ZIKV infection in the cell lines tested. Furthermore, a marked increase in the percentage of cells expressing the CD14 surface marker was observed in both GBM cell lines compared with in Vero cells; and significantly increased CD73 expression was observed particularly in U-251 cells, when compared with uninfected cells. These findings indicate that ZIKV infection could lead to reduced cell viability, elevated CD73 expression, improved cellular adherence, and higher rates of apoptosis in glioblastoma cells. Further studies are required to explore the potential use of ZIKV in the treatment of GBM.

CD73 mitigates ZEB1 expression in papillary thyroid carcinoma.

BACKGROUND: ZEB1, a core transcription factor involved in epithelial-mesenchymal transition (EMT), is associated with aggressive cancer cell behavior, treatment resistance, and poor prognosis across various tumor types. Similarly, the expression and activity of CD73, an ectonucleotidase implicated in adenosine generation, is an important marker of tumor malignancy. Growing evidence suggests that EMT and the adenosinergic pathway are intricately linked and play a pivotal role in cancer development. Therefore, this study focuses on exploring the correlations between CD73 and ZEB1, considering their impact on tumor progression. METHODS: We employed CRISPR/Cas9 technology to silence CD73 expression in cell lines derived from papillary thyroid carcinoma. These same cells underwent lentiviral transduction of a reporter of ZEB1 non-coding RNA regulation. We conducted studies on cell migration using scratch assays and analyses of cellular speed and polarity. Additionally, we examined ZEB1 reporter expression through flow cytometry and immunocytochemistry, complemented by Western blot analysis for protein quantification. For further insights, we applied gene signatures representing different EMT states in an RNA-seq expression analysis of papillary thyroid carcinoma samples from The Cancer Genome Atlas. RESULTS: Silencing CD73 expression led to a reduction in ZEB1 non-coding RNA regulation reporter expression in a papillary thyroid carcinoma-derived cell line. Additionally, it also mitigated ZEB1 protein expression. Moreover, the expression of CD73 and ZEB1 was correlated with alterations in cell morphology characteristics crucial for cell migration, promoting an increase in cell polarity index and cell migration speed. RNA-seq analysis revealed higher expression of NT5E (CD73) in samples with BRAF mutations, accompanied by a prevalence of partial-EMT/hybrid state signature expression. CONCLUSIONS: Collectively, our findings suggest an association between CD73 expression and/or activity and the post-transcriptional regulation of ZEB1 by non-coding RNA, indicating a reduction in its absence. Further investigations are warranted to elucidate the relationship between CD73 and ZEB1, with the potential for targeting them as therapeutic alternatives for cancer treatment in the near future.

The functional role of p38 MAPK pathway in malignant brain tumors.

Gliomas are extremely debilitating malignant brain tumors with very limited response to therapies. The initiation and progression of gliomas can be attributed to several molecular abnormalities, such as mutations in important regulatory networks. In this regard, the mitogen-activated protein kinases (MAPKs) arise as key signaling pathways involved in cell proliferation, survival, and differentiation. MAPK pathway has been altered in most glial tumors. In glioma cells, the activation of p38 MAPK contributes to tumor invasion and metastasis and is positively correlated with tumor grade, being considered a potential oncogenic factor contributing to brain tumorigenesis and chemotherapy resistance. Hence, a better understanding of glioma pathogenesis is essential to the advancement of therapies that provide extended life expectancy for glioma patients. This review aims to explore the role of the p38 MAPK pathway in the genesis and progression of malignant brain tumors.

P2Y12 receptor antagonism inhibits proliferation, migration and leads to autophagy of glioblastoma cells.

Glioblastoma (GBM) is the most aggressive and lethal among the primary brain tumors, with a low survival rate and resistance to radio and chemotherapy. The P2Y12 is an adenosine diphosphate (ADP) purinergic chemoreceptor, found mainly in platelets. In cancer cells, its activation has been described to induce proliferation and metastasis. Bearing in mind the need to find new treatments for GBM, this study aimed to investigate the role of the P2Y12R in the proliferation and migration of GBM cells, as well as to evaluate the expression of this receptor in patients' data obtained from the TCGA data bank. Here, we used the P2Y12R antagonist, ticagrelor, which belongs to the antiplatelet agent's class. The different GBM cells (cell line and patient-derived cells) were treated with ticagrelor, with the agonist, ADP, or both, and the effects on cell proliferation, colony formation, ADP hydrolysis, cell cycle and death, migration, and cell adhesion were analyzed. The results showed that ticagrelor decreased the viability and the proliferation of GBM cells. P2Y12R antagonism also reduced colony formation and migration potentials, with alterations on the expression of metalloproteinases, and induced autophagy in GBM cells. Changes were observed at the cell cycle level, and only the U251 cell line showed a significant reduction in the ADP hydrolysis profile. TCGA data analysis showed a higher expression of P2Y12R in gliomas samples when compared to the other tumors. These data demonstrate the importance of the P2Y12 receptor in gliomas development and reinforce its potential as a pharmacological target for glioma treatment.

Inhibition of ATP hydrolysis as a key regulator of temozolomide resistance and migratory phenotype of glioblastoma cells.

Glioblastoma (GBM) is the most lethal among malignant gliomas. The tumor invasiveness and therapy-resistance are important clinical hallmarks. Growing evidence emphasizes the purinergic signaling contributing to tumor growth. Here we exposed a potential role of extracellular ATPase activity as a key regulator of temozolomide cytotoxicity and the migration process in GBM cells. The inhibition of ATP hydrolysis was able to improve the impact of temozolomide, causing arrest mainly in S and G2 phases of the cell cycle, leading M059J and U251 cells to apoptosis. In addition to eradicating GBM cells, ATP hydrolysis exhibited a potential to modulate the invasive phenotype and the expression of proteins involved in cell migration and epithelial-to-mesenchymal-like transition in a 3D culture model. Finally, we suggest the ATPase activity as a key target to decline temozolomide resistance and the migratory phenotype in GBM cells.

P2Y12 Purinergic Receptor and Brain Tumors: Implications on Glioma Microenvironment.

Gliomas are the most common malignant brain tumors in adults, characterized by a high proliferation and invasion. The tumor microenvironment is rich in growth-promoting signals and immunomodulatory pathways, which increase the tumor's aggressiveness. In response to hypoxia and glioma therapy, the amounts of adenosine triphosphate (ATP) and adenosine diphosphate (ADP) strongly increase in the extracellular space, and the purinergic signaling is triggered by nucleotides' interaction in P2 receptors. Several cell types are present in the tumor microenvironment and can facilitate tumor growth. In fact, tumor cells can activate platelets by the ADP-P2Y12 engagement, which plays an essential role in the cancer context, protecting tumors from the immune attack and providing molecules that contribute to the growth and maintenance of a rich environment to sustain the protumor cycle. Besides platelets, the P2Y12 receptor is expressed by some tumors, such as renal carcinoma, colon carcinoma, and gliomas, being related to tumor progression. In this context, this review aims to depict the glioma microenvironment, focusing on the relationship between platelets and tumor malignancy.

Immunosuppression in Gliomas via PD-1/PD-L1 Axis and Adenosine Pathway.

Glioblastoma is the most malignant and lethal subtype of glioma. Despite progress in therapeutic approaches, issues with the tumor immune landscape persist. Multiple immunosuppression pathways coexist in the tumor microenvironment, which can determine tumor progression and therapy outcomes. Research in immune checkpoints, such as the PD-1/PD-L1 axis, has renewed the interest in immune-based cancer therapies due to their ability to prevent immunosuppression against tumors. However, PD-1/PD-L1 blockage is not completely effective, as some patients remain unresponsive to such treatment. The production of adenosine is a major obstacle for the efficacy of immune therapies and is a key source of innate or adaptive resistance. In general, adenosine promotes the pro-tumor immune response, dictates the profile of suppressive immune cells, modulates the release of anti-inflammatory cytokines, and induces the expression of alternative immune checkpoint molecules, such as PD-1, thus maintaining a loop of immunosuppression. In this context, this review aims to depict the complexity of the immunosuppression in glioma microenvironment. We primarily consider the PD-1/PD-L1 axis and adenosine pathway, which may be critical points of resistance and potential targets for tumor treatment strategies.

Anemia e índices de atividade de doença em mulheres com artrite reumatoide / Anemia and disease activity indexes in women with rheumatoid arthritis

OBJETIVOS: Avaliar os índices de atividade de doença em pacientes do sexo feminino com artrite reumatoide, anêmicas e não anêmicas, correlacionando-os com os níveis de hemoglobina. MÉTODOS: Um estudo transversal envolveu mulheres com artrite reumatoide classificadas em dois grupos: 1) Anêmicas (hemoglobina < 12g/dL) e 2) Não anêmicas. A atividade da doença foi medida pelo Disease Activity Score (DAS28), utilizando-se marcadores inflamatórios distintos: velocidade de hemossedimentação (VHS) ou proteína C reativa (PCR). Esse escore também utiliza o número de articulações edemaciadas e dolorosas e a avaliação global da doença em escala analógica visual (EVA). Realizou-se também a avaliação da capacidade funcional pelo Health Assessment Questionaire (HAQ). A estatística utilizou os testes t de Student, Mann-Whitney, Wilcoxon, Fisher, likelihood ratio e correlação de Spearman. Foi considerado significativo p<0,05. RESULTADOS: Foram incluídas 24 pacientes, sendo oito do grupo Anêmicas e 16 do grupo Não anêmicas. Os grupos foram semelhantes quanto às características clínicas, demográficas e de tratamento, diferindo apenas em relação ao fator reumatoide, positivo em todas as participantes anêmicas e em 56,2% das não anêmicas. O DAS28 VHS (mediana 6,05; intervalo interquartis [IIQ] 5,21-7,76), o DAS28 PCR (mediana 4,32; IIQ 3,98-5,92) e a EVA (mediana 66,50 mm, IIQ 54,75-80,50) foram significativamente maiores no grupo Anêmicas. O DAS28 VHS (-0,418) e a EVA (-0,426) apresentaram correlação negativa significativa com o nível de hemoglobina. Os valores de DAS28 VHS e DAS28 PCR foram diferentes no mesmo grupo, mostrando discrepância na categorização da atividade da doença. No grupo Anêmicas, o valor de DAS28 VHS (mediana 6,05; IIQ 5,21-7,76) foi maior em relação ao DAS28 PCR (mediana 4,32; IIQ 3,98-5,92). Um aumento menos discrepante de DAS28 VHS (mediana 4,01; IIQ 3,05-5,68) comparado ao DAS28 PCR (mediana 3,06; IIQ 2,18-4,66) foi observado no grupo Não anêmicas. CONCLUSÕES: A anemia foi associada a piores índices de atividade de doença nas mulheres com artrite reumatoide, estando correlacionada com maior intensidade de dor e aumento do escore DAS28 VHS.

AIMS: To evaluate disease activity indexes in female patients with rheumatoid arthritis, Anemic and Non-anemic, correlating them with hemoglobin levels. METHODS: A cross-sectional study involved women with rheumatoid arthritis classified into two groups: 1) Anemic (hemoglobin <12 g/dL) and 2) Non-anemic. Disease activity was measured by Disease Activity Index (DAS28), using different inflammatory markers: Erythrocyte Sedimentation Rate (ESR) and C-reative Protein (CRP). This score also uses the number of swollen and painful joints and an overall assessment of the disease on Visual Analogue Scale (VAS). An assessment of functional capacity by the Health Assessment Questionnaire (HAQ) was also performed. The statistic used Student's t-Test, Mann-Whitney, Wilcoxon, Fisher, likelihood ratio and Spearman correlation tests. It was considered significant p<0,05. RESULTS: Twenty-four patients were included, eight patients in the Anemic group and 16 patients in the Non-anemic group. The groups were similar in terms of clinical, demographic and treatment characteristics, differing only in relation to rheumatoid factor, positive in all anemic participants and in 56,2% of non anemic participants. DAS28 ESR (median 6,05; interquartile range [IQR] 5,21-7,76), DAS28 CRP (median 4,32; IQR 3,98-5,92) and VAS (median 66,50 mm; IQR 54,75-80,50) were significantly higher in Anemic group. DAS28 ESR (-0,418) and VAS (-0,426) showed a significant negative correlation with hemoglobin level. DAS28 ESR and DAS28 CRP values were different in the same group, showing a discrepancy in the categorization of disease activity. In Anemic group, DAS28 ESR value (median 6,05; IQR 5,21-7,76) was higher in relation to DAS28 CRP (median 4,32; IQR 3,98-5,92). A less discrepant increase of DAS28 ESR (median 4,01; IQR 3,05-5,68) compared to DAS28 CRP (median 3,06; IQR 2,18-4,66) was observed in Non-anemic group. CONCLUSIONS: Anemia was associated with worse disease activity indexes in women with rheumatoid arthritis, being correlated with greater pain intensity and increase of DAS28 ESR score.

Mitochondria and redox homoeostasis as chemotherapeutic targets of Araucaria angustifolia (Bert.) O. Kuntze in human larynx HEp-2 cancer cells.

Natural products are among one of the most promising fields in finding new molecular targets in cancer therapy. Laryngeal carcinoma is one of the most common cancers affecting the head and neck regions, and is associated with high morbidity rate if left untreated. The aim of this study was to examine the antiproliferative effect of Araucaria angustifolia on laryngeal carcinoma HEp-2 cells. The results showed that A. angustifolia extract (AAE) induced a significant cytotoxicity in HEp-2 cells compared to the non-tumor human epithelial (HEK-293) cells, indicating a selective activity of AAE for the cancer cells. A. angustifolia extract was able to increase oxidative damage to lipids and proteins, and the production of nitric oxide, along with the depletion of enzymatic antioxidant defenses (superoxide dismutase and catalase) in the tumor cell line. Moreover, AAE was able to induce DNA damage, nuclear fragmentation and chromatin condensation. A significant increase in the Apoptosis Inducing Factor (AIF), Bax, poly-(ADP-ribose) polymerase (PARP) and caspase-3 cleavage expression were also found. These effects could be related to the ability of AAE to increase the production of reactive oxygen species through inhibition of the mitochondrial electron transport chain complex I activity and ATP production by the tumor cells. The phytochemical analysis of A. angustifolia, performed using High Resolution Mass Spectrometry (HRMS) in MS and MS/MS mode, showed the presence of dodecanoic and hexadecanoic acids, and phenolic compounds, which may be associated with the chemotherapeutic effect observed in this study.

Neuroprotective and anticonvulsant effects of organic and conventional purple grape juices on seizures in Wistar rats induced by pentylenetetrazole.

Epilepsy is the most common neurological disorder worldwide. Studies have shown that recurrent seizures may increase the concentration of reactive oxygen species, which can lead to oxidative stress and neuronal damage. These seizures result in substantial deleterious effects on an individual's health. Organic and conventional grape juices are rich in polyphenols, compounds with important antioxidant activity. However, these juices could have differences in their polyphenol content. The aim of this study was to investigate the neuroprotective and anticonvulsant effects of organic and conventional grape juice treatments in Wistar rats against pentylenetetrazole (a convulsant drug)-induced damage. In addition, we evaluated potential behavioral changes in rats treated with the juices and the polyphenolic profile of those samples. Animals (n=16 in each group) received treatment with saline, organic or conventional grape juice for 17 days. On the eighteenth day, behavioral changes were evaluated by an open field test. Afterwards, half of the rats from each group received pentylenetetrazole and were observed for 30 min to evaluate possible seizure characteristics. The animals were subsequently killed by decapitation and their hippocampus, cerebellum and cerebral cortex tissues were isolated. The results of this study showed that neither organic nor conventional grape juice altered the behavior parameters, and no statistical differences were observed in the seizure characteristics of the groups. Nevertheless, both juice types were able to protect from lipid and protein oxidative damage, decrease nitric oxide content and increase enzymatic (superoxide dismutase and catalase) and non-enzymatic (sulfhydryl protein) antioxidant defenses in brain tissues following pentylenetetrazole-induced seizures. In general, organic juice showed superior results in each test, probably due to its higher polyphenol content relative to conventional juice. These results indicate that grape juices can provide further insight into natural neuroprotective compounds and may lead to the development of new therapeutic strategies for epileptic patients.