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Purines and pyrimidines are signaling molecules in the tumor microenvironment that affect cancer immunity. The purinergic signaling pathways have been shown to play an important role in the development and progression of cancer. CD39 and CD73 are ectonucleotidases responsible for breaking down ATP or ADP into adenosine, which regulates immunosuppression in various types of cancer. These enzymes have been studied as a potential therapeutic target in immunotherapy, and recent research suggests a correlation between ectonucleotidases and clinical outcomes in cancer.Prostate cancer is the most diagnosed cancer in men, after non-melanoma skin tumors, and is the second leading cause of death in men in the world. Despite having long survival periods, patients often receive excessive or insufficient treatment. Within this complex landscape, the adenosine/CD73 pathway plays a crucial role. Therefore, this review aims to highlight new findings on the potential role of purinergic signaling in cancer treatment and emphasizes the importance of anti-CD73 as a pharmacological strategy for prostate cancer therapy., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Gardani, Diz, Dondé, Rockenbach, Laufer and Morrone.)

Radiotherapy (RT) is the established noninvasive treatment for glioblastoma (GBM), a highly aggressive malignancy. However, its effectiveness in improving patient survival remains limited due to the radioresistant nature of GBM. Metal-based nanostructures have emerged as promising strategies to enhance RT efficacy. Among them, titanate nanotubes (TNTs) have gained significant attention due to their biocompatibility and cost-effectiveness. This study aimed to synthesize zinc-modified TNTs (ZnTNT) from sodium TNTs (NaTNT), in addition to characterizing the formed nanostructures and evaluating their radiosensitization effects in GBM cells (U87 and U251). Hydrothermal synthesis was employed to fabricate the TNTs, which were characterized using various techniques, including transmission electron microscopy (TEM), energy-dispersive spectroscopy, scanning-transmission mode, Fourier-transform infrared spectroscopy, ICP-MS (inductively coupled plasma mass spectrometry), X-ray photoelectron spectroscopy, and zeta potential analysis. Cytotoxicity was evaluated in healthy (Vero) and GBM (U87 and U251) cells by the MTT assay, while the internalization of TNTs was observed through TEM imaging and ICP-MS. The radiosensitivity of ZnTNT and NaTNT combined with 5 Gy was evaluated using clonogenic assays. Monte Carlo simulations using the MCNP6.2 code were performed to determine the deposited dose in the culture medium for RT scenarios involving TNT clusters and cells. The results demonstrated differences in the dose deposition values between the scenarios with and without TNTs. The study revealed that ZnTNT interfered with clonogenic integrity, suggesting its potential as a powerful tool for GBM treatment., Competing Interests: The authors declare no competing financial interest., (© 2024 The Authors. Published by American Chemical Society.)

Pterocaulon polystachyum is a species of pharmacological interest for providing volatile and non-volatile extracts with antifungal and amebicidal properties. The biological activities of non-volatile extracts may be related to the presence of coumarins, a promising group of secondary metabolites. In the present study, leaves and inflorescences previously used for the extraction of essential oils instead of being disposed of were subjected to extraction with supercritical CO 2 after pretreatment with microwaves. An experimental design was followed to seek the best extraction condition with the objective function being the maximum total extract. Pressure and temperature were statistically significant factors, and the optimal extraction condition was 240 bar, 60 °C, and pretreatment at 30 °C. The applied mathematical models showed good adherence to the experimental data. The extracts obtained by supercritical CO 2 were analyzed and the presence of coumarins was confirmed. The extract investigated for cytotoxicity against bladder tumor cells (T24) exhibited significant reduction in cell viability at concentrations between 6 and 12 μg/mL. The introduction of green technology, supercritical extraction, in the exploration of P. polystachyum as a source of coumarins represents a paradigm shift with regard to previous studies carried out with this species, which used organic solvents. Furthermore, the concept of circular bioeconomy was applied, i.e., the raw material used was the residue of a steam-distillation process. Therefore, the approach used here is in line with the sustainable exploitation of native plants to obtain extracts rich in coumarins with cytotoxic potential against cancer 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., Competing Interests: JCDC is a researcher of Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq; Federal District, Brazil), which provided funding for the present study. The other authors declare that they have no competing interests., (Copyright: © 2024 Marinowic et al.)

The danger of ionizing radiation exposure to human health is a concern. Since its wide use in medicine and industry, the development of radioprotectors has been very significant. Adenosine exerts anti-inflammatory actions and promotes tissue protection and repair, by activating the P1 receptors (A 1 , A 2A , A 2B , and A 3 ). Zebrafish (Danio rerio) is an appropriate tool in the fields of toxicology and pharmacology, including the evaluation of radiobiological outcomes and in the search for radioprotector agents. This study aims to evaluate the effect of adenosine in the toxicity induced by radiation in zebrafish. Embryos were treated with 1, 10, or 100 µM adenosine, 30 min before the exposure to 15 Gy of gamma radiation. Adenosine potentiated the effects of radiation in heart rate, body length, and pericardial edema. We evaluated oxidative stress, tissue remodeling and inflammatory. It was seen that 100 µM adenosine reversed the inflammation induced by radiation, and that A 2A2 and A 2B receptors are involved in these anti-inflammatory effects. Our results indicate that P1R activation could be a promising pharmacological strategy for radioprotection., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Prostate cancer (PC) is the most diagnosed tumor in males and ranks as the second leading cause of male mortality in the western world. The CD39 and CD73 enzymes play a crucial role in cancer regulation by degrading nucleotides and forming nucleosides. This study aimed to investigate the expression of the CD39 and CD73 enzymes as potential therapeutic targets for PC. The initial part of this study retrospectively analyzed tissue samples from 23 PC patients. Using the TissueFAXS TM cytometry platform, we found significantly higher levels of CD39-labeling its intensity compared to CD73. Additionally, we observed a correlation between the Gleason score and the intensity of CD39 expression. In the prospective arm, blood samples were collected from 25 patients at the time of diagnosis and after six months of treatment to determine the expression of CD39 and CD73 in the serum extracellular vesicles (EVs) and to analyze nucleotide hydrolysis. Notably, the expression of CD39 in the EVs was significantly increased compared to the CD73 and/or combined CD39/CD73 expression levels at initial collection. Furthermore, our results demonstrated positive correlations between ADP hydrolysis and the transurethral resection and Gleason score. Understanding the role of ectonucleotidases is crucial for identifying new biomarkers in PC.

Bladder cancer (BC) is the most common cancer of the urinary tract. Bozepinib (BZP), a purine-derived molecule, is a potential compound for the treatment of cancer. Purinergic signaling consists of the activity of nucleosides and nucleotides present in the extracellular environment, modulating a variety of biological actions. In cancer, this signaling is mainly controlled by the enzymatic cascade involving the NTPDase/E-NPP family and ecto-5'-nucleotidase/CD73, which hydrolyze extracellular adenosine triphosphate (ATP) to adenosine (ADO). The aim of this work is to evaluate the activity of BZP in the purinergic system in BC cell lines and to compare its in vitro antitumor activity with cisplatin, a chemotherapeutic drug widely used in the treatment of BC. In this study, two different BC cell lines, grade 1 RT4 and the more aggressive grade 3 T24, were used along with a human fibroblast cell line MRC-5, a cell used to predict the selectivity index (SI). BZP shows strong antitumor activity, with notable IC 50 values (8.7 ± 0.9 µM for RT4; 6.7 ± 0.7 µM for T24), far from the SI for cisplatin (SI for BZP: 19.7 and 25.7 for RT4 and T24, respectively; SI for cisplatin: 1.7 for T24). BZP arrests T24 cells in the G 2 /M phase of the cell cycle, inducing early apoptosis. Moreover, BZP increases ATP and ADP hydrolysis and gene/protein expression of the NPP1 enzyme in the T24 cell line. In conclusion, BZP shows superior activity compared to cisplatin against BC cell lines in vitro., (© 2023. The Author(s), under exclusive licence to Springer Nature B.V.)

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 P2Y 12 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 P2Y 12 R 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 P2Y 12 R 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. P2Y 12 R 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 P2Y 12 R in gliomas samples when compared to the other tumors. These data demonstrate the importance of the P2Y 12 receptor in gliomas development and reinforce its potential as a pharmacological target for glioma treatment., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

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., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Grave, Scheffel, Cruz, Rockenbach, Goettert, Laufer and Morrone.)

As alterations in purinergic signaling have been observed in bladder diseases, we aimed to assess the potential prognostic role of purinergic receptors in bladder cancer in a translational approach based on clinical databases and in vitro data. The prognostic role of purinergic receptors in the survival of patients with bladder cancer and the expression profile of the altered P2 receptors in normal and in tumor samples were determined using The Cancer Genome Atlas databank. In T24 and RT4 human bladder cancer cell lines, the P2 purinergic receptors were characterized by RT-PCR and RT-qPCR analysis including radiotherapy exposure as treatment. The cell number and the cumulative population doubling were also assessed. The expression profile of P2X6 receptor in the cancer pathological stage and in the nodal metastasis status was in agreement with Kaplan-Meier analysis, indicating that high expression of this receptor was related to an increased survival rate in patients with bladder cancer. Of all the P2 receptors expressed on T24 cell line, P2X6 presented high expression after radiotherapy, while it was not altered in RT4 cells. In addition, irradiation promoted a decrease of T24 cell number, but did not change the cell number of RT4 after the same time and radiation dose. Along 7 days after irradiation exposure, both cells regrew. However, while P2X6 receptor was downregulated in T24 cells, it was upregulated in RT4 cells. Our findings indicated that high P2X6 receptor expression induced by radiation in T24 cell line may predict a good survival prognostic factor., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

In this work, an intercomparison of sensitization effects produced by gold (GNP) and dextran-coated iron oxide (SPION-DX) nanoparticles in M059J and U87 human glioblastoma cells was performed using 6 MV-photons. Three variables were mapped: the nanoparticle material, treatment concentration, and cell radiosensitivity. For U87, GNP treatments resulted in high sensitization enhancement ratios (SER[Formula: see text] up to 2.04). More modest effects were induced by SPION-DX, but still significant reductions in survival were achieved (maximum SER[Formula: see text] ). For the radiosensitive M059J, sensitization by both NPs was poor. SER[Formula: see text] increased with the degree of elemental uptake in the cells, but not necessarily with treatment concentration. For GNP, where exposure concentration and elemental uptake were found to be proportional, SER[Formula: see text] increased linearly with concentration in both cell lines. For SPION-DX, saturation of sensitization enhancement and metal uptake occurred at high exposures. Fold change in the [Formula: see text] ratios extracted from survival curves are reduced by the presence of SPION-DX but strongly increased by GNPs , suggesting that sensitization by GNPs occurs mainly via promotion of lethal damage, while for SPION-DX repairable damage dominates. The NPs were more effective in eliminating the radioresistant glioblastoma cells, an interesting finding, as resistant cells are key targets to improve treatment outcome., (© 2022. The Author(s).)

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., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier Inc. All rights reserved.)

The zebrafish has been considered an ideal model for studies of complex behaviors since its behavioral repertoire is well described. Therefore, this study evaluated the perceived pain through behavioral changes in zebrafish larvae. Here we investigated the Acetic Acid (AA) effects on zebrafish larvae exposed in a short-time period (60 s) and the preventive effect from routinely used compounds, Dimethyl Sulfoxide (DMSO), Ethanol (EtOH), Ibuprofen (IBP), and Paracetamol (PAR). In addition, the effect of P2×7 antagonist, A740003, and pannexin channel 1 (PANX-1) inhibitor Probenecid (PROB) on AA-induced behavioral changes were evaluated. AA impaired the distance covered, acceleration, movement, and latency to the first entry in the center from 5 dpf exposed larvae. At 0.050% AA, PAR prevented alterations from the distance covered, acceleration, and movement. Surprisingly, 0.3% DMSO prevented behavioral changes induced by AA. However, the effects from 0.2% DMSO were not prominent. We used 0.2% DMSO as a PROB diluent. PROB prevented the changes in distance and movement observed at both AA concentrations (0.0025% and 0.05%) tested. Since EtOH had no analgesic properties, we used it as an A740003 vehicle to observe the analgesic effects of this compound. As noted, A740003 did not prevent the behavioral changes in the AA-induced pain model. In contrast, 0.2% DMSO and PROB prevented AA-induced behavioral changes. These data enforce that zebrafish could be used in translational studies since this species has behavioral responses related to pain in the early stages of development and responses to analgesics similar to observed in mammals., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Quercetin is a flavonoid found in a great variety of foods such as vegetables and fruits. This compound has been shown to inhibit the proliferation of various types of cancer cells, as well as the growth of tumors in animal models. In the present study, we analyze morphological and mechanical changes produced by quercetin in T24 bladder cancer cells. Decreased cell viability and cell number were observed following quercetin treatment at 40 μM and 60 μM, respectively, as observed by the MTT assay and trypan blue exclusion test, supporting the hypothesis of quercetin anticancer effect. These assays also allowed us to determine the 40, 60, and 80 μM quercetin concentrations for the following analyses, Lactate Dehydrogenase assay (LDH); Nuclear Morphometric Analysis (NMA); and atomic force microscopy (AFM). The LDH assay showed no cytotoxic effect of quercetin on T24 cancer cells. The AFM showed morphological changes following quercetin treatment, namely decreased cell body, cytoplasmic retraction, and membrane condensation. Following quercetin treatment, the NMA evidenced an increased percentage of nuclei characteristic to the apoptotic and senescence processes. Cells also presented biophysical alterations consistent with cell death by apoptosis, as increased roughness and aggregation of membrane proteins, in a dose-dependent manner. Cellular elasticity, obtained through force curves, showed increased stiffness after quercetin treatment. Data presented herein demonstrate, for the first time, in a quantitative and qualitative form, the morphological and mechanical alterations induced by quercetin on bladder cancer cells., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

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-P2Y 12 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 P2Y 12 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.

Adenine nucleotides are important signaling molecules that mediate biological functions in many conditions, including cancer. The enzymes CD39 and CD73 produce adenosine in the extracellular milieu that has a very important role in tumor development. This study aimed to evaluate nucleotide hydrolysis in the plasma blood of breast cancer elderly patients. In this prospective cohort study, we investigated the ectonucleotidases activity in breast cancer elderly patients, at the moment of diagnosis and after treatment. Control group consisted of elderly women without cancer diagnostic. The nucleotide hydrolysis assay was performed by the malachite green method and used ATP, ADP, or AMP as substrates. Paired t test or Wilcoxon rank-sum test was used. Our data showed that breast cancer patients presented high levels of ATP and AMP hydrolyses when compared to control group at the moment of diagnosis. When analyzing the differences between the samples at the time of diagnostic and 6 months after treatment, we observed a significant reduction on CD73 activity after all treatments used: surgery, chemotherapy, radiotherapy, or hormone therapy. The results with APCP, a specific CD73 inhibitor, showed that the AMP hydrolysis was inhibited in all conditions evaluated. We observed a diminished ADPase activity in the patients without metastasis when compared to metastatic breast cancer patients. The results showed that AMP hydrolysis was reduced in the blood plasma of breast cancer elderly patients after different treatments. This study strengthens the potential role of CD73 enzyme as a biomarker for breast cancer treatment response., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Hybrid nanoparticles composed of different biopolymers for delivery of enzyme/prodrug systems are of interest for cancer therapy. Hyaluronic acid-coated chitosan nanoparticles (CS/HA NP) were prepared to encapsulate individually an enzyme/pro-drug complex based on horseradish peroxidase (HRP) and indole-3-acetic acid (IAA). CS/HA NP showed size around 158 nm and increase to 170 and 200 nm after IAA and HRP encapsulation, respectively. Nanoparticles showed positive zeta potential values (between +20.36 mV and +24.40 mV) and higher encapsulation efficiencies for both nanoparticles (up to 90 %) were obtained. Electron microscopy indicated the formation of spherical particles with smooth surface characteristic. Physicochemical and thermal characterizations suggest the encapsulation of HRP and IAA. Kinetic parameters for encapsulated HRP were similar to those of the free enzyme. IAA-CS/HA NP showed a bimodal release profile of IAA with a high initial release (72 %) followed by a slow-release pattern. The combination of HRP-CS/HA NP and IAA- CS/HA NP reduced by 88 % the cell viability of human bladder carcinoma cell line (T24) in the concentrations 0.5 mM of pro-drug and 1.2 μg/mL of the enzyme after 24 h., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Indole-3-carbinol (I3C) is a plant molecule known to be active against several types of cancer, but some chemical characteristics limit its clinical applications. In order to overcome these limitations, polymeric nanoparticles can be used as carrier systems for targeted delivery of I3C. In this study, chitosan and chitosan/polyethylene glycol nanoparticles (CS NP and CS/PEG NP, respectively) were prepared to encapsulate I3C by ionic gelation method. The polymeric nanoparticles were characterized by Dynamic Scattering Light (DLS), Zeta Potential (ZP), Fourier Transform Infrared (FTIR) spetroscopy, X-Ray Diffraction (XRD), Thermogravimetric Analysis (TGA), Differential Scanning Calorimetry (DSC), and Field Emission Gun Scanning Electron Microscopy (FEG-SEM). I3C release testing was performed at an acidic media and the interactions between I3C and chitosan or PEG were evaluated by Density Functional Theory (DFT). Cytotoxicity of nanoparticles in bladder cancer T24 cell line was evaluated by the Methyl-thiazolyl-tetrazolium (MTT) colorimetric assay. The average size of the nanoparticles was observed to be in the range from 133.3 ± 3.7 nm to 180.4 ± 2.7 nm with a relatively homogeneous distribution. Samples had relatively high positive zeta potential values (between +20.3 ± 0.5 mV and + 24.3 ± 0.5 mV). Similar encapsulation efficiencies (about 80%) for both nanoparticles were obtained. Physicochemical and thermal characterizations pointed to the encapsulation of I3c. electron microscopy showed spherical particles with smooth or ragged surface characteristics, depending on the presence of PEG. The mathematical fitting of the release profile demonstrated that I3C-CS NP followed the Higuchi model whereas I3C-CS/PEG NP the Korsmeyer-Peppas model. Chemical differences between the nanoparticles as based on the I3C/CS or I3C/PEG interactions were demonstrate by computational characterization. The assessment of cell viability by the MTT test showed that the presence of both free I3C and I3C-loaded nanoparticles lead to statistically significant reduction in T24 cells viability in the concentrations from 500 to 2000 μM, when comparison to the control group after 24 h of exposure. Thus, CS and CS/PEG nanoparticles present as feasible I3C carrier systems for cancer therapy., (Copyright © 2021 Elsevier B.V. All rights reserved.)

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., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Scheffel, Grave, Vargas, Diz, Rockenbach and Morrone.)

Esophageal cancer is a prominent worldwide illness that is divided into two main subtypes: esophageal squamous cell carcinoma and esophageal adenocarcinoma. Mortality rates are alarming, and the understanding of the mechanisms involved in esophageal cancer development, becomes essential. Purinergic signaling is related to many diseases and among these various types of tumors. Here we studied the effects of the P2Y 2 receptor activation in different types of esophageal cancer. Esophageal tissue samples of healthy controls were used for P2Y 2 R expression quantification. Two human esophageal cancer cell lines Kyse-450 (squamous cell carcinoma) and OE-33 (adenocarcinoma) were used to perform in vitro analysis of cell proliferation, migration, adhesion, and the signaling pathways involved in P2Y 2 R activation. Data showed that P2Y 2 R was expressed in biopsies of patients with ESCC and adenocarcinoma, as well as in the two human esophageal cancer cell lines studied. The RT-qPCR analysis demonstrated that OE-33 cells have higher P2RY2 expression than Kyse-450 squamous cell line. Results showed that P2Y 2 R activation, induced by ATP or UTP, promoted esophageal cancer cells proliferation and colony formation. P2Y 2 R blockage with the selective antagonist, AR-C 118925XX, led to decreased proliferation, colony formation and adhesion. Treatments with ATP or UTP activated ERK 1/2 pathway in ESCC and ECA cells. The P2Y 2 R antagonism did not alter the migration of esophageal cancer cells. Interestingly, the esophageal cancer cell lines presented a distinct profile of nucleotide hydrolysis activity. The modulation of P2Y 2 receptors may be a promising target for esophageal cancer treatment., (Copyright © 2020. Published by Elsevier B.V.)

Purpose: Testicular germ cells tumor (TGCT) are associated with a high cure rate and are treated with platinum-based chemotherapy. However, a group of testicular cancer patients may have a very unfavorable evolution and insensitivity to the main therapeutic agent chemotherapy (CT) cisplatin. The aim of this study was to evaluate the risk of recurrence and overall survival related to the expression of nuclear factor kappa-B (NF-κB), transglutaminase 2 (TG2) and excision repair cross-complementation group 1 (ERCC1) in patients with TGCT treated with platinum combinations., Patients and Methods: A retrospective study was performed with TGCT patients treated with platinum-based chemotherapy. Immunohistochemical analysis was performed and the expression was correlated with clinical and laboratory data., Results: Fifty patients were included, the mean age was 28.4 years (18 to 45), and 76% were non-seminoma. All patients were treated with standard cisplatin, etoposide and bleomycin or cisplatin, and etoposide. Patient's analyzed immunodetection for NF-κB, TG2, and ERCC1 were positive in 76%, 54% and 42%, respectively. Multivariate analysis identified that positive expressions to ERCC1 and NF-κB are independent risk factors for higher recurrence TGCT after chemotherapy (RR 2.96 and 3.16, respectively). Patients with positive expression of ERCC1 presented a poor overall survival rate for 10-year follow (p=0.001)., Conclusions: The expression of ERCC1 and NF-κB give a worse prognosis for relapse, and only ERCC1 had an influence on the overall survival of TGCT patients treated with platinum-based chemotherapy. These may represent markers that predict poor clinical outcome and response to cisplatin., Competing Interests: None declared., (Copyright® by the International Brazilian Journal of Urology.)

Interest in nanostructures such as titanate nanotubes (TNT) has grown notably in recent years due to their biocompatibility and economic viability, making them promising for application in the biomedical field. Quercetin (Qc) has shown great potential as a chemopreventive agent and has been widely studied for the treatment of diseases such as bladder cancer. Motivated by the possibilities of developing a new hybrid nanostructure with potential in biomedical applications, this study aimed to investigate the incorporation of quercetin in sodium (NaTNT) and zinc (ZnTNT) titanate nanotubes, and characterize the nanostructures formed. Qc release testing was also performed and cytotoxicity in Vero and T24 cell lines evaluated by the MTT assay. The effect of TNTs on T24 bladder cancer cell radiosensitivity was also assessed, using cell proliferation and a clonogenic assay. The TNT nanostructures were synthesized and characterized by FESEM, EDS, TEM, FTIR, XRD and TGA. The results showed that the nanostructures have a tubular structure and that the exchange of Na + ions for Zn 2+ and incorporation of quercetin did not alter this morphology. In addition, interaction between Zn and Qc increased the thermal stability of the nanostructures. The release test showed that maximum Qc delivery occurred after 24 h and the presence of Zn controlled its release. Biological assays indicated that the NaTNTQc and ZnTNTQc nanostructures decreased the viability of T24 cells after 48 h at high concentrations. Furthermore, the clonogenic assay showed that NaTNT, NaTNTQc, ZnTNT and ZnTNTQc combined with 5 Gy reduced the formation of polyclonal colonies of T24 cells after 48 h. The results suggest that the nanostructures synthesized in this study interfere in cell proliferation and can therefore be a powerful tool in the treatment of bladder cancer., (Copyright © 2020 Elsevier B.V. All rights reserved.)

This study investigated the effects of systemic treatment with a new formulation of resveratrol (RSV) vehicled in rice oil (RSVO) in experimental rat models of inflammation. Male Wistar rats were evaluated in the following in vivo models: carrageenan-induced acute edema, complete Freund's adjuvant (CFA)-evoked sub-chronic edema, and CFA-induced polyarthritis. The animals were treated orally with RSVO (10-15 mg/kg) or RSV (100-200 mg/kg), depending on the experimental protocol. RSV was more effective than RSVO in carrageenan-elicited acute edema when dosed in either prophylactic or therapeutic schemes of administration. However, the repeated RSVO administration, at 10-fold lower doses, exhibited superior anti-inflammatory actions in either the sub-chronic edema or the chronic polyarthritis model elicited by CFA, when compared with RSV. The novel formulation RSVO displayed a lower plasma biotransformation when compared with the RSV-treated group-46% versus 88% of metabolites, respectively. RSVO also prevented polyarthritis-related cartilage destruction, an effect that might rely on the inhibition of the pro-inflammatory cytokine interleukin-6 (IL-6), associated with an increase of the anti-inflammatory cytokine interleukin-10 (IL-10). Noteworthy, the long-term administration of RSVO did not elicit any gastrointestinal harm. Our study revealed that RSVO was notably effective in the long-term inflammatory and degenerative responses triggered by CFA. This innovative formulation might well represent a promising alternative for treating chronic inflammatory diseases, such as arthritis.

Adenosine is a nucleoside that acts as a signaling molecule by activating P1 purinergic receptors (A 1 , A 2A , A 2B and A 3 ). This activation is involved in immune responses, inflammation, and tissue remodeling and tumor progression. Gamma rays are a type of ionizing radiation widely adopted in radiotherapy of tumors. Although it brings benefits to the success of the therapeutic scheme, it can trigger cellular damages, inducing a perpetual inflammatory response that culminates in adverse effects and severe toxicity. Our study aims to characterize the adenosinergic system in a zebrafish embryo radiotherapy model, relating the adenosine signaling to the changes elicited by radiation exposure. To standardize the radiotherapy procedure, we established a toxicological profile after exposure. Zebrafish were irradiated with different doses of gamma rays (2, 5, 10, 15 and 20 Gy) at 24 hpf. Survival, hatching rate, heartbeats, locomotor activity and morphological changes were determined during embryos development. Although without significant difference in survival, gamma-irradiated embryos had their heartbeats increased and presented decreased hatching time, changes in locomotor activity and important morphological alterations. The exposure to 10 Gy disrupted the ecto-5'-nucleotidase/CD73 and adenosine deaminase/ADA enzymatic activity, impairing adenosine metabolism. We also demonstrated that radiation decreased A 2B receptor gene expression, suggesting the involvement of extracellular adenosine in the changes prompted by radiotherapy. Our results indicate that the components of the adenosinergic system may be potential targets to improve radiotherapy and manage the tissue damage and toxicity of ionizing radiation., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Copper is a metal that participates in several essential reactions in living organisms, and it has been used as an inflammatory inducing agent in zebrafish larvae. In this study, we evaluated the effect P2X7 receptor and/or pannexin channel 1 (PANX-1) blockage in this inflammation model. To perform the experiments, 7 dpf larvae were exposed to 10 μM of copper and treated with 100 μM probenecid, PANX-1 inhibitor, and/or 300 nM A740003, a P2X7R selective antagonist. Larvae survival was assessed up to 24 h after treatments. The evaluation of larvae behavior was evaluated after acute (4 h) and chronic (24 h) exposure. The parameters of locomotor activity measured were: mobile time, average speed, distance and turn angle. We analyzed the gene expression of the P2X7 receptor, PANX1a and PANX1b channels and interleukins IL-10 and IL-1b after 24 h of treatment. Treatments did not decrease larval survival in the time interval studied. Changes in larvae locomotion were observed after the longest time of exposure to copper and the treatment with probenecid was able to reverse part of the effects caused by copper. No significant difference was observed in the oxidative stress assays and probenecid and copper treatment decrease partially PANX1a gene expression groups. The data presented herein shows the relevance of the blockage of P2X7-PANX-1 in copper-induced inflammation., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Prostate cancer is among the major malignancies that affect men around the world. Adenine nucleotides are important signaling molecules that mediate innumerous biological functions in pathophysiological conditions, including cancer. These molecules are degraded by several ectoenzymes named ectonucleotidases that produce adenosine in the extracellular medium. Some of these ecto-enzymes can be found in soluble in the blood stream. Thus, the present study aimed to evaluate the hydrolysis of adenine nucleotides (ATP, ADP, and AMP) in the plasma blood of patients with prostate cancer. Peripheral blood samples were collected, and questionnaires were filled based on the clinical data of the medical records. The nucleotide hydrolysis was performed by Malachite Green method using ATP, ADP, and AMP as substrates. Plasma from prostate cancer patients presented an elevated hydrolysis of all nucleotides evaluated when compared to healthy individuals. NTPDase inhibitor (ARL67156) and the alkaline phosphatase inhibitor (levamisole) did not alter ATP hydrolysis. However, AMP hydrolysis was reduced by the CD73 inhibitor, APCP, and by levamisole, suggesting the action of a soluble form of CD73 and alkaline phosphatase. On microvesicles, it was observed that there was a low expression and activity of CD39 and almost absent of CD73. The correlation of ATP, ADP, and AMP hydrolysis with clinic pathological data demonstrated that patients who received radiotherapy showed a higher AMP hydrolysis than those who did not, and patients with lower clinical stage (CS-IIA) presented an elevated ATP hydrolysis when compared to those with more advanced clinical stages (CS-IIB and CS-III). Patients of all clinical stages presented an elevated AMPase activity. Therefore, we can suggest that the nucleotide hydrolysis might be attributed to soluble ecto-enzymes present in the plasma, which, in a coordinate manner, produce adenosine in the blood stream, favoring prostate cancer progression.

The release of damage-associated molecular patterns, including uridine triphosphate (UTP) and adenosine triphosphate (ATP) to the extracellular milieu is a key component of innate immune response to infection. Previously, we showed that macrophage infection by the protozoan parasite Leishmania amazonensis -the etiological agent of cutaneous leishmaniasis-can be controlled by ATP- and UTP-mediated activation of P2Y and P2X7 receptors (activated by UTP/ATP and ATP, respectively), which provided comparable immune responses against the parasite. Interestingly, in context of Leishmania amazonensis infection, UTP/P2Y triggered apoptosis, reactive oxygen species, and oxide nitric (NO) production, which are characteristic of P2X7 receptor activation. Here, we examined a possible "cross-talk" between P2Y 2 and P2X7 receptors, and the requirement for pannexin-1 (PANX-1) in the control of L. amazonensis infection in mouse peritoneal macrophages and in vivo . UTP treatment reduced L. amazonensis parasite load, induced extracellular ATP release [which was pannexin-1 (PANX-1) dependent], and triggered leukotriene B 4 (LTB 4 ) production in macrophages. UTP-induced parasite control was blocked by pharmacological antagonism of P2Y 2 or P2X7 receptors and was absent in macrophages lacking P2X7 or PANX-1. In addition, ATP release induced by UTP was also inhibited by PANX-1 blocker carbenoxolone, and partially reversed by inhibitors of vesicle traffic and actin cytoskeleton dynamics. In vivo , UTP treatment reduced footpad and popliteal lymph node parasite load, and the lesion in wild-type (WT) mice; fact not observed in P2X7 -/- mice. Our data reveal that P2Y 2 and P2X7 receptors cooperate to trigger potent innate immune responses against L. amazonensis infection.

Purpose: Trimodal therapy is a reasonable bladder-preserving option to radical cystectomy. However, many tumors are radioresistive. In this sense, the identification of new prognostic and predictive biomarkers that allow the selection of patients with better responses to radiation therapy would improve outcomes. With the aim of using ecto-5'-nucleotidase/CD73 as a predictive biomarker, the role of this enzyme in the context of radiotherapy in T24 human bladder cancer cell line was investigated., Methods: T24 cell line was exposure to a single dose of radiation (4 Gray) and trypan blue assay (pharmacological assays of viability/cumulative population doubling), flow cytometry (cell cycle/cell death/active caspase-3/ecto-5'-nucleotidase/CD73 protein staining), DAPI staining (nuclear morphometric assay), RT-PCR and real-time PCR, malachite green method (ectonucleotidase enzymatic assay), and HPLC (analysis of AMP metabolism) were carried out. T24 cell line in which ecto-5'-nucleotidase/CD73 has been completely silenced (5'KO) was also used., Results: The exposure of T24 cell line to a single dose (4 Gray) of radiation-induced cell death and triggered a transitory increase in ecto-5'-nucleotidase/CD73 expression, increased ectonucleotidase activity, and led to adenosine and inosine accumulation in the extracellular medium. Pharmacological inhibition or knocking out ecto-5'-nucleotidase/CD73 rescued cells' proliferative capacity, reducing their sensitivity to radiation., Conclusion: Our findings show that the induction of ecto-5'-nucleotidase/CD73 by radiation contributes to the radiosensitivity of T24 cell line.

This study investigated the role of kinins and their receptors in malignant brain tumors. As a first approach, GL-261 glioma cells were injected (2 × 10 5 cells in 2 μl/2 min) into the right striatum of adult C57/BL6 wild-type, kinin B 1 and B 2 receptor knockout (KOB 1 R and KOB 2 R) and B 1 and B 2 receptor double knockout mice (KOB 1 B 2 R). The animals received the selective B 1 R (SSR240612) and/or B 2 R (HOE-140) antagonists by intracerebroventricular (i.c.v.) route at 5, 10, and 15 days. The tumor size quantification, mitotic index, western blot analysis, quantitative autoradiography, immunofluorescence, and confocal microscopy were carried out in brain tumor samples, 20 days after tumor induction. Our results revealed an uncontrolled tumor growing in KOB 1 R or SSR240612-treated mice, which was blunted by B 2 R blockade with HOE-140, suggesting a crosstalk between B 1 R and B 2 R in tumor growing. Combined treatment with B 1 R and B 2 R antagonists normalized the upregulation of tumor B 1 R and decreased the tumor size and the mitotic index, as was seen in double KOB 1 B 2 R. The B 1 R was detected on astrocytes in the tumor, indicating a close relationship between this receptor and astroglial cells. Noteworthy, an immunohistochemistry analysis of tumor samples from 16 patients with glioma diagnosis revealed a marked B 1 R immunopositivity in low-grade gliomas or in older glioblastoma individuals. Furthermore, the clinical data revealed a significantly higher immunopositivity for B 1 R, when compared to a lower B 2 R immunolabeling. Taken together, our results show that blocking simultaneously both kinin receptors or alternatively stimulating B 1 R may be of therapeutic value in the treatment of brain glioblastoma growth and malignancy.

Esophageal cancer is an aggressive tumor and is the sixth leading cause of cancer death worldwide. ATP is well known to regulate cancer progression in a variety of models by different mechanisms, including P2X7R activation. This study aimed to evaluate the role of P2X7R in esophageal squamous cell carcinoma (ESCC) proliferation. Our results show that treatment with high ATP concentrations induced a decrease in cell number, cell viability, number of polyclonal colonies, and reduced migration of ESCC. The treatment with the selective P2X7R antagonist A740003 or siRNA for P2X7 reverted this effect in the KYSE450 cell line. In addition, results showed that P2X7R is highly expressed, at mRNA and protein levels, in KYSE450 lineage. Additionally, KYSE450, KYSE30, and OE21 cells express P2X3R, P2X4R, P2X5R, P2X6R, and P2X7R genes. P2X1R is expressed by KYSE30 and KYSE450, and only KYSE450 expresses the P2X2R gene. Furthermore, esophageal cancer cell line KYSE450 presented higher expression of E-NTPDases 1 and 2 and of Ecto-5'-NT/CD73 when compared to normal cells. This cell line also exhibits ATPase, ADPase, and AMPase activity, although in different levels, and the co-treatment of apyrase was able to revert the antiproliferative effects of ATP. Moreover, results showed high immunostaining for P2X7R in biopsies of patients with esophageal carcinoma, indicating the involvement of this receptor in the growth of this type of cancer. The results suggest that P2X7R may be a potential pharmacological target to treat ESCC and can lead us to further investigate the effect of this receptor in cancer cell progression.

This study investigated the effects of caffeine in the behavioral and inflammatory alterations caused by copper in zebrafish larvae, attempting to correlate these changes with the modulation of adenosine receptors. To perform a survival curve, 7dpf larvae were exposed to 10μM CuSO 4 , combined to different concentrations of caffeine (100μM, 500μM and 1mM) for up to 24h. The treatment with copper showed lower survival rates only when combined with 500μM and 1mM of caffeine. We selected 4 and 24h as treatment time-points. The behavior evaluation was done by analyzing the traveled distance, the number of entries in the center, and the length of permanence in the center and the periphery of the well. The exposure to 10μM CuSO 4 plus 500μM caffeine at 4 and 24h changed the behavioral parameters. To study the inflammatory effects of caffeine, we assessed the PGE 2 levels by using UHPLC-MS/MS, and TNF, COX-2, IL-6 and IL-10 gene expression by RT-qPCR. The expression of adenosine receptors was also evaluated with RT-qPCR. When combined to copper, caffeine altered inflammatory markers depending on the time of exposure. Adenosine receptors expression was significantly increased, especially after 4h exposure to copper and caffeine together or separately. Our results demonstrated that caffeine enhances the inflammation induced by copper by decreasing animal survival, altering inflammatory markers and promoting behavioral changes in zebrafish larvae. We also conclude that alterations in adenosine receptors are related to those effects., (Copyright © 2017 Elsevier Inc. All rights reserved.)

This study investigated the effects of P/Q- and N-type voltage-gated calcium channel (VGCC) blockers derived from P. nigriventer in glioma progression, by means of in vitro and in vivo experiments. Glioma cells M059J, U-138MG and U-251MG were used to evaluate the antiproliferative effects of P/Q- and N-type VGCC inhibitors PhTx3-3 and Phα1β from P. nigriventer (0.3-100 pM), in comparison to MVIIC and MVIIA from C. magus (0.3-100 pM), respectively. The toxins were also analyzed in a glioma model induced by implantation of GL261 mouse cells. PhTx3-3, Phα1β and MVIIA displayed significant inhibitory effects on the proliferation and viability of all tested glioma cell lines, and evoked cell death mainly with apoptosis characteristics, as indicated by Annexin V/propidium iodide (PI) positivity. The antiproliferative effects of toxins were confirmed by flow cytometry using Ki67 staining. None of the tested toxins altered the proliferation rates of the N9 non-tumor glial cell line. Noteworthy, the administration of the preferential N-type VGCC inhibitors, Phα1β (50 pmol/site; i.c.v.), its recombinant form CTK 01512-2 (50 pmol/site; i.c.v. and i.t.), or MVIIA (10 pmol/site; i.c.v.) caused significant reductions of tumor areas in vivo. N-type VGCC inhibition by Phα1β, CTK 01512-2, and MVIIA led to a marked increase of GFAP-activated astrocytes, and Iba-1-positive microglia, in the peritumoral region, which might explain, at least in part, the inhibitory effects of the toxins in tumor development. This study provides novel evidence on the potential effects of P. nigriventer-derived P/Q-, and mainly, N-type VGCC inhibitors, in glioma progression., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Malignant brain tumors are highly lethal and aggressive. Despite recent advances in the current therapies, which include the combination of surgery and radio/chemotherapy, the average survival rate remains poor. Altered regulation of ion channels is part of the neoplastic transformation, which suggests that ion channels are involved in cancer. Distinct classes of calcium-permeable channels are abnormally expressed in cancer and are likely involved in the alterations underlying malignant growth. Specifically, cytosolic Ca(2+) activity plays an important role in the regulation of cell proliferation, and Ca(2+) signaling is altered in proliferating tumor cells. A series of previous studies emphasized the importance of the T-type low-voltage-gated calcium channels (VGCC) in different cancer types, including gliomas, and remarkably, pharmacologic inhibition of T-type VGCC caused antiproliferative effects and triggered apoptosis of human glioma cells. Other calcium permeable channels, such as transient receptor potential (TRP) channels, contribute to changes in Ca(2+) by modulating the driving force for Ca(2+) entry, and some TRP channels are required for proliferation and migration in gliomas. Furthermore, recent evidence shows that TRP channels contribute to the progression and survival of the glioblastoma patients. Likewise, the purinergic P2X7 receptor acts as a direct conduit for Ca(2+)-influx and an indirect activator of voltage-gated Ca(2+)-channel. Evidence also shows that P2X7 receptor activation is linked to elevated expression of inflammation promoting factors, tumor cell migration, an increase in intracellular mobilization of Ca(2+), and membrane depolarization in gliomas. Therefore, this review summarizes the recent findings on calcium channels and associated receptors as potential targets to treat malignant gliomas., (Copyright © 2016 by The American Society for Pharmacology and Experimental Therapeutics.)

The objective of this study was to explore the association between the P2X7 purinergic receptor (P2X7R) and neuroinflammation using a preclinical model of acute bipolar mania. We analyzed the modulatory effects of P2X7R agonist (3'-O-(4-benzoyl)benzoyl-adenosine 5'-triphosphate, BzATP) and antagonists (brilliant blue, BBG and 3-[[5-(2,3 dichlorophenyl)-1H-tetrazol-1-yl]methyl]pyridine hydrochloride, A438079) on assessments related to behavior (locomotor activity), neuroinflammation (interleukin-1 beta, IL-1β; tumor necrosis factor alpha, TNF-α; and interleukin- 6, IL-6), oxidative stress (thiobarbituric acid reactive substances, TBARS) and neuroplasticity (brain-derived neurotrophic factor, BDNF) markers in a pharmacological model of mania induced by acute and chronic treatment with D-amphetamine (AMPH) (2 mg/kg) in mice. An apparent lack of responsiveness to AMPH was observed in terms of the locomotor activity in animals with blocked P2X7R or with genetic deletion of P2X7R in knockout (P2X7R(-/-)) mice. Likewise, P2X7R participated in the AMPH-induced increase of the proinflammatory and excitotoxic environment, as demonstrated by the reversal of IL-1β, TNF-α, and TBARS levels caused by P2X7R blocking. Our results support the hypothesis that P2X7R plays a role in the neuroinflammation induced by AMPH in a preclinical model of mania, which could explain the altered behavior. The present data suggest that P2X7R may be a therapeutic target related to the neuroinflammation reported in bipolar disorder.

A cross-sectional, population-based study was conducted on a random sample of 427 elderly individuals living in Porto Alegre, Brazil, to establish the nutrient consumption profile and verify its association with sociodemographic and health variables. Dietary intake was assessed using the 24-hour Food Recall Survey and the Dietetic Research Investigation technique. Seventy percent of the elderly respondents were women: 48.5% were between 60 and 69 years old; 68.8% had less than 8 years of schooling; 39% had a family income of between 2 and 5 minimum wages and 58.4% took no physical exercise. Hypertension was the most prevalent disease among the elderly and 54.9% were underweight. Men consumed more calories, protein, fiber, minerals and vitamins than women. Carbohydrate and calcium intake increases with advancing age, while zinc intake decreases. Physical exercise increased the intake of calories, magnesium, potassium and phosphorus. The higher the schooling the greater the intake of vitamins B6 and B12; the higher the family income, the greater the consumption of vitamin B6 and folic acid. The results show that there are nutritional deficiencies in the daily diet of the Brazilian elderly population, especially among women and individuals over 80 years of age.

Elevated plasma levels of homocysteine (Hcy) are associated with the development of coronary artery disease (CAD), peripheral vascular disease, and atherosclerosis. Hyperhomocysteinemia is likely related to the enhanced production of pro-inflammatory cytokines including IL-1β. However, the mechanisms underlying the effects of Hcy in immune cells are not completely understood. Recent studies have established a link between macrophage accumulation, cytokine IL-1β, and the advance of vascular diseases. The purpose of the present study is to investigate the effects of Hcy on IL-1β secretion by murine macrophages. Hcy (100 μM) increases IL-1β synthesis via enhancement of P2X7 expression and NF-ĸB and ERK activation in murine macrophages. In addition, the antioxidant agent N-acetylcysteine (NAC) reduces NF-κB activation, ERK phosphorylation, and IL-1β production in Hcy-exposed macrophages, indicating the importance of ROS in this pro-inflammatory process. In summary, our results show that Hcy may be involved in the synthesis and secretion of IL-1β via NF-ĸB, ERK, and P2X7 stimulation in murine macrophages.

Glioblastoma multiforme (GBM) is considered the most lethal intracranial tumor and the median survival time is approximately 14 months. Although some glioma cells present radioresistance, radiotherapy has been the mainstay of therapy for patients with malignant glioma. The activation of P2X7 receptor (P2X7R) is responsible for ATP-induced death in various cell types. In this study, we analyzed the importance of ATP-P2X7R pathway in the radiotherapy response P2X7R silenced cell lines, in vivo and human tumor samples. Both glioma cell lines used in this study present a functional P2X7R and the P2X7R silencing reduced P2X7R pore activity by ethidium bromide uptake. Gamma radiation (2Gy) treatment reduced cell number in a P2X7R-dependent way, since both P2X7R antagonist and P2X7R silencing blocked the cell cytotoxicity caused by irradiation after 24h. The activation of P2X7R is time-dependent, as EtBr uptake significantly increased after 24h of irradiation. The radiotherapy plus ATP incubation significantly increased annexin V incorporation, compared with radiotherapy alone, suggesting that ATP acts synergistically with radiotherapy. Of note, GL261 P2X7R silenced-bearing mice failed in respond to radiotherapy (8Gy) and GL261 WT-bearing mice, that constitutively express P2X7R, presented a significant reduction in tumor volume after radiotherapy, showing in vivo that functional P2X7R expression is essential for an efficient radiotherapy response in gliomas. We also showed that a high P2X7R expression is a good prognostic factor for glioma radiosensitivity and survival probability in humans. Our data revealed the relevance of P2X7R expression in glioma cells to a successful radiotherapy response, and shed new light on this receptor as a useful predictor of the sensitivity of cancer patients to radiotherapy and median survival., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Uncaria tomentosa is widely used in folk medicine for the treatment of numerous diseases, such as urinary tract disease. Hemorrhagic cystitis (HE) is an inflammatory condition of the bladder associated with the use of anticancer drugs such as cyclophosphamide (CYP). Sodium 2-mercaptoethanesulfonate (Mesna) has been used to prevent the occurrence of HE, although this compound is not effective in established lesions. It has been demonstrated that the purinergic system is involved in several pathophysiological events. Among purinergic receptors, P2X7 deserves attention because it is involved in HE induced by CYP and, therefore, can be considered a therapeutic target. The objective of this study was to investigate the potential therapeutic effect of the quinovic acid glycosides purified fraction (QAPF) from U. tomentosa in the mouse model of CYP-induced HE. Pretreatment with QAPF not only had a protective effect on HE-induced urothelial damage (edema, hemorrhage and bladder wet weight) but was also able to control visceral pain, decrease IL-1β levels and down-regulates P2X7 receptors, most likely by inhibit the neutrophils migration to the bladder. This research clearly demonstrates the promising anti-inflammatory properties of QAPF, supporting its use as complementary therapy. QAPF represents a promising therapeutic option for this pathological condition.

Extracellular adenosine 5'-triphosphate (ATP) has significant effects on a variety of pathological conditions and it is the main physiological agonist of P2X7 purinergic receptor (P2X7R). It is known that ATP acting via purinergic receptors plays a relevant role on skin inflammation, and P2X7R is required to neutrophil recruitment in a mice model of irritant contact dermatitis (ICD).The present study investigated the effects of chemical irritant croton oil (CrO) upon ATP, ADP, and AMP hydrolysis in mice blood serum, and the potential involvement of P2X7R. The topical application CrO induced a decrease on soluble ATP/ADPase activities (~50 %), and the treatment with the selective P2X7R antagonist, A438079, reversed these effects to control level. Furthermore, we showed that CrO decreased cellular viability (52.6 % ± 3.9) in relation to the control and caused necrosis in keratinocytes (PI positive cells). The necrosis induced by CrO was prevented by the pre-treatment with the selective P2X7R antagonist A438079. The results presented herein suggest that CrO exerts an inhibitory effect on the activity of ATPDase in mouse serum, reinforcing the idea that ICD has a pathogenic mechanism dependent of CD39. Furthermore, it is tempting to suggest that P2X7R may act as a controller of the extracellular levels of ATP.

Background and Purpose: Spinal voltage-gated calcium channels (VGCCs) are pivotal regulators of painful and inflammatory alterations, representing attractive therapeutic targets. We examined the effects of epidural administration of the P/Q- and N-type VGCC blockers Tx3-3 and Phα1β, respectively, isolated from the spider Phoneutria nigriventer, on symptomatic, inflammatory and functional changes allied to mouse cyclophosphamide (CPA)-induced haemorrhagic cystitis (HC). The effects of P. nigriventer-derived toxins were compared with those displayed by MVIIC and MVIIA, extracted from the cone snail Conus magus., Experimental Approach: HC was induced by a single i.p. injection of CPA (300 mg·kg(-1) ). Dose- and time-related effects of spinally administered P/Q and N-type VGCC blockers were assessed on nociceptive behaviour and macroscopic inflammation elicited by CPA. The effects of toxins were also evaluated on cell migration, cytokine production, oxidative stress, functional cystometry alterations and TRPV1, TRPA1 and NK1 receptor mRNA expression., Key Results: The spinal blockage of P/Q-type VGCC by Tx3-3 and MVIIC or N-type VGCC by Phα1β attenuated nociceptive and inflammatory events associated with HC, including bladder oxidative stress and cytokine production. CPA produced a slight increase in bladder TRPV1 and TRPA1 mRNA expression, which was reversed by all the toxins tested. Noteworthy, Phα1β strongly prevented bladder neutrophil migration, besides HC-related functional alterations, and its effects were potentiated by co-injecting the selective NK1 receptor antagonist CP-96345., Conclusions and Implications: Our results shed new light on the role of spinal P/Q and N-type VGCC in bladder dysfunctions, pointing out Phα1β as a promising alternative for treating complications associated with CPA-induced HC., (© 2014 The British Pharmacological Society.)

Adenosine triphosphate (ATP) plays a role in cell signaling. It was soon proposed that ATP activates ionotropic P2X receptors, exerting an influence on neurons as well as on glial cells. In addition to the fact that the activation of P2X and P2Y receptors can stimulate or inhibit the release of glutamate from rat hippocampal neurons, the release of ATP has been implicated in hippocampal long-term potentiation (LTP). Through different behavioral paradigms, this study aimed to investigate the participation of P2X7R in genetically modified (knockout (KO)) mice with the suppressed expression of this receptor and in the pharmacological blockage of this receptor in rats, as well as to evaluate the effect of environmental enrichment on potential mnemonic deficits. The results suggest that P2X7R participates in aversive memory processes: pharmacological blockage with the selective P2X7R antagonist, A-740003, in different time frames elicited dose-dependent impairments in memory acquisition, consolidation and retrieval in rats that were submitted to the contextual fear-conditioning (FC) task, and the deletion of P2X7R hampered the aversive memory processes of mice that were subjected to the FC paradigm. Experiments using mice that were subjected to environmental enrichment suggest that this form of stimulation reverses mnemonic impairments that are ascribed to the absence of the P2X7R, suggesting that these receptors do not participate on such a reversal. Finally, no alterations were observed in the habituation memory of P2X7KO mice., (Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.)

Gliomas are the most common malignant brain tumors in adults. Bradykinin (BK) displays an important role in cancer, although the exact role of kinin receptors in the glioma biology remains unclear. This study investigated the role of kinin B1 and B2 receptors (B1R and B2R) on cell proliferation in human glioblastoma cell lineages. The mRNA expression of B1R and B2R was verified by RT-qPCR, whereas the effects of kinin agonists (des-Arg(9)-BK and BK) were analyzed by cell counting, MTT assay and annexin-V/PI determination. The PI3K/Akt and ERK1/2 signaling activation was assessed by flow cytometry. Our results demonstrated that both human glioblastoma cell lines U-138MG and U-251MG express functional B1R and B2R. The proliferative effects induced by the incubation of des-Arg(9)-BK and BK are likely related to the activation of PI3K/Akt and ERK 1/2 pathways. Moreover, the pre-incubation of the selective PI3Kγ blocker AS252424 markedly prevented kinin-induced AKT phosphorylation. Noteworthy, the selective B1R and B2R antagonists SSR240612 and HOE-140 were able to induce cell death of either lineages, with mixed apoptosis/necrosis characteristics. Taken together, the present results show that activation of B1R and B2R might contribute to glioblastoma progression in vitro. Furthermore, PI3K/Akt and ERK 1/2 signaling may be a target for adjuvant treatment of glioblastoma with a possible impact on tumor proliferation.

According to the World Health Organization, bladder cancer is the seventh most common cancer among men in the world. The current treatments for this malignancy are not efficient to prevent the recurrence and progression of tumors. Then, researches continue looking for better therapeutic targets which can end up in new and more efficient treatments. One of the recent findings was the identification that the purinergic system was involved in bladder tumorigenesis. The ectonucleotidases, mainly ecto-5'-nucleotidase/CD73 have been revealed as new players in cancer progression and malignity. In this work, we investigated the NTPDase3 and ecto-5'-nucleotidase/CD73 expression in cancer progression in vivo. Bladder tumor was induced in mice by the addition of 0.05 % of N-butyl-N-(hydroxybutyl)-nitrosamine (BBN) in the drinking water for 4, 8, 12, 18, and 24 weeks. After this period, mice bladders were removed for histopathology analysis and immunofluorescence assays. The bladder of animals which has received BBN had alterations, mainly inflammation, in initial times of tumor induction. After 18 weeks, mice's bladder has developed histological alterations similar to human transitional cell carcinoma. The cancerous urothelium, from mice that received BBN for 18 and 24 weeks, presented a weak immunostaining to NTPDase3, in contrast to an increased expression of ecto-5'-nucleotidase/CD73. The altered expression of NTPDase3 and ecto-5'-nucleotidase/CD73 presented herein adds further evidence to support the idea that alterations in ectonucleotidases are involved in bladder tumorigenesis and reinforce the ecto-5'-nucleotidase/CD73 as a future biomarker and/or a target for pharmacological therapy of bladder cancer.