RESUMO
Glioblastoma (GBM) is the most common and malignant primary brain cancer in adults. Without treatment the mean patient survival is approximately 6 months, which can be extended to 15 months with the use of multimodal therapies. The low effectiveness of GBM therapies is mainly due to the tumor infiltration into the healthy brain tissue, which depends on GBM cells' interaction with the tumor microenvironment (TME). The interaction of GBM cells with the TME involves cellular components such as stem-like cells, glia, endothelial cells, and non-cellular components such as the extracellular matrix, enhanced hypoxia, and soluble factors such as adenosine, which promote GBM's invasiveness. However, here we highlight the role of 3D patient-derived glioblastoma organoids cultures as a new platform for study of the modeling of TME and invasiveness. In this review, the mechanisms involved in GBM-microenvironment interaction are described and discussed, proposing potential prognosis biomarkers and new therapeutic targets.
Assuntos
Neoplasias Encefálicas , Glioblastoma , Adulto , Humanos , Glioblastoma/terapia , Glioblastoma/patologia , Neoplasias Encefálicas/terapia , Neoplasias Encefálicas/patologia , Células Endoteliais/patologia , Encéfalo/patologia , Matriz Extracelular/patologia , Microambiente Tumoral , Linhagem Celular TumoralRESUMO
Glioblastoma (GBM) is the most common and aggressive type of brain tumor due to its elevated recurrence following treatments. This is mainly mediated by a subpopulation of cells with stemness traits termed glioblastoma stem-like cells (GSCs), which are extremely resistant to anti-neoplastic drugs. Thus, an advancement in the understanding of the molecular processes underlying GSC occurrence should contribute significantly towards progress in reducing aggressiveness. High levels of endothelin-converting enzyme-1 (ECE1), key for endothelin-1 (ET-1) peptide activation, have been linked to the malignant progression of GBM. There are four known isoforms of ECE1 that activate ET-1, which only differ in their cytoplasmic N-terminal sequences. Isoform ECE1c is phosphorylated at Ser-18 and Ser-20 by protein kinase CK2, which increases its stability and hence promotes aggressiveness traits in colon cancer cells. In order to study whether ECE1c exerts a malignant effect in GBM, we designed an ECE1c mutant by switching a putative ubiquitination lysine proximal to the phospho-serines Lys-6-to-Arg (i.e., K6R). This ECE1cK6R mutant was stably expressed in U87MG, T98G, and U251 GBM cells, and their behavior was compared to either mock or wild-type ECE1c-expressing clone cells. ECE1cK6R behaved as a highly stable protein in all cell lines, and its expression promoted self-renewal and the enrichment of a stem-like population characterized by enhanced neurospheroid formation, as well as increased expression of stem-like surface markers. These ECE1cK6R-derived GSC-like cells also displayed enhanced resistance to the GBM-related chemotherapy drugs temozolomide and gemcitabine and increased expression of the ABCG2 efflux pump. In addition, ECE1cK6R cells displayed enhanced metastasis-associated traits, such as the modulation of adhesion and the enhancement of cell migration and invasion. In conclusion, the acquisition of a GSC-like phenotype, together with heightened chemoresistance and invasiveness traits, allows us to suggest phospho-ECE1c as a novel marker for poor prognosis as well as a potential therapeutic target for GBM.
Assuntos
Glioblastoma , Humanos , Glioblastoma/metabolismo , Enzimas Conversoras de Endotelina/genética , Enzimas Conversoras de Endotelina/metabolismo , Linhagem Celular Tumoral , Células-Tronco Neoplásicas/patologia , FenótipoRESUMO
Cryptocarya alba (Peumo; CA) and Laurelia sempervirens (Laurel; LS) are herbs native to the Chilean highlands and have historically been used for medicinal purposes by the Huilliches people. In this work, the essential oils were extracted using hydrodistillation in Clevenger apparatus and analyzed by GC-MS to determine their composition. The antioxidant capacity (AC) was evaluated in vitro. The cytotoxicity was determined using cell line cultures both non tumoral and tumoral. The toxicity was determined using the nematode Caenorhabditis elegans. The antimicrobial activity was evaluated against 52 bacteria using the agar disc diffusion method and the minimum inhibitory concentrations (MICs) were determined. The principal compounds found in C. alba essential oil (CA_EO) were α-terpineol (24.96%) and eucalyptol (21.63%) and were isazafrol (91.9%) in L. sempervirens essential oil (LS_EO). Both EOs showed antioxidant capacity in vitro. Both EO showed antibacterial activity against bacteria using. LS_EO showed more inhibitory effect on these cell lines respect to CA_EO. Both EOs showed toxicity against the nematode C.elegans at 3.12-50 mg/mL. The essential oils of CA and LS have an important bioactive potential in their antioxidant, antibacterial and cytotoxicity activity. Both essential oils could possibly be used in the field of natural medicine, natural food preservation, cosmetics, sanitation and plaguicides among others.
Assuntos
Antibacterianos/farmacologia , Antifúngicos/farmacologia , Antineoplásicos/farmacologia , Antioxidantes/farmacologia , Cryptocarya/química , Óleos Voláteis/farmacologia , Extratos Vegetais/farmacologia , Animais , Bactérias/efeitos dos fármacos , Bactérias/crescimento & desenvolvimento , Caenorhabditis elegans/efeitos dos fármacos , Caenorhabditis elegans/crescimento & desenvolvimento , Proliferação de Células , Humanos , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Células Tumorais CultivadasRESUMO
Diabetic nephropathy (DN) is considered the main cause of kidney disease in which myofibroblasts lead to renal fibrosis. Macrophages were recently identified as the major source of myofibroblasts in a process known as macrophage-myofibroblast transition (MMT). Adenosine levels increase during DN and in vivo administration of MRS1754, an antagonist of the A2B adenosine receptor (A2BAR), attenuated glomerular fibrosis (glomerulosclerosis). We aimed to investigate the association between A2BAR and MMT in glomerulosclerosis during DN. Kidneys/glomeruli of non-diabetic, diabetic, and MRS1754-treated diabetic (DM+MRS1754) rats were processed for histopathologic, transcriptomic, flow cytometry, and cellular in vitro analyses. Macrophages were used for in vitro cell migration/transmigration assays and MMT studies. In vivo MRS1754 treatment attenuated the clinical and histopathological signs of glomerulosclerosis in DN rats. Transcriptomic analysis demonstrated a decrease in chemokine-chemoattractants/cell-adhesion genes of monocytes/macrophages in DM+MRS1754 glomeruli. The number of intraglomerular infiltrated macrophages and MMT cells increased in diabetic rats. This was reverted by MRS1754 treatment. In vitro cell migration/transmigration decreased in macrophages treated with MRS1754. Human macrophages cultured with adenosine and/or TGF-ß induced MMT, a process which was reduced by MRS1754. We concluded that pharmacologic blockade of A2BAR attenuated some clinical signs of renal dysfunction and glomerulosclerosis, and decreased intraglomerular macrophage infiltration and MMT in DN rats.
Assuntos
Nefropatias Diabéticas/metabolismo , Nefropatias Diabéticas/patologia , Macrófagos/patologia , Monócitos/patologia , Miofibroblastos/patologia , Receptor A2B de Adenosina/metabolismo , Acetamidas/farmacologia , Antagonistas do Receptor A2 de Adenosina/farmacologia , Animais , Biomarcadores/metabolismo , Moléculas de Adesão Celular/metabolismo , Quimiocinas/metabolismo , Fatores Quimiotáticos/farmacologia , Fibrose , Humanos , Glomérulos Renais/efeitos dos fármacos , Glomérulos Renais/patologia , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Miofibroblastos/efeitos dos fármacos , Miofibroblastos/metabolismo , Purinas/farmacologia , Ratos Sprague-Dawley , Transcrição Gênica/efeitos dos fármacosRESUMO
Progressive diabetic nephropathy (DN) and loss of renal function correlate with kidney fibrosis. Crosstalk between TGF-ß and adenosinergic signaling contributes to the phenotypic transition of cells and to renal fibrosis in DN models. We evaluated the role of TGF-ß on NT5E gene expression coding for the ecto-5`-nucleotidase CD73, the limiting enzyme in extracellular adenosine production. We showed that high d-glucose may predispose HK-2 cells towards active transcription of the proximal promoter region of the NT5E gene while additional TGF-ß results in full activation. The epigenetic landscape of the NT5E gene promoter was modified by concurrent TGF-ß with occupancy by the p300 co-activator and the phosphorylated forms of the Smad2/3 complex and RNA Pol II. Transcriptional induction at NT5E in response to TGF-ß was earlier compared to the classic responsiveness genes PAI-1 and Fn1. CD73 levels and AMPase activity were concomitantly increased by TGF-ß in HK-2 cells. Interestingly, we found increased CD73 content in urinary extracellular vesicles only in diabetic patients with renal repercussions. Further, CD73-mediated AMPase activity was increased in the urinary sediment of DN patients. We conclude that the NT5E gene is a target of the profibrotic TGF-ß cascade and is a traceable marker of progressive DN.
Assuntos
5'-Nucleotidase/genética , Nefropatias Diabéticas/genética , Fibrose/genética , Fator de Crescimento Transformador beta/genética , Adenosina/biossíntese , Biomarcadores/metabolismo , Linhagem Celular , Nefropatias Diabéticas/patologia , Proteína p300 Associada a E1A/genética , Epigênese Genética/genética , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Fibrose/patologia , Proteínas Ligadas por GPI/genética , Regulação da Expressão Gênica , Humanos , Rim/metabolismo , Rim/patologia , Túbulos Renais Proximais/metabolismo , Túbulos Renais Proximais/patologia , Nucleotidases/genética , Regiões Promotoras Genéticas/genética , RNA Polimerase II/genéticaRESUMO
It has been argued that the central nervous system relies on combining simple movement elements (i.e. motor primitives) to generate complex motor outputs. However, how movement elements are generated and combined during the acquisition of new motor skills is still a source of debate. Herein, we present results providing new insights into the role of movement elements in the acquisition of motor skills that we obtained by analysing kinematic data collected while healthy subjects learned a new motor task. The task consisted of playing an interactive game using a platform with embedded sensors whose aggregate output was used to control a virtual object in the game. Subjects learned the task over multiple blocks. The analysis of the kinematic data was carried out using a recently developed technique referred to as "movement element decomposition." The technique entails the decomposition of complex multi-dimensional movements in one-dimensional elements marked by a bell-shaped velocity profile. We computed the number of movement elements during each block and measured how closely they matched a theoretical velocity profile derived by minimizing a cost function accounting for the smoothness of movement and the cost of time. The results showed that, in the early stage of motor skill acquisition, two mechanisms underlie the improvement in motor performance: 1) a decrease in the number of movement elements composing the motor output and 2) a gradual change in the movement elements that resulted in a shape matching the velocity profile derived by using the above-mentioned theoretical model.
Assuntos
Destreza Motora , Movimento , Fenômenos Biomecânicos , AprendizagemRESUMO
Glioblastoma is the brain tumor with the worst prognosis. This is mainly due to a cell subpopulation with an extremely aggressive potential, called glioblastoma stem-like cells (GSCs). These cells produce high levels of extracellular adenosine, which are increased even more under hypoxic conditions. Under hypoxia, adenosine signaling is related to HIF-2α expression, enhancing cell aggressiveness. Adenosine can be degraded using recombinant adenosine deaminase (ADA) to revert its pathological effects. The aim of this study was to degrade adenosine using ADA in order to decrease malignancy of GSCs. Adenosine depletion was performed using recombinant ADA. Migration and invasion were measured by transwell and matrigel-coated transwell assay, respectively. HIF-2α-dependent cell migration/invasion decreased in GSCs treated with ADA under hypoxia. MRPs-mediated chemoresistance and colony formation decreased in treatment with ADA. In conclusion, adenosine depletion using adenosine deaminase decreases GSCs aggressiveness.
Assuntos
Adenosina/deficiência , Neoplasias Encefálicas/patologia , Movimento Celular , Proliferação de Células , Resistencia a Medicamentos Antineoplásicos , Glioblastoma/patologia , Células-Tronco Neoplásicas/patologia , Adenosina/metabolismo , Antineoplásicos Fitogênicos/farmacologia , Apoptose , Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/metabolismo , Adesão Celular , Glioblastoma/tratamento farmacológico , Glioblastoma/metabolismo , Humanos , Hipóxia , Invasividade Neoplásica , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/metabolismo , Células Tumorais Cultivadas , Vincristina/farmacologiaRESUMO
Diabetic nephropathy (DN) is the main cause of end-stage renal disease, which remains incurable. The progression of DN is associated with progressive and irreversible renal fibrosis and also high levels of adenosine. Our aim was to evaluate the effects of ADORA3 antagonism on renal injury in streptozotocin-induced diabetic rats. An ADORA3 antagonist that was administered in diabetic rats greatly inhibited the levels of inflammatory interleukins IL-1ß and IL-18, meanwhile when adenosine deaminase was administered, there was a non-selective attenuation of the inflammatory mediators IL-1ß, IL-18, IL-6, and induction of IL-10. The ADORA3 antagonist attenuated the high glucose-induced activation of caspase 1 in HK2 cells in vitro. Additionally, ADORA3 antagonisms blocked the increase in caspase 1 and the nuclear localization of NFκB in the renal tubular epithelium of diabetic rats, both events that are involved in regulating the production and activation of IL-1ß and IL-18. The effects of the A3 receptor antagonist resulted in the attenuation of kidney injury, as evidenced by decreased levels of the pro-fibrotic marker α-SMA at histological levels and the restoration of proteinuria in diabetic rats. We conclude that ADORA3 antagonism represents a potential therapeutic target that mechanistically works through the selective blockade of the NLRP3 inflammasome.
Assuntos
Antagonistas do Receptor A3 de Adenosina/administração & dosagem , Caspase 1/metabolismo , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/tratamento farmacológico , Nefropatias Diabéticas/prevenção & controle , Antagonistas do Receptor A3 de Adenosina/farmacologia , Adenosina Desaminase/efeitos adversos , Animais , Linhagem Celular , Diabetes Mellitus Experimental/enzimologia , Nefropatias Diabéticas/induzido quimicamente , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Injeções Intraperitoneais , Interleucina-18/metabolismo , Interleucina-1beta/metabolismo , Túbulos Renais/efeitos dos fármacos , Túbulos Renais/enzimologia , Masculino , Ratos , EstreptozocinaRESUMO
Glioblastoma is a highly aggressive brain tumor, characterized by the formation of dysfunctional blood vessels and a permeable endothelial barrier. S-nitrosylation, a post-translational modification, has been identified as a regulator of endothelial function. In this work we explored whether S-nitrosylation induced by glioblastoma tumors regulates the endothelial function. As proof of concept, we observed that S-nitrosylation is present in the tumoral microenvironment of glioblastoma in two different animal models. Subsequently, we measured S nitrosylation and microvascular permeability in EAhy296 endothelial cells and in cremaster muscle. In vitro, conditioned medium from the human glioblastoma cell line U87 activates endothelial nitric oxide synthase, causes VE-cadherin- S-nitrosylation and induces hyperpermeability. Blocking Interleukin-8 (IL-8) in the conditioned medium inhibited S-nitrosylation of VE-cadherin and hyperpermeability. Recombinant IL-8 increased endothelial permeability by activating eNOS, S-nitrosylation of VE-cadherin and p120, internalization of VE-cadherin and disassembly of adherens junctions. In vivo, IL-8 induced S-nitrosylation of VE-cadherin and p120 and conditioned medium from U87 cells caused hyperpermeability in the mouse cremaster muscle. We conclude that eNOS signaling induced by glioma cells-secreted IL-8 regulates endothelial barrier function in the context of glioblastoma involving S-nitrosylation of VE-cadherin and p120. Our results suggest that inhibiting S-nitrosylation may be an effective way to control and/or block damage to the endothelial barrier and prevent cancer progression.
RESUMO
OBJECTIVE: To assess the impact of tDCS on posture, gait and coordination of movements in subjects with cerebellar ataxia. PATIENTS AND METHODS: This is a two-phase, double blind, auto matched, pilot study. Seven people were selected to participate in the study aged from 14 to 57. tDCS and sham-tDCS were applied at different times to all participants for 40 min over five consecutive days so that they were blind to which of the two techniques was applied at any one time. The area stimulated was the bilateral motor cortex. Subjects were evaluated before and after the interventions using the Scale for Assessment and Rating of Ataxia (SARA) and specific tests to measure posture and balance were carried out using the Wii Fit platform and CvMob software. RESULTS: The study indicates a statistically significant improvement in respect of gait parameters and the total score of the SARA scale and Wii Fit platform after tDCS when compared with data obtained from sham-tDCS trials (p: 0,03). The adverse events relating to tDCS were all self-limiting and from mild to moderate intensity. CONCLUSION: Despite the small sample size, tDCS showed positive results in some motor parameters and could be considered a valuable new option for the treatment of cerebellar ataxias.