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PorGal8, a porphyrin conjugated with dendritic galactose units, binds to galectin-1 in bladder cancer cells and induces localized cell death after light activation. Although it has been previously shown that photodynamic treatment (PDT) affects the cytoskeleton of cancer cells, it is still unclear how this change contributes to PDT-induced cell death. In this work, the association between changes in the cytoskeletal constituents and cell death triggered by PDT with PorGal8 was investigated in two bladder cancer cell lines derived from transitional cell carcinoma (UM-UC-3 and HT-1376 cells). Photoactivated PorGal8 did not change [Formula: see text]-tubulin protein levels in UM-UC-3 cells but reduced [Formula: see text]-tubulin in HT-1376 cells. A significant decrease in vimentin protein levels was exhibited in both cell lines 24 hours after irradiation. In the initial post-irradiation stage, both cell lines showed changes in actin filaments, but only recovery was apparent in HT-1376 cells 24 hours after treatment. In cells expressing higher levels of galectin-1 (UM-UC-3), PDT did not significantly affect these protein levels. Interestingly, 24 hours after irradiation, there was a robust increase in galectin-1 levels in HT-1376 cells. A small GTPases family protein, RhoA, involved in the galectin-1 expression, was also evaluated, indicating an increase in HT-1376 cells 24 hours after therapy. Overall, our results bring new insights into the relationship between the phototoxic effects of PorGal8 and the disorganization of the cytoskeleton. Clarifying the mechanisms underlying PDT efficiency might contribute to envisaging new potential therapeutic adjuvants for PDT, acting on the cytoskeleton, to treat resistant cancers.

Lung metastasis represents the leading cause of osteosarcoma-related death. Progress in preventing lung metastasis is pretty modest due to the inherent complexity of the metastatic process and the lack of suitable models. Herein, we provide mechanistic insights into how osteosarcoma systemically reprograms the lung microenvironment for metastatic outgrowth using metastatic mouse models and a multi-omics approach.We found that osteosarcoma-bearing mice or those preconditioned with cell-secretome harbour profound lung structural alteration with airways damage, inflammation, neutrophil infiltration, and remodelling of the extracellular matrix with deposition of fibronectin and collagen by stromal activated fibroblasts for tumour cell adhesion. These changes, supported by transcriptomic and histological data, promoted and accelerated the development of lung metastasis. Comparative proteome profiling of the cell secretome and mouse plasma identified a large number of proteins engaged in the extracellular-matrix organization, cell-matrix adhesion, neutrophil degranulation, and cytokine-mediated signalling, which were consistent with the observed lung microenvironmental changes. Moreover, we identified EFEMP1, a secreted extracellular matrix glycoprotein, as a potential risk factor for lung metastasis and a poor prognosis factor in osteosarcoma patients.

Objectives: To analyze, in patients with myelodysplastic syndrome (MDS) hypoplastic variant of the Bone Marrow Transplant service, the response to the transplant and perform a comparison with data from the MDS Latin American database. Methodology: Analytical, cross-sectional and retrospective study by analyzing medical records, including all patients diagnosed with MDS undergoing bone marrow transplantation, evaluation of post-transplantation follow-up, overall survival and complications. Comparison with data from the database of the MDS Latin American Society. Statistical analysis using the JAMOVI and MedCalc programs. Results: A total of 13 MDS patients received an allogenic bone marrow transplant, of which 6 had hypoplastic variant, corresponding to 46.15%. Compared to data from the Latin American database, this percentage is only 12.75%. Conclusions: Our service presented a higher frequency of hypoplastic variant MDS than that described in the medical literature, which corresponds to only 10-15%. Of the 6 patients with hypoplastic MDS, two died, reaching 33.3% against 13.5% in the Latin American database. The other transplanted patients are in remission so far with an overall 2-year survival rate of 83.3%, showing good results compared to the world literature. Objetivos: Analisar, nos pacientes com síndrome mielodisplásica (SMD) variante hipoplásica do serviço de transplante de medula óssea, a resposta ao transplante e realizar comparação com os dados do banco de dados Latino-Americano de SMD. Metodologia: Estudo analítico, corte transversal e retrospectivo por análise de prontuários, incluídos todos os pacientes com diagnóstico de SMD submetidos a transplante de medula óssea, avaliação do seguimento pós-transplante, sobrevida global e complicações. Comparação com dados do banco de dados da Sociedade Latino-Americana de SMD. Análise estatística pelo programa JAMOVI e MedCalc. Resultados: Um total de 13 pacientes com SMD receberam transplante de medula óssea alogênico, dos quais 6 apresentavam variante hipoplásica, correspondendo a 46,15%. Comparado aos dados do banco de dados latino-americano, essa porcentagem é de apenas 12,75%. Conclusões: Nosso serviço apresentou uma frequência maior de SMD variante hipoplásica que a descrita na literatura médica, que corresponde a apenas 10-15%. Dos 6 pacientes portadores de SMDh, dois evoluíram a óbito, chegando a 33,3% contra os 13,5% do banco de dados Latino-Americano. Os outros pacientes transplantados encontram-se em remissão até o presente momento com taxa de sobrevida global em 2 anos de 83,3%, apresentando bons resultados comparados à literatura mundial.

Lung metastases represent the most adverse clinical factor and rank as the leading cause of osteosarcoma-related death. Nearly 80% of patients present lung micrometastasis at diagnosis not detected with current clinical tools. Herein, an exosome (EX)-based imaging tool is developed for lung micrometastasis by positron emission tomography (PET) using osteosarcoma-derived EXs as natural nanocarriers of the positron-emitter copper-64 (

The pandemic of coronavirus disease (COVID)-19 is a global threat, causing high mortality, especially in the elderly. The main symptoms and the primary cause of death are related to interstitial pneumonia. Viral entry also into myocardial cells mainly via the angiotensin converting enzyme type 2 (ACE2) receptor and excessive production of pro-inflammatory cytokines, however, also make the heart susceptible to injury. In addition to the immediate damage caused by the acute inflammatory response, the heart may also suffer from long-term consequences of COVID-19, potentially causing a post-pandemic increase in cardiac complications. Although the main cause of cardiac damage in COVID-19 remains coagulopathy with micro- (and to a lesser extent macro-) vascular occlusion, open questions remain about other possible modalities of cardiac dysfunction, such as direct infection of myocardial cells, effects of cytokines storm, and mechanisms related to enhanced coagulopathy. In this opinion paper, we focus on these lesser appreciated possibilities and propose experimental approaches that could provide a more comprehensive understanding of the cellular and molecular bases of cardiac injury in COVID-19 patients. We first discuss approaches to characterize cardiac damage caused by possible direct viral infection of cardiac cells, followed by formulating hypotheses on how to reproduce and investigate the hyperinflammatory and pro-thrombotic conditions observed in the heart of COVID-19 patients using experimental in vitro systems. Finally, we elaborate on strategies to discover novel pathology biomarkers using omics platforms.

Cardiovascular diseases (CVDs) are the leading cause of mortality worldwide and, together with associated risk factors such as diabetes, hypertension, and dyslipidaemia, greatly impact patients’ quality of life and health care systems. This burden can be alleviated by fomenting lifestyle modifications and/or resorting to pharmacological approaches. However, due to several side effects, current therapies show low patient compliance, thus compromising their efficacy and enforcing the need to develop more amenable preventive/therapeutic strategies. In this scenario, medicinal and aromatic plants are a potential source of new effective agents. Specifically, plants from the Allioideae subfamily (formerly Alliaceae family), particularly those from the genus Allium and Tulbaghia, have been extensively used in traditional medicine for the management of several CVDs and associated risk factors, mainly due to the presence of sulphur-containing compounds. Bearing in mind this potential, the present review aims to gather information on traditional uses ascribed to these genera and provide an updated compilation of in vitro and in vivo studies validating these claims as well as clinical trials carried out in the context of CVDs. Furthermore, the effect of isolated sulphur-containing compounds is presented, and whenever possible, the relation between composition and activity and the mechanisms underlying the beneficial effects are pointed out.

The maintenance of tissue, organ, and organism homeostasis relies on an intricate network of players and mechanisms that assist in the different forms of cell-cell communication. Myocardial infarction, following heart ischemia and reperfusion, is associated with profound changes in key processes of intercellular communication, involving gap junctions, extracellular vesicles, and tunneling nanotubes, some of which have been implicated in communication defects associated with cardiac injury, namely arrhythmogenesis and progression into heart failure. Therefore, intercellular communication players have emerged as attractive powerful therapeutic targets aimed at preserving a fine-tuned crosstalk between the different cardiac cells in order to prevent or repair some of harmful consequences of heart ischemia and reperfusion, re-establishing myocardial function.

Summary Pulmonary arterial hypertension (PAH) is a rare, chronic disease of the pulmonary vasculature that is associated with poor outcomes. Its pathogenesis is multifactorial and includes micro-RNA (miRNA) deregulation. The understanding of the role of miRNAs in PAH is expanding quickly, and it is increasingly difficult to identify which miRNAs have the highest translational potential. This review summarizes the current knowledge of miRNA expression in PAH, discusses the challenges in miRNA analysis and interpretation, and highlights 4 promising miRNAs in this field (miR-29, miR-124, miR-140, and miR-204).

Glaucoma is a degenerative disease that causes irreversible loss of vision and is characterized by retinal ganglion cell (RGC) loss. Others and we have demonstrated that chronic neuroinflammation mediated by reactive microglial cells plays a role in glaucomatous pathology. Exosomes are extracellular vesicles released by most cells, including microglia, that mediate intercellular communication. The role of microglial exosomes in glaucomatous degeneration remains unknown. Taking the prominent role of microglial exosomes in brain neurodegenerative diseases, we studied the contribution of microglial-derived exosomes to the inflammatory response in experimental glaucoma. Microglial cells were exposed to elevated hydrostatic pressure (EHP), to mimic elevated intraocular pressure, the main risk factor for glaucoma. Naïve microglia (BV-2 cells or retinal microglia) were exposed to exosomes derived from BV-2 cells under EHP conditions (BV-Exo-EHP) or cultured in control pressure (BV-Exo-Control). We found that BV-Exo-EHP increased the production of pro-inflammatory cytokines, promoted retinal microglia motility, phagocytic efficiency, and proliferation. Furthermore, the incubation of primary retinal neural cell cultures with BV-Exo-EHP increased cell death and the production of reactive oxygen species. Exosomes derived from retinal microglia (MG-Exo-Control or MG-Exo-EHP) were injected in the vitreous of C57BL/6J mice. MG-Exo-EHP sustained activation of retinal microglia, mediated cell death, and impacted RGC number. Herein, we show that exosomes derived from retinal microglia have an autocrine function and propagate the inflammatory signal in conditions of elevated pressure, contributing to retinal degeneration in glaucomatous conditions.

Acute myocardial infarction (AMI) and the heart failure (HF) that often result remain the leading causes of death and disability worldwide. As such, new therapeutic targets need to be discovered to protect the myocardium against acute ischaemia/reperfusion (I/R) injury in order to reduce myocardial infarct (MI) size, preserve left ventricular function and prevent the onset of HF. Mitochondrial dysfunction during acute I/R injury is a critical determinant of cell death following AMI, and therefore, ion channels in the inner mitochondrial membrane, which are known to influence cell death and survival, provide potential therapeutic targets for cardioprotection. In this article, we review the role of mitochondrial ion channels, which are known to modulate susceptibility to acute myocardial I/R injury, and we explore their potential roles as therapeutic targets for reducing MI size and preventing HF following AMI.

New treatments are needed to protect the myocardium against the detrimental effects of acute ischaemia/reperfusion (IR) injury following an acute myocardial infarction (AMI), in order to limit myocardial infarct (MI) size, preserve cardiac function and prevent the onset of heart failure (HF). Given the critical role of mitochondria in energy production for cardiac contractile function, prevention of mitochondrial dysfunction during acute myocardial IRI may provide novel cardioprotective strategies. In this regard, the mitochondrial fusion and fissions proteins, which regulate changes in mitochondrial morphology, are known to impact on mitochondrial quality control by modulating mitochondrial biogenesis, mitophagy and the mitochondrial unfolded protein response. In this article, we review how targeting these inter‐related processes may provide novel treatment targets and new therapeutic strategies for reducing MI size, preventing the onset of HF following AMI.

PurposeTear fluid biomarkers may offer a non-invasive strategy for detecting diabetic patients with increased risk of developing diabetic retinopathy (DR) or increased disease progression, thus helping both improving diagnostic accuracy and understanding the pathophysiology of the disease. Here, we assessed the tear fluid of nondiabetic individuals, diabetic patients with no DR, and diabetic patients with nonproliferative DR (NPDR) or with proliferative DR (PDR) to find putative biomarkers for the diagnosis and staging of DR.MethodsTear fluid samples were collected using Schirmer test strips from a cohort with 12 controls and 54 Type 2 Diabetes (T2D) patients, and then analyzed using mass spectrometry (MS)-based shotgun proteomics and bead-based multiplex assay. Tear fluid-derived small extracellular vesicles (EVs) were analyzed by transmission electron microscopy, Western Blotting, and nano tracking.ResultsProteomics analysis revealed that among the 682 reliably quantified proteins in tear fluid, 42 and 26 were differentially expressed in NPDR and PDR, respectively, comparing to the control group. Data are available via ProteomeXchange with identifier PXD033101. By multicomparison analyses, we also found significant changes in 32 proteins. Gene ontology (GO) annotations showed that most of these proteins are associated with oxidative stress and small EVs. Indeed, we also found that tear fluid is particularly enriched in small EVs. T2D patients with NPDR have higher IL-2/-5/-18, TNF, MMP-2/-3/-9 concentrations than the controls. In the PDR group, IL-5/-18 and MMP-3/-9 concentrations were significantly higher, whereas IL-13 was lower, compared to the controls.ConclusionsOverall, the results show alterations in tear fluid proteins profile in diabetic patients with retinopathy. Promising candidate biomarkers identified need to be validated in a large sample cohort.

Heat shock protein 90 (HSP90) facilitates folding and stability and prevents the degradation of multiple client proteins. One of these HSP90 clients is BCR-ABL, the oncoprotein characteristic of chronic myeloid leukemia (CML) and the target of tyrosine kinase inhibitors, such as imatinib. Alvespimycin is an HSP90 inhibitor with better pharmacokinetic properties and fewer side effects than other similar drugs, but its role in overcoming imatinib resistance is not yet clarified. This work studied the therapeutic potential of alvespimycin in imatinib-sensitive (K562) and imatinib-resistant (K562-RC and K562-RD) CML cell lines. Metabolic activity was determined by the resazurin assay. Cell death, caspase activity, mitochondrial membrane potential, and cell cycle were evaluated by means of flow cytometry. Cell death was also analyzed by optical microscopy. HSPs expression levels were assessed by western blotting. Alvespimycin reduced metabolic activity in a time-, dose-, and cell line-dependent manner. Resistant cells were more sensitive to alvespimycin with an IC50 of 31 nM for K562-RC and 44 nM for K562-RD, compared to 50 nM for K562. This drug induced apoptosis via the mitochondrial pathway. In K562 cells, alvespimycin induced cell cycle arrest in G0/G1. As a marker of HSP90 inhibition, a significant increase in HSP70 expression was observed. Our results suggest that alvespimycin might be a new therapeutic approach to CML treatment, even in cases of resistance to imatinib.

Background Idiopathic pulmonary arterial hypertension (iPAH) is a rare and chronic disease associated with poor outcomes. Previously considered a disease restricted to the pulmonary circulation, PAH is now being recognized as a systemic disorder that is associated with metabolic dysfunction. The aim of this study is to analyze the metabolic reprogramming in the lung and peripheral blood mononuclear cell (PBMCs) of iPAH patients and explore their potential roles in PAH pathophysiology. Methods Five independent datasets, containing transcriptomic data of human PBMCs (GSE22356 and GSE33463) and lung (GSE48149 GSE113439 and GSE117261) samples, from 139 iPAH patients and 96 healthy controls, were downloaded at the GEO database. In each dataset, the samples were normalized and a pair-wise comparison between control and iPAH samples was performed using limma package, for the R programming language. Genes with a p-value lower than 0.05 were considered differentially expressed between the two groups. A subset of metabolism related genes was selected, and their expression was compared across the datasets. Results Among the 13 genes with differential expression identified, only 10 had a coherent expression across all datasets (Figure 1). Firstly, we report an association with insulin resistance through impairment of PI3K signaling in iPAH patients, by expressing lower levels of the heterodimer PIK3CD and regulatory PIK3IP1 and PIKR1 subunits in PBMCs, and by expressing higher levels of its downstream targets in the lung (TBC1D4). However, more extensive metabolic dysfunction was observed. A significant glycolytic shift in the lung and PBMCs was present, as a consequence of deregulation in genes involved in aerobic glycolysis and decreased fatty acid oxidation, namely increased expression of PD1K and lower levels of expression of LDHB. The findings of decreased SLC25A1 protein in both PBMCs and lung suggest impairment of the tricarboxylic acid (TCA) cycle flux in PAH. Additionally, SLC1A5 highlights the involvement of glutamine metabolism and glutaminolysis derangements in PAH. Conversely, SREBP1 is involved in sterol biosynthesis and lower levels in PMBCs results in impaired resolution of inflammatory responses. Finally, although the role of autophagy in iPAH is complex, higher levels of expression of ATG13 in PBMCs and lower levels in the lung confirm autophagy deregulation in iPAH. Interestingly, all the metabolic pathways identified (Figure 2) are hallmarks of the metabolic reprogramming seen in cancer cells, a finding already suggested by the clonal proliferation of pulmonary artery smooth muscle cells described in plexiform lesions. Conclusion Our results provide novel insights into the metabolic regulation in iPAH. Molecularly, these cells exhibit many features common to cancer cells, suggesting the opportunity to exploit therapeutic strategies used in cancer to treat iPAH. Funding Acknowledgement Type of funding sources: None.

Essential oils’ therapeutic potential is highly recognized, with many applications rising due to reported anti-inflammatory, cardioprotective, neuroprotective, anti-aging, and anti-cancer effects. Nevertheless, clinical translation still remains a challenge, mainly due to essential oils’ volatility and low water solubility and stability. The present review gathers relevant information and postulates on the potential application of plant nanovesicles to effectively deliver essential oils to target organs. Indeed, plant nanovesicles are emerging as alternatives to mammalian vesicles and synthetic carriers due to their safety, stability, non-toxicity, and low immunogenicity. Moreover, they can be produced on a large scale from various plant parts, enabling an easier, more rapid, and less costly industrial application that could add value to waste products and boost the circular economy. Importantly, the use of plant nanovesicles as delivery platforms could increase essential oils’ bioavailability and improve chemical stability while reducing volatility and toxicity issues. Additionally, using targeting strategies, essential oils’ selectivity, drug delivery, and efficacy could be improved, ultimately leading to dose reduction and patient compliance. Bearing this in mind, information on current pharmaceutical technologies available to enable distinct routes of administration of loaded vesicles is also discussed.

Background: Cardiovascular diseases are the leading cause of global mortality with a tendency to increase due to population ageing as well as an increase in associated risk factors. Although current therapies improve survival rates, they are associated with several side effects, thus justifying the development of novel preventive and/or therapeutic approaches. In this way, plant metabolites such as essential oils have emerged as promising agents due to their biological effects. Objective: Bearing in mind that several essential oils are characterized by high amounts of phenylpropanoids, which may play a crucial role in the activity of these volatile extracts, a comprehensive and systematic review focusing on the cardiovascular effects of phenylpropanoid-rich essential oils is presented. Methods: Popular search engines including PubMed, Science Direct, Scopus and Google Scholar were consulted and papers from 2000 onwards were selected. Non-volatile phenylpropanoids were not considered in this review. Results: A compilation of the current knowledge on this thematic pointed out beneficial effects for volatile phenylpropanoids namely hypotensive, vasorelaxant, antiplatelet aggregation, antidyslipidaemic and antidiabetic, as well as protective properties against ischemia/reperfusion injury and heart hypertrophy. Conclusion: A better understanding of the protective effects of phenylpropanoids on the cardiovascular system is presented, thus paving the way towards future research on plant-based therapies for cardiovascular diseases.

For the first time, the present study unravels a cardiospecific therapeutic approach for Pulmonary Arterial Hypertension (PAH), a disease with a very poor prognosis and high mortality rates due to right ventricle (RV) dysfunction. We first established a new in vitro model of high-pressure-induced hypertrophy that closely resembles heart defects associated with PAH and validated our in vitro findings on a preclinical in vivo model of monocrotaline (MCT)-induced PAH. Our results showed the in vitro antihypertrophic effect of 1,8-cineole, a monoterpene widely found in several essential oils. Also, a decrease in RV hypertrophy and fibrosis, and an improvement in heart function in vivo was observed, when 1,8-cineole was applied topically. Furthermore, 1,8-cineole restored gap junction protein connexin43 distribution at the intercalated disks and mitochondrial functionality, suggesting it may act by preserving cardiac cell-to-cell communication and bioenergetics. Overall, our results point out a promising therapeutic compound that can be easily applied topically, thus paving the way for the development of effective cardiac-specific therapies to greatly improve PAH outcomes.

A well-balanced intercellular communication between the different cells within the heart is vital for the maintenance of cardiac homeostasis and function. Despite remarkable advances on disease management and treatment, acute myocardial infarction remains the major cause of morbidity and mortality worldwide. Gold standard reperfusion strategies, namely primary percutaneous coronary intervention, are crucial to preserve heart function. However, reestablishment of blood flow and oxygen levels to the infarcted area are also associated with an accumulation of reactive oxygen species (ROS), leading to oxidative damage and cardiomyocyte death, a phenomenon termed myocardial reperfusion injury. In addition, ROS signaling has been demonstrated to regulate multiple biological pathways, including cell differentiation and intercellular communication. Given the importance of cell-cell crosstalk in the coordinated response after cell injury, in this review, we will discuss the impact of ROS in the different forms of inter- and intracellular communication, as well as the role of gap junctions, tunneling nanotubes and extracellular vesicles in the propagation of oxidative damage in cardiac diseases, particularly in the context of ischemia/reperfusion injury.

Simple Summary The gap junction protein connexin 43 (Cx43) facilitates direct intercellular communication and displays a complex dichotomous role in cancer. Here, we used breast cancer cell lines and CRISPR/Cas9 technology to dissect some of the poorly understood multimodal functions of Cx43 in this setting. Our data show that Cx43 directly modulates the formation of pro-tumorigenic tunneling nanotubes (TNTs), which are cellular protrusions that connect non-adjacent cells to facilitate cellular communication, including the direct cell–cell transfer of organelles. We identify several important cancer signaling pathways that affect TNT formation and show that this effect is strongly influenced by the presence or absence of Cx43. Finally, we demonstrate that Cx43 can affect TNT formation by modulating the cellular secretome. This work provides important insight into the pro-tumorigenic role of Cx43 and its interconnections with TNTs. Abstract Connexin 43 (Cx43) forms gap junctions that mediate the direct intercellular diffusion of ions and small molecules between adjacent cells. Cx43 displays both pro- and anti-tumorigenic properties, but the mechanisms underlying these characteristics are not fully understood. Tunneling nanotubes (TNTs) are long and thin membrane projections that connect cells, facilitating the exchange of not only small molecules, but also larger proteins, organelles, bacteria, and viruses. Typically, TNTs exhibit increased formation under conditions of cellular stress and are more prominent in cancer cells, where they are generally thought to be pro-metastatic and to provide growth and survival advantages. Cx43 has been described in TNTs, where it is thought to regulate small molecule diffusion through gap junctions. Here, we developed a high-fidelity CRISPR/Cas9 system to knockout (KO) Cx43. We found that the loss of Cx43 expression was associated with significantly reduced TNT length and number in breast cancer cell lines. Notably, secreted factors present in conditioned medium stimulated TNTs more potently when derived from Cx43-expressing cells than from KO cells. Moreover, TNT formation was significantly induced by the inhibition of several key cancer signaling pathways that both regulate Cx43 and are regulated by Cx43, including RhoA kinase (ROCK), protein kinase A (PKA), focal adhesion kinase (FAK), and p38. Intriguingly, the drug-induced stimulation of TNTs was more potent in Cx43 KO cells than in wild-type (WT) cells. In conclusion, this work describes a novel non-canonical role for Cx43 in regulating TNTs, identifies key cancer signaling pathways that regulate TNTs in this setting, and provides mechanistic insight into a pro-tumorigenic role of Cx43 in cancer.

Ubiquitin signals the sorting of Cx43 into EVs, which is impaired during ischemia, resulting in decreased levels of Cx43 in cardiomyocyte-derived and circulating EVs from myocardial infarcted patients., Ischemic heart disease has been associated with an impairment on intercellular communication mediated by both gap junctions and extracellular vesicles. We have previously shown that connexin 43 (Cx43), the main ventricular gap junction protein, assembles into channels at the extracellular vesicle surface, mediating the release of vesicle content into target cells. Here, using a comprehensive strategy that included cell-based approaches, animal models and human patients, we demonstrate that myocardial ischemia impairs the secretion of Cx43 into circulating, intracardiac and cardiomyocyte-derived vesicles. In addition, we show that ubiquitin signals Cx43 release in basal conditions but appears to be dispensable during ischemia, suggesting an interplay between ischemia-induced Cx43 degradation and secretion. Overall, this study constitutes a step forward for the characterization of the signals and molecular players underlying vesicle protein sorting, with strong implications on long-range intercellular communication, paving the way towards the development of innovative diagnostic and therapeutic strategies for cardiovascular disorders.

Introduction Age-related macular degeneration (AMD) is a degenerative retinal disease that affects central vision. Most of their phenotypical features are believed to be associated with the dysfunction of retinal pigment epithelium (RPE). The accumulation of damaged proteins in aged RPE is associated with disruption of proteolytic pathways and exocytic activity, with release of intracellular proteins via exosomes (Exo), that are important players in intercellular communication and can contribute to disease progression. However, the impact of their secretion by polarized RPE on outer blood retinal barrier (oBRB) breakdown remains largely elusive Objectives Our aim was to explore the role of inflammation on the loss of RPE integrity and to understand the relative role of directional secretion of Exo by RPE in the loss of polarity and barrier disruption Methodology We used a human RPE cell line (ARPE-19), highly polarized RPE primary cultures (pRPE) and porcine eyecups. To mimic the inflammatory conditions present in AMD, cells were treated with two inflammatory stimuli, TNF (10 ng/mL) or LPS (100 ng/mL) Results TNF and LPS do not affect the viability of the RPE cells. RPE cells developed a confluent monolayer and reached a relatively constant TER of about 40 Ω/cm2 (ARPE-19) or higher than 150 Ω/cm2 (pRPE). Treatment with TNF significantly reduces the TER, decreased immunoreactivity and co-localization of the TJ proteins ZO-1 and occludin and increases MMP-2/-9 activity in the medium. Apical Exo isolated from the RPE cells are enriched in CD63 compared to the basolateral Exo, that are enriched in CD81. The Exo isolated from porcine eyecups, especially with the LPS stimulus, are enriched in CD81 and MMP-2 but have similar levels of CD63 Conclusion Overall, our results show that inflammation induces loss of RPE integrity and release of different populations of Exo. The unravelling of novel drug targets paves the way for development of new therapeutic strategies for AMD.

Photodynamic therapy (PDT) has demonstrated encouraging anticancer therapeutic results, but the current clinically approved photosensitizers (PSs) are not ideal in the treatment of bladder cancer. Conventional PSs have low selectivity to the bladder tumor tissue and induce toxicity or bystander effects on nontumor urothelium. Previous studies demonstrated that the use of galactose-photosensitizer (PS) conjugates is a more selective method of delivering PDT-mediated toxicity due to their ability to recognize carbohydrate-binding domains overexpressed in bladder tumors. Using patient-derived bladder tumor specimens cultured ex vivo and bladder cancer cell lines with different PDT sensitivity, we find that a galactose-phthalocyanine (PcGal16) accumulates in bladder tumors expressing galactose-binding proteins and internalizes through an endocytic process. The endocytosis mechanism is cell line-dependent. In HT-1376 bladder cancer lines resistant to PDT, depletion of caveolin-1-the main structural protein of caveolae structures-increased the amount of sugar-binding proteins, i.e. GLUT1, at the cell membrane resulting in an improved PcGal16 uptake and PDT efficacy. These data show the potential of ex vivo cultures of bladder cancer, that ideally could mimic the original microenvironment, in screening galacto-PDT agents. Additionally, our studies demonstrate that PDT efficacy in bladder cancer depends on the endocytic mechanisms that regulate PS accumulation and internalization in cancer cells.

Rationale: Efficient communication between heart cells is vital to ensure the anisotropic propagation of electrical impulses, a function mainly accomplished by gap junctions (GJ) composed of Cx43 (connexin 43). Although the molecular mechanisms remain unclear, altered distribution and function of gap junctions have been associated with acute myocardial infarction and heart failure. Objective: A recent proteomic study from our laboratory identified EHD1 (Eps15 [endocytic adaptor epidermal growth factor receptor substrate 15] homology domain-containing protein 1) as a novel interactor of Cx43 in the heart. Methods and Results: In the present work, we demonstrate that knockdown of EHD1 impaired the internalization of Cx43, preserving gap junction-intercellular coupling in cardiomyocytes. Interaction of Cx43 with EHD1 was mediated by Eps15 and promoted by phosphorylation and ubiquitination of Cx43. Overexpression of wild-type EHD1 accelerated internalization of Cx43 and exacerbated ischemia-induced lateralization of Cx43 in isolated adult cardiomyocytes. In addition, we show that EHDs associate with Cx43 in human and murine failing hearts. Conclusions: Overall, we identified EHDs as novel regulators of endocytic trafficking of Cx43, participating in the pathological remodeling of gap junctions, paving the way to innovative therapeutic strategies aiming at preserving intercellular communication in the heart.

Background: Despite evidence supporting the overall efficacy of mechanical thrombectomy (MT) in acute ischemic stroke (AIS) due to large vessel occlusion (LVO) of the anterior circulation, it is unclear whether the treatment effect of MT differs by sex in different populations. We assessed the impact of sex differences in the treatment effect of MT in the RESILIENT trial. Methods: RESILIENT was a prospective, multicenter, randomized phase III trial that was designed to assess the safety, efficacy, and cost-effectiveness of mechanical thrombectomy as compared to medical treatment alone in patients treated under the less than ideal conditions typically found in the public healthcare system of a developing country. Results: Among 221 patients enrolled in the trial, 104 (47,1%) were female. Baseline characteristics were well balanced between sexes, except for a higher prevalence of hypertension (76% vs. 57.4%; p=0.004) and diabetes (34.3% vs. 21.7%; p=0.039) and a lower frequency of alcohol abuse (4% vs. 28.9%; p=0.001) in females. After adjustment for baseline characteristics, we found a significant interaction (p=0.026) between sex and the effect of MT with a lower efficacy of MT for functional independency at 90 days among women (aOR=1.13;95%CI:0.42-3.02) compared to men (aOR=4.78; 95%CI:1.88-12.15). Conclusions: In our study population of patients with AIS caused by LVO of the anterior circulation, women were less likely to benefit from MT than men. Further studies are necessary to investigate these findings.

Given the low mitotic activity of cardiomyocytes, the contractile unit of the heart, these cells strongly rely on efficient and highly regulated mechanisms of protein degradation to eliminate unwanted potentially toxic proteins. This is particularly important in the context of disease, where an impairment of protein quality control mechanisms underlies the onset and development of diverse cardiovascular maladies. One of the biological processes which is tightly regulated by proteolysis mechanisms is intercellular communication. The different types of cells that form the heart, including cardiomyocytes, endothelial cells, fibroblasts, and macrophages, can communicate directly, through gap junctions (GJ) or tunneling nanotubes (TNT), or at long distances, via extracellular vesicles (EV) or soluble factors.

O propósito deste artigo é demonstrar, de forma sucinta, os caminhos a trilhar pela comunicação de ciência, mais especificamente pela comunicação em saúde, em termos institucionais e no contexto de uma Escola Médica. Entende-se que esta área tem sido desvalorizada, mas assume, atualmente, uma enorme preponderância no sentido de uma responsabilização por parte da sociedade, dado que é o elo entre comunicação científica e o público não especializado. Assim, seguindo algumas tendências recentes, propõe-se que uma Escola Médica não deve descurar esta área de estudo, mas antes reforçar a aposta, assentando-a em quarto valências fundamentais - formação, educação cívica, promoção e divulgação, e inovação e investigação, - formando melhores profissionais e aproximando a sociedade daquilo que são as suas áreas de investigação.

Acute myeloid leukaemia (AML) comprises a heterogeneous group of hematologic neoplasms characterized by diverse combinations of genetic, phenotypic and clinical features representing a major challenge for the development of targeted therapies. Metabolic reprogramming, mainly driven by deregulation of the nutrient-sensing pathways as AMPK, mTOR and PI3K/AKT, has been associated with cancer cells, including AML cells, survival and proliferation. Nevertheless, the role of these metabolic adaptations on the AML pathogenesis is still controversial. In this work, the metabolic status and the respective metabolic networks operating in different AML cells (NB-4, HL-60 and KG-1) and their impact on autophagy and survival was characterized. Data show that whereas KG-1 cells exhibited preferential mitochondrial oxidative phosphorylation metabolism with constitutive co-activation of AMPK and mTORC1 associated with increased autophagy, NB-4 and HL-60 cells displayed a dependent glycolytic profile mainly associated with AKT/mTORC1 activation and low autophagy flux. Inhibition of AKT is disclosed as a promising therapeutical target in some scenarios while inhibition of AMPK and mTORC1 has no major impact on KG-1 cells' survival. The results highlight an exclusive metabolic profile for each tested AML cells and its impact on determination of the anti-leukaemia efficacy and on personalized combinatory therapy with conventional and targeted agents., This work was developed under the scope of the project NORTE‐01‐0145‐FEDER‐000013, supported by the Northern Portugal Regional Operational Programme (NORTE 2020), under the Portugal 2020 Partnership Agreement, through the European Regional Development Fund (FEDER), and by FEDER, through the Competitiveness Factors Operational Programme (COMPETE), and by National funds, through the Foundation for Science and Technology (FCT), under the scope of the project POCI‐01‐0145‐FEDER‐007038. OP and BSM are supported by fellowships from the Fundação para a Ciência e Tecnologia (FCT, Portugal) (SFRH/BD/52292/2013 and SFRH/BPD/90533/2012, respectively), info:eu-repo/semantics/publishedVersion

Glioblastoma (GBM) is the most malignant primary brain tumor, with an average survival rate of 15 months. GBM is highly refractory to therapy, and such unresponsiveness is due, primarily, but not exclusively, to the glioma stem-like cells (GSCs). This subpopulation express stem-like cell markers and is responsible for the heterogeneity of GBM, generating multiple differentiated cell phenotypes. However, how GBMs maintain the balance between stem and non-stem populations is still poorly understood. We investigated the GBM ability to interconvert between stem and non-stem states through the evaluation of the expression of specific stem cell markers as well as cell communication proteins. We evaluated the molecular and phenotypic characteristics of GSCs derived from differentiated GBM cell lines by comparing their stem-like cell properties and expression of connexins. We showed that non-GSCs as well as GSCs can undergo successive cycles of gain and loss of stem properties, demonstrating a bidirectional cellular plasticity model that is accompanied by changes on connexins expression. Our findings indicate that the interconversion between non-GSCs and GSCs can be modulated by extracellular factors culminating on differential expression of stem-like cell markers and cell-cell communication proteins. Ultimately, we observed that stem markers are mostly expressed on GBMs rather than on low-grade astrocytomas, suggesting that the presence of GSCs is a feature of high-grade gliomas. Together, our data demonstrate the utmost importance of the understanding of stem cell plasticity properties in a way to a step closer to new strategic approaches to potentially eliminate GSCs and, hopefully, prevent tumor recurrence.

Introduction and Objectives: Cardiovascular disease is the leading cause of mortality and morbidity associated with diabetes. Although impairment of the cell response to hypoxia due to destabilization of the transcription factor hypoxia-inducible factor-1α (HIF-1α), which regulates the expression of genes that help cells to cope with low oxygen tension, has been implicated in diabetes-associated disease, the molecular mechanisms involved remain elusive. It is known that hyperglycemia leads to the enhanced production of methylglyoxal (MGO). Therefore, the main objective of this study was to establish whether MGO leads to the degradation of HIF-1α in cardiomyocytes subjected to hypoxia. Methods: The mouse atrial cardiomyocyte cell line, HL-1, was exposed to chemical hypoxia with CoCl2 in the absence or presence of MGO. Cell viability was assessed by MTT assay, and levels of HIF-1α and endogenous ubiquitin conjugates were determined by western blotting. Proteasome activity was analyzed using a specific chymotrypsin-like fluorogenic substrate. Results: The results obtained indicate that MGO induces time- and dose-dependent degradation of HIF-1α accumulated under hypoxia. Additionally, we show that accumulation of endogenous ubiquitin conjugates in the presence of MGO is associated with decreased proteasome activity. Conclusion: Taken together, the results obtained in this study suggest that MGO compromises the ability of cells to adapt to low oxygen tensions, by stimulating the degradation of HIF-1α, likely contributing to the development of diabetes-associated cardiac dysfunction. Resumo: Introdução e objetivos: As complicações cardiovasculares constituem a principal causa de morbimortalidade em doentes diabéticos. A destabilização do fator de transcrição induzido pela hipoxia (HIF-1α), que regula a expressão de genes de adaptação celular a condições de baixos níveis de oxigénio, parece comprometer a resposta celular de diversos tecidos à hipoxia, em condições de hiperglicemia. No entanto, os mecanismos moleculares subjacentes à destabilização do HIF-1α estão ainda por estabelecer. Está bem estabelecido que em situações de hiperglicemia ocorre o aumento da produção de metilglioxal (MGO). Assim, o objetivo deste trabalho foi estabelecer o impacto do MGO na degradação do HIF-1α em cardiomiócitos sujeitos a condições de hipoxia. Métodos: Uma linha celular de cardiomiócitos da aurícula de ratos, HL-1, foi sujeita a hipoxia química com CoCl2, na ausência e na presença de MGO. De seguida, avaliou-se a viabilidade celular pelo ensaio de MTT, e determinaram-se os níveis de HIF-1α e de conjugados endógenos de ubiquitina por western blotting. A atividade do proteosoma foi avaliada utilizando um substrato fluorogénico específico do tipo quimotripsina. Resultados: Os resultados obtidos mostram que o MGO induz degradação do HIF-1α em condições de hipoxia de uma forma dependente da dose de MGO utilizada, assim como do tempo de tratamento. Por outro lado, a acumulação de conjugados endógenos de ubiquitina na presença de MGO associa-se a um decréscimo na atividade do proteosoma. Conclusão: No conjunto, os resultados sugerem que o MGO compromete a capacidade de adaptação celular a condições de hipoxia através da degradação do HIF-1α, o que pode contribuir para o desenvolvimento da disfunção cardíaca associada à diabetes. Keywords: Diabetic heart disease, Hypoxia, Hypoxia-inducible factor-1α, Cardiomyocytes, Palavras-chave: Doenças cardiovasculares e diabetes, Hipoxia, HIF-1α, Cardiomiócitos

Glioblastoma (GBM) is a grade IV astrocytoma. GBM patients show resistance to chemotherapy such as temozolomide (TMZ), the gold standard treatment. In order to simulate the molecular mechanisms behind the different chemotherapeutic responses in GBM patients we compared the cellular heterogeneity and chemotherapeutic resistance mechanisms in different GBM cell lines. We isolated and characterized a human GBM cell line obtained from a GBM patient, named GBM11. We studied the GBM11 behaviour when treated with Tamoxifen (TMX) that, among other functions, is a protein kinase C (PKC) inhibitor, alone and in combination with TMZ in comparison with the responses of U87 and U118 human GBM cell lines. We evaluated the cell death, cell cycle arrest and cell proliferation, mainly through PKC expression, by flow cytometry and western blot analysis and, ultimately, cell migration capability and F-actin filament disorganization by fluorescence microscopy. We demonstrated that the constitutive activation of p-PKC seems to be one of the main metabolic implicated on GBM malignancy. Despite of its higher resistance, possibly due to the overexpression of P-glycoprotein and stem-like cell markers, GBM11 cells presented a subtle different chemotherapeutic response compared to U87 and U118 cells. The GBM11, U87, U118 cell lines show subtle molecular differences, which clearly indicate the characterization of GBM heterogeneity, one of the main reasons for tumor resistance. The adding of cellular heterogeneity in molecular behaviour constitutes a step closer in the understanding of resistant molecular mechanisms in GBM, and can circumvents the eventual impaired therapy.

Aims Connexin-based gap junctions are crucial for electrical communication in the heart; they are each composed of two docked hemichannels (HCs), supplied as unpaired channels via the sarcolemma. When open, an unpaired HC forms a large pore, high-conductance and Ca2+-permeable membrane shunt pathway that may disturb cardiomyocyte function. HCs composed of connexin 43 (Cx43), a major cardiac connexin, can be opened by electrical stimulation but only by very positive membrane potentials. Here, we investigated the activation of Cx43 HCs in murine ventricular cardiomyocytes voltage-clamped at −70 mV. Methods and results Using whole-cell patch-clamp, co-immunoprecipitation, western blot analysis, immunocytochemistry, proximity ligation assays, and protein docking studies, we found that stimulation of ryanodine receptors (RyRs) triggered unitary currents with a single-channel conductance of ∼220 pS, which were strongly reduced by Cx43 knockdown. Recordings under Ca2+-clamp conditions showed that both RyR activation and intracellular Ca2+ elevation were necessary for HC opening. Proximity ligation studies indicated close Cx43-RyR2 apposition ( Conclusion Our results demonstrate that Cx43 HCs are intimately linked to RyRs, allowing them to open at negative diastolic membrane potential in response to RyR activation.

Connexin43 (Cx43) is commonly associated with direct cell-cell communication through gap junctions (GJs). However, recent groundbreaking studies have challenged this dogma, implicating Cx43 in other biological processes, such as transcription, metabolism, autophagy, and ion channel trafficking. How Cx43 participates in these processes remains largely unknown, although its high turnover rate, capacity to bind to myriad proteins, and the discovery of truncated isoforms of Cx43, ascribe to this protein unanticipated roles in chief processes that require fine-tuned regulation. Accordingly, Cx43 can be regarded as a central integrative hub to which diverse cues converge to be processed in a concerted manner. In this review, we examine the noncanonical roles of Cx43 and discuss the implications of these functions in human diseases and future therapeutic strategies.

Deregulation of proteostasis is a main feature of many age-related diseases, often leading to the accumulation of toxic oligomers and insoluble protein aggregates that accumulate intracellularly or in the extracellular space. To understand the mechanisms whereby toxic or otherwise unwanted proteins are secreted to the extracellular space, we inactivated the quality-control and proteostasis regulator ubiquitin ligase STUB1/CHIP. Data indicated that STUB1 deficiency leads both to the intracellular accumulation of protein aggregates and to an increase in the secretion of small extracellular vesicles (sEVs), including exosomes. Secreted sEVs are enriched in ubiquitinated and/or undegraded proteins and protein oligomers. Data also indicates that oxidative stress induces an increase in the release of sEVs in cells depleted from STUB1. Overall, the results presented here suggest that cells use exosomes to dispose of damaged and/or undegraded proteins as a means to reduce intracellular accumulation of proteotoxic material. publishersversion published

(1) Aim: This study aims to analyze the in vitro infiltration of a silicate root canal sealer into dentinal tubules after using different endodontic irrigating solutions. (2) Methods: Twenty-nine teeth with single roots were separated into three groups according to the final irrigation protocol: G1 n = 10) = 17% EDTA (ethylenediaminetetraacetic acid) + 3.0% sodium hypochlorite (NaOCl), G2 (n = 10) = 17% EDTA + 2.0% chlorhexidine and G3 (Control group, n = 9) = 17% EDTA + saline solution. Root canals were filled using cold lateral compaction technique with MTA Fillapex sealer and gutta-percha. The sealer was labeled with rhodamine B. The teeth were segmented at the middle and third apical sections, which were visualized using 10&times, confocal laser microscopy to determine the sealer penetration percentage. (3) Results: In the apical section, no statistically significant differences were found between the groups regarding sealer penetration. In the middle section, Group 1 obtained the highest percentage, and Group 2 the lowest (p = 0.004). Group 1 also presented statistically significant differences in the Control Group (p = 0.031) and had close sealer penetration values. Meanwhile, the Control Group (p = 0.023) and Group 2 (p = 0.029) revealed a significant decrease of sealer penetration between the apical and middle sections. (4) Conclusion: The obtained results support that final irrigation with NaOCl promoted similar sealer penetration in the apical and middle sections. On the other hand, a significant decrease in the sealer penetration of the middle section was observed for the chlorhexidine and saline groups. Compared to other irrigant solutions, NaOCl promotes more uniform sealer penetration, which can correlate with better sealing and, consequently, higher endodontic treatment success.

Conventional photodynamic agents used in clinic are porphyrin-based photosensitizers. However, they have low tumour selectivity, which may induce unwanted side-effects and damage to healthy tissues. In this study, we used a porphyrin with dendritic units of galactose (PorGal8) developed by us, which can target the galactose-binding protein, galectin-1, known to be overexpressed in many tumour tissues. In vitro and in vivo studies had been conducted for the validation of PorGal8 effectiveness. We showed a specific uptake of PorGal8 and induction of apoptotic cell death by generating oxidative stress and alterations in the cytoskeleton of bladder cancer cells overexpressing galectin-1. We further validated the photodynamic efficiency of PorGal8 in athymic nude mice (Balb/c nu/nu) bearing subcutaneously implanted luciferase-positive bladder cancer xenografts, overexpressing galectin-1 protein. PorGal8 (5 μmol/kg, intraperitoneal), injected 24 h before light delivery (50.4 J/cm2), inhibited tumour growth. We conclude that the use of PorGal8 enables selective target and cytotoxicity by photodynamic therapy in cancer cells overexpressing galectin-1, preventing undesired phototoxicity in the surrounding healthy tissues.

Heart proteostasis relies on a complex and integrated network of molecular processes surveilling organ performance under physiological and pathological conditions. For this purpose, cardiac cells depend on the correct function of their proteolytic systems, such as the ubiquitin-proteasome system (UPS), autophagy and the calpain system. Recently, the role of protein SUMOylation (an ubiquitin-like modification), has emerged as important modulator of cardiac proteostasis, which will be the focus of this review.

Myocardium infarction is one of the most deathly cardiovascular diseases. It is characterized by myocardium ischemia as a result of nutrients depletion and hypoxia. The cell can respond to this injury by autophagy or apoptosis, which determines the evolution and possible recovery of the myocardium infarction. Lipids play an important role in cardiovascular disease. However reports stating lipidome variations in cardiovascular disease are scarce and the role that lipids play in this pathological condition is not completely understood. The aim of this work was to identify changes in lipid profile of a myoblast H9c2 cell line under starvation and ischemia, to better understand and recognize new biomarkers for myocardial infarction. Lipidomic profile was evaluated by HILIC-LC-MS and GC-MS. Cardiac cells showed alterations in phosphatidylcholines PC (34:1) and PC (36:2), lysophosphatidylcholines lyso PC(16:0), lysoPC(18:1) and lysoPC(18:0), phosphatidylethanolamine PE (34:1), phosphatidylserine PS (36:1), phosphatidylinositol PI (36:2), PI (38:3) and PI (38:5), sphingomyelin SM (34:1) and cardiolipins CL(68:4), CL(72:5) and CL(74:7) in ischemia and/or starvation, in comparison with control. Specific differences observed only in starvation were decrease of SM (34:1) and FA (20:4), and increase of PS (36:1). Differences observed only in ischemia were decrease of PC (36:2), lyso PC (16:0) and FA (18:1) and simultaneous increase of FA (16:0), and FA (18:0). Interestingly, PC (34:1) increased in ischemia and decreased in starvation. In conclusion, our work suggests that lipids are potential markers for evaluation of cell fate, either cell death or recovery, which will be useful to improve diagnosis and prognostic of cardiovascular diseases. J. Cell. Physiol. 231: 2266-2274, 2016. © 2016 Wiley Periodicals, Inc.

Efficient contraction of the heart relies on a highly regulated communication network between cardiac cells. Direct intercellular communication is mediated by gap junctions but can also occur through tubular structures named tunnelling nanotubes (TNTs), which connect the cytoplasm of neighbouring cells and facilitate the transport of various cargoes. Although the formation of TNTs between cardiomyocytes has been reported, the effect of ischaemia on this process remains unclear. In this work, we assessed the impact of ischaemia and oxidative stress on TNT-mediated communication between cardiac cells. We found that cardiac cell lines and neonatal primary cultures of cardiomyocytes subjected to in vitro ischaemia form more TNTs than control cells. Moreover, antioxidants prevented ischaemia-induced TNT formation, suggesting that oxidative stress regulates this process. Furthermore, we identified troponin T as a new specific marker of cardiomyocyte-derived TNTs, which allows for the identification of heterocellular TNT connections between cardiomyocytes and other resident cells in the heart, such as fibroblasts. We also determined the presence of TNT-like structures in rat and human hearts. Rat hearts subjected to global ischaemia in the ex vivo Langendorff system showed increased formation of TNTs. Altogether, this study demonstrates that ischaemia affects the formation of TNTs in the heart and sheds new light on the regulation of TNT-mediated communication between cardiomyocytes.

Background/aim Breast cancer 1, early onset (BRCA1) gene is expressed in the cells of the breast and other tissues, where it plays a role in cell-cycle regulation, transcription, repair of DNA double-stranded breaks, ubiquitination, transcriptional regulation as well as other functions, such as cell response regulation to mitogenic signals triggered by estrogens. Considering that meningioma shows greater tumor growth during pregnancy, can express estrogen receptors and proliferate in response to estrogenic stimulation, the hypothesis that this type of tumor may share molecular mechanisms that involve exposure to estrogen should be investigated. Therefore, the aim of the present study was to investigate the BRCA1 gene methylation profile in meningioma. Materials and methods Methylation-specific polymerase chain reaction (PCR) assay was performed on 50 meningioma samples from male and female patients. Statistical analysis was carried out using Fisher's exact test. Results The most important finding of this study was that 100% of the male patients over 55 years with meningioma showed BRCA1 methylated in their tumor cells. Conclusion The silencing of BRCA1 through hypermethylation seems to play an important role in meningioma.

Breast cancer 1, early onset (BRCA1) gene is expressed in the cells of the breast and other tissues, where it plays a role in cell-cycle regulation, transcription, repair of DNA double-stranded breaks, ubiquitination, transcriptional regulation as well as other functions, such as cell response regulation to mitogenic signals triggered by estrogens. Considering that meningioma shows greater tumor growth during pregnancy, can express estrogen receptors and proliferate in response to estrogenic stimulation, the hypothesis that this type of tumor may share molecular mechanisms that involve exposure to estrogen should be investigated. Therefore, the aim of the present study was to investigate the BRCA1 gene methylation profile in meningioma.Methylation-specific polymerase chain reaction (PCR) assay was performed on 50 meningioma samples from male and female patients. Statistical analysis was carried out using Fisher's exact test.The most important finding of this study was that 100% of the male patients over 55 years with meningioma showed BRCA1 methylated in their tumor cells.The silencing of BRCA1 through hypermethylation seems to play an important role in meningioma.

Connexins (Cxs) are transmembrane proteins that form channels which allow direct intercellular communication (IC) between neighbouring cells via gap junctions. Mechanisms that modulate the amount of channels at the plasma membrane have emerged as important regulators of IC and their de-regulation has been associated with various diseases. Although Cx-mediated IC can be modulated by different mechanisms, ubiquitination has been described as one of the major post-translational modifications involved in Cx regulation and consequently IC. In this review, we focus on the role of ubiquitin and its effect on gap junction intercellular communication.

Background Glioblastoma (GBM) is a highly proliferative, angiogenic grade IV astrocytoma that develops resistance to the alkylating agents used in chemotherapy, such as temozolomide (TMZ), which is considered the gold standard. The mean survival time for GBM patients is approximately 12 months, increasing to 14.6 months after TMZ treatment. The resistance of GBM to chemotherapy seems to be associated to genetic alterations and to the constitutive activation of several signaling pathways. Therefore, the combination of different drugs with different mechanisms of action may contribute to circumvent the chemoresistance of glioma cells. Here we describe the potential synergistic behavior of the therapeutic combination of tamoxifen (TMX), a known inhibitor of PKC, and TMZ in GBM. Methods We used two GBM cell lines incubated in absence and presence of TMX and/or TMZ and measured cell viability, proliferation, apoptosis, cell cycle, migration ability, cytoskeletal organization and the phosphorylated amount of the p-PKC-pan. Results The combination of low doses of TMX with increasing doses of TMZ shows an increased antiproliferative and apoptotic effect compared to the effect with TMX alone. Conclusions The combination of TMX and TMZ seems to potentiate the effect of each other. These alterations seem to be associated to a decrease in the phosphorylation status of PKC. General significance We emphasize that TMX is an inhibitor of the p-PKC-pan and that these combination is more effective in the reduction of proliferation and in the increase of apoptosis than each drug alone, which presents a new therapeutic strategy in GBM treatment.