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In glioblastoma (GBM), tumor-associated macrophages (TAM) represent up to one half of the cells of the tumor mass, including both infiltrating macrophages and resident brain microglia. In an effort to delineate the temporal and spatial dynamics of TAM composition during gliomagenesis, we used genetically engineered and GL261-induced mouse models in combination with CX3CR1GFP/WT;CCR2RFP/WT double knock-in mice. Using this approach, we demonstrated that CX3CR1LoCCR2Hi monocytes were recruited to the GBM, where they transitioned to CX3CR1HiCCR2Lo macrophages and CX3CR1HiCCR2− microglia-like cells. Infiltrating macrophages/monocytes constituted approximately 85% of the total TAM population, with resident microglia accounting for the approximately 15% remaining. Bone marrow–derived infiltrating macrophages/monocytes were recruited to the tumor early during GBM initiation, where they localized preferentially to perivascular areas. In contrast, resident microglia were localized mainly to peritumoral regions. RNA-sequencing analyses revealed differential gene expression patterns unique to infiltrating and resident cells, suggesting unique functions for each TAM population. Notably, limiting monocyte infiltration via genetic Ccl2 reduction prolonged the survival of tumor-bearing mice. Our findings illuminate the unique composition and functions of infiltrating and resident myeloid cells in GBM, establishing a rationale to target infiltrating cells in this neoplasm. Cancer Res; 77(9); 2266–78. ©2017 AACR.

Dynamic modulation of CCR2 and CX3CR1 in myeloid cells isolated from tumors. With the decrease of Ly6C intensity, the cells present increased CX3CR1-GFP intensity but decreased CCR2-RFP intensity.

Figure S1. Circulating Ly6CHi inflammatory monocytes diminish at terminal illness. Figure S2. Enhanced BM-derived cell infiltration in Cx3cr1GFP/GFP;Ccr2RFP/WT mice. Figure S3. RFP+ cells are only observed in the meninges in healthy naïve Cx3cr1GFP/WT;Ccr2RFP/WT mice. Figure S4. Bone marrow reconstitution efficiency as examined by flow cytometry. Figure S5. Significant up-regulated (red) and down-regulated (green) genes based on the pairwise comparisons. Figure S6. Loss of Ccl2 in stroma does not prolong survival of tumor-bearing mice. Figure S7. Immunohistochemistry staining and quantification of phosphor-histone 3 proliferating cells. Figure S8. Immunohistochemistry staining and quantification of Ki67+ proliferating cells. Figure S9. TUNEL staining of apoptotic cells. Figure S10. Survival curve of PDGF-driven tumor in WT, Ccl2+/- and Ccl2-/- mice. Supplementary Table S1. Combined Cx3cr1 and Ccr2 heterozygous knockout does not affect tumor development or survival. Supp. Table S2. Significantly enriched functional pathways in tumor-associated microglia. Supp. Table S3. Significantly enriched functional pathways in tumor-associated microglia and tumor-associate macrophages. Supp. Table S4. Significantly enriched functional pathways in tumor-associated macrophages.

Background: Metabolic syndrome (MS) is a complex and prevalent disorder. Oxidative stress and inflammation might contribute to the progression of MS. Soluble ST2 (sST2) is an attractive and druggable molecule that sits at the interface between inflammation, oxidative stress and fibrosis. This study aims to analyze the relationship among sST2, oxidative stress, inflammation and echocardiographic parameters in MS patients. Methods: Fifty-eight patients with MS were recruited and underwent physical, laboratory and transthoracic echocardiography examinations. Commercial ELISA and appropriate colorimetric assays were used to quantify serum levels of oxidative stress and inflammation markers and sST2. Results: Circulating sST2 was increased in MS patients and was significantly correlated with the oxidative stress markers nitrotyrosine and 8-hydroxy-2′-deoxyguanosine as well as with peroxide levels. The inflammatory parameters interleukin-6, intercellular adhesion molecule-1 and myeloperoxidase were positively correlated with sST2. Noteworthy, sST2 was positively correlated with left ventricular mass, filling pressures and pulmonary arterial pressures. Conclusion: Circulating levels of sST2 are associated with oxidative stress and inflammation burden and may underlie the pathological remodeling and dysfunction of the heart in MS patients. Our results suggest that sST2 elevation precedes diastolic dysfunction, emerging as an attractive biotarget in MS.

PIK3CA is one of the two most frequently mutated genes in breast cancers, occurring in 30–40% of cases. Four frequent ‘hotspot’ PIK3CA mutations (E542K, E545K, H1047R and H1047L) account for 80–90% of all PIK3CA mutations in human malignancies and represent predictive biomarkers. Here we describe a PIK3CA mutation specific nuclease-based enrichment assay, which combined with a low-cost real-time qPCR detection method, enhances assay detection sensitivity from 5% for E542K and 10% for E545K to 0.6%, and from 5% for H1047R to 0.3%. Moreover, we present a novel flexible prediction method to calculate initial mutant allele frequency in tissue biopsy and blood samples with low mutant fraction. These advancements demonstrated a quick, accurate and simple detection and quantitation of PIK3CA mutations in two breast cancer cohorts (first cohort n = 22, second cohort n = 25). Hence this simple, versatile and informative workflow could be applicable for routine diagnostic testing where quantitative results are essential, e.g. disease monitoring subject to validation in a substantial future study.

Introduction Aortic stenosis (AS) is the most common heart valve disease and it is strongly prevalent with elderly. AS is a progressive, degenerative disease associated with fibrosis and calcification of the valve leaflets. Surgical valve replacement is the only treatment available. Molecular, cellular and interstitial events activate multifactorial and complex cues with a significant contribution by valve interstitial cells (VICs). Despite male sex is a risk factor for developing AS, there is scant information on sex-specific differences in aortic valve (AV) biology or pathology. Purpose The aim of our study was to analyse sex-specific differences in aortic valves from AS patients. Methods 185 patients with severe AS undergoing surgical valve replacement were recruited. 149 AVs (66 women; 83 men) were used for ex vivo analyses. Human VICs were isolated from 36 AVs (12 women; 24 men) for in vitro experiments. AVs structure were evaluated by haematoxylin-eosin, Movat, Alizarin Red, Congo red and Alcian blue/Sirius Red staining and immunohistochemistry. Western blot, ELISA and zymography were used for molecular biology studies. Results AVs from men presented increased inflammatory infiltrates (CD68 and CD45 positive cells) as compared to women. Complementarily, AVs from men exhibited higher levels of the inflammatory molecules interleukin (IL)-6 and IL-1b and RANTES. In line with these results, oxidative stress markers (eNOS, myeloperoxidase, malondialdehyde and nitrotyrosine) were upregulated in male AVs. Concerning, fibrosis, increased levels of collagen type I, fibronectin and syndecan-1 were found in AVs from men. Extracellular matrix (ECM) remodelling was characterized by reduced metalloproteinase-1 expression and increased tissue inhibitor of metalloproteinase-2 expression in male AVs. Importantly, calcification and osteogenic markers (bone morphogenetic protein-9, periostin, osteocalcin and Sox-9) was greatly enhanced in men AVs as compared to women. These findings were confirmed in isolated VICs. At baseline, male VICs presented higher myofibroblast-like phenotype than female VICs. In line with our ex vivo results, male VICs exhibited increased inflammatory, oxidative stress, fibrotic and osteogenic differentiation markers. Conclusions Our results suggest that the mechanisms driving the AV pathogenesis could be different in men and women patients with the same AS severity. Male AVs and isolated VICs presented more inflammation, oxidative stress, fibrosis and ECM remodelling including extensive calcification as compared to female. A better knowledge of the pathophysiological pathways in AVs and VICs will allow developing sex-specific options for AS treatment. Funding Acknowledgement Type of funding sources: Public grant(s) – National budget only. Main funding source(s): Instituto de Salud Carlos III

PTEN is a phosphatase which metabolises PIP3, the lipid product of PI 3-Kinase, directly opposing the activation of the oncogenic PI3K/AKT/mTOR signalling network. Accordingly, loss of function of the PTEN tumour suppressor is one of the most common events observed in many types of cancer. Although the mechanisms by which PTEN function is disrupted are diverse, the most frequently observed events are deletion of a single gene copy of PTEN and gene silencing, usually observed in tumours with little or no PTEN protein detectable by immunohistochemistry. Accordingly, with the exceptions of glioblastoma and endometrial cancer, mutations of the PTEN coding sequence are uncommon (

Background: Soluble ST2 (interleukin 1 receptor-like 1) (sST2) is involved in inflammatory diseases and increased in heart failure (HF). We herein investigated sST2 effects on oxidative stress and inflammation in human cardiac fibroblasts and its pathological role in human aortic stenosis (AS). Methods and results: Using proteomics and immunodetection approaches, we have identified that sST2 down-regulated mitofusin-1 (MFN-1), a protein involved in mitochondrial fusion, in human cardiac fibroblasts. In parallel, sST2 increased nitrotyrosine, protein oxidation and peroxide production. Moreover, sST2 enhanced the secretion of pro-inflammatory cytokines interleukin (IL)-6, IL-1β and monocyte chemoattractant protein-1 (CCL-2). Pharmacological inhibition of transcriptional factor nuclear factor κB (NFκB) restored MFN-1 levels and improved oxidative status and inflammation in cardiac fibroblasts. Mito-Tempo, a mitochondria-specific superoxide scavenger, as well as Resveratrol, a general antioxidant, attenuated oxidative stress and inflammation induced by sST2. In myocardial biopsies from 26 AS patients, sST2 up-regulation paralleled a decrease in MFN-1. Cardiac sST2 inversely correlated with MFN-1 levels and positively associated with IL-6 and CCL-2 in myocardial biopsies from AS patients. Conclusions: sST2 affected mitochondrial fusion in human cardiac fibroblasts, increasing oxidative stress production and inflammatory markers secretion. The blockade of NFκB or mitochondrial reactive oxygen species restored MFN-1 expression, improving oxidative stress status and reducing inflammatory markers secretion. In human AS, cardiac sST2 levels associated with oxidative stress and inflammation. The present study reveals a new pathogenic pathway by which sST2 promotes oxidative stress and inflammation contributing to cardiac damage.

Rationale: Mitral valve prolapse (MVP) is one of the most common valvular disorders. However, the molecular and cellular mechanisms involved in fibromyxomatous changes in the mitral leaflet tissue have not been elucidated. Aldosterone (Aldo) promotes fibrosis in myocardium, and MR (mineralocorticoid receptor) antagonists (MRAs) improve cardiac function by decreasing cardiac fibrosis. Objective: We investigated the role of the Aldo/MR in the fibromyxomatous modifications associated with MVP. Methods and Results: Aldo enhanced valvular interstitial cell activation markers and induced endothelial-mesenchymal transition in valvular endothelial cells, resulting in increased proteoglycan secretion. MRA blocked all the above effects. Cytokine arrays showed CT-1 (cardiotrophin-1) to be a mediator of Aldo-induced valvular interstitial cell activation and proteoglycan secretion and CD (cluster of differentiation) 14 to be a mediator of Aldo-induced endothelial-mesenchymal transition and proteoglycan secretion in valvular endothelial cells. In an experimental mouse model of MVP generated by nordexfenfluramine administration, MRA treatment reduced mitral valve thickness and proteoglycan content. Endothelial-specific MR deletion prevented fibromyxomatous changes induced by nordexfenfluramine administration. Moreover, proteoglycan expression was slightly lower in the mitral valves of MVP patients treated with MRA. Conclusions: These findings demonstrate, for the first time, that the Aldo/MR pathway regulates the phenotypic, molecular, and histological changes of valvular interstitial cells and valvular endothelial cells associated with MVP development. MRA treatment appears to be a promising option to reduce fibromyxomatous alterations in MVP.

Objective: Aortic valve (AV) calcification plays an important role in the progression of aortic stenosis (AS). MMP-10 (matrix metalloproteinase-10 or stromelysin-2) is involved in vascular calcification in atherosclerosis. We hypothesize that MMP-10 may play a pathophysiological role in calcific AS. Approach and Results: Blood samples (n=112 AS and n=349 controls) and AVs (n=88) from patients undergoing valve replacement were analyzed. Circulating MMP-10 was higher in patients with AS compared with controls ( P S =0.451; P S =0.5045, P S =0.5003, P P Conclusions: MMP-10, which is overexpressed in aortic valve from patients with AS, seems to play a central role in calcification in AS through Akt phosphorylation. MMP-10 could be a new therapeutic target for delaying the progression of aortic valve calcification in AS.

Aortic valve surgery in non-elderly patients represents a very challenging patient population. The younger the patient is at the point of aortic valve intervention, the longer their anticipated life expectancy will be, with longer exposure to valve-related complications and risk for re-operation. Although the latest international guidelines recommend aortic valve repair in patients with aortic valve insufficiency, what we see in the real world is that the vast majority of these aortic valves are replaced. However, current prosthetic valves has now been shown to lead to significant loss of life expectancy for non-elderly patients up to 50% for patients in their 40s undergoing mechanical aortic valve replacement. Bioprostheses carry an even worse long-term survival, with higher rates of re-intervention. The promise of trans-catheter valve-in-valve technology is accentuating the trend of bioprosthetic implantation in younger patients, without yet the appropriate evidence. In contrast, aortic valve repair has shown excellent outcomes in terms of quality of life, freedom from re-operation and freedom from major adverse valve-related events with similar life expectancy to general population as it is also found for the Ross procedure, the only available living valve substitute. We are at a time when the paradigm of aortic valve surgery needs to change for the better. To better serve our patients, we must acquire high quality real-world evidence from multiple centers globally - this is the vision of the AVIATOR registry and our common responsibility.

Galectin-3 (Gal-3) is increased in heart failure (HF) and promotes cardiac fibrosis and inflammation. We investigated whether Gal-3 modulates oxidative stress in human cardiac fibroblasts, in experimental animal models and in human aortic stenosis (AS). Using proteomics and immunodetection approaches, we have identified that Gal-3 down-regulated the antioxidant peroxiredoxin-4 (Prx-4) in cardiac fibroblasts. In parallel, Gal-3 increased peroxide, nitrotyrosine, malondialdehyde, and N-carboxymethyl-lysine levels and decreased total antioxidant capacity. Gal-3 decreased prohibitin-2 expression without modifying other mitochondrial proteins. Prx-4 silencing increased oxidative stress markers. In Gal-3-silenced cells and in heart from Gal-3 knockout mice, Prx-4 was increased and oxidative stress markers were decreased. Pharmacological inhibition of Gal-3 with modified citrus pectin restored cardiac Prx-4 as well as prohibitin-2 levels and improved oxidative status in spontaneously hypertensive rats. In serum from 87 patients with AS, Gal-3 negatively correlated with total antioxidant capacity and positively correlated with peroxide. In myocardial biopsies from 26 AS patients, Gal-3 up-regulation paralleled a decrease in Prx-4 and in prohibitin-2. Cardiac Gal-3 inversely correlated with Prx-4 levels in myocardial biopsies. These data suggest that Gal-3 decreased Prx-4 antioxidant system in cardiac fibroblasts, increasing oxidative stress. In pathological models presenting enhanced cardiac Gal-3, the decrease in Prx-4 expression paralleled increased oxidative stress. Gal-3 blockade restored Prx-4 expression and improved oxidative stress status. In AS, circulating levels of Gal-3 could reflect oxidative stress. The alteration of the balance between antioxidant systems and reactive oxygen species production could be a new pathogenic mechanism by which Gal-3 induces cardiac damage in HF.

Consumers increasingly prefer to use mobile phones to surf the internet and make purchases. Mobile applications are also being more used in the area of trade between consumers. This study seeks to understand how satisfaction is generated in the context of consumer-to-consumer (C2C) commerce via mobile applications for buying and selling second-hand fashion products (e.g., clothing, accessories). The research model was empirically evaluated using data collected by an online survey and analysed through partial least square structural equation modelling (PLS-SEM). The results show that trust, perceived usefulness and mobility have significant and positive impacts on consumer satisfaction with C2C mobile applications in fashion sales. Conversely, factors such as security, privacy and enjoyment do not have significant influence.

Mitral valve disease (MVD) is a frequent cause of heart failure and death worldwide, but its etiopathogenesis is not fully understood. Interleukin (IL)-33 regulates inflammation and thrombosis in the vascular endothelium and may play a role in the atherosclerotic process, but its role in mitral valve has not been investigated. We aim to explore IL-33 as a possible inductor of myxomatous degeneration in human mitral valves. We enrolled 103 patients suffering from severe mitral regurgitation due to myxomatous degeneration undergoing mitral valve replacement. Immunohistochemistry of the resected leaflets showed IL-33 and ST2 expression in both valve interstitial cells (VICs) and valve endothelial cells (VECs). Positive correlations were found between the levels of IL-33 and molecules implicated in the development of myxomatous MVD, such as proteoglycans, extracellular matrix remodeling enzymes (matrix metalloproteinases and their tissue inhibitors), inflammatory and fibrotic markers. Stimulation of single cell cultures of VICs and VECs with recombinant human IL-33 induced the expression of activated VIC markers, endothelial–mesenchymal transition of VECs, proteoglycan synthesis, inflammatory molecules and extracellular matrix turnover. Our findings suggest that the IL-33/ST2 system may be involved in the development of myxomatous MVD by enhancing extracellular matrix remodeling.

The PI3K/AKT/mTOR signaling pathway is one of the most frequently mutated pathways in human cancers, including breast cancer. Given the current urge for more personalized approaches, there is an increasing need for a rigorous list of biomarkers and detection methods with advanced, less-invasive, reliable, and low-cost technologies. Despite limited clinical results to date, the detection of PIK3CA mutations shows great potential to contribute in cancer patient management in the future at many different levels, including diagnosis, treatment choice and monitoring, and identification of drug resistance. However, current molecular methods still lack analytical sensitivity to detect mutations of low abundance in high wild-type DNA background. Makrigiorgos et al. recently developed a nuclease-assisted minor-allele enrichment assay with overlapping probes, which is practical and cost-effective and allows detection of very-low-abundance mutations that could have relevance in the clinic. Thus we aimed to develop PIK3CA mutation-specific nuclease-based enrichment assay and validate it with tissue biopsy and blood samples using crystal dPCR as well as combine it with our developed Sybr Green real-time qPCR detection method (Alvarez-Garcia et al. 2018), which allows a quick detection of PIK3CA mutations in clinical samples at low cost. Citation Format: Ieva Keraite, Virginia Alvarez-Garcia, Jurgen Del Favero, Maiwenn Kersaudy-Kerhoas, Nicholas Leslie. PIK3CA mutation enrichment and detection in clinical samples [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B55.

The phosphatase and tensin homologue deleted on chromosome 10 (PTEN) phosphatase dephosphorylates PIP3, the lipid product of the class I PI 3-kinases, and suppresses the growth and proliferation of many cell types. It has been heavily studied, in large part due to its status as a tumour suppressor, the loss of function of which is observed through diverse mechanisms in many tumour types. Here we present a concise review of our understanding of the PTEN protein and highlight recent advances, particularly in our understanding of its localization and regulation by ubiquitination and SUMOylation.

PIK3CA mutations are seemingly the most common driver mutations in breast cancer with H1047R and E545K being the most common of these, accounting together for around 60% of all PIK3CA mutations and have promising therapeutic implications. Given the low sensitivity and the high cost of current genotyping methods we sought to develop fast, simple and inexpensive assays for PIK3CA H1047R and E545K mutation screening in clinical material. The methods we describe are based on a real-time PCR including a mutation specific primer combined with a non-productive oligonucleotide which inhibits wild-type amplification and a parallel internal control reaction. We demonstrate consistent detection of PIK3CA H1047R mutant DNA in genomic DNA extracted from frozen breast cancer biopsies, FFPE material or cancer cell lines with a detection sensitivity of approximately 5% mutant allele fraction and validate these results using both Sanger sequencing and deep next generation sequencing methods. The detection sensitivity for PIK3CA E545K mutation was approximately 10%. We propose these methods as simple, fast and inexpensive diagnostic tools to determine PIK3CA mutation status.

// Xi Feng 1 , Frank Szulzewsky 2,* , Alexan Yerevanian 1,3,* , Zhihong Chen 1 , David Heinzmann 1,4 , Rikke Darling Rasmussen 1 , Virginia Alvarez-Garcia 1 , Yeonghwan Kim 5 , Bingcheng Wang 6 , Ilaria Tamagno 1 , Hao Zhou 7 , Xiaoxia Li 7 , Helmut Kettenmann 2 , Richard M. Ransohoff 1,8 and Dolores Hambardzumyan 1 1 Department of Neurosciences at Cleveland Clinic, Cleveland, Ohio, USA 2 Cellular Neurosciences, Max Delbruck Center for Molecular Medicine, Berlin, Germany 3 Case Western Reserve University School of Medicine, Cleveland, Ohio, USA 4 Department of Cardiology at Tubingen University School of Medicine, Tubingen, Germany 5 Department of Stem Cell Biology and Regenerative Medicine, Cleveland, Ohio, USA 6 Rammelkamp Center for Research, MetroHealth Center, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA 7 Department of Immunology at Cleveland Clinic, Cleveland, Ohio, USA 8 Neuroinflammation Research Center, Cleveland Clinic, Cleveland, Ohio, USA * These authors have contributed equally to this work Correspondence to: Dolores Hambardzumyan, email: // Keywords : CX3CR1/CX3CL1 signaling, glioblastoma, microglia, monocyte Received : January 19, 2015 Accepted : March 10, 2015 Published : March 30, 2015 Abstract The most abundant populations of non-neoplastic cells in the glioblastoma (GBM) microenvironment are resident microglia, macrophages and infiltrating monocytes from the blood circulation. The mechanisms by which monocytes infiltrate into GBM, their fate following infiltration, and their role in GBM growth are not known. Here we tested the hypothesis that loss of the fractalkine receptor CX3CR1 in microglia and monocytes would affect gliomagenesis. Deletion of Cx3cr1 from the microenvironment resulted in increased tumor incidence and shorter survival times in glioma-bearing mice. Loss of Cx3cr1 did not affect accumulation of microglia/macrophages in peri-tumoral areas, but instead indirectly promoted the trafficking of CD11b + CD45 hi CX3CR1 low Ly-6C hi Ly-6G - F4/80 -/low circulating inflammatory monocytes into the CNS, resulting in their increased accumulation in the perivascular area. Cx3cr1-deficient microglia/macrophages and monocytes demonstrated upregulation of IL1β expression that was inversely proportional to Cx3cr1 gene dosage. The Proneural subgroup of the TCGA GBM patient dataset with high IL1β expression showed shorter survival compared to patients with low IL1β. IL1β promoted tumor growth and increased the cancer stem cell phenotype in murine and human Proneural glioma stem cells (GSCs). IL1β activated the p38 MAPK signaling pathway and expression of monocyte chemoattractant protein (MCP-1/CCL2) by tumor cells. Loss of Cx3cr1 in microglia in a monocyte-free environment had no impact on tumor growth and did not alter microglial migration. These data suggest that enhancing signaling to CX3CR1 or inhibiting IL1β signaling in intra-tumoral macrophages can be considered as potential strategies to decrease the tumor-promoting effects of monocytes in Proneural GBM.

To analyze the risk profile of patients with diabetes who seek care from hospital emergency departments and emergency department involvement in preventing cardiovascular complications in these patients.Cross-sectional analysis of case series from 2 Spanish hospital emergency departments. We included all patients with a history or final diagnosis of diabetes mellitus who were treated in the emergency department between November 1, 2010, and June 30, 2011. Each patient's cardiovascular risk profile was analyzed. The main outcome was the appropriate of prescribed treatment to prevent cardiovascular complications according to the 2012 guidelines of the American Diabetes Association on the patient's discharge from emergency care.A total of 298 patients were included; 275 (92%) had type II diabetes. Ninety percent of the series (269 patients) had at least 1 cardiovascular risk factor and 147 (49%) had prior target organ damage; target organ damage was newly diagnosed in 41 (14%). Fifty-eight percent (172 patients) were discharged home from the emergency department. Although 215 patients (72%) were not adhering to at least 1 previously prescribed preventive treatment and 30 (10%) were not adhering to any prescribed treatment, drug prescriptions were modified only in 1.1% to 3.3% of patients and no follow-up was recommended in 42 cases (24%).Although diabetic patients treated in emergency departments are at high risk for cardiovascular complications, their visit is not used to optimize preventive treatment for these complications or ensure appropriate follow-up.Analizar el perfil de riesgo y la prescripción de tratamiento para la prevención de complicaciones cardiovasculares en los pacientes diabéticos que acuden a los servicios de urgencias hospitalarios (SUH).Estudio de series de casos con análisis transversal llevado a cabo en dos SUH españoles. Se incluyeron de forma consecutiva todos los pacientes con antecedentes o diagnóstico final de diabetes mellitus entre 1 de noviembre de 2010 y 30 de junio de 2011. Se analizó su perfil de riesgo cardiovascular y se consideró como variable de resultado principal la prescripción de tratamiento para la prevención de las complicaciones cardiovasculares al alta del SUH de acuerdo a las recomendaciones de la American Diabetes Association de 2012.Se incluyeron 298 pacientes diabéticos, 275 (92%) tipo 2. De ellos 269 (90%) presentaban algún otro factor de riesgo cardiovascular, 147 (49%) lesión previa de órgano diana y 41 (14%) lesión nueva de órgano diana. Ciento setenta y dos pacientes (58%) fueron dados de alta desde el SUH. Aunque 215 pacientes (72%) no cumplían previamente alguna y 30 (10%) ninguna de las recomendaciones de prevención, sólo se modificó el tratamiento farmacológico en un rango de un 1,1% a 3,3% de los pacientes y no se recomendó seguimiento posterior en 42 casos (24%).Aunque los pacientes diabéticos atendidos en los SUH presentan un elevado riesgo cardiovascular, no se aprovecha esta oportunidad para optimizar el tratamiento de la prevención de las complicaciones cardiovasculares ni garantizar un seguimiento adecuado.

Aims Galectin-3 (Gal-3), a β-galactoside-binding lectin involved in cardiac inflammation and fibrosis, could regulate oxidative stress, although the mechanisms have not been elucidated. We herein investigated the changes in oxidative stress-related mediators induced by Gal-3 in human cardiac fibroblasts and in pathological animal and human models of cardiac diseases. Results Using quantitative proteomics and immunodetection approaches, we have identified that Gal-3 down-regulated fumarate hydratase (FH) in human cardiac fibroblasts. In parallel, Gal-3 increased fumarate production in a time-dependent manner. Gal-3 treatment enhanced carbonylated proteins detected through OxyBlot technique. Interestingly, treatment of cells with fumarate induced oxidative stress, enhanced fibroblast activation markers and increased collagen and interleukin-6 secretion. In Gal-3-silenced cells and in heart from Gal-3 knock-out mice, FH was increased and fumarate was decreased. In myocardial biopsies from patients with aortic stenosis (AS, n =26), FH levels were decreased as compared to Controls ( n =13). Cardiac Gal-3 inversely correlated with FH levels in myocardial biopsies. In an experimental model of AS rats, pharmacological inhibition of Gal-3 restored cardiac FH, decreased fumarate concentration and improved oxidative status. Conclusion In human cardiac fibroblasts, Gal-3 decreased FH expression increasing fumarate concentration and promoting oxidative stress. In human AS, cardiac levels of Gal-3 inversely associated with FH. Gal-3 blockade restored FH and improved fumarate and oxidative stress status in AS rats. FH is therefore a key molecule mediating Gal-3-induced oxidative stress in cardiac cells.

Endothelial cells represent one of the critical cellular elements in tumor microenvironment playing a crucial role in the growth and progression of cancer through controlling angiogenesis. Vascular endothelial growth factor (VEGF) produced from tumor cells is essential for the expansion of breast cancer and may function in both paracrine and autocrine manners to promote proliferation, growth, survival and migration of endothelial cells. Since melatonin regulates tumor microenvironment by decreasing the secretion of VEGF by malignant epithelial cells and also regulates VEGF expression in human breast cancer cells, the aim of the present study was to investigate the anti-angiogenic activity of melatonin against the pro-angiogenic effects of breast cancer cells. In this work, we demonstrate that melatonin strongly inhibited the proliferation as well as invasion/migration of human umbilical vein endothelial cells (HUVECs). Melatonin disrupted tube formation and counteracted the VEGF-stimulated tubular network formation by HUVEC. In addition, conditioned media collected from human breast cancer cells were angiogenically active and stimulated tubule length formation and this effect was significantly counteracted by the addition of anti-VEGF or melatonin. Melatonin also disintegrated preformed capillary network. All these findings demonstrate that melatonin may play a role in the paracrine interactions that take place between malignant epithelial cells and proximal endothelial cells. Melatonin could be important in reducing endothelial cell proliferation, invasion, migration and tube formation, through a downregulatory action on VEGF. Taken together, our findings suggest that melatonin could potentially be beneficial as an antiangiogenic agent in breast cancer with possible future clinical applications.

In glioblastoma (GBM), tumor-associated macrophages (TAM) represent up to one half of the cells of the tumor mass, including both infiltrating macrophages and resident brain microglia. In an effort to delineate the temporal and spatial dynamics of TAM composition during gliomagenesis, we used genetically engineered and GL261-induced mouse models in combination with CX3CR1GFP/WT;CCR2RFP/WT double knock-in mice. Using this approach, we demonstrated that CX3CR1LoCCR2Hi monocytes were recruited to the GBM, where they transitioned to CX3CR1HiCCR2Lo macrophages and CX3CR1HiCCR2− microglia-like cells. Infiltrating macrophages/monocytes constituted approximately 85% of the total TAM population, with resident microglia accounting for the approximately 15% remaining. Bone marrow–derived infiltrating macrophages/monocytes were recruited to the tumor early during GBM initiation, where they localized preferentially to perivascular areas. In contrast, resident microglia were localized mainly to peritumoral regions. RNA-sequencing analyses revealed differential gene expression patterns unique to infiltrating and resident cells, suggesting unique functions for each TAM population. Notably, limiting monocyte infiltration via genetic Ccl2 reduction prolonged the survival of tumor-bearing mice. Our findings illuminate the unique composition and functions of infiltrating and resident myeloid cells in GBM, establishing a rationale to target infiltrating cells in this neoplasm. Cancer Res; 77(9); 2266–78. ©2017 AACR.

Galectin-3 (Gal-3) is involved in cardiovascular fibrosis and aortic valve (AV) calcification. We hypothesized that Gal-3 pharmacological inhibition with modified citrus pectin (MCP) could reduce aortic and AV remodeling in normotensive rats with pressure overload (PO). Six weeks after aortic constriction, vascular Gal-3 expression was up-regulated in male Wistar rats. Gal-3 overexpression was accompanied by an increase in the aortic media layer thickness, enhanced total collagen, and augmented expression of fibrotic mediators. Further, vascular inflammatory markers as well as inflammatory cells content were greater in aorta from PO rats. MCP treatment (100 mg/kg/day) prevented the increase in Gal-3, media thickness, fibrosis, and inflammation in the aorta of PO rats. Gal-3 levels were higher in AVs from PO rats. This paralleled enhanced AV fibrosis, inflammation, as well as greater expression of calcification markers. MCP treatment prevented the increase in Gal-3 as well as fibrosis, inflammation, and calcification in AVs. Overall, Gal-3 is overexpressed in aorta and AVs from PO rats. Gal-3 pharmacological inhibition blocks aortic and AV remodeling in experimental PO. Gal-3 could be a new therapeutic approach to delay the progression and the development of aortic remodeling and AV calcification in PO.

Melatonin exerts oncostatic effects on breast cancer by interfering with the estrogen signaling pathways. Melatonin inhibits aromatase enzyme in breast cancer cells and fibroblasts. In addition, melatonin stimulates the adipogenic differentiation of fibroblasts. Our objective was to study whether melatonin interferes in the desmoplastic reaction by regulating some factors secreted by malignant cells, tumor necrosis factor (TNF)-α, interleukin (IL)-11, and interleukin (IL)-6. To accomplish this, we co-cultured 3T3-L1 cells with MCF-7 cells. The addition of breast cancer cells to the co-cultures inhibited the differentiation of 3T3-L1 preadipocytes to mature adipocytes, by reducing the intracytoplasmic triglyceride accumulation, an indicator of adipogenic differentiation, and also stimulated their aromatase activity. Melatonin counteracted the inhibitory effect on adipocyte differentiation and aromatase activity induced by MCF-7 cells in 3T3-L1 cells. The levels of cytokines in the co-culture media were 10 times those found in culture of 3T3-L1 cells alone. Melatonin decreased the concentrations of cytokines in the media and counteracted the stimulatory effect induced by MCF-7 cells on the cytokine levels. One millimolar melatonin induced a reduction in TNF-α, IL-6, and IL-11 mRNA expression in MCF-7 and 3T3-L1 cells. The findings suggest that melatonin may play a role in the desmoplastic reaction in breast cancer through a downregulatory action on the expression of antiadipogenic cytokines, which decrease the levels of these cytokines. Lower levels of cytokines stimulate the differentiation of fibroblasts and decrease both aromatase activity and expression, thereby reducing the number of estrogen-producing cells proximal to malignant cells.