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  • Background: Antibody validation for tissue staining is required for reproducibility and criteria to ensure validity have been published recently. The majority of these recommendations imply the use of routinely processed tissue (FFPE). Materials & Methods: We applied to lightly fixed frozen sections a panel of 126 antibodies validated for FFPE with extended criteria. Results: Less than 30% performed conservatively with all fixations, 35% preferred one fixation over another, 13% gave non-specific staining, 23% did not stain at all. Conclusions: Individual antibody variability of the paratope fitness for the fixed antigen may be the cause. Re-validation of established antibody panels is required when applied to sections whose fixation and processing is different from the tissue where they have been initially validated. These are supplementary Data to the manuscript. In addition Tabel 1 and Supplementary Table 1 are provided in Excel format.
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  • Personalized immune intervention to release and redirect the T cells against a tumor has shown substantial progress in aggressive tumors such as melanoma and lung cancer, despite the fact that predictors of sustained response are still unclear. Data on less common histotypes are scanty. Among soft tissue sarcomas, uterine leiomyosarcomas (ULMS) have a dire prognosis, yet therapeutic advances are needed in order to improve the actual treatment. Immune checkpoint inhibitor therapy has been applied to exceptionally few cases, of which the immune cell composition was not examined in detail. We analyzed in situ the inflammatory infiltrate of 21 untreated ULMS in high-dimensional, single cell phenotyping on routinely processed tissue, directed at the characterization of lymphoid cells and macrophages. T-lymphoid cells displayed a composite phenotype common to all tumors, suggestive of antigen-exposure; in about half of the cases containing sufficient lymphocytes, we found evidence of exhaustion and a CD8+ TCF7+ phenotype, this latter associated with T-cell reactivation. To the contrary, myelomonocytic cells had case-specific individual combinations of phenotypes and subsets. We identified five distinct monocyte-macrophage cell types: histiocytes, phagocytes, tumor-associated macrophages, inflammatory monocytes and myelomonocytic cells of undefined phenotype. Immunosuppressive molecules (TIM3, B7H3, VISTA, PD1, PDL1) were heterogeneously expressed in inflammatory and endothelial cells. The heterogeneity and phenotype of the monocyte-macrophage population may represents a challenge for which we provide an initial understanding.
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  • Dataset and materials for the rTMS study "Keeping order in the brains: the supramarginal gyrus and serial order in short-term memory". In the "Experiment folders" there are the E-Prime script of the tasks and the stimuli used. In the "Database and all anlysis" folder, there is the anonymus excel database of all 4 experiments and the Statistica workbook with all the analyses conducted and reported in the paper.
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  • Pipelines for CellProfiler mask generation, creation of heatmaps with R from Phenograph .csv data, .csv data exported from HistoCAT after segmentation and analysis. Refers to The landscape of S100B+ and HLA-DR+ dendritic cell subsets in tonsils at the single cell level via high-parameter mapping. by Bolognesi MM et al.
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  • The correlation of clinical, radiological and laboratory findings of patients at admission in the Emergency Department (ED) with clinical severity and risk of mortality was investigated. Adult coronavirus disease 2019 (COVID-19) patients hospitalized in March 2020 in Desio Hospital, Lombardy, were retrospectively included in the study, and categorized in terms of disease severity and adverse outcome. Out of the 175 patients enrolled, 79% presented one or more comorbidities, with cardiovascular disease being the most frequent (62%). More than half of the patients showed lymphocytopenia and 20% thrombocytopenia. The patients in the severe group presented higher absolute neutrophil count (ANC), C-reactive protein (CRP), AST, LDH, procalcitonin (PCT) and BUN values compared to the non-severe group (p 2)/fraction of inspired oxygen (FIO2) ratio < 200 (OR = 4.97; 95% CI 1.55–15.84), clinical severity (OR = 21.32; 95% CI 2.27–200.13), creatinine > 106.08 µmol/L (OR = 2.87; 95% CI 1.04–7.92) and creatine kinase > 2.90 µkat/L (OR = 3.80; 95% CI 1.31–10.9) were observed on admission (p
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  • This record contains raw data related to the article "Coronary artery mechanics induces human saphenous vein remodelling via recruitment of adventitial myofibroblast-like cells mediated by Thrombospondin-1". Rationale: Despite the preferred application of arterial conduits, the greater saphenous vein (SV) remains indispensable for coronary bypass grafting (CABG), especially in multi-vessel coronary artery disease (CAD). The objective of the present work was to address the role of mechanical forces in the activation of maladaptive vein bypass remodeling, a process determining progressive occlusion and recurrence of ischemic heart disease. Methods: We employed a custom bioreactor to mimic the coronary shear and wall mechanics in human SV vascular conduits and reproduce experimentally the biomechanical conditions of coronary grafting and analyzed vein remodeling process by histology, histochemistry and immunofluorescence. We also subjected vein-derived cells to cyclic uniaxial mechanical stimulation in culture, followed by phenotypic and molecular characterization using RNA and proteomic methods. We finally validated our results in vitro and using a model of SV carotid interposition in pigs. Results: Exposure to pulsatile flow determined a remodeling process of the vascular wall involving reduction in media thickness. Smooth muscle cells (SMCs) underwent conversion from contractile to synthetic phenotype. A time-dependent increase in proliferating cells expressing mesenchymal (CD44) and early SMC (SM22α) markers, apparently recruited from the SV adventitia, was observed especially in CABG-stimulated vessels. Mechanically stimulated SMCs underwent transition from contractile to synthetic phenotype. MALDI-TOF-based secretome analysis revealed a consistent release of Thrombospondin-1 (TSP-1), a matricellular protein involved in TGF-β-dependent signaling. TSP-1 had a direct chemotactic effect on SV adventitia resident progenitors (SVPs); this effects was inhibited by blocking TSP-1 receptor CD47. The involvement of TSP-1 in adventitial progenitor cells differentiation and graft intima hyperplasia was finally contextualized in the TGF-β-dependent pathway, and validated in a saphenous vein into carotid interposition pig model. Conclusions: Our results provide the evidence of a matricellular mechanism involved in the human vein arterialization process controlled by alterations in tissue mechanics, and open the way to novel potential strategies to block VGD progression based on targeting cell mechanosensing-related effectors.
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  • Fibroblast growth factor receptor 2 (FGFR2) might have an important role in the pathogenesis and biology of cholangiocarcinoma (CCA). We examined FGFR expression in CCA tumor specimens obtained from patients and CCA cell lines, and then determined the effects of the novel FGFR inhibitor, derazantinib (DZB; formally, ARQ 087), which is currently in clinical phase 2 trials for intrahepatic CCA. DZB inhibited the growth of CCA cell lines in a dose-dependent manner, and extracellular signal-regulated kinase 1/2 and AKT. It also activated apoptotic and cell growth arrest signaling. DZB reduced the in vitro invasiveness and the expression of key epithelial-mesenchymal transition genes. The in vitro data correlated with the expression of FGFRs in human CCA specimens by immunohistochemistry (FGFR1, 30% positive; and FGFR2, 65% positive) and the CCA cell lines assayed by Western blot analysis. These correlated in vitro studies suggest that FGFR may play an important role in the pathogenesis and biology of CCA. Our findings support the notion that FGFR inhibitors, like DZB, should be further evaluated at the clinical stage as targeted therapy for CCA treatment.
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  • Bone marrow Mesenchymal Stem Cells (BM-MSCs), due to their strong protective and anti-inflammatory abilities, have been widely investigated in the context of several diseases for their possible therapeutic role, based on the release of a highly proactive secretome composed of soluble factors and Extracellular Vesicles (EVs). BM-MSC-EVs, in particular, convey many of the beneficial features of parental cells, including direct and indirect β-amyloid degrading-activities, immunoregulatory and neurotrophic abilities. Therefore, EVs represent an extremely attractive tool for therapeutic purposes in neurodegenerative diseases, including Alzheimer's disease (AD). We examined the therapeutic potential of BM-MSC-EVs injected intracerebrally into the neocortex of APPswe/PS1dE9 AD mice at 3 and 5 months of age, a time window in which the cognitive behavioral phenotype is not yet detectable or has just started to appear. We demonstrate that BM-MSC-EVs are effective at reducing the Aβ plaque burden and the amount of dystrophic neurites in both the cortex and hippocampus. The presence of Neprilysin on BM-MSC-EVs, opens the possibility of a direct β-amyloid degrading action. Our results indicate a potential role for BM-MSC-EVs already in the early stages of AD, suggesting the possibility of intervening before overt clinical manifestations.
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