62 research outputs found

    A chemokine network of T cell exhaustion and metabolic reprogramming in renal cell carcinoma

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    Renal cell carcinoma (RCC) is frequently infiltrated by immune cells, a process which is governed by chemokines. CD8+ T cells in the RCC tumor microenvironment (TME) may be exhausted which most likely influence therapy response and survival. The aim of this study was to evaluate chemokine-driven T cell recruitment, T cell exhaustion in the RCC TME, as well as metabolic processes leading to their functional anergy in RCC. Eight publicly available bulk RCC transcriptome collectives (n=1819) and a single cell RNAseq dataset (n=12) were analyzed. Immunodeconvolution, semi-supervised clustering, gene set variation analysis and Monte Carlo-based modeling of metabolic reaction activity were employed. Among 28 chemokine genes available, CXCL9/10/11/CXCR3, CXCL13/CXCR5 and XCL1/XCR1 mRNA expression were significantly increased in RCC compared to normal kidney tissue and also strongly associated with tumor-infiltrating effector memory and central memory CD8+ T cells in all investigated collectives. M1 TAMs, T cells, NK cells as well as tumor cells were identified as the major sources of these chemokines, whereas T cells, B cells and dendritic cells were found to predominantly express the cognate receptors. The cluster of RCCs characterized by high chemokine expression and high CD8+ T cell infiltration displayed a strong activation of IFN/JAK/STAT signaling with elevated expression of multiple T cell exhaustion-associated transcripts. Chemokinehigh RCCs were characterized by metabolic reprogramming, in particular by downregulated OXPHOS and increased IDO1-mediated tryptophan degradation. None of the investigated chemokine genes was significantly associated with survival or response to immunotherapy. We propose a chemokine network that mediates CD8+ T cell recruitment and identify T cell exhaustion, altered energy metabolism and high IDO1 activity as key mechanisms of their suppression. Concomitant targeting of exhaustion pathways and metabolism may pose an effective approach to RCC therapy

    A Single cis Element Maintains Repression of the Key Developmental Regulator Gata2

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    In development, lineage-restricted transcription factors simultaneously promote differentiation while repressing alternative fates. Molecular dissection of this process has been challenging as transcription factor loci are regulated by many trans-acting factors functioning through dispersed cis elements. It is not understood whether these elements function collectively to confer transcriptional regulation, or individually to control specific aspects of activation or repression, such as initiation versus maintenance. Here, we have analyzed cis element regulation of the critical hematopoietic factor Gata2, which is expressed in early precursors and repressed as GATA-1 levels rise during terminal differentiation. We engineered mice lacking a single cis element −1.8 kb upstream of the Gata2 transcriptional start site. Although Gata2 is normally repressed in late-stage erythroblasts, the −1.8 kb mutation unexpectedly resulted in reactivated Gata2 transcription, blocked differentiation, and an aberrant lineage-specific gene expression pattern. Our findings demonstrate that the −1.8 kb site selectively maintains repression, confers a specific histone modification pattern and expels RNA Polymerase II from the locus. These studies reveal how an individual cis element establishes a normal developmental program via regulating specific steps in the mechanism by which a critical transcription factor is repressed

    EpCAM expression varies significantly and is differentially associated with prognosis in the luminal B HER2+, basal-like, and HER2 intrinsic subtypes of breast cancer

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    BACKGROUND: Epithelial cell adhesion molecule (EpCAM) is frequently expressed in breast cancer, and its expression has been associated with poor prognosis. Breast cancer can be subdivided into intrinsic subtypes, differing in prognosis and response to therapy. METHODS: To investigate the association between EpCAM expression and prognosis in the intrinsic subtypes of breast cancer, we performed immunohistochemical studies on a tissue microarray encompassing a total of 1365 breast cancers with detailed clinicopathological annotation and outcomes data. RESULTS: We observed EpCAM expression in 660 out of 1365 (48%) cases. EpCAM expression varied significantly in the different intrinsic subtypes. In univariate analyses of all cases, EpCAM expression was associated with a significantly worse overall survival. In the intrinsic subtypes, EpCAM expression was associated with an unfavourable prognosis in the basal-like and luminal B HER2(+) subtypes but associated with a favourable prognosis in the HER2 subtype. Consistently, specific ablation of EpCAM resulted in increased cell viability in the breast cancer cell line SKBR3 (ER(−), PR(−), and HER2(+)) but decreased viability in the breast cancer cell line MDA-MB-231 (ER(−), PR(−), and HER2(−) ). CONCLUSION: The differential association of EpCAM expression with prognosis in intrinsic subtypes has important implications for the development of EpCAM-targeted therapies in breast cancer

    Effects of EpCAM overexpression on human breast cancer cell lines

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    <p>Abstract</p> <p>Background</p> <p>Recently, EpCAM has attracted major interest as a target for antibody- and vaccine-based cancer immunotherapies. In breast cancer, the EpCAM antigen is overexpressed in 30-40% of all cases and this increased expression correlates with poor prognosis. The use of EpCAM-specific monoclonal antibodies is a promising treatment approach in these patients.</p> <p>Methods</p> <p>In order to explore molecular changes following EpCAM overexpression, we investigated changes of the transcriptome upon EpCAM gene expression in commercially available human breast cancer cells lines Hs578T and MDA-MB-231. To assess cell proliferation, a tetrazolium salt based assay was performed. A TCF/LEF Reporter Kit was used to measure the transcriptional activity of the Wnt/β-catenin pathway. To evaluate the accumulation of β-catenin in the nucleus, a subcellular fractionation assay was performed.</p> <p>Results</p> <p>For the first time we could show that expression profiling data of EpCAM transfected cell lines Hs578T<sup>EpCAM </sup>and MDA-MB-231<sup>EpCAM </sup>indicate an association of EpCAM overexpression with the downregulation of the Wnt signaling inhibitors SFRP1 and TCF7L2. Confirmation of increased Wnt signaling was provided by a TCF/LEF reporter kit and by the finding of the nuclear accumulation of ß-catenin for MDA-MB-231<sup>EpCAM </sup>but not Hs578T<sup>EpCAM </sup>cells. In Hs578T cells, an increase of proliferation and chemosensitivity to Docetaxel was associated with EpCAM overexpression.</p> <p>Conclusions</p> <p>These data show a cell type dependent modification of Wnt signaling components after EpCAM overexpression in breast cancer cell lines, which results in marginal functional changes. Further investigations on the interaction of EpCAM with SFRP1 and TCF7L2 and on additional factors, which may be causal for changes upon EpCAM overexpression, will help to characterize unique molecular properties of EpCAM-positive breast cancer cells.</p

    In vivo single cell analysis reveals Gata2 dynamics in cells transitioning to hematopoietic fate

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    Cell fate is established through coordinated gene expression programs in individual cells. Regulatory networks that include the Gata2 transcription factor play central roles in hematopoietic fate establishment. Although Gata2 is essential to the embryonic development and function of hematopoietic stem cells that form the adult hierarchy, little is known about the in vivo expression dynamics of Gata2 in single cells. Here, we examine Gata2 expression in single aortic cells as they establish hematopoietic fate in Gata2Venus mouse embryos. Time-lapse imaging reveals rapid pulsatile level changes in Gata2 reporter expression in cells undergoing endothelial-to-hematopoietic transition. Moreover, Gata2 reporter pulsatile expression is dramatically altered in Gata2+/- aortic cells, which undergo fewer transitions and are reduced in hematopoietic potential. Our novel finding of dynamic pulsatile expression of Gata2 suggests a highly unstable genetic state in single cells concomitant with their transition to hematopoietic fate. This reinforces the notion that threshold levels of Gata2 influence fate establishment and has implications for transcription factor-related hematologic dysfunctions

    Modelowanie nielokalne w mechanice kontinuum

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    The objective of the paper is to provide an overview of nonlocal formulations for models of elastic solids. The author presents the physical foundations for nonlocal theońes of continuum mechanics, followed by vanous analytical and numeńcal techniques. The characteristics and range of practical applications for the presented approaches are discussed. The results of numerical simulations for the selected case studies are provided to demonstrate the properties of the described methods. The paper is illustrated with outcomes from peridynamic analyses. Fatigue and axial stretching were simulated to show the capabilities of the developed numeńcal tools.W artykule dokonano przeglądu nielokalnych sformułowań dla mechaniki bryły odkształcalnej. Autor przedstawia podstawy fizyczne nielokalnych teorii mechaniki kontinuum oraz dokonuje przeglądu technik analitycznych i numerycznych stosowanych w modelach matematycznych. W pracy przedyskutowano charakterystyki oraz zakres praktycznych zastosowań wspomnianych technik modelowania. Ocena własności nielokalnych modeli została przeprowadzana na podstawie wyników symulacji numerycznych dla wybranych typów analiz z zastosowaniem perydynamiki. Przedstawiono wyniki symulacji zmęczeniowych oraz jednoosiowego rozciągania, uzyskane przy użyciu opracowanych narzędzi analiz symulacyjnych

    Perydynamika w modelowaniu uszkodzeń i symulacji ich propagacji w ujęciu numerycznym

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    The present work provides an overview on the selected applications of peridynamics to damage modelling and simulations of its propagation in mechanical structures. The theoretical fundamentals of the method are briefly described to highlight its advantages and the scope of practical use in the field of computational mechanics. Selected results of numerical analyses are provided to illustrate demanded capabilities. The perspectives of nonlocal and integral based problem formulations for dynamics are discussed.Artykuł stanowi przegląd wybranych zastosowań perydynamiki w modelowaniu uszkodzeń i symulacji zjawisk ich propagacji w konstrukcjach mechanicznych. Przedstawiono podstawy teoretyczne metody ze szczególnym uwzględnieniem specyfiki zalet perydynamiki w zastosowaniach w mechanice komputerowej. Opisywane zalety metody są zilustrowane wybranymi przykładami analiz numerycznych. Artykuł przedstawia możliwości obliczeniowe nielokalnych sformułowań dla dynamiki bazujących na całkowych równaniach ruchu

    Materiały z pamięcią kształtu jako człony wykonawcze w siłowniku Braillea

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    The objective of this paper is to show a designing process for actuators that utilizes shape-memory materials. Analytical studies are carried out to design a dot actuator to be used in an electronic Braille display. Shape-memory materials are a relatively new group of smart materials that exhibit the transformation of their internal structures in solid state phases resulting from external mechanical or thermal excitation. This unique property shows up as three phenomena: one- and two-way memory effects as well as superelasticity, which all significantly influence the change of both the mateńal properties and geometry. Hence, shape-memory mateńals are used as actuators effectively controlled by the above-mentioned types of excitation. The work presents the design guidelines for a proposed technical solution as well as the phenomenon utilized in the elaborated model of the actuator. The authors derived the mathematical descriptions for the proposed design configuration and an example of the calculations is shown. Moreover, the characteristics of the elaborated configuration are also discussed, taking into account the range of application areas.W artykule przedstawiono proces projektowania siłowników zbudowanych z materiałów z pamięcią kształtu z zastosowaniem metod analitycznych na przykładzie członu wykonawczego punktu Braille'a stosowanego w elektronicznych wyświetlaczach alfabetu Braille'a. Materiały z pamięcią kształtu są stosunkowo nową grupą materiałów inteligentnych, które charakteryzują się przebudową struktury wewnętrznej w fazach stałych na skutek zewnętrznego wymuszenia mechanicznego lub termicznego. Ta szczególna własność widoczna jest w trzech zjawiskach: jednokierunkowego oraz dwukierunkowego efektu pamięci kształtu oraz supersprężystości, które istotnie wpływają na zmianę wartości parametrów materiałowych oraz geometrię. W efekcie materiały z pamięcią kształtu znajdują zastosowanie jako napędy w członach wykonawczych, w których skok siłownika jest funkcją jednego z wyżej wymienionych wymuszeń. W artykule przedstawiono założenia projektowe zaproponowanego rozwiązania technicznego oraz omówiono zjawisko wykorzystywane w opracowanym modelu. Autorzy wyprowadzili odpowiednie zależności ilościowe dla przyjętej konfiguracji siłownika, których użycie zaprezentowano na przykładzie obliczeniowym. Poddano dyskusji charakterystyki zaproponowanej konfiguracji projektowej, z uwzględnieniem obszaru jej zastosowania
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