Growth delay of human bladder cancer cells by Prostate Stem Cell Antigen downregulation is associated with activation of immune signaling pathways

<p>Abstract</p> <p>Background</p> <p>Prostate stem cell antigen (PSCA) is a glycosylphosphatidylinositol (GPI) anchored protein expressed not only in prostate but also in pancreas and bladder cancer as shown by immunohistochemistry and mRNA analysis. It has been targeted by monoclonal antibodies in preclinical animal models and more recently in a clinical trial in prostate cancer patients. The biological role played in tumor growth is presently unknown. In this report we have characterized the contribution of PSCA expression to tumor growth.</p> <p>Methods</p> <p>A bladder cell line was engineered to express a doxycycline (dox) regulated shRNA against PSCA. To shed light on the PSCA biological role in tumor growth, microarray analysis was carried out as a function of PSCA expression. Expression of gene set of interest was further analyzed by qPCR</p> <p>Results</p> <p>Down regulation of the PSCA expression was associated with reduced cell proliferation <it>in vitro </it>and <it>in vivo</it>. Mice bearing subcutaneous tumors showed a reduced tumor growth upon treatment with dox, which effectively induced shRNA against PSCA as revealed by GFP expression. Pathway analysis of deregulated genes suggests a statistical significant association between PSCA downregulation and activation of genes downstream of the IFNα/β receptor.</p> <p>Conclusions</p> <p>These experiments established for the first time a correlation between the level of PSCA expression and tumor growth and suggest a role of PSCA in counteracting the natural immune response.</p

Rabies Virus Infection Induces Type I Interferon Production in an IPS-1 Dependent Manner While Dendritic Cell Activation Relies on IFNAR Signaling

As with many viruses, rabies virus (RABV) infection induces type I interferon (IFN) production within the infected host cells. However, RABV has evolved mechanisms by which to inhibit IFN production in order to sustain infection. Here we show that RABV infection of dendritic cells (DC) induces potent type I IFN production and DC activation. Although DCs are infected by RABV, the viral replication is highly suppressed in DCs, rendering the infection non-productive. We exploited this finding in bone marrow derived DCs (BMDC) in order to differentiate which pattern recognition receptor(s) (PRR) is responsible for inducing type I IFN following infection with RABV. Our results indicate that BMDC activation and type I IFN production following a RABV infection is independent of TLR signaling. However, IPS-1 is essential for both BMDC activation and IFN production. Interestingly, we see that the BMDC activation is primarily due to signaling through the IFNAR and only marginally induced by the initial infection. To further identify the receptor recognizing RABV infection, we next analyzed BMDC from Mda-5−/− and RIG-I−/− mice. In the absence of either receptor, there is a significant decrease in BMDC activation at 12h post infection. However, only RIG-I−/− cells exhibit a delay in type I IFN production. In order to determine the role that IPS-1 plays in vivo, we infected mice with pathogenic RABV. We see that IPS-1−/− mice are more susceptible to infection than IPS-1+/+ mice and have a significantly increased incident of limb paralysis

Challenging local realism with human choices

A Bell test is a randomized trial that compares experimental observations against the philosophical worldview of local realism 1, in which the properties of the physical world are independent of our observation of them and no signal travels faster than light. A Bell test requires spatially distributed entanglement, fast and high-efficiency detection and unpredictable measurement settings 2,3 . Although technology can satisfy the first two of these requirements 4-7, the use of physical devices to choose settings in a Bell test involves making assumptions about the physics that one aims to test. Bell himself noted this weakness in using physical setting choices and argued that human 'free will' could be used rigorously to ensure unpredictability in Bell tests 8 . Here we report a set of local-realism tests using human choices, which avoids assumptions about predictability in physics. We recruited about 100,000 human participants to play an online video game that incentivizes fast, sustained input of unpredictable selections and illustrates Bell-test methodology 9 . The participants generated 97,347,490 binary choices, which were directed via a scalable web platform to 12 laboratories on five continents, where 13 experiments tested local realism using photons 5,6, single atoms 7, atomic ensembles 10 and superconducting devices 11 . Over a 12-hour period on 30 November 2016, participants worldwide provided a sustained data flow of over 1,000 bits per second to the experiments, which used different human-generated data to choose each measurement setting. The observed correlations strongly contradict local realism and other realistic positions in bipartite and tripartite 12 scenarios. Project outcomes include closing the 'freedom-of-choice loophole' (the possibility that the setting choices are influenced by 'hidden variables' to correlate with the particle properties 13 ), the utilization of video-game methods 14 for rapid collection of human-generated randomness, and the use of networking techniques for global participation in experimental science

Challenging local realism with human choices

A Bell test is a randomized trial that compares experimental observations against the philosophical worldview of local realism. A Bell test requires spatially distributed entanglement, fast and high-efficiency detection and unpredictable measurement settings. Although technology can satisfy the first two of these requirements, the use of physical devices to choose settings in a Bell test involves making assumptions about the physics that one aims to test. Bell himself noted this weakness in using physical setting choices and argued that human `free will' could be used rigorously to ensure unpredictability in Bell tests. Here we report a set of local-realism tests using human choices, which avoids assumptions about predictability in physics. We recruited about 100,000 human participants to play an online video game that incentivizes fast, sustained input of unpredictable selections and illustrates Bell-test methodology. The participants generated 97,347,490 binary choices, which were directed via a scalable web platform to 12 laboratories on five continents, where 13 experiments tested local realism using photons, single atoms, atomic ensembles, and superconducting devices. Over a 12-hour period on 30 November 2016, participants worldwide provided a sustained data flow of over 1,000 bits per second to the experiments, which used different human-generated data to choose each measurement setting. The observed correlations strongly contradict local realism and other realistic positions in bipartite and tripartite scenarios. Project outcomes include closing the `freedom-of-choice loophole' (the possibility that the setting choices are influenced by `hidden variables' to correlate with the particle properties), the utilization of video-game methods for rapid collection of human generated randomness, and the use of networking techniques for global participation in experimental science.Comment: This version includes minor changes resulting from reviewer and editorial input. Abstract shortened to fit within arXiv limit

Generation in vivo of peptide-specific cytotoxic T cells and presence of regulatory T cells during vaccination with hTERT (class I and II) peptide-pulsed DCs

Optimal techniques for DC generation for immunotherapy in cancer are yet to be established. Study aims were to evaluate: (i) DC activation/maturation milieu (TNF-α +/- IFN-α) and its effects on CD8+ hTERT-specific T cell responses to class I epitopes (p540 or p865), (ii) CD8+ hTERT-specific T cell responses elicited by vaccination with class I alone or both class I and II epitope (p766 and p672)-pulsed DCs, prepared without IFN-α, (iii) association between circulating T regulatory cells (Tregs) and clinical responses

IFN-α boosts epitope cross-presentation by dendritic cells via modulation of proteasome activity

Wehave investigated the molecular mechanisms underlying the peculiar cross-presentation efficiency of human dendritic cells (DCs) differentiated from monocytes in the presence of Granulocyte-Macrophage Colony-Stimulating Factor (GM-CSF) and Interferon (IFN)- (IFN-DCs). To this end, we evaluated the capability of IFN-DCs to present and cross-present epitopes derived from Epstein–Barr Virus (EBV) or human melanoma-associated antigens after exposure to cell lysates or apoptotic cells. In an autologous setting, IFN-DCs loaded with Lymphoblastoid Cell Lines (LCL) lysates or apoptotic LCL were highly efficient in expanding, respectively, EBV-specific class II- or class I-restricted memory T cell responses. Of note, IFN-DCs loaded with apoptotic LCL were more potent than fully mature DCs in triggering the cytotoxicity of CD8+ T lymphocytes recognizing a subdominant HLA-A*0201-restricted epitope derived from EBV latent membrane protein 2 (LMP2). In addition, IFN-DCs loaded with apoptotic human melanoma cells were highly efficient in cross-presenting the MART-127–35 epitope to a specific CD8+ cytotoxic T cell clone, and this functional activity was proteasome-dependent. These IFN-DC properties were associated with an enhanced expression of all the immunoproteasome subunits as well as of TAP-1, TAP-2 and tapasin, and, interestingly, to a higher proteasome proteolytic activity as compared to immature or mature DCs. Altogether, these results highlight new mechanisms by which IFN- influences antigen processing and cross-presentation ability of monocyte-derived DCs, with potentially important implications for the development of DC-based therapeutic vaccines

Interferon-alpha and viral triggers promote functional maturation of human monocyte-derived dendritic cells

BACKGROUND: Type I interferons (IFNs) play an important role in the pathogenesis of many autoimmune disorders including psoriasis. In the presence of IFN-α and granulocyte/macrophage colony-stimulating factor (GM-CSF), monocytes differentiate into dendritic cells (DCs) referred to as IFN-DCs. IFN-DCs potentially mimic DC populations involved in psoriasis and express a wide range of Toll-like receptor (TLR) subtypes. OBJECTIVES: Recently, it was shown that single-stranded RNA (ssRNA) triggers TLR7 and TLR8; therefore we studied ssRNA, as a surrogate for ssRNA viruses and their impact on IFN-DCs. METHODS: We established culture conditions for IFN-DCs, generated from plastic adherent monocytes using GM-CSF plus IFN-α. For DC stimulation ssRNA40, a 20-mer ssRNA oligonucleotide was used. The phenotypic analysis of DC preparations was performed using flow cytometry. The production of various cytokines was analysed by enzyme-linked immunosorbent assay, and real-time quantitative polymerase chain reaction was used to quantify TLR and cytokine gene expression. The ability of IFN-DCs to stimulate allogeneic T-cell proliferation was evaluated in a mixed leucocyte reaction. RESULTS: We found that IFN-DCs express mRNA for TLR7 and TLR8 and that ssRNA stimulation significantly improves their costimulatory molecule expression, stabilizes their phenotype and enhances their capacity to stimulate naive T-cell proliferation. Unstimulated IFN-DCs did not produce bioactive interleukin (IL)-12 and produced low levels of other proinflammatory cytokines. In contrast, ssRNA stimulation led to a significant production of IL-12p70, IL-1β, IL-6 and tumour necrosis factor α. IFN-DCs contained mRNA for IL-12p35, IL-12p40, IL-23p19, IL-27p28 and IL-27EBI, which was further increased by incubation with ssRNA. CONCLUSIONS: Our study sheds light on a potential role for IFN-α and viral infections in triggering DC populations in psoriasis. These results provide additional data for the better understanding of human autoimmune and antiviral responses and may also have implications for strategies developing cancer immunotherapy