Correction: F11-Mediated Inhibition of RhoA Signalling Enhances the Spread of Vaccinia Virus In Vitro and In Vivo in an Intranasal Mouse Model of Infection

Bystander activation, i.e., activation of T cells specific for an antigen X during an immune response against antigen Y may occur during viral infections. However, the low frequency of bystander-activated T cells has rendered it difficult to define the mechanisms and possible in vivo relevance of this nonspecific activation. This study uses transgenic mice expressing a major histocompatibility complex class I–restricted TCR specific for glycoprotein peptide 33-41 of lymphocytic choriomeningitis virus (LCMV) to overcome this limitation. CD8+ T cells from specific pathogen-free maintained, unimmunized “naive” TCR transgenic mice can differentiate into LCMV-specific cytolytic effector CTL during infections with vaccinia virus or Listeria monocytogenes in vivo or mixed lymphocyte culture in vitro. We show that in these model situations (a) nonspecifically activated CTL are able to confer antiviral protection in vivo, (b) bystander activation is largely independent of the expression of a second T cell receptor of different specificity, (c) bystander activation is not mediated by a broadly cross-reactive TCR, but rather by cytokines, (d) bystander activation can be mediated by cytokines such as IL-2, but not α/β-IFN in vitro; (e) bystander activation is, overall, a rare event, occuring in vivo in roughly 1 in 200 of the LCMV-specific CTL during infection of TCR transgenic mice with vaccinia virus; (f) bystander activation does not have a significant functional impact on nontransgenic CTL memory under the conditions tested; and (g) even in the TCR transgenic situation, where unphysiologically high numbers of T cells of a single specificity are present, bystander activation is not sufficient to cause clinically manifest autoimmune disease in a transgenic mouse model of diabetes. We conclude that although bystander activation via cytokines may generate cytolytically active CTL from naive precursors, quantitative considerations suggest that this is usually not of major biological consequence.

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Treating Influenza and SARS-CoV-2 via mRNA-encoded Cas13a

10.1101/2020.04.24.060418 ◽

2020 ◽

Cited By ~ 2

Author(s):

Emmeline L. Blanchard ◽

Daryll Vanover ◽

Swapnil Subhash Bawage ◽

Pooja Munnilal Tiwari ◽

Laura Rotolo ◽

...

Keyword(s):

Influenza Virus ◽

Mouse Model ◽

Post Infection ◽

Cytopathic Effect ◽

Respiratory Infections ◽

Synthetic Mrna ◽

Mouse Model Of Infection ◽

Over Time

ABSTRACTHere, Cas13a has been used to target and mitigate influenza virus A (IAV) and SARS-CoV-2 using a synthetic mRNA-based platform. CRISPR RNAs (crRNA) against PB1 and highly conserved regions of PB2 were screened in conjunction with mRNA-encoded Cas13a. Screens were designed such that only guides that decreased influenza RNA levels in a Cas13-mediated fashion, were valid. Cas13a mRNA and validated guides, delivered post-infection, simulating treatment, were tested in combination and across multiplicities of infection. Their function was also characterized over time. Similar screens were performed for guides against SARS-CoV-2, yielding multiple guides that significantly impacted cytopathic effect. Last, the approach was utilized in vivo, demonstrating the ability to degrade influenza RNA in a mouse model of infection, using polymer-formulated, nebulizer-based mRNA delivery. Our findings demonstrate the applicability of Cas13a in mitigating respiratory infections both in vitro and in a mouse model, paving the way for future therapeutic use.

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In Vivo Investigations of Selected Diamidine Compounds against Trypanosoma evansi Using a Mouse Model

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00422-09 ◽

2009 ◽

Vol 53 (12) ◽

pp. 5074-5079 ◽

Cited By ~ 7

Author(s):

Kirsten Gillingwater ◽

Arvind Kumar ◽

Mariappan Anbazhagan ◽

David W. Boykin ◽

Richard R. Tidwell ◽

...

Keyword(s):

Mouse Model ◽

Trypanosoma Evansi ◽

Chemotherapeutic Agents ◽

Control Measures ◽

Lead Compounds ◽

Protozoan Infection ◽

Drug Doses ◽

Mouse Model Of Infection

ABSTRACT Surra is an animal pathogenic protozoan infection, caused by Trypanosoma evansi, that develops into a fatal wasting disease. Control measures rely on diagnosis and treatment. However, with the continuous emergence of drug resistance, this tactic is failing, and the pressing need for new chemotherapeutic agents is becoming critical. With the introduction of novel aromatic diamidines, a new category of antitrypanosomal drugs was discovered. Nevertheless, their efficacy within a T. evansi-infected mouse model was not known. In total, 30 compounds previously selected based on their in vitro activity were tested in a T. evansi mouse model of infection. Six of the compounds were capable of curing T. evansi-infected mice at drug doses as low as 0.5 and 0.25 mg/kg of body weight administered for 4 consecutive days, and they were more effective than the standard drugs suramin, diminazene, and quinapyramine. After all selection criteria were applied, three diamidine compounds (DB 75, DB 867, and DB 1192) qualified as lead compounds and were considered to have the potential to act as preclinical candidates against T. evansi infection.

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A novel phosphoramide compound, DCZ0805, shows potent anti-myeloma activity via the NF-κB pathway

Cancer Cell International ◽

10.1186/s12935-021-01973-1 ◽

2021 ◽

Vol 21 (1) ◽

Author(s):

Xuejie Gao ◽

Bo Li ◽

Anqi Ye ◽

Houcai Wang ◽

Yongsheng Xie ◽

...

Keyword(s):

Mouse Model ◽

Tumor Burden ◽

High Rate ◽

Cell Counting ◽

Tumor Xenograft ◽

Luciferase Reporter ◽

Therapeutic Effects ◽

Xenograft Mouse Model

Abstract Background Multiple myeloma (MM) is a highly aggressive and incurable clonal plasma cell disease with a high rate of recurrence. Thus, the development of new therapies is urgently needed. DCZ0805, a novel compound synthesized from osalmide and pterostilbene, has few observed side effects. In the current study, we intend to investigate the therapeutic effects of DCZ0805 in MM cells and elucidate the molecular mechanism underlying its anti-myeloma activity. Methods We used the Cell Counting Kit-8 assay, immunofluorescence staining, cell cycle assessment, apoptosis assay, western blot analysis, dual-luciferase reporter assay and a tumor xenograft mouse model to investigate the effect of DCZ0805 treatment both in vivo and in vitro. Results The results showed that DCZ0805 treatment arrested the cell at the G0/G1 phase and suppressed MM cells survival by inducing apoptosis via extrinsic and intrinsic pathways. DCZ0805 suppressed the NF-κB signaling pathway activation, which may have contributed to the inhibition of cell proliferation. DCZ0805 treatment remarkably reduced the tumor burden in the immunocompromised xenograft mouse model, with no obvious toxicity observed. Conclusion The findings of this study indicate that DCZ0805 can serve as a novel therapeutic agent for the treatment of MM.

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Selective Pseudo-irreversible Butyrylcholinesterase Inhibitors Transferring Antioxidant Moieties to the Enzyme Show Pronounced Neuroprotective Efficacy In Vitro and In Vivo in an Alzheimer’s Disease Mouse Model

Journal of Medicinal Chemistry ◽

10.1021/acs.jmedchem.1c00534 ◽

2021 ◽

Author(s):

Matthias Scheiner ◽

Matthias Hoffmann ◽

Feng He ◽

Eleonora Poeta ◽

Arnaud Chatonnet ◽

...

Keyword(s):

Alzheimer’S Disease ◽

Alzheimer's Disease ◽

Mouse Model ◽

Neuroprotective Efficacy

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MODL-12. DEVELOPMENT OF A NOVEL IMMUNOCOMPETENT MOUSE MODEL FOR DIFFUSE INTRINSIC PONTINE GLIOMA

Neuro-Oncology ◽

10.1093/neuonc/noaa222.586 ◽

2020 ◽

Vol 22 (Supplement_3) ◽

pp. iii413-iii413

Author(s):

Maggie Seblani ◽

Markella Zannikou ◽

Katarzyna Pituch ◽

Liliana Ilut ◽

Oren Becher ◽

...

Keyword(s):

Flow Cytometry ◽

Mouse Model ◽

Growth Factor Receptor ◽

Diffuse Intrinsic Pontine Glioma ◽

Pontine Glioma ◽

Time Of Application ◽

Tumor Associated Antigen ◽

Sarcoma Virus

Abstract Diffuse intrinsic pontine glioma (DIPG) is a devastating brain tumor affecting young children. Immunotherapies hold promise however the lack of immunocompetent models recreating a faithful tumor microenvironment (TME) remains a challenge for development of targeted immunotherapeutics. We propose to generate an immunocompetent DIPG mouse model through induced overexpression of interleukin 13 receptor alpha 2 (IL13Rα2), a tumor-associated antigen overexpressed by glioma cells. A model with an intact TME permits comprehensive preclinical assessment of IL13Rα2-targeted immunotherapeutics. Our novel model uses the retroviral avian leucosis and sarcoma virus (RCAS) for in vivo gene delivery leading to IL13Rα2 expression in proliferating progenitor cells. Transfected cells expressing IL13Rα2 and PDGFB, a ligand for platelet derived growth factor receptor, alongside induced p53 loss via the Cre-Lox system are injected in the fourth ventricle in postnatal pups. We validated the expression of PDGFB and IL13Rα2 transgenes in vitro and in vivo and will characterize the TME through evaluation of the peripheral and tumor immunologic compartments using immunohistochemistry and flow cytometry. We confirmed expression of transgenes via flow cytometry and western blotting. Comparison of survival dynamics in mice inoculated with PDGFB alone with PDGFB+IL13Rα2 demonstrated that co-expression of IL13Rα2 did not significantly affect mice survival compared to the PDGFB model. At time of application, we initiated experiments to characterize the TME. Preliminary data demonstrate establishment of tumors within and adjacent to the brainstem and expression of target transgenes. Preclinical findings in a model recapitulating the TME may provide better insight into outcomes upon translation to clinical application.

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Neutrophil subtypes shape HIV-specific CD8 T-cell responses after vaccinia virus infection

npj Vaccines ◽

10.1038/s41541-021-00314-7 ◽

2021 ◽

Vol 6 (1) ◽

Author(s):

Mauro Di Pilato ◽

Miguel Palomino-Segura ◽

Ernesto Mejías-Pérez ◽

Carmen E. Gómez ◽

Andrea Rubio-Ponce ◽

...

Keyword(s):

T Cell ◽

Immune Responses ◽

Vaccinia Virus ◽

Adaptive Immune System ◽

T Cell Responses ◽

Cd8 T Cell ◽

Cell Responses ◽

Adaptive Immune

AbstractNeutrophils are innate immune cells involved in the elimination of pathogens and can also induce adaptive immune responses. Nα and Nβ neutrophils have been described with distinct in vitro capacity to generate antigen-specific CD8 T-cell responses. However, how these cell types exert their role in vivo and how manipulation of Nβ/Nα ratio influences vaccine-mediated immune responses are not known. In this study, we find that these neutrophil subtypes show distinct migratory and motility patterns and different ability to interact with CD8 T cells in the spleen following vaccinia virus (VACV) infection. Moreover, after analysis of adhesion, inflammatory, and migration markers, we observe that Nβ neutrophils overexpress the α4β1 integrin compared to Nα. Finally, by inhibiting α4β1 integrin, we increase the Nβ/Nα ratio and enhance CD8 T-cell responses to HIV VACV-delivered antigens. These findings provide significant advancements in the comprehension of neutrophil-based control of adaptive immune system and their relevance in vaccine design.

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TMOD-24. GENERATION AND CHARACTERIZATION OF A NOVEL TRANSGENIC IDO REPORTER MOUSE FOR IDO POSTTRANSLATIONAL MODIFICATION ANALYSIS IN SITU

Neuro-Oncology ◽

10.1093/neuonc/noaa215.974 ◽

2020 ◽

Vol 22 (Supplement_2) ◽

pp. ii233-ii233

Author(s):

April Bell ◽

Lijie Zhai ◽

Erik Ladomersky ◽

Kristen Lauing ◽

Lakshmi Bollu ◽

...

Keyword(s):

Mouse Model ◽

Tumor Cell ◽

Central Nervous System Tumor ◽

Post Translational Modifications ◽

Reporter Mouse ◽

C Terminus ◽

Human Gbm

Abstract Glioblastoma (GBM) is the most common and aggressive primary central nervous system tumor in adults with a median survival of 14.6 months. GBM is a potently immunosuppressive cancer due in-part to the prolific expression of immunosuppressive indoleamine 2,3 dioxygenase 1 (IDO). Tumor cell IDO facilitates the intratumoral accumulation of regulatory T cells (Tregs; CD4+CD25+FoxP3+). Although immunosuppressive IDO activity is canonically characterized by the conversion of tryptophan into kynurenine, we have utilized transgenic and syngeneic mouse models and mutant glioma lines to demonstrate that tumor cell IDO increases Treg accumulation independent of tryptophan metabolism. Here, we address the gap in our understanding of IDO signaling activity in vivo. Subcutaneously-engrafted human GBM expressing human IDO-GFP cDNA was isolated from immunodeficient humanized NSG-SGM3 mice. The tumor was immunoprecipitated for the GFP tag using GFP-TRAP followed by mass spectrometry which revealed a novel methylation site on a lysine residue at amino acid 373 in the IDO C-terminus region. Western blot analysis of IDO protein also revealed the presence of tyrosine phosphorylation. Additionally, we recently created a new transgenic IDO reporter mouse model whereby endogenous IDO is fused to GFP via a T2A linker (IDO→GFP). This model allows for the isolation of IDO+ cells in real-time and without causing cell death, thereby creating the opportunity for downstream molecular analysis of in situ-isolated GFP+ cells. Collectively, our work suggests that IDO non-enzyme activity may involve the post-translational modifications we recently identified. As IDO activity may differ between in vitro and in vivo modeling systems, we will use the new IDO→GFP reporter mouse model for an improved mechanistic understanding of how immunosuppressive IDO facilitates Treg accumulation in vivo.

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Exosome-transported circRNA_0001236 enhances chondrogenesis and suppress cartilage degradation via the miR-3677-3p/Sox9 axis

Stem Cell Research & Therapy ◽

10.1186/s13287-021-02431-5 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Guping Mao ◽

Yiyang Xu ◽

Dianbo Long ◽

Hong Sun ◽

Hongyi Li ◽

...

Keyword(s):

Bone Marrow ◽

Mouse Model ◽

Chondrogenic Differentiation ◽

Cartilage Degradation ◽

Human Bone ◽

Human Bone Marrow ◽

Luciferase Reporter ◽

Gene And Protein Expression

Abstract Objectives Aberrations in exosomal circular RNA (circRNA) expression have been identified in various human diseases. In this study, we investigated whether exosomal circRNAs could act as competing endogenous RNAs (ceRNAs) to regulate the pathological process of osteoarthritis (OA). This study aimed to elucidate the specific MSC-derived exosomal circRNAs responsible for MSC-mediated chondrogenic differentiation using human bone marrow-derived MSCs (hMSCs) and a destabilization of the medial meniscus (DMM) mouse model of OA. Methods Exosomal circRNA deep sequencing was performed to evaluate the expression of circRNAs in human bone marrow-derived MSCs (hMSCs) induced to undergo chondrogenesis from day 0 to day 21. The regulatory and functional roles of exosomal circRNA_0001236 were examined on day 21 after inducing chondrogenesis in hMSCs and were validated in vitro and in vivo. The downstream target of circRNA_0001236 was also explored in vitro and in vivo using bioinformatics analyses. A luciferase reporter assay was used to evaluate the interaction between circRNA_0001236 and miR-3677-3p as well as the target gene sex-determining region Y-box 9 (Sox9). The function and mechanism of exosomal circRNA_0001236 in OA were explored in the DMM mouse model. Results Upregulation of exosomal circRNA_0001236 enhanced the expression of Col2a1 and Sox9 but inhibited that of MMP13 in hMSCs induced to undergo chondrogenesis. Moreover, circRNA_0001236 acted as an miR-3677-3p sponge and functioned in human chondrocytes via targeting miR-3677-3p and Sox9. Intra-articular injection of exosomal circRNA_0001236 attenuated OA in the DMM mouse model. Conclusions Our results reveal an important role for a novel exosomal circRNA_0001236 in chondrogenic differentiation. Overexpression of exosomal circRNA_0001236 promoted cartilage-specific gene and protein expression through the miR-3677-3p/Sox9 axis. Thus, circRNA_0001236-overexpressing exosomes may alleviate cartilage degradation, suppressing OA progression and enhancing cartilage repair. Our findings provide a potentially effective therapeutic strategy for treating OA.

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