Intracranial Injection of Dengue Virus Induces Interferon Stimulated Genes and CD8+ T Cell Infiltration by Sphingosine Kinase 1 Independent Pathways

Copyright: © 2017 Al-Shujairi et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.We have previously reported that the absence of sphingosine kinase 1 (SK1) affects both dengue virus (DENV) infection and innate immune responses in vitro. Here we aimed to define SK1-dependancy of DENV-induced disease and the associated innate responses in vivo. The lack of a reliable mouse model with a fully competent interferon response for DENV infection is a challenge, and here we use an experimental model of DENV infection in the brain of immunocompetent mice. Intracranial injection of DENV-2 into C57BL/6 mice induced body weight loss and neurological symptoms which was associated with a high level of DENV RNA in the brain. Body weight loss and DENV RNA level tended to be greater in SK1-/- compared with wildtype (WT) mice. Brain infection with DENV-2 is associated with the induction of interferon-β (IFN-β) and IFN-stimulated gene (ISG) expression including viperin, Ifi27l2a, IRF7, and CXCL10 without any significant differences between WT and SK1-/- mice. The SK2 and sphingosine-1-phosphate (S1P) levels in the brain were unchanged by DENV infection or the lack of SK1. Histological analysis demonstrated the presence of a cellular infiltrate in DENV-infected brain with a significant increase in mRNA for CD8 but not CD4 suggesting this infiltrate is likely CD8+ but not CD4+ T-lymphocytes. This increase in T-cell infiltration was not affected by the lack of SK1. Overall, DENV-infection in the brain induces IFN and T-cell responses but does not influence the SK/S1P axis. In contrast to our observations in vitro, SK1 has no major influence on these responses following DENV-infection in the mouse brain

Intracranial injection of dengue virus induces interferon stimulated genes and CD8(+) T cell infiltration by sphingosine kinase 1 independent pathways

We have previously reported that the absence of sphingosine kinase 1 (SK1) affects both dengue virus (DENV) infection and innate immune responses in vitro. Here we aimed to define SK1-dependancy of DENV-induced disease and the associated innate responses in vivo. The lack of a reliable mouse model with a fully competent interferon response for DENV infection is a challenge, and here we use an experimental model of DENV infection in the brain of immunocompetent mice. Intracranial injection of DENV-2 into C57BL/6 mice induced body weight loss and neurological symptoms which was associated with a high level of DENV RNA in the brain. Body weight loss and DENV RNA level tended to be greater in SK1-/- compared with wildtype (WT) mice. Brain infection with DENV-2 is associated with the induction of interferon-β (IFN-β) and IFN-stimulated gene (ISG) expression including viperin, Ifi27l2a, IRF7, and CXCL10 without any significant differences between WT and SK1-/- mice. The SK2 and sphingosine-1-phosphate (S1P) levels in the brain were unchanged by DENV infection or the lack of SK1. Histological analysis demonstrated the presence of a cellular infiltrate in DENV-infected brain with a significant increase in mRNA for CD8 but not CD4 suggesting this infiltrate is likely CD8+ but not CD4+ T-lymphocytes. This increase in T-cell infiltration was not affected by the lack of SK1. Overall, DENV-infection in the brain induces IFN and T-cell responses but does not influence the SK/S1P axis. In contrast to our observations in vitro, SK1 has no major influence on these responses following DENV-infection in the mouse brain.Wisam H. Al-Shujairi, Jennifer N. Clarke, Lorena T. Davies, Mohammed Alsharifi, Stuart M. Pitson, Jillian M. Car

Dual Role of Bacteria in Carcinoma: Stimulation and Inhibition

Although what unifies the carcinogenic microorganisms has not been determined by multiple studies, the role of bacteria in the development of neoplasms has not been properly elucidated. In this review, we discuss links between the bacterial species and cancer, with focus on immune responses for the stimulation of tumor cells such as induction of inflammation. Finally, we will describe the potential therapeutic strategies of bacteria on target tumors to improve treatment while mitigating adverse reactions. Cancer is a series of genetic changes that transform normal cells into tumor cells. These changes come from several reasons, including smoking, drinking alcohol, sunlight, exposure to chemical or physical factors, and finally chronic infection with microorganisms, including bacteria. In fact, bacterial infections are not carcinogenic, but recently it was discovered that the association between bacteria and cancer is through two mechanisms, the first stimulating chronic inflammation and the second producing carcinogenic metabolites. While bacteria are carcinogenic agents also, they have a dual role eliminating and removing tumor cells. However, the traditional cancer treatments that include chemotherapy, radiotherapy, surgery, and immunotherapy increase the chances of survival, and there are many side effects of these therapies, including the high toxicity of tissues and normal cells, could not penetrate the tumor cells, and resistance of these therapies by tumor cells. Therefore, the world has turned to an alternative solution, which is the use of genetically engineered microorganisms; thus, the use of living bacteria targeting cancerous cells is the unique option to overcome these challenges. Bacterial therapies, whether used alone or combination with chemotherapy, give a positive effect to treat multiple conditions of cancer. Also, bacteria can be used as vectors for drug, gene, or therapy, and this is a great step to treat cancer. Thus, we review the mechanisms underlying the interaction of the microbiota residents with cancer. Cancer-associated bacteria differ from those in healthy human and are linked with gene-expression profile. We also discuss how live bacteria interact with tumor microenvironments to induce tumor regression through colonization and spread. Finally, we provide past and ongoing clinical trials that include bacteria targeting tumors

Intracranial Injection of Dengue Virus Induces Interferon Stimulated Genes and CD8+ T Cell Infiltration by Sphingosine Kinase 1 Independent Pathways.

We have previously reported that the absence of sphingosine kinase 1 (SK1) affects both dengue virus (DENV) infection and innate immune responses in vitro. Here we aimed to define SK1-dependancy of DENV-induced disease and the associated innate responses in vivo. The lack of a reliable mouse model with a fully competent interferon response for DENV infection is a challenge, and here we use an experimental model of DENV infection in the brain of immunocompetent mice. Intracranial injection of DENV-2 into C57BL/6 mice induced body weight loss and neurological symptoms which was associated with a high level of DENV RNA in the brain. Body weight loss and DENV RNA level tended to be greater in SK1-/- compared with wildtype (WT) mice. Brain infection with DENV-2 is associated with the induction of interferon-β (IFN-β) and IFN-stimulated gene (ISG) expression including viperin, Ifi27l2a, IRF7, and CXCL10 without any significant differences between WT and SK1-/- mice. The SK2 and sphingosine-1-phosphate (S1P) levels in the brain were unchanged by DENV infection or the lack of SK1. Histological analysis demonstrated the presence of a cellular infiltrate in DENV-infected brain with a significant increase in mRNA for CD8 but not CD4 suggesting this infiltrate is likely CD8+ but not CD4+ T-lymphocytes. This increase in T-cell infiltration was not affected by the lack of SK1. Overall, DENV-infection in the brain induces IFN and T-cell responses but does not influence the SK/S1P axis. In contrast to our observations in vitro, SK1 has no major influence on these responses following DENV-infection in the mouse brain

Primer sequences used in this study for qRT-PCR.

<p>Primer sequences used in this study for qRT-PCR.</p

T-cell infiltration in the brain of WT mice following ic infection with DENV-2.

<p>WT mice were ic infected with DENV, as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0169814#pone.0169814.g001" target="_blank">Fig 1</a> and brain tissue was harvested. <b>A.</b> at 3 and 6 dpi, tissue was fixed and processed for H&E staining. Images are representative of n = 3 mice. Arrows indicate sites of cellular infiltrate; <b>B.</b> at the indicated time point RNA was extracted and CD4 and CD8 mRNA determined by qRT-PCR with n = 3 mice at each time point. Data represent average PCR values from individual mice and normalized against GAPDH by ΔCt method. Statistical significance was assessed by unpaired student <i>t</i>-test. * = p < 0.05.</p

The time course of induction of IFN-β and ISGs in WT mice following ic infection with DENV-2.

<p>WT mice (n = 3 at each time point) were ic infected with DENV, as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0169814#pone.0169814.g001" target="_blank">Fig 1</a>. At the time point indicated RNA was isolated from infected mice brain tissues and analysed by real time qRT-PCR for <b>A.</b> IFN-β; <b>B.</b> ISGs viperin, Ifi27l2a, IRF7 and CXCL10. Data represent average PCR values from individual mice and normalized against GAPDH by ΔCt method. Statistical significance was assessed by unpaired student <i>t</i>-test * = p < 0.05, ** = p < 0.005, *** = p < 0.0005.</p

DENV-2 RNA levels increase WT and SK1-/- mice following ic DENV infection.

<p>WT and SK1^-/- mice were ic infected with DENV, as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0169814#pone.0169814.g001" target="_blank">Fig 1</a>. RNA was isolated from infected mice brain tissues and analysed by real time qRT-PCR for DENV. A. Total DENV-2 RNA increases with time in WT mice, n = 3 at each time point; B. RNA was isolated from infected WT and SK1^-/- mice brain tissues at 3 dpi, n = 8 for each strain; C. RNA was isolated from infected mice brain tissues at the time of humane sacrifice, representing 7 (n = 7), 8 (n = 5), 9 (n = 4) or 14 (n = 1) dpi for WT and 7 (n = 7), 8 (n = 3), 9 (n = 2) or 14 (n = 1) dpi for SK1^-/- mice. Each symbol represents an individual mouse sample. Data represent average PCR values from individual mice. Statistical significance was assessed by unpaired Student <i>t</i>-test.</p