小鼠FICZ暴露与病毒感染

Taisho  Yamada; Akinori Takaoka

doi:10.21769/BioProtoc.2096

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Peer-reviewed

FICZ Exposure and Viral Infection in Mice

小鼠FICZ暴露与病毒感染

Taisho Yamada

Akinori Takaoka email

发布: 2017年01月05日第7卷第1期 DOI: 10.21769/BioProtoc.2096 浏览次数: 8269

评审: Ivan ZanoniMeenal SinhaBenoit Chassaing

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Cover of Nature Immunology, featuring study using the protocol.

May 2016

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Abstract

The aryl hydrocarbon receptor (AHR) is known as a sensor for dioxins that mediates their toxicity, and also has important biophysiological roles such as circadian rhythms, cell differentiation and immune responses. 6-formylindolo(3,2-b)carbazole (FICZ), which is derived through the metabolism of L-tryptophan by ultraviolet B irradiation, is one of putative physiological ligands for AHR (Smirnova et al., 2016). It has recently been shown that endogenously-activated AHR signaling modulates innate immune response during viral infection (Yamada et al., 2016). This section describes how to treat mice with FICZ and to infect them with virus.

Keywords: Aryl hydrocarbon receptor (芳烃受体)

6-formylindolo(3,2-b)carbazole (6-甲酰基吲哚并（3,2-b）咔唑)

Viral infection (病毒感染)

Innate immunity (先天免疫)

Interferon response (干扰素反应)

Background

The role of AHR had so far been investigated mainly on the basis of experiments with dioxin treatment. On the other hand, it has been shown that AHR-mediated signaling is activated by endogenous tryptophan metabolites (FICZ, Kynurenine, etc.), heme metabolites (bilirubin etc.), and eicosanoids (Lipoxin A etc.). In particular, it was demonstrated that FICZ is a physiological high affinity ligand for AHR, and many accumulating reports have shown that FICZ is involved in various basic biological processes including adaptive responses to ultraviolet light, immune responses, genomic instability, and homeostasis of stem cells. Recently, Yamada et al. (2016) demonstrated its role in innate immune response: FICZ treatment in vivo suppresses type I interferon (IFN) production in response to viral infection and promotes levels of viral titer in sera of mice.

Materials and Reagents

0.1-10 μl pipet tips (Thermo Fisher Scientific, Thermo Scientific^TM, catalog number: QSP#TF104 )
1-200 μl pipet tips (Corning, catalog number: 4845 )
100-1,000 μl pipet tips (Corning, catalog number: 4846 )
1.5 ml microcentrifuge tubes (Corning, Axygen^®, catalog number: MCT-150-A )
1 ml syringe (NIPRO, catalog number: 4987458080104 )
27 G hypodermic needle (TERUMO, catalog number: NN-2719S )
Feather disposable scalpel (Sigma-Aldrich, catalog number: Z692395 )
Falcon 6-well tissue culture plate (Corning, Falcon^®, catalog number: 353046 )
C57BL/6NJcl mice (male, 6 weeks of age, purchasable from CLEA Japan, model: C57BL/6NJcl mice )
Vero cells (purchasable from ATCC, catalog number: CCL-81 )
6-formylindolo(3,2-b)carbazole (FICZ) (Enzo Life Sciences, catalog number: BML-GR206-0100 )
Corn oil (stored at room temperature) (Sigma-Aldrich, catalog number: C8267 )
Vesicular stomatitis virus (VSV) (Indiana strain: Kuroda et al., 2016)
Phosphate-buffered saline (PBS) (pH 7.4) (NISSUI PHARMACEUTICAL, catalog number: 05913 )
VeriKine-HS^TM mouse IFN beta serum ELISA kit (Pestka Biomedical Laboratories, catalog number: 42410 )
Dulbecco’s modified Eagle’s medium (DMEM) (‘Nissui’ ②) (NISSUI PHARMACEUTICAL, catalog number: 05919 )
Methyl cellulose, 4,000 cps (Sigma-Aldrich, catalog number: 19-2930 )
Fetal bovine serum (FBS) (Thermo Fisher Scientific, Gibco^TM, catalog number: 10437-028 )
Acetic acids (KANTO CHEMICAL, catalog number: 01021-70 )
Methanol (NACALAI TESQUE, catalog number: 21915-93 )
Crystal violet (Sigma-Aldrich, catalog number: C6158 )
Formaldehyde (Wako Pure Chemical Industries, catalog number: 063-04815 )
FICZ solution (see Recipes)
VSV stock solution (see Recipes)
Fixation solution (see Recipes)
Crystal violet staining solution (see Recipes)

Equipment

Pipettes (PIPETMAN P2) (Gilson, catalog number: F144801 )
Pipettes (PIPETMAN P20) (Gilson, catalog number: F123600 )
Pipettes (PIPETMAN P1000) (Gilson, catalog number: F123602 )
Centrifuge
Deep freezer
Biosafety hood in a biosafety level 3 (BSL3) facility (HITACHI, model: SCV-1303 ECII-AG )
Labnet VX100 vortex (Labnet International, catalog number: 13111-LV2 )