Contribution of IPS-1 to polyI:C-induced cytokine production in conjunctival epithelial cells
Contribution of IPS-1 to polyI:C-induced cytokine production in conjunctival epithelial cells
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DOI：
10.1016/j.bbrc.2010.11.136
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发表时间：
2011-01-07
影响因子：
3.1
通讯作者：
Kinoshita, Shigeru
中科院分区：
文献类型：
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作者：
Ueta, Mayumi;Kawai, Taro;Kinoshita, Shigeru
我们以前证明，眼表上皮细胞表达TLR 3和它的配体，polyI：C，刺激诱导的炎性细胞因子和I型IFN的分泌。最近报道RIG-I和MDA 5也识别模拟polyI：C的病毒dsRNA。在这项研究中，我们调查是否RIG-I和/或MDA 5有助于polyI：C诱导的反应，结膜上皮。通过RT-PCR检测人结膜上皮中RIG-I、MDA 5和TLR 3的表达，并通过定量RT-PCR和免疫印迹分析检测polyI：C刺激后RIG-I、MDA 5和TLR 3的上调。人结膜上皮细胞也表达RIG-I、MDA 5和TLR 3 mRNA和蛋白。在polyI：C刺激后，RIG-I和MDA 5的表达显著上调，但TLR 3的表达不上调。我们还使用IPS-1 KO和TLR 3 KO小鼠检查了IPS-1（RIG-I和/或MDA 5共有的衔接分子）和TLR 3在结膜上皮中的功能。为了在体内分析鼠结膜上皮细胞，将10 μ l的100 μ g/ml polyI：C溶液以滴眼液形式结膜下递送，然后对结膜上皮细胞进行基因表达分析。我们专注于10转录上调后，在小鼠结膜上皮polyI：C刺激。Cxcl 10、Mx 1、Ifi 44、Ifi 203、Iigp 2和Rtp 4主要受IPS-1调节，Ccl 5主要受TLR 3调节，Rsad 2主要受IPS-1调节。Mx2和Cmpk 2受TLR 3和IPS-1调节。我们的研究结果表明，结膜上皮细胞表达RIG-I和MDA 5，IPS-1，一种RIG-I和MDA 5共同的衔接分子，有助于结膜上皮细胞中polyI：C诱导的细胞因子的产生。(C)2010年爱思唯尔公司All rights reserved.
We previously demonstrated that ocular surface epithelium expressed TLR3 and that its ligand, polyI:C, stimulation induced the secretion of inflammatory cytokines and type I IFN. It was recently reported that RIG-I and MDA5 also recognize viral dsRNA mimicking polyI:C. In this study, we investigated whether RIG-I and/or MDA5 contribute to polyI:C-inducible responses in conjunctival epithelium. The expression of RIG-I, MDA5, and TLR3 in human conjunctival epithelium was examined by RT-PCR and their up-regulation after polyI:C stimulation by quantitative RT-PCR and immunoblot analysis. Human conjunctival epithelial cells also expressed RIG-I, MDA5 and TLR3 mRNA and protein. The expression of RIG-I and MDA5, but not of TLR3, was markedly up-regulated upon polyI:C stimulation. We also examined the function of IPS-1 (an adaptor molecule common to RIG-I and/or MDA5) and TLR3 in conjunctival epithelium using IPS-1 KO and TLR3 KO mice. To analyze in vivo murine conjunctival epithelial cells, 10 mu l of a 100 mu g/ml polyI:C solution were delivered subconjunctivally and as eye drops, then conjunctival epithelial cells were subjected to gene expression analysis. We focused on 10 transcripts up-regulated in murine conjunctival epithelium upon polyI:C stimulation. Cxcl10, Mx1, Ifi44, Ifi203, Iigp2 and Rtp4 were dominantly regulated by IPS-1, Ccl5 by TLR3, and Rsad2. Mx2 and Cmpk2 were regulated by TLR3 and IPS-1. Our results showed that conjunctival epithelial cells express RIG-I and MDA5, and IPS-1, an adaptor molecule common to RIG-I and MDA5, contributes to polyI:C-inducible cytokine production in conjunctival epithelial cells. (C) 2010 Elsevier Inc. All rights reserved.