Let-7b is involved in the inflammation and immune responses associated with Helicobacter pylori infection by targeting Toll-like receptor 4.
Let-7b is involved in the inflammation and immune responses associated with Helicobacter pylori infection by targeting Toll-like receptor 4.
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Let-7b 通过靶向 Toll 样受体 4 参与与幽门螺杆菌感染相关的炎症和免疫反应
DOI：
10.1371/journal.pone.0056709
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发表时间：
2013
期刊：
影响因子：
3.7
通讯作者：
Li J
中科院分区：
文献类型：
--
作者：
Teng GG;Wang WH;Dai Y;Wang SJ;Chu YX;Li J
目的Toll样受体（Toll like receptor，TLR）是机体对微生物感染的天然免疫应答的重要启动因子。TLR 4在胃上皮细胞中响应于幽门螺杆菌感染而上调。然而，TLR 4在幽门螺杆菌感染中表达的调控机制尚未明确。本研究的目的是证实microRNA let-7 B在人胃上皮细胞中直接调控TLR 4的表达，进而影响NF-κB的活化及下游基因的表达。方法采用定量RT-PCR方法检测let-7 b在正常胃黏膜组织和两种胃上皮细胞系中的表达。采用免疫组化和RT-PCR方法检测TLR 4的表达。通过荧光素酶报告基因分析和Western blot鉴定let-7 b的潜在靶点。使用Let-7 B模拟物和抑制剂检测let-7 B对NF-κB活性的影响。还测定了感染幽门螺杆菌26695的细胞中NF-κB下游基因的表达。结果Hp 26695（cagA+）感染的胃粘膜标本和胃上皮细胞株（AGS、GES-1）中Let-7 b表达明显降低。Let-7 b与TLR 4 mRNA的3′-UTR互补，通过转录后抑制调节胃上皮细胞TLR 4的表达。H.pylori感染可诱导AGS和GES-1细胞TLR 4的表达和NF-κB的活化。过表达let-7 B可下调TLR 4表达，随后减弱NF-κB、MyD 88、NF-κB1/p50、RelA/p65。let-7 b过表达的细胞IL-8、考克斯-2和CyclinD 1的表达受到抑制。TAK-242（TLR 4抑制剂）和SN 50（NF-κB抑制剂）均显著抑制幽门螺杆菌诱导的let-7 B下调。结论Let-7 B靶向TLR 4 mRNA，调控NF-κB B的活化及下游炎症和免疫相关基因的表达。
Objectives Toll-like receptors (TLRs) are important initiators in native immune responses to microbial infections. TLR4 is up-regulated in response to H.pylori infection in gastric epithelial cells. However, the regulatory mechanisms for the expression of TLR4 in H.pylori infection have not been clearly defined. The aims of this study are to present the evidence that microRNA let-7b directly regulates TLR4 expression in human gastric epithelial cells, and subsequently influences the activation of NF-κB and the expression of the downstream genes in H.pylori infection. Methods The expression of let-7b was determined in gastric mucosa specimens and in two gastric epithelial cell lines using quantitative RT-PCR. The expression of TLR4 was determined by immunohistochemistry staining and RT-PCR. The potential target of let-7b was identified by luciferase reporter assay and Western blot. Let-7b mimics and inhibitors were used to examine the effects of let-7b on NF-κB activity. The expression of the downstream genes of NF-κB was also determined in cells infected with H.pylori 26695. Results Let-7b was significantly decreased in gastric mucosa specimens and in gastric epithelial cell lines (AGS, GES-1) infected with H.pylori 26695 (cagA+). Let-7b was complementary to the 3′-UTR of TLR4 mRNA and regulated TLR4 expression via post-transcriptional suppression in gastric epithelium. Infection of H.pylori induced the expression of TLR4 and activated NF-κB in AGS and GES-1 cells. Overexpression of let-7b by mimics downregulated TLR4, and subsequently attenuated NF-κB, MyD88, NF-κB1/p50, RelA/p65. The expression of IL-8, COX-2 and CyclinD1 was inhibited in H.pylori infected cells with let-7b overexpression. Both TAK-242 (TLR4 inhibitor) and SN50 (NF-κB inhibitor) significantly inhibited the H.pylori induced downregulation of let-7b. Conclusions Let-7b targets at TLR4 mRNA, and regulates the activation of NF-κB and the expression of the downstream genes related to the inflammation and immune responses in H.pylori infection.
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