Overexpression of Tim-3 reduces Helicobacter pylori-associated inflammation through TLR4/NFκB signaling in vitro

Wang,Fucai; Mao,Zhirong; Liu,Dongsheng; Yu,Jing; Wang,Youhua; Ye,Wen; Lin,Dongjia; Zhou,Nanjin; Xie,Yong

doi:10.3892/mmr.2017.6346

May-2017 Volume 15 Issue 5

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May-2017 Volume 15 Issue 5

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Article

Overexpression of Tim-3 reduces Helicobacter pylori-associated inflammation through TLR4/NFκB signaling in vitro

Authors:
- Fucai Wang
- Zhirong Mao
- Dongsheng Liu
- Jing Yu
- Youhua Wang
- Wen Ye
- Dongjia Lin
- Nanjin Zhou
- Yong Xie
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Affiliations: Department of Gastroenterology, The First Affiliated Hospital of Nanchang University, Gastroenterology Institute of Jiangxi, Key Laboratory of Digestive Diseases of Jiangxi, Nanchang, Jiangxi 330006, P.R. China, Department of Immunology, Medical College of Nanchang University, Nanchang, Jiangxi 330006, P.R. China, Institute of Immunology and Biological Therapy, Jiangxi Academy of Medical Sciences, Nanchang, Jiangxi 330006, P.R. China
Pages: 3252-3258
|
Published online on: March 21, 2017

https://doi.org/10.3892/mmr.2017.6346
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Abstract

The present study aimed to investigate the interaction between T-cell immunoglobulin and mucin-domain-containing molecule-3 (Tim-3) and Toll-like receptor 4 (TLR4)/nuclear factor κB (NF‑κB) signaling in Helicobacter pylori-infected RAW264.7 macrophage cells. RAW264.7 cells were co‑cultured with H. pylori SS1 at different bacteria/cell ratios, and subsequently the mRNA expression of Tim‑3, TLR4, and myeloid differentiation factor 88 (MyD88) was measured by reverse transcription-quantitative polymerase chain reaction (RT‑qPCR). Furthermore, the effect of Tim‑3 overexpression was examined by transfection of RAW264.7 with pLVX-IRES-ZsGreen-Tim-3 and co‑culturing with H. pylori. mRNA and protein expression levels were then analyzed for Tim‑3, TLR4, MyD88, and phosphorylated (p‑) NF‑κB by RT‑qPCR and western blot analysis respectively. The concentrations of pro‑inflammatory cytokines [tumor necrosis factor‑α (TNF‑α), interleukin 6 (IL-6), interferon‑γ (IFN‑γ) and interleukin 10 (IL‑10)] released in the culture supernatants were measured by ELISA. H. pylori stimulation resulted in a significant increase of Tim‑3, TLR4, and MyD88 mRNA expression in RAW264.7 cells. H. pylori stimulation upregulated Tim‑3 expression even in the Tim‑3‑overexpressing RAW264.7 cells compared with unstimulated cells. TLR4, MyD88, and pNF‑κB protein expression and pro‑inflammatory cytokines (TNF‑α, IL‑6, and IFN‑γ) release levels were increased in the control RAW264.7 cells following H. pylori infection, but not in the Tim-3-overexpressing RAW264.7 cells. By contrast, IL‑10 levels were decreased following H. pylori infection in both control and Tim‑3‑overexpressing RAW264.7 cells. Overexpression of Tim-3 reduced H. pylori-associated inflammation in RAW264.7 macrophages, by downregulating expression of proteins in the TLR4 pathway and release of pro‑inflammatory cytokines. These findings suggest that Tim‑3 serves a crucial role in the negative regulation of H. pylori-associated inflammation and may be a novel therapeutic target for H. pylori infection.

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