A decoy peptide derived from TIR domain of TIRAP inhibits TLR signaling pathway
A decoy peptide derived from TIR domain of TIRAP inhibits TLR signaling pathway
- 주제(키워드) TLR inhibiting peptide , innate immune , Toll-like receptor 4 , Toll-like receptor 3
- 발행기관 아주대학교
- 지도교수 최상돈
- 발행년도 2017
- 학위수여년월 2017. 2
- 학위명 석사
- 학과 및 전공 일반대학원 분자과학기술학과
- 실제URI http://www.dcollection.net/handler/ajou/000000024263
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
ABSTRACT Toll-like receptors (TLRs) belong to the family of type I transmembrane proteins. These receptors are capable of recognizing specific parts of microbes and are able of activating different groups of signaling molecules. The activation of these receptors leads to a protective innate immune response against invading microbial and viral pathogens along with the production of inflammatory cytokines, chemokines, and type I interferons. However, an excessive TLRs-mediated immune response is associated with the onset and/or the aggravation of various inflammatory and auto-immune diseases. Thus, targeting TLRs or TLRs adaptor molecules offers a promising therapeutic approach for the inhibition of inflammation-related diseases. TIRAP (Toll/interleukin-1 receptor (TIR) domain containing adaptor protein) is an important adaptor molecule that is involved in TLR4 signaling spread. Herein, a TIRAP derived peptide, TIP1, has been assessed to hinder the TLR4-mediated signaling pathway. TIP1 inhibited lipopolysaccharide (LPS)-induced activation of NF-κB and MAPKs, and diminished the TLR4-mediated production of pro-inflammatory cytokines (i.e. IL-6 and TNF-α). It also reduced the expression levels of iNOS, COX2, and suppressed intracellular NO production, NO secretion and intracellular ROS generation. TIP1 also potently inhibited TLR3-mediated TNF-α and IFN-β secretion. Cell imaging by confocal microscopy, western blotting, and in vitro studies demonstrated that TIP1 preferentially inhibits TLR4, 3-mediated signaling and it also slightly hinders TLR2/1, 2/6, 7, 8, 9-mediated signaling. Finally, TIP1 diminished systemic cytokine responses (i.e. TNF-α, IL-12p40 and IL-6) elicited in vivo by LPS at 2 h challenge. Thus, TIP1 represents a novel inhibitory agent that majorly suppresses TLR4- and TLR3-mediated inflammatory response and blocks protein-protein interaction of the downstream mediators.
more목차
INTRODUCTION 1
RESULTS 5
A decoy peptide, TIP1, inhibits LPS-induced NF-kB activation in HEK-Blue™ hTLR4 cells. 5
TIP1 majorly suppresses TLR4 and TLR3-mediated signaling pathway in mouse leukemic monocyte macrophage cell line (RAW 264.7 cell). 6
TIP1 effectively suppresses TLR4 and TLR3 signaling pathway in mBMDM and hMNC-PB cell lines. 9
TIP1 blocks TLR4-mediated pro-inflammatory cytokines in mice 10
TIP1 interacts with BB- and DD-loop of TLR4 TIR domain. 11
DISCCUSION 22
MATERIALS & METHODS 22
Cell culture and animals 26
Cell viability analysis. 27
SEAP activity analysis. 28
Western blot analysis. 28
Confocal microscopy. 29
ELISA analysis. 30
Intracellular NO and ROS analysis. 31
NO secretion analysis. 31
Construction of peptide and TLR4 32
Protein-Protein docking 32
Statistical analysis. 32
REFERENCES 33
