Effect of Cysteinyl Leukotriene D4 on Chemokine Expression via Cysteinyl Leukotriene Receptor 1 in Human Lung Epithelial Cells, A549

BACKGROUND: Cysteinyl leukotrienes (CysLTs) such as LTC4, LTD4 and LTE4 are proinflammtory lipid mediators synthesized by the 5-lipoxygenase pathway from arachidonic acid and play important roles in eosinophilic chemotactic activity to airway inflammation characterized by bronchoconstriction, mucus secretion and airway hyperresponsiveness through G-protein coupled receptor, cysteinyl leukotriene receptor 1(CysLTR1). CCL2 (MCP-1, Monocyte Chemoattractant protein-1) is one of the major chemokines responsible for attracting monocytes. CXCL10 (IP-10, IFN-γ-inducible protein 10kDa) is an IFN-γ-inducible chemokine that preferentially attracts activated Th1 lymphocytes. OBJECTIVE: The aim of this study is to investigate transcriptional regulation of MCP-1 and IP-10 by LTD4 stimulation via CysLTR1 in human lung epithelial A549 cells. METHOD: To investigate the effect of LTD4 and CysLTR1 on MCP-1 and IP-10, Two different kinds of epithelial cells, A549 cells and CysLTR1 over-expressed A549 cells, were prepared and treated with LTD4 100nM for 0.5, 1, 2, 4 and 8hours. After LDT4 stimulation, mRNA levels of MCP-1 and IP-10 were examined by real-time RT-PCR. To make sure whether MCP-1 and IP-10 are involved with CysLTR1, prior to treatment of LTD4, cells were pre-treated with CysLTR1 antagonist, MK-571 100nM for 3hrs. To confirm the effect of LTD4 and/or CysLTR1 on releasing MCP-1 and IP-10, ELISA was performed with cell supernatant. RESULT: mRNA expression level of MCP-1 was augmented as soon as treated with LTD4. mRNA expression level of IP-10 was increased 1hr after LTD4 100nM stimulation in A549cells. After transfection of CysLTR1, the mRNA expression of both MCP-1 and IP-10 were enhanced about more than 500-fold and 300-fold respectively. In case of released protein, IP-10 were increased about more than 100 times whereas MCP-1 was not affected. Although the tendency of the effect of LTD4 on IP-10 after transfection of CysLTR1 in mRNA level was similar with one before transfection, protein level of IP-10 was increased preferentially at 1hr after LTD4 treatment in CysLTR1-transfected A549 cells. MK571, CysLTR1 antagonist inhibited the expression of IP-10 in mRNA level while couldn’t block MCP-1 expression. IP-10 blocked by MK-571 was recovered immediately by LTD4 treatment. CONCLUSION: IP-10 and MCP-1 are up-regulated by LTD4 stimulus both in transcriptional and translational levels. Up-regulation of IP-10 by LTD4 was specifically inhibited by MK-571, cysLTR1 specific antagonist while the increased expression of MCP-1 was not. These data suggest that IP-10 expression may be involved in the pathogenic mechanism of asthma via CysLTR1.

Author dc:creator

• 김, 설화

Contributors dc:contributor

• 박, 해심
• 대학원 의학과
• 200524156

• asthma
• cysteiny
• leukotriene D4
• cysteinyl leukotriene receptor 1
• cytokines
• chemokines

Language dc:language

en

Identifier

http://dcoll.ajou.ac.kr:9080/dcollection/jsp/common/DcLoOrgPer.jsp?sItemId=000000002540
000000002540

OAI identifier oai:identifier

oai:repository.ajou.ac.kr:201003/1361