Lipopolysaccharide pretreatment increases protease-activated receptor-2 expression and monocyte chemoattractant protein-1 secretion in vascular endothelial cells |
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Authors: | Hung-Hsing Chao Po-Yuan Chen Wen-Rui Hao Wei-Ping Chiang Tzu-Hurng Cheng Shih-Hurng Loh Yuk-Man Leung Ju-Chi Liu Jin-Jer Chen Li-Chin Sung |
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Institution: | 1.Division of Cardiovascular Surgery, Department of Surgery,Shin Kong Wu Ho-Su Memorial Hospital,Taipei,Taiwan;2.Department of Surgery,School of Medicine, Taipei Medical University,Taipei,Taiwan;3.Department of Biological Science and Technology,College of Biopharmaceutical and Food Sciences, China Medical University,Taichung,Taiwan;4.Division of Cardiology, Department of Internal Medicine,Shuang Ho Hospital, Taipei Medical University,New Taipei City,Taiwan;5.Department of Biochemistry,School of Medicine, China Medical University,Taichung,Taiwan;6.Department of Pharmacology & Graduate Institute of Pharmacology,National Defense Medical Center,Taipei,Taiwan;7.Department of Physiology,School of Medicine, China Medical University,Taichung,Taiwan;8.Department of Internal Medicine,School of Medicine, College of Medicine, Taipei Medical University,Taipei,Taiwan;9.Graduate Institute of Clinical Medicine,College of Medicine, China Medical University,Taichung,Taiwan;10.Institute of Biomedical Sciences,Academia Sinica,Taipei,Taiwan |
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Abstract: | BackgroundThis study investigated whether lipopolysaccharide (LPS) increase protease-activated receptor-2 (PAR-2) expression and enhance the association between PAR-2 expression and chemokine production in human vascular endothelial cells (ECs).MethodsThe morphology of ECs was observed through microphotography in cultured human umbilical vein ECs (EA. hy926 cells) treated with various LPS concentrations (0, 0.25, 0.5, 1, and 2 μg/mL) for 24 h, and cell viability was assessed using the MTT assay. Intracellular calcium imaging was performed to assess agonist (trypsin)-induced PAR-2 activity. Western blotting was used to explore the LPS-mediated signal transduction pathway and the expression of PAR-2 and adhesion molecule monocyte chemoattractant protein-1 (MCP-1) in ECs.ResultsTrypsin stimulation increased intracellular calcium release in ECs. The calcium influx was augmented in cells pretreated with a high LPS concentration (1 μg/mL). After 24 h treatment of LPS, no changes in ECs viability or morphology were observed. Western blotting revealed that LPS increased PAR-2 expression and enhanced trypsin-induced extracellular signal-regulated kinase (ERK)/p38 phosphorylation and MCP-1 secretion. However, pretreatment with selective ERK (PD98059), p38 mitogen-activated protein kinase (MAPK) (SB203580) inhibitors, and the selective PAR-2 antagonist (FSLLRY-NH2) blocked the effects of LPS-activated PAR-2 on MCP-1 secretion.ConclusionsOur findings provide the first evidence that the bacterial endotoxin LPS potentiates calcium mobilization and ERK/p38 MAPK pathway activation and leads to the secretion of the pro-inflammatory chemokine MCP-1 by inducing PAR-2 expression and its associated activity in vascular ECs. Therefore, PAR-2 exerts vascular inflammatory effects and plays an important role in bacterial infection-induced pathological responses. |
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