期刊
JOURNAL OF ALLERGY AND CLINICAL IMMUNOLOGY
卷 111, 期 1, 页码 113-116出版社
MOSBY, INC
DOI: 10.1067/mai.2003.27
关键词
chemotaxis; eosinophil; eotaxin; mitogen-activated protein kinase; myosin light chain kinase
资金
- NIAID NIH HHS [P01 AI46004, R01 AI50179] Funding Source: Medline
- NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES [P01AI046004, R01AI050179] Funding Source: NIH RePORTER
Background: Eosinophil migration in the tissue is one characteristic feature of allergic diseases. The CC chemokine eotaxin plays a pivotal role in local accumulation of eosinophils. Myosin light chain kinase (MLCK) is known to regulate cytoskeletal rearrangement and cell motility by means of phosphorylation of myosin light chain (MLC). Objective: We have previously shown that mitogen-activated protein (MAP) kinases are important for eosinophil migration. In the present study we hypothesized that MLCK is downstream of MAP kinases, thereby linking the MAP kinase pathway to the activation of cytoskeletal components required for eosinophil chemotaxis. Methods: Blood eosinophils were purified by using Percoll and anti-CD16 antibody-coated magnetic beads. We investigated the phosphorylation of MLCK and MLC by using the phosphorous 32-orthophosphates-labeled eosinophils. The kinase activity of MLCK was determined by measuring the phosphotransferase activity for the MLCK-specific peptide substrate. The chemotaxis assay was performed in a 48-well Boyden microchamber. Results: The phosphotransferase activity of MLCK for a substrate peptide was enhanced in eotaxin-stimulated eosinophils. We also found that eotaxin induced phosphorylation of MLCK in vivo in phosphorous 32-orthophosphate-labeled eosinophils. PD98059 (MAP/extracellular signal-regulated kinase inhibitor) or SB202190 (p38 MAP kinase inhibitor) abrogated the eotaxin-induced phosphorylation of MLCK. The phosphorylation of MLC was upregulated by eotaxin. Eosinophil chemotaxis was inhibited by means of pretreatment of the MLCK inhibitor ML-7. Conclusion: These results suggest that eotaxin regulates MLCK through both extracellular signal-regulated kinase 1 and p38 MAP kinase. MLCK activation is a critical step in the cytoskeletal rearrangements leading to eosinophil migration.
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