| Autor: |
Zhu Xiangdong, Osan Christopher M, Binder David C, Meliton Lucille N, Muñoz Nilda M, Meliton Angelo Y, Dudek Steven M, Leff Alan R |
| Jazyk: |
angličtina |
| Rok vydání: |
2010 |
| Předmět: |
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| Zdroj: |
Journal of Inflammation, Vol 7, Iss 1, p 14 (2010) |
| Druh dokumentu: |
article |
| ISSN: |
1476-9255 |
| DOI: |
10.1186/1476-9255-7-14 |
| Popis: |
Abstract Background Cytosolic gIVaPLA2 is a critical enzyme in the generation of arachidonate metabolites and in induction of β2-integrin adhesion in granulocytes. We hypothesized that gIVaPLA2 activation also is an essential downstream step for post adhesive migration of PMN in vitro. Methods Migration of PMNs caused by IL-8/CXCL8 was assessed using a transwell migration chamber. PMNs were pretreated with two structurally unrelated inhibitors of gIVaPLA2, arachidonyl trifluoromethylketone (TFMK) or pyrrophenone, prior to IL-8/CXCL8 exposure. The fraction of migrated PMNs present in the lower chamber was measured as total myeloperoxidase content. GIVaPLA2 enzyme activity was analyzed using [14C-PAPC] as specific substrate F-actin polymerization and cell structure were examined after rhodamine-phalloidin staining. Results IL-8/CXCL8-induced migration of PMNs was elicited in concentration- and time-dependent manner. Time-related phosphorylation and translocation of cytosolic gIVaPLA2 to the nucleus was observed for PMNs stimulated with IL-8/CXCL8 in concentration sufficient to cause upstream phosphorylation of MAPKs (ERK-1/2 and p38) and Akt/PKB. Inhibition of gIVaPLA2 corresponded to the magnitude of blockade of PMN migration. Neither AA nor LTB4 secretion was elicited following IL-8/CXCL8 activation. In unstimulated PMNs, F-actin was located diffusely in the cytosol; however, a clear polarized morphology with F-actin-rich ruffles around the edges of the cell was observed after activation with IL-8/CXCL8. Inhibition of gIVaPLA2 blocked change in cell shape and migration caused by IL-8/CXCL8 but did not cause F-actin polymerization or translocation of cytosolic F-actin to inner leaflet of the PMN membrane. Conclusion We demonstrate that IL-8/CXCL8 causes a) phosphorylation and translocation of cytosolic gIVaPLA2 to the nucleus, b) change in cell shape, c) polymerization of F-actin, and d) chemoattractant/migration of PMN in vitro. Inhibition of gIVaPLA2 blocks the deformability and subsequent migration of PMNs caused by IL-8/CXCL8. Our data suggest that activation of gIVaPLA2 is an essential step in PMN migration in vitro. |
| Databáze: |
Directory of Open Access Journals |
| Externí odkaz: |
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