Cargando…
Cytosolic group IVa phospholipase A(2 )mediates IL-8/CXCL8-induced transmigration of human polymorphonuclear leukocytes in vitro
BACKGROUND: Cytosolic gIVaPLA(2 )is a critical enzyme in the generation of arachidonate metabolites and in induction of β(2)-integrin adhesion in granulocytes. We hypothesized that gIVaPLA(2 )activation also is an essential downstream step for post adhesive migration of PMN in vitro. METHODS: Migrat...
Autores principales: | , , , , , , , |
---|---|
Formato: | Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2010
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2848033/ https://www.ncbi.nlm.nih.gov/pubmed/20298597 http://dx.doi.org/10.1186/1476-9255-7-14 |
_version_ | 1782179635650363392 |
---|---|
author | Meliton, Angelo Y Muñoz, Nilda M Meliton, Lucille N Binder, David C Osan, Christopher M Zhu, Xiangdong Dudek, Steven M Leff, Alan R |
author_facet | Meliton, Angelo Y Muñoz, Nilda M Meliton, Lucille N Binder, David C Osan, Christopher M Zhu, Xiangdong Dudek, Steven M Leff, Alan R |
author_sort | Meliton, Angelo Y |
collection | PubMed |
description | BACKGROUND: Cytosolic gIVaPLA(2 )is a critical enzyme in the generation of arachidonate metabolites and in induction of β(2)-integrin adhesion in granulocytes. We hypothesized that gIVaPLA(2 )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 gIVaPLA(2), 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. GIVaPLA(2 )enzyme activity was analyzed using [(14)C-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 gIVaPLA(2 )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 gIVaPLA(2 )corresponded to the magnitude of blockade of PMN migration. Neither AA nor LTB(4 )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 gIVaPLA(2 )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 gIVaPLA(2 )to the nucleus, b) change in cell shape, c) polymerization of F-actin, and d) chemoattractant/migration of PMN in vitro. Inhibition of gIVaPLA(2 )blocks the deformability and subsequent migration of PMNs caused by IL-8/CXCL8. Our data suggest that activation of gIVaPLA(2 )is an essential step in PMN migration in vitro. |
format | Text |
id | pubmed-2848033 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2010 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-28480332010-04-01 Cytosolic group IVa phospholipase A(2 )mediates IL-8/CXCL8-induced transmigration of human polymorphonuclear leukocytes in vitro Meliton, Angelo Y Muñoz, Nilda M Meliton, Lucille N Binder, David C Osan, Christopher M Zhu, Xiangdong Dudek, Steven M Leff, Alan R J Inflamm (Lond) Research BACKGROUND: Cytosolic gIVaPLA(2 )is a critical enzyme in the generation of arachidonate metabolites and in induction of β(2)-integrin adhesion in granulocytes. We hypothesized that gIVaPLA(2 )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 gIVaPLA(2), 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. GIVaPLA(2 )enzyme activity was analyzed using [(14)C-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 gIVaPLA(2 )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 gIVaPLA(2 )corresponded to the magnitude of blockade of PMN migration. Neither AA nor LTB(4 )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 gIVaPLA(2 )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 gIVaPLA(2 )to the nucleus, b) change in cell shape, c) polymerization of F-actin, and d) chemoattractant/migration of PMN in vitro. Inhibition of gIVaPLA(2 )blocks the deformability and subsequent migration of PMNs caused by IL-8/CXCL8. Our data suggest that activation of gIVaPLA(2 )is an essential step in PMN migration in vitro. BioMed Central 2010-03-18 /pmc/articles/PMC2848033/ /pubmed/20298597 http://dx.doi.org/10.1186/1476-9255-7-14 Text en Copyright ©2010 Meliton et al; licensee BioMed Central Ltd. http://creativecommons.org/licenses/by/2.0 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Meliton, Angelo Y Muñoz, Nilda M Meliton, Lucille N Binder, David C Osan, Christopher M Zhu, Xiangdong Dudek, Steven M Leff, Alan R Cytosolic group IVa phospholipase A(2 )mediates IL-8/CXCL8-induced transmigration of human polymorphonuclear leukocytes in vitro |
title | Cytosolic group IVa phospholipase A(2 )mediates IL-8/CXCL8-induced transmigration of human polymorphonuclear leukocytes in vitro |
title_full | Cytosolic group IVa phospholipase A(2 )mediates IL-8/CXCL8-induced transmigration of human polymorphonuclear leukocytes in vitro |
title_fullStr | Cytosolic group IVa phospholipase A(2 )mediates IL-8/CXCL8-induced transmigration of human polymorphonuclear leukocytes in vitro |
title_full_unstemmed | Cytosolic group IVa phospholipase A(2 )mediates IL-8/CXCL8-induced transmigration of human polymorphonuclear leukocytes in vitro |
title_short | Cytosolic group IVa phospholipase A(2 )mediates IL-8/CXCL8-induced transmigration of human polymorphonuclear leukocytes in vitro |
title_sort | cytosolic group iva phospholipase a(2 )mediates il-8/cxcl8-induced transmigration of human polymorphonuclear leukocytes in vitro |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2848033/ https://www.ncbi.nlm.nih.gov/pubmed/20298597 http://dx.doi.org/10.1186/1476-9255-7-14 |
work_keys_str_mv | AT melitonangeloy cytosolicgroupivaphospholipasea2mediatesil8cxcl8inducedtransmigrationofhumanpolymorphonuclearleukocytesinvitro AT munoznildam cytosolicgroupivaphospholipasea2mediatesil8cxcl8inducedtransmigrationofhumanpolymorphonuclearleukocytesinvitro AT melitonlucillen cytosolicgroupivaphospholipasea2mediatesil8cxcl8inducedtransmigrationofhumanpolymorphonuclearleukocytesinvitro AT binderdavidc cytosolicgroupivaphospholipasea2mediatesil8cxcl8inducedtransmigrationofhumanpolymorphonuclearleukocytesinvitro AT osanchristopherm cytosolicgroupivaphospholipasea2mediatesil8cxcl8inducedtransmigrationofhumanpolymorphonuclearleukocytesinvitro AT zhuxiangdong cytosolicgroupivaphospholipasea2mediatesil8cxcl8inducedtransmigrationofhumanpolymorphonuclearleukocytesinvitro AT dudekstevenm cytosolicgroupivaphospholipasea2mediatesil8cxcl8inducedtransmigrationofhumanpolymorphonuclearleukocytesinvitro AT leffalanr cytosolicgroupivaphospholipasea2mediatesil8cxcl8inducedtransmigrationofhumanpolymorphonuclearleukocytesinvitro |