Investigation of vascular endothelial growth factor effects on pulmonary endothelial monolayer permeability and neutrophil transmigration.
This study sought to determine whether vascular endothelial growth factor (VEGF)-induced permeabilisation of pulmonary endothelium to macromolecules could be related to a permissive role for neutrophil-derived VEGF in neutrophil transmigration. Treatment of human pulmonary artery endothelial cell (HPAEC) monolayers with 1, 10 or 100 ng/ml VEGF for 15 min or 1, 10 ng/ml for 90 min significantly increased endothelial permeability to trypan blue-labelled albumin (TB-BSA). These increases were correlated with changes in the cellular distribution of F-actin, as visualised by rhodamine-phalloidin staining: increased stress fibre formation, cellular elongation and formation of intercellular gaps after 15 min; at 90 min, there was also evidence of microspike formation and extension of spindle processes from the cell surface. Treatment of human neutrophil suspensions with 200 nM phorbol myristyl acetate (PMA), n-formyl-methionyl leucylphenylalanine (fMLP, 10 nM), interleukin-8 (IL-8, 10 nM) (but not with leukotriene B(4) (LTB(4)) 100 nM), for 30 min caused significant extracellular release of neutrophil VEGF stores. A permissive role for neutrophil-derived VEGF in facilitating migration across HPAEC monolayers was assessed in experiments using a functional blocking antihuman VEGF antibody. In the presence of this antibody (10 microg/ml), neutrophil migration in response to fMLP (10 nM), IL-8 (10 nM) or LTB(4) (100 nM) was not significantly different to that in the absence of antibody. We conclude that neutrophil-derived VEGF does not play a functional role in facilitating neutrophil migration across pulmonary vascular endothelium, despite its ability to induce cytoskeletal changes and enhance endothelial macromolecular permeability.
Cullen
V C
VC
Department of Pharmacology, Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Belfield, Dublin 4, Ireland.
Mackarel
A J
AJ
Hislip
S J
SJ
O'Connor
C M
CM
Keenan
A K
AK
D000368:Aged; D002199:Capillary Permeability / Q000502:physiology*; D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D002469:Cell Separation; D004305:Dose-Response Relationship, Drug; D016228:Endothelial Growth Factors / Q000096:biosynthesis / Q000378:metabolism / Q000502:physiology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D005260:Female; D006801:Humans; D008222:Lymphokines / Q000096:biosynthesis / Q000378:metabolism / Q000502:physiology*; D009504:Neutrophils / Q000166:cytology* / Q000378:metabolism; D011651:Pulmonary Artery / Q000166:cytology / Q000378:metabolism; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D008222:Lymphokines; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors