BACKGROUND: Granulocyte-macrophage colony-stimulating factor (GM-CSF) is an important mediator in the differentiation, maturation, and survival of inflammatory cells. It might play a significant role in the pathogenesis of asthma. We have shown previously that cytosolic phospholipase A2 group IV A (cPLA2alpha) is able to activate gene expression, through peroxisome proliferator-activated receptor (PPAR)-gamma response elements (PPRE). In the promoter regions of GM-CSF gene (CSF2), we have found potential PPRE. The goal of the current study was to investigate the influence of cPLA2 overexpression and activation on CSF2 gene expression in human lung cells. MATERIAL/ METHODS: Subconfluent A549 cells were transfected with GM-CSF reporter gene and cPLA2alpha overexpression vector. Transfected cells were treated with or without calcium ionophore, A23187, or epidermal growth factor (EGF). Cell lysates were collected and assayed for dual luciferase activity. Some cultures were preincubated with a potent cPLA2alpha inhibitor, methyl arachidonyl fluorophosphonate (MAFP); a secretory phospholipase A2 inhibitor, thioetheramide-PC; a calcium-independent phospholipase A2 inhibitor, bromoenol lactone (BEL); or vehicle. After preincubation, cells were treated with A23187. Expression of GM-CSF messenger RNA (mRNA) was measured using real-time polymerase chain reaction mRNA quantification. RESULTS: Overexpression of cPLA2alpha or activation of endogenous cPLA2alpha by calcium ionophore or EGF caused a significant increase in GM-CSF relative luciferase activity. Calcium ionophore significantly increased GM-CSF mRNA in A549 human lung cells. These effects were at least in part inhibited by MAFP treatment, but not by BEL or thioetheramide-PC. CONCLUSIONS: These preliminary data might suggest an influence of cPLA2alpha on GM-CSF gene expression in human lung cells, which could play a role in the pathogenesis of asthma and other inflammatory diseases.
BACKGROUND:Granulocyte-macrophage colony-stimulating factor (GM-CSF) is an important mediator in the differentiation, maturation, and survival of inflammatory cells. It might play a significant role in the pathogenesis of asthma. We have shown previously that cytosolic phospholipase A2 group IV A (cPLA2alpha) is able to activate gene expression, through peroxisome proliferator-activated receptor (PPAR)-gamma response elements (PPRE). In the promoter regions of GM-CSF gene (CSF2), we have found potential PPRE. The goal of the current study was to investigate the influence of cPLA2 overexpression and activation on CSF2 gene expression in human lung cells. MATERIAL/ METHODS: Subconfluent A549 cells were transfected with GM-CSF reporter gene and cPLA2alpha overexpression vector. Transfected cells were treated with or without calcium ionophore, A23187, or epidermal growth factor (EGF). Cell lysates were collected and assayed for dual luciferase activity. Some cultures were preincubated with a potent cPLA2alpha inhibitor, methyl arachidonyl fluorophosphonate (MAFP); a secretory phospholipase A2 inhibitor, thioetheramide-PC; a calcium-independent phospholipase A2 inhibitor, bromoenol lactone (BEL); or vehicle. After preincubation, cells were treated with A23187. Expression of GM-CSF messenger RNA (mRNA) was measured using real-time polymerase chain reaction mRNA quantification. RESULTS: Overexpression of cPLA2alpha or activation of endogenous cPLA2alpha by calcium ionophore or EGF caused a significant increase in GM-CSF relative luciferase activity. Calcium ionophore significantly increased GM-CSF mRNA in A549 human lung cells. These effects were at least in part inhibited by MAFP treatment, but not by BEL or thioetheramide-PC. CONCLUSIONS: These preliminary data might suggest an influence of cPLA2alpha on GM-CSF gene expression in human lung cells, which could play a role in the pathogenesis of asthma and other inflammatory diseases.
Authors: Milena Sokolowska; Valerie F J Quesniaux; Cezmi A Akdis; Kian Fan Chung; Bernhard Ryffel; Dieudonnée Togbe Journal: Front Immunol Date: 2019-09-13 Impact factor: 7.561
Authors: Clare S Hardman; Yi-Ling Chen; Maryam Salimi; Rachael Jarrett; David Johnson; Valtteri J Järvinen; Raymond J Owens; Emmanouela Repapi; David J Cousins; Jillian L Barlow; Andrew N J McKenzie; Graham Ogg Journal: Sci Immunol Date: 2017-12-22