Cytosolic phospholipase A2 group IVA influence on GM-CSF expression in human lung cells: a pilot study.
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.


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.
Sokolowska
Milena
M
Department of Immunopathology, Chair of Allergology, Immunology and Dermatology, Faculty of Medical Sciences and Postgraduate Training, Medical University of Lodz, Lodz, Poland.
Stefanska
Joanna
J
Wodz-Naskiewicz
Karolina
K
Pawliczak
Rafal
R
D000818:Animals; D045744:Cell Line, Tumor; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D017930:Genes, Reporter; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics* / Q000378:metabolism; D054513:Group IV Phospholipases A2 / Q000037:antagonists & inhibitors / Q000172:deficiency / Q000502:physiology*; D006801:Humans; D008168:Lung / Q000201:enzymology* / Q000473:pathology; D008175:Lung Neoplasms / Q000201:enzymology / Q000235:genetics / Q000473:pathology; D010865:Pilot Projects; D051381:Rats; D013997:Time Factors
D004791:Enzyme Inhibitors; D004815:Epidermal Growth Factor; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D054513:Group IV Phospholipases A2; C516389:PLA2G4A protein, human