Proteasome inhibition augments cigarette smoke-induced GM-CSF expression in trophoblast cells via the epidermal growth factor receptor.
Maternal cigarette smoking has adverse effects on pregnancy outcomes. The granulocyte-macrophage colony-stimulating factor (GM-CSF) is an essential cytokine for a normal pregnancy. We investigated the impact of cigarette smoke extract (CSE) on GM-CSF expression in human cytotrophoblast cells and suggested a cellular mechanism underlying the CSE-induced GM-CSF expression. An immortalized normal human trophoblast cell line (B6Tert-1) was treated with CSE. The viability and proliferation of the CSE-treated B6Tert-1 cells were evaluated, and the expression of GM-CSF in these cells was quantified at the mRNA and the protein levels by means of reverse-transcription and quantitative polymerase chain reaction (RT-qPCR); and enzyme-linked immunosorbent assay (ELISA), respectively. Human trophoblast cells treated with CSE had an increased expression of GM-CSF at both the mRNA and the protein levels. The CSE-induced GM-CSF expression was synergistically enhanced by the addition of the proteasome inhibitor MG-132, but inhibited by AG-1478, an inhibitor of the epidermal growth factor receptor (EGFR) kinase. Furthermore, CSE treatment increased the phosphorylation of the extracellular-signal regulated kinases (ERK1/2) in the trophoblast cells. The expression of other growth factors such as heparin-binding epidermal growth factor-like growth factor (HB-EGF) and vascular endothelial growth factor (VEGF) was also evaluated. Our data suggested that cigarette smoking and proteasome inhibition synergistically up-regulate GM-CSF cytokine expression by activating the EGFR signaling pathway.
State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing, People's Republic of China.
Ya-Yuan
YY
Fu
Jennifer C
JC
Nergard
Nicole K
NK
Barnette
Yan-Ling
YL
Wang
Karl X
KX
Chai
Li-Mei
LM
Chen
D015153:Blotting, Western; D002460:Cell Line; D004797:Enzyme-Linked Immunosorbent Assay; D066246:ErbB Receptors / Q000037:antagonists & inhibitors; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000378:metabolism*; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology; D008856:Microscopy, Fluorescence; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D010766:Phosphorylation; D011247:Pregnancy; D061988:Proteasome Inhibitors / Q000494:pharmacology*; D011799:Quinazolines / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012906:Smoke / Q000009:adverse effects*; D014026:Nicotiana / Q000737:chemistry*; D014327:Trophoblasts / Q000378:metabolism*; D020032:Tyrphostins / Q000494:pharmacology
D036341:Intercellular Signaling Peptides and Proteins; D007976:Leupeptins; D061988:Proteasome Inhibitors; D011799:Quinazolines; D012333:RNA, Messenger; D012906:Smoke; D020032:Tyrphostins; C101044:RTKI cpd; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D066246:ErbB Receptors; D048052:Mitogen-Activated Protein Kinase 3; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde