Dysfunctional interaction of C/EBPalpha and the glucocorticoid receptor in asthmatic bronchial smooth-muscle cells.
BACKGROUND
Increased proliferation of bronchial smooth-muscle cells may lead to increased muscle mass in the airways of patients with asthma. The antiproliferative effect of glucocorticoids in bronchial smooth-muscle cells in subjects without asthma is mediated by a complex of the glucocorticoid receptor and the CCAAT/enhancer binding protein alpha (C/EBPalpha). We examined the signaling pathway controlling the inhibitory effect of glucocorticoids on cell proliferation and interleukin-6 synthesis in bronchial smooth-muscle cells of subjects with asthma and those without asthma.


METHODS
Lines of bronchial smooth-muscle cells were established from cells from 20 subjects with asthma, 8 subjects with emphysema, and 26 control subjects. Cell proliferation was determined by means of cell counts and [3H]thymidine incorporation. Signal transduction was studied by means of an electrophoretic DNA mobility-shift assay, a supershift electrophoretic-mobility assay, immunoblotting, use of C/EBPalpha antisense oligonucleotides, and use of a human C/EBPalpha expression vector. Interleukin-6 release was determined by means of an enzyme-linked immunosorbent assay.


RESULTS
Glucocorticoids activated the glucocorticoid receptor and inhibited serum-induced secretion of interleukin-6 in bronchial smooth-muscle cells from both subjects with asthma and those without asthma; however, glucocorticoids inhibited proliferation only in bronchial smooth-muscle cells from subjects without asthma. C/EBPalpha protein was detected by immunoblotting in all bronchial smooth-muscle cells from subjects without asthma but not in those with asthma, whereas the protein was expressed in lymphocytes from both groups of subjects. C/EBPalpha antisense oligonucleotides or the glucocorticoid-receptor inhibitor mifepristone reversed the antiproliferative effect of glucocorticoids in bronchial smooth-muscle cells from subjects without asthma. When bronchial smooth-muscle cells from subjects with asthma were transiently transfected with an expression vector for human C/EBPalpha, two forms of the protein were expressed, and subsequent administration of glucocorticoids inhibited cell proliferation.


CONCLUSIONS
We hypothesize that a cell-type-specific absence of C/EBPalpha is responsible for the enhanced proliferation of bronchial smooth-muscle cells derived from subjects with asthma and that it explains the failure of glucocorticoids to inhibit proliferation in vitro.
Department of Pharmacology and the Woolcock Institute of Medical Research, University of Sydney, Sydney, Australia.
Michael
M
Roth
Peter R A
PR
Johnson
Peter
P
Borger
Michel P
MP
Bihl
Jochen J
JJ
Rüdiger
Gregory G
GG
King
Qi
Q
Ge
Katrin
K
Hostettler
Janette K
JK
Burgess
Judith L
JL
Black
Michael
M
Tamm
D001249:Asthma / Q000378:metabolism*; D001980:Bronchi / Q000166:cytology*; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000235:genetics / Q000378:metabolism*; D016022:Case-Control Studies; D002455:Cell Division / Q000187:drug effects; D002478:Cells, Cultured; D015536:Down-Regulation; D005822:Genetic Vectors; D005938:Glucocorticoids / Q000494:pharmacology*; D006727:Hormone Antagonists / Q000494:pharmacology; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis; D015735:Mifepristone / Q000494:pharmacology; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009841:Oligonucleotides / Q000494:pharmacology; D011656:Pulmonary Emphysema / Q000378:metabolism; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects; D014162:Transfection
D022763:CCAAT-Enhancer-Binding Protein-alpha; D005938:Glucocorticoids; D006727:Hormone Antagonists; D015850:Interleukin-6; D009841:Oligonucleotides; D011965:Receptors, Glucocorticoid; D015735:Mifepristone