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.
Roth
Michael
M
Department of Pharmacology and the Woolcock Institute of Medical Research, University of Sydney, Sydney, Australia.
Johnson
Peter R A
PR
Borger
Peter
P
Bihl
Michel P
MP
Rüdiger
Jochen J
JJ
King
Gregory G
GG
Ge
Qi
Q
Hostettler
Katrin
K
Burgess
Janette K
JK
Black
Judith L
JL
Tamm
Michael
M
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