Differential regulation of chemokine expression by Th1 and Th2 cytokines and mechanisms of eotaxin/CCL-11 expression in human airway smooth muscle cells.
BACKGROUND
Airway smooth muscle (ASM) cells may contribute to the pathogenesis of asthma including airway inflammation and remodeling. We focused our study on the regulation of chemokine expression by cytokines and analyzed the mechanisms of eotaxin/CCL-11 expression in ASM cells.


METHODS
Human ASM cells were cultured in vitro and treated with IL-4, interferon-gamma (IFNgamma), and tumor necrosis factor-alpha (TNFalpha). Secretion of chemokines into the culture medium was analyzed by ELISA. Expression of eotaxin mRNA was analyzed by reverse transcription-polymerase chain reaction (RT-PCR). Binding of transcription factor signal transducer activator of transcription (STAT) 6 to the eotaxin promoter-derived DNA was analyzed by pull-down Western blot. To assess transcriptional regulation of eotaxin, cells were transfected with eotaxin promoter-luciferase reporter plasmids, and activity was determined by dual luciferase assay.


RESULTS
The Th2 cytokine IL-4 preferentially stimulated the expression of the CC chemokine receptor (CCR) 3-ligand chemokines eotaxin, eotaxin-3, and MCP-4. The Th1 cytokine IFNgamma stimulated the expression of chemokines IP-10 and RANTES. IL-4 stimulated nuclear translocation of signal transducer activator of transcription 6 (STAT6) and its binding to the eotaxin promoter region. IL-4 activated the eotaxin promoter and its activity was inhibited by mutation of the binding site for STAT6 in the promoter.


CONCLUSIONS
The Th2 cytokine IL-4 preferentially stimulated the expression of CCR3 ligand chemokines including eotaxin in ASM cells. The transcription factor STAT6 may play a pivotal role in the activation of eotaxin transcription in response to IL-4.
Odaka
Miho
M
First Department of Internal Medicine, Showa University School of Medicine, Shinagawa, Tokyo, Japan. miho-odk@gk9.so-net.ne.jp
Matsukura
Satoshi
S
Kuga
Hideki
H
Kokubu
Fumio
F
Kasama
Tsuyoshi
T
Kurokawa
Masatsugu
M
Kawaguchi
Mio
M
Ieki
Koushi
K
Suzuki
Shintaro
S
Watanabe
Shin
S
Homma
Tetsuya
T
Takeuchi
Hiroko
H
Nohtomi
Kyoko
K
Schleimer
Robert P
RP
Adachi
Mitsuru
M
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D054413:Chemokine CCL11; D000075906:Chemokine CCL26; D018946:Chemokine CCL5 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D054357:Chemokine CXCL10; D019742:Chemokines, CC / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism; D019743:Chemokines, CXC / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D004357:Drug Synergism; D004797:Enzyme-Linked Immunosorbent Assay; D005786:Gene Expression Regulation / Q000187:drug effects; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology* / Q000502:physiology; D015847:Interleukin-4 / Q000494:pharmacology* / Q000502:physiology; D018945:Monocyte Chemoattractant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011485:Protein Binding / Q000187:drug effects; D012333:RNA, Messenger; D011994:Recombinant Proteins / Q000494:pharmacology; D012137:Respiratory System / Q000166:cytology*; D050800:STAT6 Transcription Factor / Q000502:physiology*; D018417:Th1 Cells / Q000502:physiology*; D018418:Th2 Cells / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology* / Q000502:physiology; D015854:Up-Regulation / Q000187:drug effects
C515456:CCL11 protein, human; C099677:CCL13 protein, human; C120429:CCL26 protein, human; C114753:CXCL10 protein, human; D054413:Chemokine CCL11; D000075906:Chemokine CCL26; D018946:Chemokine CCL5; D054357:Chemokine CXCL10; D019742:Chemokines, CC; D019743:Chemokines, CXC; C508598:IL4 protein, human; D018945:Monocyte Chemoattractant Proteins; D012333:RNA, Messenger; D011994:Recombinant Proteins; D050800:STAT6 Transcription Factor; C494101:STAT6 protein, human; D014409:Tumor Necrosis Factor-alpha; D015847:Interleukin-4; D007371:Interferon-gamma