IFN-gamma, IL-4 and IL-13 modulate responsiveness of human airway smooth muscle cells to IL-13.
BACKGROUND
IL-13 is a critical mediator of allergic asthma and associated airway hyperresponsiveness. IL-13 acts through a receptor complex comprised of IL-13Ralpha1 and IL-4Ralpha subunits with subsequent activation of signal transducer and activator of transcription 6 (STAT6). The IL-13Ralpha2 receptor may act as a decoy receptor. In human airway smooth muscle (HASM) cells, IL-13 enhances cellular proliferation, calcium responses to agonists and induces eotaxin production. We investigated the effects of pre-treatment with IL-4, IL-13 and IFN-gamma on the responses of HASM cells to IL-13.


METHODS
Cultured HASM were examined for expression of IL-13 receptor subunits using polymerase chain reaction, immunofluorescence microscopy and flow cytometry. Effects of cytokine pre-treatment on IL-13-induced cell responses were assessed by looking at STAT6 phosphorylation using Western blot, eotaxin secretion and calcium responses to histamine.


RESULTS
IL-13Ralpha1, IL-4Ralpha and IL-13Ralpha2 subunits were expressed on HASM cells. IL-13 induced phosphorylation of STAT6 which reached a maximum by 30 minutes. Pre-treatment with IL-4, IL-13 and, to a lesser degree, IFN-gamma reduced peak STAT6 phosphorylation in response to IL-13. IL-13, but not IFN-gamma, pre-treatment abrogated IL-13-induced eotaxin secretion. Pre-treatment with IL-4 or IL-13 abrogated IL-13-induced augmentation of the calcium transient evoked by histamine. Cytokine pre-treatment did not affect expression of IL-13Ralpha1 and IL-4Ralpha but increased expression of IL-13Ralpha2. An anti-IL-13Ralpha2 neutralizing antibody did not prevent the cytokine pre-treatment effects on STAT6 phosphorylation. Cytokine pre-treatment increased SOCS-1, but not SOCS-3, mRNA expression which was not associated with significant increases in protein expression.


CONCLUSIONS
Pre-treatment with IL-4 and IL-13, but not IFN-gamma, induced desensitization of the HASM cells to IL-13 as measured by eotaxin secretion and calcium transients to histamine. The mechanism of IL-4 and IL-13 induced desensitization does not appear to involve either downregulation of receptor expression or induction of the IL-13Ralpha2 or the SOCS proteins.
Moynihan
Barry J
BJ
Department of Medicine, McGill University, Montreal, Canada. barry.moynihan@gmail.com
Tolloczko
Barbara
B
El Bassam
Souad
S
Ferraro
Pascale
P
Michoud
Marie-Claire
MC
Martin
James G
JG
Laberge
Sophie
S
D002118:Calcium / Q000378:metabolism; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D054413:Chemokine CCL11 / Q000378:metabolism; D006632:Histamine / Q000502:physiology; D006801:Humans; D007371:Interferon-gamma / Q000502:physiology*; D018793:Interleukin-13 / Q000502:physiology*; D053661:Interleukin-13 Receptor alpha1 Subunit / Q000378:metabolism; D053663:Interleukin-13 Receptor alpha2 Subunit / Q000378:metabolism; D015847:Interleukin-4 / Q000502:physiology*; D008168:Lung / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D019948:Receptors, Interleukin-4 / Q000378:metabolism; D050800:STAT6 Transcription Factor / Q000378:metabolism; D000071222:Suppressor of Cytokine Signaling 1 Protein; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins / Q000378:metabolism
C515456:CCL11 protein, human; D054413:Chemokine CCL11; D018793:Interleukin-13; D053661:Interleukin-13 Receptor alpha1 Subunit; D053663:Interleukin-13 Receptor alpha2 Subunit; D012333:RNA, Messenger; D019948:Receptors, Interleukin-4; C108944:SOCS1 protein, human; C106644:SOCS3 protein, human; D050800:STAT6 Transcription Factor; C494101:STAT6 protein, human; D000071222:Suppressor of Cytokine Signaling 1 Protein; D000071223:Suppressor of Cytokine Signaling 3 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D015847:Interleukin-4; D006632:Histamine; D007371:Interferon-gamma; D002118:Calcium