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Post‐transcriptional silencing of CCR3 downregulates IL‐4 stimulated release of CCL26 in alveolar type II cells
Author(s) -
Taka Equar,
Errahali Younes J,
Heiman Ann S
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.762.4
Subject(s) - small interfering rna , chemistry , transfection , ccr3 , a549 cell , gene silencing , microbiology and biotechnology , eotaxin , chemokine , receptor , cell , chemokine receptor , biology , biochemistry , gene
Trafficking and activation of cells in diseases such as asthma are, in part, modulated by eotaxin‐2 (CCL24) and eotaxin‐3 (CCL26) chemokines that signal via CCR3. Short‐interfering RNAs (siRNA) are novel tools to inhibit gene functions in human diseases. In this context, we hypothesized that alveolar type II cells can be transfected with siRNAs targeting CCR3 (CCR3‐siRNA) which will down‐regulate the receptor and decrease synthesis and release of its ligands CCL24 and CCL26. To test our hypothesis, human A549 alveolar type II epithelium‐like cells were transfected with four separate and combined CCR3‐siRNA duplexes and effects on CCR3, CCL26 and CCL24 were studied. FACS analysis showed decreased cell surface CCR3. Densitometry of lysate immunoblots indicated up to 75–84% reduction in CCR3. RT‐PCR showed an 80% decrease in CCR3 mRNA. IL‐4 stimulated CCL26 release (959 ± 201 pg/ml) was reduced up to 65% and constitutive CCL24 release (285 ± 64 pg/ml) was reduced by 80% in CCR3‐siRNA treated cells. Activation of eosinophils, assessed as superoxide anion generation, was reduced up to 80% when treated with supernatants of A549 cells pretreated with CCR3‐siRNAs. These findings suggest that CCR3‐siRNA treatment of alveolar type II cells may be useful for development of novel therapies for controlling airway inflammation. Support for this research was provided in part by NIH grants RR08111 and RR03020.