Post-transcriptional silencing of CCR3 downregulates IL-4 stimulated release of eotaxin-3 (CCL26) and other CCR3 ligands in alveolar type II cells.
Trafficking and inflammation in airway diseases are, in part, modulated by members of the CC chemokine family, eotaxin-1 (CCL11), eotaxin-2 (CCL24), and eotaxin-3 (CCL26), which transduce signals through their CCR3 receptor. In this context, we hypothesized that transfecting alveolar type II epithelial cells with CCR3-targeted siRNA or antisense (AS-ODN) sequences will downregulate cellular synthesis and release of the primary CCR3 ligands CCL26 and CCL24 and will modulate other CCR3 ligands. The human A549 alveolar type II epithelium-like cell culture model was used for transfection and subsequent effects on CCR3 agonists. siRNAs were particularly effective. PCR showed a 60-80% decrease in mRNA and immunoblots showed up to 75-84% reduction of CCR3 in siRNA treated cells. CCR3-siRNA treatments reduced IL-4 stimulated CCL26 release and constitutive CCL24 release by 65% and 80%, respectively. Release of four additional CCR3 agonists RANTES, MCP-2, MCP-3 and MCP-4 was also significantly reduced by CCR3-siRNA treatments of the alveolar type II cells. Activation of eosinophils, assessed as superoxide anion generation, was reduced when eosinophils were treated with supernatants of A549 cells pretreated with CCR3-targeted siRNAs or AS-ODNs. Collectively, the data suggest that post-transcriptional regulation of CCR3 receptors may be a potential therapeutic approach for interrupting proinflammatory signaling.
Taka
Equar
E
College of Pharmacy and Pharmaceutical Sciences, Florida A & M University, Tallahassee, FL 32307, USA. equar.taka@famu.edu
Errahali
Younes J
YJ
Abonyo
Barack O
BO
Bauer
David M
DM
Heiman
Ann S
AS
D045744:Cell Line, Tumor; D054423:Chemokine CCL24 / Q000276:immunology; D000075906:Chemokine CCL26; D019742:Chemokines, CC / Q000378:metabolism*; D015536:Down-Regulation / Q000235:genetics*; D006801:Humans; D015847:Interleukin-4 / Q000276:immunology / Q000494:pharmacology*; D008024:Ligands; D011650:Pulmonary Alveoli / Q000166:cytology* / Q000187:drug effects* / Q000276:immunology / Q000378:metabolism; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D054397:Receptors, CCR3 / Q000172:deficiency* / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism
C120429:CCL26 protein, human; C515350:CCR3 protein, human; D054423:Chemokine CCL24; D000075906:Chemokine CCL26; D019742:Chemokines, CC; D008024:Ligands; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D054397:Receptors, CCR3; D013481:Superoxides; D015847:Interleukin-4