Effect of MIF siRNA on the expression of Toll-like receptors and down stream signaling components in A549 cells

Abstract

Abstract: AIM: To investigate the effect and mechanism of the small interfering RNAs (siRNA) for macrophage migration inhibitory factor (MIF) on expression of TLRs and downstream signaling components in alveolar epithial cell line A549 stimulated by lipopolysaccharide (LPS).METHODS: MIF siRNA and nonspecific siRNA were transfected into A549 cells by liposome respectively, then these cells were stimulated with LPS. The expression of MIF, TLR2, TLR4 mRNA were measured by RT-PCR. The proteins expression of MyD88 and IRF3 of TLR's downstream signaling components were analyzed by Western Blot. The nuclear translocation of NF-κB was observed by cell immunofluorescence. The cells supernatant was collected and the inflammatory factors such as TNF-α, IL-1β, IL-6, IFN-β were measured by ELISA.RESULTS: LPS could stimulated the high expression of MIF, TLR2, TLR4 mRNA, and MIF siRNA could down regulate them and partially blocked NF-κB (p65) nuclear translocation. MIF siRNA also decreased significantly the high expression of MyD88 and the secretion levels of TNF-α, IL-1β, IL-6, but MIF siRNA did not work to IRF3 and IFN-β.CONCLUSION: MIF siRNA can inhibit the over-secretion of the inflammatory mediators TNF-α, IL-1β, IL-6 in A549 cells stimulated by LPS. The mechanism may be MIF siRNA reduced the highly expression of MIF, TLR2, and TLR4 mRNA, and blocked MyD88 protein of TLR downstream signal transduction pathway and the migration of NF-κB nuclear.

Key words: Macrophage migration inhibitory factor (MIF), siRNA, Toll-like receptors, Signaling pathway, Lipopolysaccharide (LPS)

CLC Number:

R965.2

MO Hong-ying, JIANG Yong-nan, LI Yi-min, LAI Le, XIAO Zheng-lun. Effect of MIF siRNA on the expression of Toll-like receptors and down stream signaling components in A549 cells[J]. Chinese Journal of Clinical Pharmacology and Therapeutics, 2013, 18(4): 382-387.

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