Effects of 3-phenyl-propenal on the expression of toll-like receptors and downstream signaling components on raw264.7 murine macrophages.
3-phenyl-propenal is one of the principle compounds isolated from Guizhi (Ramulus Cinnamomi), the principal drug in Guizhi-Tang (GZT), a famous traditional Chinese medical formula. The aim of the present study was to investigate the effects of 3-phenyl-propenal on the expression of toll-like receptor 3 (TLR3), TLR4 and the downstream signaling components on Raw264.7 murine microphages. Raw264.7 cells were cultured in RPMI-1640 medium containing LPS (lipopolysaccharide) or poly (I:C) in the presence or absence of 3-phenyl-propenal. After 24-hour incubation, the medium was collected and the amount of TNF-alpha and IFN-beta was measured by ELISA. mRNA expression of TLR3, TLR4, myeloid differentiation factor (MyD88), TRAF-6 (tumor necrosis factor receptor-associated), TRAM (toll-like receptor-associated molecule) and TRIF (TIR domain-containing adaptor inducing IFN-beta) were analyzed by real-time PCR with SYBR green dye. Protein expression of TLR3 and TLR4 was analyzed by Western blotting and that of MyD88 and TRAF-6 was analyzed by immunofluorescence assay. The results indicate that LPS increased the expression of TLR4, MyD88, TRAF-6, TRAM and TRIF, but had no influence on TLR3, while poly (I:C) up-regulated the expression of TLR3, MyD88, TRAM and TRIF. 3-phenyl-propenal significantly decreased the expression of LPS-induced TLR4, MyD88, TRAF-6, while possessing no effect on LPS-induced TRAM and TRIF expression in Raw264.7 cells. When cells were stimulated by poly (I:C), 3-phenyl-propenal significantly decreased TLR3 and MyD88 expression. In conclusion, 3-phenyl-propenal blocked the over-expression of TLR3, TLR4, their downstream signaling components MyD88 and TRAF-6, which indicate that it had an antagonistic effect on TLR3 and TLR4.
Zhao
Bao-Sheng
BS
Tang Center for Herbal Medicine Research, Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing 100700, China.
Huo
Hai-Ru
HR
Ma
Yue-Ying
YY
Liu
Hong-Bin
HB
Li
Lan-Fang
LF
Sui
Feng
F
Li
Cang-Hai
CH
Guo
Shu-Ying
SY
Jiang
Ting-Liang
TL
D000171:Acrolein / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D018929:Cell Culture Techniques; D002460:Cell Line; D016899:Interferon-beta / Q000378:metabolism; D008264:Macrophages / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D016133:Polymerase Chain Reaction; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D051196:Toll-Like Receptor 3 / Q000187:drug effects / Q000235:genetics; D051197:Toll-Like Receptor 4 / Q000187:drug effects / Q000235:genetics; D051193:Toll-Like Receptors / Q000235:genetics*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
C519703:TLR3 protein, mouse; C493487:Tlr4 protein, mouse; D051196:Toll-Like Receptor 3; D051197:Toll-Like Receptor 4; D051193:Toll-Like Receptors; D014409:Tumor Necrosis Factor-alpha; D016899:Interferon-beta; D000171:Acrolein; C012843:cinnamaldehyde