SM905, an artemisinin derivative, inhibited NO and pro-inflammatory cytokine production by suppressing MAPK and NF-kappaB pathways in RAW 264.7 macrophages.
OBJECTIVE
To elucidate the anti-inflammatory potentials and underlying mechanisms of SM905, a novel artemisinin derivative, in lipopolysaccharide (LPS)-stimulated murine macrophage RAW 264.7 cells.


METHODS
Nitric oxide (NO) generation, cytokine production, and the protein expression levels of inducible nitric-oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) were examined using a Griess assay, an enzyme-linked immunosorbent assay (ELISA) and a Western blotting assay, respectively. The mRNA expression was measured using real-time PCR. The phosphorylation of extracellular signal-regulated kinase1/2 (ERK1/2), p38, c-jun N-terminal kinase (JNK), and the degradation of IkappaBalpha were assessed by Western blotting analysis. The nuclear translocation of nuclear factor-kappaB (NF-kappaB) was observed using confocal microscopy.


RESULTS
Pretreatment with SM905 (0, 0.1, 1, and 10 micromol/L) suppressed LPS-induced NO, TNF-alpha, IL-1beta, and IL-6 production, and decreased both protein and mRNA levels of iNOS and COX-2. The mRNA expression of LPS receptor Toll-like receptor 4 (TLR4) and myeloid differentiation protein-2 (MD-2) was not changed, while LPS-induced CD14 expression was slightly reduced after SM905 treatment. SM905 markedly decreased the activation of ERK1/2, p38 and JNK suppressed the degradation of IkappaBalpha, but did not modify the expression of interferon regulatory factor-1 (IRF-1), signal transducer and activator of transcription 1 (STAT1) or interferon-inducible protein-10 (IP-10). By using confocal microscopy, we further observed that NF-kappaB was correspondingly inhibited in SM905-treated cells.


CONCLUSIONS
SM905 inhibited NO and pro-inflammatory cytokine production in LPS-stimulated RAW 264.7 cells and these effects are at least partially mediated through suppression of the MAPK and NF-kappaB signaling pathways.
Laboratory of Immunopharmacology, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China.
Jun-Xia
JX
Wang
Li-Fei
LF
Hou
Yang
Y
Yang
Wei
W
Tang
Ying
Y
Li
Jian-Ping
JP
Zuo
D000818:Animals; D037621:Artemisinins / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D002460:Cell Line; D051546:Cyclooxygenase 2 / Q000378:metabolism; D016207:Cytokines / Q000096:biosynthesis*; D004305:Dose-Response Relationship, Drug; D051550:I-kappa B Kinase / Q000378:metabolism; D007249:Inflammation / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors*; D008970:Molecular Weight; D016328:NF-kappa B / Q000037:antagonists & inhibitors*; D009569:Nitric Oxide / Q000037:antagonists & inhibitors*; D052247:Nitric Oxide Synthase Type II / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D012333:RNA, Messenger / Q000032:analysis; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
D037621:Artemisinins; D016207:Cytokines; D008070:Lipopolysaccharides; D016328:NF-kappa B; D012333:RNA, Messenger; C522065:SM905; D009569:Nitric Oxide; D052247:Nitric Oxide Synthase Type II; D051546:Cyclooxygenase 2; D051550:I-kappa B Kinase; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases