Dibenzylbutyrolactone Lignans from Forsythia koreana Fruits Attenuate Lipopolysaccharide-Induced Inducible Nitric Oxide Synthetase and Cyclooxygenase-2 Expressions through Activation of Nuclear Factor-κB and Mitogen-Activated Protein Kinase in RAW264.7 Cells

Previously, we reported that dibenzylbutyrolactone lignans (DBLLs) from the fruit of Forsythia koreana NAKAI (Oleaceae) has anti-inflammatory, antioxidant, and anti-asthmatic effects. In this study, to clarify the anti-inflammatory mechanisms of DBLL, we evaluated the effects of DBLLs on lipopolysaccharide-stimulated inducible nitric oxide synthetase (iNOS) and cyclooxygenase-2 (COX-2) expressions, nitric oxide (NO) and prostaglandin E-2 (PGE(2)) productions, nuclear factor-kappa B (NF-kappa B) and mitogen-activated protein kinase (MAPK) activations, inhibitor of kappa B (I kappa B) and inhibitor of kappa B kinase (IKK) phosphorylations in cytosolic proteins, and cytotoxicity in Raw264.7 cells. DBLLs potently suppressed both the enzyme expression and DNA-binding activity of NF-kappa B. Arctiin, arctigenin (1.0 mu m) and matairesinol (10 mu M) inhibited the expression of iNOS by 37.71 +/- 2.86%, 32.51 +/- 4.28%, and 27.44 +/- 2.65%, respectively, and arctiin, arctigenin (0.1 pm) and matairesinol (1.0 gm) inhibited COX-2 expression by 37.93 +/- 7.81%, 26.70 +/- 4.61% and 29.37 +/- 5.21%, respectively. The inhibitory effects of DBLLs on NO and PGE(2) productions were the same patterns as those seen for the reductions in iNOS and COX-2 expression, respectively. Arctiin, arctigenin (1.0 mu M) and matairesinol (10 mu M) significantly (p<0.05) inhibited NF-kappa B DNA binding by 44.85 +/- 6.67%, 44.16 +/- 6.61%, and 44.79 +/- 5.62%, respectively, and arctiin (0.1 gm) and arctigenin (1.0 mu M) significantly (p<0.05) inhibited the phosphorylation of I kappa B by 20.58 +/- 3.86% and 25.99 +/- 6.18%, respectively. Furthermore, arctiin, matairesinol (1.0 gm) and arctigenin (10 pi) inhibited the phosphorylation of IKK by 38.80 +/- 6.64%, 38.33 +/- 6.65%, and 38.57 +/- 8.14%, respectively. In addition, DBLLs potently inhibited the lipopolysaccharide (LPS)-induced activation of MAPKs (SAPK/c-Jun NH2-terminal kinase (JNK), p38, and extracellular signal receptor-activated kinase (ERK)1/2). Overall, arctiin was the most effective; its effect was nearly the same as that of 10 gm helenalin. These findings suggest that treatment with non-toxic DBLLs inhibits not only NF-kappa B and NF-kappa B-regulated protein activation, but also potently inhibits the activations of specific MAPKs.