Tumor Necrosis Factor-Producing Activity of Wogonin in RAW 264.7 Murine Macrophage Cell Line

Abstract

Wogonin from Scutellaria baicalensis, was demonstrated to increase nitric oxide (NO) in the murine macrophage cell line RAW 264.7. It is our aim to investigate the modulatory effect of wogonin on tumor necrosis factor-α (TNF-α) gene expression. By using RAW 264.7 as an in vitro model, the effects of wogonin on inducible nitric oxide synthase (NOS2) and TNF-α gene expression were evaluated by ELISA and reverse transcribed polymerase chain reaction (RT-PCR). Aspirin and H7 were used to determine the possible signal transduction pathways. The results showed that wogonin at the concentration of 10^-5 M and 10^-6 M up-regulated NOS2 gene expression in RAW 264.7 cells. Besides, wogonin up-regulated the gene expression of TNF-α, in terms of TNF-α secretion and transcription, in a dose dependent manner. The fact that aspirin but not H7 blocks the enhancing effect suggests that NF-κB might be involved in wogonin-enhanced TNF-α gene expression. We conclude that a low concentration of wogonin up-regulates NOS2 and TNF-α gene expression through NF-κB pathway.

References

• 1 Currier A R, Ziegler M H, Riley M M, Babcock T A, Telbis V P, Carlin J M. Tumor necrosis factor-alpha and lipopolysaccharide enhance interferon-induced antichlamydial indoleamine dioxygenase activity independently.  J Interf Cyto Res. 2000;  20 369-76
• 2 Murphey E D, Traber D L. Pretreatment with tumor necrosis factor-alpha attenuates arterial hypotension and mortality induced by endotoxin in pigs.  Crit Care Med. 2000;  28 2015-21
• 3 Daubener W, Remscheid C, Nockemann S, Pilz K, Seghrouchni S, Mackenzie C, Hadding U. Anti-parasitic effector mechanisms in human brain tumor cells: role of interferon-gamma and tumor necrosis factor-alpha.  Eur J Immunol. 1996;  26 487-92
• 4 Ten Hagen T L, Van Der Veen A H, Nooijen P T, Van Tiel S T, Seynhaeve A L, Eggermont A M. Low-dose tumor necrosis factor-alpha augments antitumor activity of stealth liposomal doxorubicin (DOXIL) in soft tissue sarcoma-bearing rats.  Int J Cancer. 2000;  87 829-37
• 5 Chen Y C, Shen S C, Chen L G, Lee T J, Yang L L. Wogonin, baicalin, and baicalein inhibition of inducible nitric oxide synthase and cyclooxygenase-2 gene expressions induced by nitric oxide synthase inhibitors and lipopolysaccharide.  Biochem Pharmacol. 2001;  61 1417-27
• 6 Huang R L, Chen C C, Huang H L, Chang C G, Chen C F, Chang C, Hsieh M T. Anti-hepatitis B virus effects of wogonin isolated from Scutellaria baicalensis .  Planta Medica. 2000;  66 694-8
• 7 Gao Z, Huang K, Yang X, Xu H. Free radical scavenging and antioxidant activities of flavonoids extracted from the radix of Scutellaria baicalensis Georgi.  Biochim Biophysic Acta. 1999;  1472 643-50
• 8 Ohtsuka M, Fukuda K, Yano H, Kojiro M. Effects of nine active ingredients in Chinese herbal medicine sho-saiko-to on 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide mutagenicity.  Jap J Cancer Res. 1995;  86 1131-5
• 9 Chi Y S, Jong H G, Son K H, Chang H W, Kang S S, Kim H P. Effects of naturally occurring prenylated flavonoids on enzymes metabolizing arachidonic acid: cyclooxygenases and lipoxygenases.  Biochem Pharmacol. 2001;  62 185-91
• 10 Kim H K, Cheon B S, Kim Y H, Kim S Y, Kim H P. Effects of naturally occurring flavonoids on nitric oxide production in the macrophage cell line RAW 264.7 and their structure-activity relationships.  Biochem Pharmacol. 1999;  58 59-65
• 11 Wakabayashi I. Inhibitory effects of baicalein and wogonin on lipopolysaccharide-induced nitric oxide production in macrophages.  Pharmacol Toxicol . 1999;  84 88-91
• 12 Kim H, Kim Y S, Kim S Y, Suk K. The plant flavonoid wogonin suppresses death of activated C6 rat glial cells by inhibiting nitric oxide production.  Neurosci Let. 2001;  309 67-71
• 13 Kim H M, Moon E J, Li E, Kim K M, Nam S Y, Chung C K. The nitric oxide-producing activities of Scutellaria baicalensis .  Toxicology. 1999;  135 109-15
• 14 Wu K K. Aspirin and salicylate: An old remedy with a new twist.  Circulation. 2000;  102 2022-3
• 15 Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays.  J Immunol Met. 1983;  65 55-63
• 16 Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction.  Anal Biochem. 1987;  162 156-9
• 17 Wasser S, Ho J M, Ang H K, Tan C E. Salvia miltiorrhiza reduces experimentally-induced hepatic fibrosis in rats.  J Hepatol. 1998;  29 760-71