Enhancing Effects of Scutellaria baicalensis and Some of its Constituents on TGF-β1 Gene Expression in RAW 264.7 Murine Macrophage Cell Line

Abstract

Transforming growth factor β1 (TGF-β1) has been implicated as an inhibitor of cell proliferation and a potent inducer of apoptosis. Scutellaria baicalensis Georgi (SbG) has been widely used in Asia and recent investigations have shown that SbG has anticancer, antiviral, and anti-inflammatory effects. The aim of this study is to investigate the modulatory effect of SbG on TGF-β1 gene expression. By using RAW 264.7 cell line as an in vitro model, the effects of SbG on TGF-β1 gene expression were evaluated by ELISA, reverse-transcription polymerase chain reaction (RT-PCR) and quantitative PCR. Many inhibitors such as mitogen-activated protein kinase (MAPK) inhibitor (PD98059), p38-MAPK inhibitor (SB203580), NF-κB inhibitor (aspirin) and protein kinase C inhibitor (H7) were used to determine the possible signal transduction pathways. The results showed that crude extracts of SbG as well as its pure compounds, baicalin, baicalein and chrysin up-regulated TGF-β1 gene expression on RAW 264.7 cells in a concentration-dependent manner. However, the flavonoid of SbG, wogonin, did not up-regulate TGF-β1 expression on gene and protein levels on RAW264.7 cells. The facts that aspirin and H7 but not PD98059 and SB203580 blocked the enhancing effect suggested that NF-κB and PKC might be involved in baicalin-enhanced TGF-β1 gene expression. We conclude that SbG up-regulates TGF-β1 gene expression on RAW264.7 cells through NF-κB and PKC pathways and this might provide evidence to explain the therapeutic effect and potential adverse effects on the clinical use of Scutellaria baicalensis Georgi.

Scutellaria baicalensis Georgi - Lamiaceae - TGF-β1 - macrophages - alternative medicine

References

• 1 Yue J, Mulder K M. Transforming growth factor-beta signal transduction in epithelial cells.  Pharmacol Ther. 2001;  91 1-34
• 2 Shimizu I. Sho-saiko-to: Japanese herbal medicine for protection against hepatic fibrosis and carcinoma.  J Gastroenterol Hepatol. 2000;  15 D84-90
• 3 Oka H, Yamamoto S, Kuroki T, Harihara S, Marumo T, Kim S R. et al . Prospective study of chemoprevention of hepatocellular carcinoma with Sho-saiko-to (TJ-9).  Cancer. 1995;  76 743-9
• 4 Chitturi S, Farrell G C. Herbal hepatotoxicity: an expanding but poorly defined problem.  J Gastroenterol Hepatol. 2000;  15 1093-9
• 5 Itoh S, Marutani K, Nishijima T, Matsuo S, Itabashi M. Liver injuries induced by herbal medicine, syo-saiko-to (xiao-chai-hu-tang).  Dig Dis Sci. 1995;  40 1845-8
• 6 Ankoma-Sey V. Hepatic regeneration - revisiting the myth of Prometheus.  News Physiol Sci. 1999;  14 149-55
• 7 Stout R D, Suttles J. T cell signaling of macrophage function in inflammatory disease.  Front Biosci. 1997;  2 d197-206
• 8 Krakauer T, Li B Q, Young H A. The flavonoid baicalin inhibits superantigen-induced inflammatory cytokines and chemokines.  FEBS Lett. 2001;  500 52-5
• 9 Chiu J H, Lay I S, Su M Y, Chiu H L, Chiu A C, Lui W Y. et al . Tumor necrosis factor-producing activity of wogonin in RAW 264.7 murine macrophage cell line.  Planta Med. 2002;  68 1036-9
• 10 Kopp E, Ghosh S. Inhibition of NF-kappa B by sodium salicylate and aspirin.  Science. 1994;  265 956-9
• 11 Hidaka H, Inagaki M, Kawamoto S, Sasaki Y. Isoquinolinesulfonamides, novel and potent inhibitors of cyclic nucleotide dependent protein kinase and protein kinase C.  Biochemistry. 1984;  23 5036-41
• 12 Alessi D R, Cuenda A, Cohen P, Dudley D T, Saltiel A R. PD 098 059 is a specific inhibitor of the activation of mitogen-activated protein kinase in vitro and in vivo .  J Biol Chem. 1995;  270 27 489-94
• 13 Young P R, McLaughlin M M, Kumar S, Kassis S, Doyle M L, McNulty D. et al . Pyridinyl imidazole inhibitors of p38 mitogen-activated protein kinase bind in the ATP site.  J Biol Chem. 1997;  272 12 116-21
• 14 Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays.  J Immunol. 1983;  65 55-63
• 15 Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction.  Anal Biochem. 1987;  162 156-9
• 16 Akita K, Okuno M, Enya M, Imai S, Moriwaki H, Kawada N, Suzuki Y, Kojima S. Impaired liver regeneration in mice by lipopolysaccharide via TNF-alpha/kallikrein-mediated activation of latent TGF-beta.  Gastroenterology. 2002;  123 352-64
• 17 Zhang D Y, Wu J, Ye F, Xue L, Jiang S, Yi J. et al . Inhibition of cancer cell proliferation and prostaglandin E2 synthesis by Scutellaria baicalensis .  Cancer Res. 2003;  63 4037-43
• 18 Tang W, Hemm I, Bertram B. Recent development of antitumor agents from Chinese herbal medicines; part I. Low molecular compounds.  Planta Med. 2003;  69 97-108
• 19 Gohongi T, Fukumura D, Boucher Y, Yun C O, Soff G A, Compton C. et al . Tumor-host interactions in the gallbladder suppress distal angiogenesis and tumor growth: involvement of transforming growth factor beta1.  Nat Med. 1999;  5 1203-8
• 20 Weigert C, Sauer U, Brodbeck K, Pfeiffer A, Haring H U, Schleicher E D. AP-1 proteins mediate hyperglycemia-induced activation of the human TGF-beta1 promoter in mesangial cells.  J Am Soc Nephrol. 2000;  11 2007-16
• 21 Cohen M P, Shea E, Chen S, Shearman C W. Glycated albumin increases oxidative stress, activates NF-kappa B and extracellular signal-regulated kinase (ERK), and stimulates ERK-dependent transforming growth factor-beta 1 production in macrophage RAW cells.  J Lab Clin Med. 2003;  141 242-9
• 22 Rameshwar P, Narayanan R, Qian J, Denny T N, Colon C, Gascon P. NF-kappa B as a central mediator in the induction of TGF-beta in monocytes from patients with idiopathic myelofibrosis: an inflammatory response beyond the realm of homeostasis.  J Immunol. 2000;  165 2271-7
• 23 Akashi S, Shimazu R, Ogata H, Nagai Y, Takeda K, Kimoto M. et al . Cutting edge: cell surface expression and lipopolysaccharide signaling via the toll-like receptor 4-MD-2 complex on mouse peritoneal macrophages.  J Immunol. 2000;  164 3471-5