Andrographolide Inhibits IFN-γ and IL-2 Cytokine Production and Protects Against Cell Apoptosis

Abstract

Andrographolide is the main labdane diterpene present in Andrographis paniculata. Two lines of evidence report immunostimulant and anti-inflamatory properties for andrographolide in different models. Using murine T-cells in vitro we demonstrated that andrographolide and to a lesser extent, 14-deoxyandrographolide (14-DAP), reduced significantly, in a dose-dependent manner, the IFN-γ production induced by concanavaline A (CON-A), with an IC₅₀ of 1.7 ± 0.07 μM and 35.8 ± 0.50 μM, respectively. Andrographolide, but not 14-DAP, inhibited partially the IL-2 production induced by CON-A. Andrographolide at doses of 5 and 10 μM reduced the extracellular-signal-regulated protein kinase (ERK1/2) phosphorylation induced by CON-A, whereas 14-DAP only reduced ERK1 and partially the ERK2 phosphorylation. The inhibition of ERK1/2 phosphorylation was associated to a decrease in the IFN-γ production, due that UO126, a specific ERK1/2 inhibitor, also reduced the IFN-γ production in murine T-cells induced by CON-A. Additionally, andrographolide and to a lesser extent 14-DAP, at doses of 50 μM and 100 μM, respectively, reduced the apoptosis induced by hydrocortisone and PMA in thymocytes, which was associated to a decrease in caspase-3 like activity. We conclude that both diterpenic labdanes isolated from A. paniculata can exert potent immunosuppressant effects without affecting the viability of the cells.

Key words

Andrographolide - 14-deoxyandrographolide - apoptosis - cytokine - immunosuppressant - Acanthaceae

References

• 1 Poolsup N, Suthisisang C, Prathanturarug S, Asawamekin A, Chanchareon U. Andrographis paniculata in the symptomatic treatment of uncomplicated upper respiratory tract infection: systematic review of randomized controlled trials.  J Clin Pharm Ther. 2004;  29 37-45
• 2 Chiou W F, Lin J J, Chen C F. Andrographolide suppresses the expression of inducible nitric oxide synthase in macrophage and restores the vasoconstriction in rat aorta treated with lipopolysaccharide.  Br J Pharmacol. 1998;  125 327-334
• 3 Shen Y C, Chen C F, Chiou W F. Andrographolide prevents oxygen radical production by human neutrophils: possible mechanism(s) involved in its anti-inflammatory effect.  Br J Pharmacol. 2002;  135 399-406
• 4 Wang T, Liu B, Zhang W, Wilson B, Hong J S. Andrographolide reduces inflammation-mediated dopaminergic neurodegeneration in mesencephalic neuron-glia cultures by inhibiting microglial activation.  J Pharmacol Exp Ther. 2004;  308 975-83
• 5 Dimas K, Demetzos C, Vaos V, Ioannidis P, Trangas T. Labdane type diterpenes down-regulate the expression of c-Myc protein, but not of Bcl-2, in human leukemia T-cells undergoing apoptosis.  Leuk Res. 2001;  25 449-54
• 6 Ojeda F, Guarda M I, Maldonado C, Folch H. Protein kinase-C involvement in thymocyte apoptosis induced by hydrocortisone.  Cell Immunol. 1990;  125 535-9
• 7 Hidalgo M A, Ojeda F, Eyre P, LaBranche T P, Smith C, Hancke J L, Burgos R A. Platelet-activating factor increases pH(i) in bovine neutrophils through the PI3K-ERK1/2 pathway.  Br J Pharmacol. 2004;  141 311-21
• 8 Ojeda F, Guarda M I, Maldonado C, Folch H. A flow-cytometric method to study DNA fragmentation in lymphocytes.  J Immunol Methods. 1992;  152 171-6
• 9 Matsuda T, Kuroyanagi M, Sugiyama S, Umehara K, Ueno A, Nishi K. Cell differentiation-inducing diterpenes from Andrographis paniculata Nees.  Chem Pharm Bull. 1994;  42 1216-25
• 10 Kumar R A, Sridevi K, Kumar N V, Nanduri S, Rajagopal S. Anticancer and immunostimulatory compounds from Andrographis paniculata .  J Ethnopharmacol. 2004;  92 291-5
• 11 Panossian A, Davtyan T, Gukassyan N, Gukasova G, Mamikonyan G, Gabrielian E, Wikman G. Effect of andrographolide and Kan Jang - fixed combination of extract SHA-10 and extract SHE-3 on proliferation of human lymphocytes, production of cytokines and immune activation markers in the whole blood cells culture.  Phytomedicine. 2002;  9 598-605
• 12 Puri A, Saxena R, Saxena R P, Saxena K C, Srivastava V, Tandon J S. Immunostimulant agents from Andrographis paniculata .  J Nat Prod. 1993;  56 995-9
• 13 Chiou W F, Chen C F, Lin J J. Mechanisms of suppression of inducible nitric oxide synthase (iNOS) expression in RAW 264.7 cells by andrographolide.  Br J Pharmacol. 2000;  129 1553-60
• 14 Habtemariam S. Andrographolide inhibits the tumour necrosis factor-alpha-induced upregulation of ICAM-1 expression and endothelial-monocyte adhesion.  Phytother Res. 1998;  12 37-40
• 15 Amrani Y, Ammit A J, Panettieri R A, Jr. Tumor necrosis factor receptor (TNFR) 1, but not TNFR2, mediates tumor necrosis factor-alpha-induced interleukin-6 and RANTES in human airway smooth muscle cells: role of p38 and p42/44 mitogen-activated protein kinases.  Mol Pharmacol. 2001;  60 646-55
• 16 Kim S H, Kim J, Sharma R P. Inhibition of p38 and ERK MAP kinases blocks endotoxin-induced nitric oxide production and differentially modulates cytokine expression.  Pharmacol Res. 2004;  49 433-9
• 17 Lo C J. MAPK regulation of prostaglandin E2 production by lipopolysaccharide-stimulated macrophages is not dependent on nuclear factor kappaB.  J Surg Res. 2003;  113 89-94
• 18 Dumont F J, Staruch M J, Fischer P, DaSilva C, Camacho R. Inhibition of T cell activation by pharmacologic disruption of the MEK1/ERK MAP kinase or calcineurin signaling pathways results in differential modulation of cytokine production.  J Immunol. 1998;  160 2579-89
• 19 Tsai H R, Yang L M, Tsai W J, Chiou W F. Andrographolide acts through inhibition of ERK1/2 and Akt phosphorylation to suppress chemotactic migration.  Eur J Pharmacol. 2004;  498 45-52
• 20 Egerton M, Fitzpatrick D R, Kelso A. Activation of the extracellular signal-regulated kinase pathway is differentially required for TCR-stimulated production of six cytokines in primary T lymphocytes.  Int Immunol. 1998;  10 223-9
• 21 Navarro A, de las Heras B, Villar A. Immunomodulating properties of the diterpene andalusol.  Planta Med. 2000;  66 289-91
• 22 Souza-Fagundes E M, Gazzinelli G, Parreira G G, Martins-Filho O A, Amarante-Mendes G P, Correa-Oliveira R, Zani C L. In vitro activity of labdane diterpene from Alomia myriadenia (Asteraceae): immunosuppression via induction of apoptosis in monocytes.  Int Immunopharmacol. 2003;  3 383-92
• 23 Cifone M G, Migliorati G, Parroni R, Marchetti C, Millimaggi D, Santoni A, Riccardi C. Dexamethasone-induced thymocyte apoptosis: apoptotic signal involves the sequential activation of phosphoinositide-specific phospholipase C, acidic sphingomyelinase, and caspases.  Blood. 1999;  93 2282-96
• 24 Chen J H, Hsiao G, Lee A R, Wu C C, Yen M H. Andrographolide suppresses endothelial cell apoptosis via activation of phosphatidyl inositol-3-kinase/Akt pathway.  Biochem Pharmacol. 2004;  67 1337-45
• 25 Bommhardt U, Scheuring Y, Bickel C, Zamoyska R, Hunig T. MEK activity regulates negative selection of immature CD4+CD8+ thymocytes.  J Immunol. 2000;  164 2326-37
• 26 Skurkovich B, Skurkovich S. Anti-interferon-gamma antibodies in the treatment of autoimmune diseases.  Curr Opin Mol Ther. 2003;  5 52-7