Activation of α_1A-Adrenoceptors by Genistein at Concentrations Lower than that to Inhibit Tyrosine Kinase in Cultured C₂C₁₂ Cells

Abstract

Genistein, an isoflavonoid natural product, is widely used to inhibit protein tyrosine kinase (PTK). In the present study, we investigated the possible influence of genistein on α₁-adrenoceptors (AR) in cultured C₂C₁₂ cells. Genistein enhanced the uptake of radioactive glucose into C₂C₁₂ cells in a concentration-dependent manner. Similar results were also observed in samples treated with daidzein, the inactive congener for PTK inhibition. The effect of genistein on α₁-AR was further characterized using the displacement of [³ H]prazosin binding in C₂C₁₂ cells. The increase in radioactive glucose uptake by genistein was abolished by RS17053 at a concentration sufficient to block α_1A-AR. The pharmacological inhibition of phospholipase C (PLC) by U73122 resulted in a concentration-dependent reduction of genistein-stimulated glucose uptake in C₂C₁₂ cells. This inhibition by U73122 was specific because the inactive congener, U73343, failed to modify the action of genistein. Moreover, genistein can activate α_1A-AR at a concentration (1 μmol/L) lower than that (50 μmol/L) needed to abolish the insulin-stimulated phosphorylation of PTK. The obtained data indicate an activation of α_1A-AR by genistein to increase the glucose uptake into C₂C₁₂ cells and this supports the application of genistein as a TK inhibitor.

Genistein - C₂C₁₂ cells - α_1A-adrenoceptor - phospholipase C - protein tyrosine kinase

References

• 1 Akiyama T, Ishida J, Nakagawa S, Ogawara H, Watanabe S, Itoh N, Shibuya M, Fukami Y. Genistein, a specific inhibitor of tyrosine-specific protein kinase.  J Biol Chem. 1987;  262 5592-5
• 2 Shuba L M, McDonald T F. Synergistic activation of guinea-pig cardiac cystic fibrosis transmembrane conductance regulator by the tyrosine kinase inhibitor genistein and cAMP.  J Physiol. 1997;  505 23-40
• 3 Kim D H, Yu K U, Bae E A, Han M J. Metabolism of puerarin and daidzein by human intestinal bacteria and their relation to in vitro cytotoxicity.  Biol Pharm Bull. 1998;  21 628-30
• 4 Hsu H H, Chang C K, Su H C, Liu I M, Cheng J T. Stimulatory effect of puerarin on alpha1A-adrenoceptor to increase glucose uptake into cultured C2C12 cells of mice.  Planta Med. 2002;  68 999-1003
• 5 Sheriff S, Fischer J E, Balasubramaniam A. Amylin inhibits insulin-stimulated glucose uptake in C₂C₁₂ muscle cell line through a cholera-toxin-sensitive mechanism.  Biochim Biophys Acta. 1992;  1136 219-22
• 6 Cheng J T, Liu I M. Stimulatory effect of caffeic acid on α_1A-adrenoceptors to increase glucose uptake into cultured C₂C₁₂ cells.  Naunyn Schmiedebergs Arch Pharmacol. 2000;  362 122-7
• 7 Christ C Y, Hunt D, Hancock J, Garcia-Macedo R, Mandarino L J, Ivy J L. Exercise training improves muscle insulin resistance but not insulin receptor signaling in obese Zucker rats.  J Appl Physiol. 2002;  92 736-44
• 8 Marshall I, Burt R P, Green G M, Hussain M B, Chapple C R. Different subtypes of alpha 1A-adrenoceptor mediating contraction of rat epididymal vas deferens, rat hepatic portal vein and human prostate distinguished by the antagonist RS 17 053.  Br J Pharmacol. 1996;  119 407-15
• 9 Zhong H, Minneman K P. α₁-Adrenoceptor subtypes.  Eur J Pharmacol. 1999;  375 261-76
• 10 Hieble J P, Bylund D B, Clarke D E, Eikenburg D C, Langer S Z, Lefkowitz R J, Minneman K P, Ruffolo J r. International Union of Pharmacology: X. Recommendation for nomenclature of alpha 1-adrenoceptors: consensus update.  Pharmacol Rev. 1995;  47 267-70
• 11 Ishizuka T, Cooper D R, Hernandez H, Buckley D, Standaert M, Farese R V. Effects of insulin on diacylglycerol-protein kinase C signaling in rat diaphragm and soleus muscles and relationship to glucose transport.  Diabetes. 1990;  39 181-90
• 12 Van Epps-Fung M, Gupta K, Hatdy R W, Wells A. A role for phospholipase C activity in GLUT4-mediated glucose transport.  Endocrinology. 1997;  138 5170-5
• 13 Smallridge R C, Kiang J G, Gist I D, Fein H G, Gallowat R J. U-73 122, an aminosteroid phospholipase C antagonist, non-competitively inhibits thyrotropin-releasing hormone effects in GH3 rat pituitary cell.  Endocrinology. 1992;  131 1883-8
• 14 Muto Y, Nagao T, Urushidani T. The putative phospholipase C inhibitor U73122 and its negative control, U73343, elicit unexpected effects on the rabbit parietal cell.  J Pharmacol Exp Ther. 1997;  282 1379-88
• 15 Myers MG J r, White M F. Insulin signal transduction and the IRS proteins.  Ann Rev Pharmacol Toxicol. 1996;  36 615-58
• 16 Drejer K. The bioactivity of insulin analogues from in vitro receptor binding to in vivo glucose uptake.  Diabetes Metab Rev. 1992;  8 259-85