Effect of genistein on agonist-induced airway smooth muscle contraction

Abstract

Male Dunkin–Hartley and Tricolour guinea-pigs (250–400 g) were used according to local ethical committee and Home Offi ce regulations. In experiments with an immunological stimulant, animals were sensitised by a single subcutaneous injection of 0.5 ml saline containing 10 μg ovalbumin (chicken) and 100 mg aluminium hydroxide, before being used 3 weeks later. Isolated tracheal rings (GPTR, 1–3 per preparation) were suspended in Krebs–Henseleit (KH) buffer gassed with 95 % O2/ 5 % CO2 , at 37 °C and attached to an isometric transducer, which was connected to an amplifi er and in turn to a Picolog analog-digital converter (ADC-42) and computer. They were put under a basal tension of 0.5–1.0 g. Contraction of GPTR was induced by carbachol (0.5–50 μM, acting via M3-receptors), histamine (5–100 μM, via H1-receptors), NaF (1– 10 mM, G-protein activator), or antigen (1–10,000 ng ml). We tested the following compounds for inhibition of these contractile responses: genistein, tyrphostin 51 (both best known as PTK inhibitors), daidzein (PTK-inactive analogue of genistein) and benzalkonium chloride (1 μM, G-protein Gi-type inhibitor). Agents tested for their ability to reverse the blockade (15 min pre-incubation) by the latter 4 substances of the contractile response to histamine, carbachol or NaF were: indomethacin (up to 10 M), aminoguanidine (10 M), methylene blue (20 μM), Ntosyl-L-phenylalanine chloromethyl ketone (TPCK, a-chymotrypsin inhibitor, 500 μM), or cyclosporin-A (CsA 0.5–1 μM). Other details of the methods and materials were as previously described [2, 3]. Results and discussion