Brazilein-induced contraction of rat arterial smooth muscle involves activation of Ca2+ entry and ROK, ERK pathways

Abstract

Brazilein (6a,7-dihydro-3,6a,10-trihydroxy-benz[b]indeno[1,2-d]pyran-9(6H)-one) is a compound isolated from Caesalpinia sappan. The vasoactivities of brazilein were evaluated in isolated rat thoracic aorta. The results showed that brazilein can dose-dependently induce contraction of rat thoracic aorta in the resting and phenylephrine pre-evoked state. The average response to 100 μM of brazilein was 30% of the 50 mM KCl contraction, 26% of the 10 μM phenylephrine and 116% of the 20 mM caffeine contraction in comparison. The effects of vasocontraction were proved not to be endothelial dependent and could not be inhibited by α-adrenergic receptor blocker phentolamine, β-adrenergic receptor blocker propranolol, M-adrenaline receptor blocker atropine, angiotensin II receptor blocker losartan or the non-selective nitric oxide synthase (NOS) inhibitor NG-Nitro-l-Arginine Methyl Ester (l-NAME). However the influx of extracellular calcium seemed to be required for this action, because depletion of extracellular calcium and the addition of L-type calcium ion channel antagonist (nimodipine and diltiazem), calcium ion channel activator (BAY-K8644) and potassium ion channel opener (pinacidil) could significantly affect the contraction induced by brazilein. We also investigated the possible signal mechanisms underlying brazilein-induced contraction using selective inhibitors. The inhibitors of myosin light chain kinase (MLCK), Rho-kinase (ROK) and extracellular signal regulated kinase (ERK) can suppress the effect of brazilein respectively, whereas inhibitors of other signaling or receptor molecules such as protein kinase C (PKC) and inositol 1,4,5-triphosphate (IP3) receptor had no effect. All these results demonstrated that brazilein can induce contraction of rat aorta, that the Ca2+ influx, ROK and ERK signal pathways and MLCK activation must be involved in the contractile processes.