| Budget Amount *help |
¥3,600,000 (Direct Cost: ¥3,600,000)
Fiscal Year 2004: ¥800,000 (Direct Cost: ¥800,000)
Fiscal Year 2003: ¥2,800,000 (Direct Cost: ¥2,800,000)
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| Research Abstract |
This study was designed to determine involvement of Rho in the pathophysiology of bronchial asthma such as bronchial hyperreactivity, β-adrenergic desensitization, and airway remodeling. After exposure of the fura-2 loaded strip of tracheal smooth muscle to sphingosine 1-phosphate (S1IP,10 μM) for more than 15 min, contraction by methacholine(MCh), a muscarinic receptor agonist, was markedly augmented in a time-dependent fashion. However, concentration of intracellular Ca^<2+>([Ca^<2+>]i) by MCh was not affected. The increased response to MCh (bronchial hyperreactivity) by S1P was antagonized in the presence of 0.01-1 μM Y-27632, a selective inhibitor of Rho-kinase, in a concentration-dependent fashion. Pre exposure to pertussis toxin over 6 hours also prohibited the increased response to MCh by S1P. Next, after exposure of the tissues to S1P (<1 μM) for an equivalent time, the inhibitory effect of isoproterenol (ISO), a β-adrenergic receptor agonist, against MCh-induced contraction wa
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s markedly attenuated without no change in [Ca^<2+>]i. The reduced responsiveness to ISO (desensitization of β-adrenergic receptors) by S1P was inhibited by Y-27632 in a concentration-dependent fashion. Pre exposure to pertussis toxin over an equivalent time also prohibited the reduced response to ISO by S1P. On the other hand, after repeated exposure to ISO every 30 min, response to ISO was gradually attenuated with an increase in [Ca^<2+>]i. This reduced responsiveness to ISO was inhibited in the presence of verapamil, a selective antagonist of voltage-dependent Ca^<2+> channels. In cultured bronchial smooth muscle cells, S1P phosphorylated myosin phosphatase targeting protein (MYPT1) that is a specific protein in myosin phosphatase affected by Rho-kinase. Proliferation of the bronchial smooth muscle cells (airway remodeling) was reduced by Y-27632 in a concentration-dependent fashion. In conclusion, Rho/Rho-kinase processes play an important role in the pathophysiology of bronchial asthma. Less
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