Interleukin-13 (IL-13) is one of the central mediators for development of airway hyperresponsiveness in asthma. However, its effect on bronchial smooth muscle (BSM) is not well known. Recent studies revealed an involvement of RhoA/Rho-kinase in BSM contraction, and this pathway has now been proposed as a new target for asthma therapy. To elucidate the role of IL-13 on the induction of BSM hyperresponsiveness, effects of IL-13 on contractility and RhoA expression in BSMs were investigated. Male BALB/c mice were sensitized and repeatedly challenged with ovalbumin antigen. In the repeatedly antigen-challenged mice, marked airway inflammation and BSM hyperresponsiveness with an up-regulation of IL-13 in bronchoalveolar lavage fluids were observed. In cultured human BSM cells, IL-13 caused an up-regulation of RhoA. The IL-13-induced up-regulation of RhoA was inhibited by leflunomide, an inhibitor of signal transducer and activator of transcription 6 (STAT6). In isolated BSM tissues of naive mice, the contractility was significantly enhanced by organ culture in the presence of IL-13. Moreover, in vivo treatment of airways with IL-13 by intranasal instillation caused a BSM hyperresponsiveness with an up-regulation of RhoA in naive mice. These findings suggest that IL-13/STAT6 signaling is critical for development of antigen-induced BSM hyperresponsiveness and that agents that specifically inhibit this pathway in BSM may provide a novel strategy for the treatment of asthma.

These findings suggest that RhoA expression is negatively regulated by miR-133a in BSMs. IL-13 might, at least in part, contribute to the reduction of miR-133a.

Background: It has recently been suggested that RhoA plays an important role in the enhancement of the Ca 2+ sensitization of smooth muscle contraction. In the present study, a participation of RhoA-mediated Ca 2+ sensitization in the augmented bronchial smooth muscle (BSM) contraction in a murine model of allergic asthma was examined.

Abstract-Nitric oxide/cGMP pathway induces vasodilatation, yet the underlying mechanism is obscure. In the present study, we studied the mechanism of cGMP-induced relaxation of the smooth muscle contractile apparatus using permeabilized rabbit femoral arterial smooth muscle. 8-Br-cGMP-induced relaxation was accompanied with a decrease in myosin light chain (MLC) phosphorylation. MLC phosphatase (MLCP) activity, once decreased by agoniststimulation, recovered to the resting level on addition of 8-Br-cGMP. Because MLCP activity is regulated by the phosphorylation of a MLCP-specific inhibitor, CPI17 at Thr38 and MBS (myosin binding subunit of MLCP) at Thr696, we examined the effect of 8-Br-cGMP on the phosphorylation of these MLCP modulators. Whereas CPI17 phosphorylation was unchanged after addition of 8-Br-cGMP, MBS phosphorylation at Thr696 was significantly decreased by 8-Br-cGMP. We found that 8-Br-cGMP markedly increased MBS phosphorylation at Ser695 in the fiber pretreated with phenylephrine. MBS phosphorylation of Thr696 phosphorylated MBS at Ser695 partially resumed MLCP activity inhibited by Thr696 phosphorylation. Whereas Ser695 phosphorylation was markedly increased, the extent of diphosphorylated MBS at Ser695 and Thr696 in fibers was unchanged after cGMP-stimulation. We found that MBS phosphatase activity in arteries for both diphosphorylated MBS and monophosphorylated MBS at Thr696 significantly increased by 8-Br-cGMP, whereas MBS kinase activity was unchanged. These results suggest that the phosphorylation at Ser640 induced by cGMP shifted the equilibrium of the Thr641 phosphorylation toward dephosphorylation, thus increasing MLCP activity. This results in the decrease in MLC phosphorylation and smooth muscle relaxation. Key Words: cGMP Ⅲ myosin light chain phosphatase Ⅲ vasodilation Ⅲ phosphorylation Ⅲ smooth muscle I t has been known that endothelial-derived nitric oxide (NO) acts as a vasodilator, 1 and the pharmacological NO producing drugs have been used to prevent acute heart failure. NO has been defined as the activator of soluble guanylate cyclase, 2 thus increasing cGMP. A number of studies have indicated that cGMP induces relaxation of various smooth muscles contracted either by receptor-coupling agonists or depolarization 3 ; therefore, NO-induced vasodilation is thought to be attributable to the cGMP-induced relaxation of vascular smooth muscle. The key question is how cGMP triggers the vascular smooth muscle relaxation.Smooth muscle contraction is controlled by the phosphorylation of the regulatory light chain (RLC) of myosin at Ser19 4,5 by Ca 2ϩ /calmodulin-dependent protein kinase, called myosin light chain kinase (MLCK). 4 -6 On the other hand, MLC phosphatase (MLCP) activity is also regulated during the agonist-induced contraction of smooth muscle, thus contributing to the increase in RLC phosphorylation, but in contrast to the regulation of MLCK, the mechanism is Ca 2ϩ independent. 6 MLCP consists of 3 subunits, a myosin binding large subunit (MBS), 7,8 a 20-kDa small subunit (M2...

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