Theophylline and cAMP inhibit lysophosphatidic acid‐induced hyperresponsiveness of bovine tracheal smooth muscle cells

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Abstract
We have established an in vitro model of airway hyperresponsiveness, using a bovine tracheal smooth muscle cell (BTSMC)‐embedded collagen gel lattice. When the gel was pretreated with lysophosphatidic acid (LPA), which activates the small G protein RhoA, ATP‐ and high K+ solution‐induced gel contraction was significantly augmented. This was not due to the modulation of Ca2+ mobilizing properties, since ATP‐ and high K+‐induced Ca2+ transients were not significantly different between control and LPA‐treated BTSMC. Y‐27632, an inhibitor of Rho‐kinase, suppressed the LPA‐induced augmentation of gel contraction, whereas it did not inhibit the contraction of control gels. Theophylline (> 1 μm) reversed the LPA‐induced augmentation of gel contraction, whereas it inhibited control gel contraction only with a very high concentration (100 μm). We confirmed that theophylline increased the intracellular concentration of cAMP ([cAMP]i) in BTSMC. Elevation of [cAMP]i with dibutyryl cAMP or forskolin also reversed the LPA‐induced augmentation of gel contraction. Furthermore, theophylline, as well as dibutyryl cAMP and forskolin, suppressed the LPA‐induced membrane translocation of RhoA, indicating that they prevented airway hyperresponsiveness by inhibiting RhoA. We conclude from these results that theophylline inhibits LPA‐induced, RhoA/Rho‐kinase‐mediated hyperresponsiveness of tracheal smooth muscle cells due to the accumulation of cAMP.
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