h-89 and Asthma

H89 is a potent inhibitor of Protein Kinase A (PKA) and Mitogen- and Stress-Activated protein Kinase 1 (MSK1) with some inhibitory activity on other members of the AGC kinase family. H89 has been extensively used in vitro but its anti-inflammatory potential in vivo has not been reported to date. To assess the anti-inflammatory properties of H89 in mouse models of asthma.. Mice were sensitized intraperitoneally (i.p.) to ovalbumin (OVA) with or without alum, and challenged intranasally with OVA. H89 (10 mg/kg) or vehicle was given i.p. two hours before each OVA challenge. Airway hyperresponsiveness (AHR) was assessed by whole-body barometric plethysmography. Inflammation was assessed by the total and differential cell counts and IL-4 and IL-5 levels in bronchoalveolar lavage (BAL) fluid. Lung inflammation, mucus production and mast cell numbers were analyzed after histochemistry. We show that treatment with H89 reduces AHR, lung inflammation, mast cell numbers and mucus production. H89 also inhibits IL-4 and IL-5 production and infiltration of eosinophils, neutrophils and lymphocytes in BAL fluid.. Taken together, our findings implicate that blockade of AGC kinases may have therapeutic potential for the treatment of allergic airway inflammation.

Topics: Animals; Asthma; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Immunoglobulin E; Immunoglobulin G; Inflammation; Isoquinolines; Lung; Male; Mast Cells; Mice; Mucus; Ovalbumin; Protein Kinase Inhibitors; Sulfonamides; Th2 Cells

Beta2-adrenergic receptor (beta2AR) agonists acutely relieve bronchoconstriction via cAMP-mediated relaxation of airway smooth muscle (ASM). Airway constrictor responsiveness may be significantly heightened, however, following protracted exposure to these agents, presumably reflecting the effects of beta2AR desensitization in ASM accompanying prolonged cAMP signaling. Because cAMP phosphodiesterase (PDE) activity can significantly modulate ASM contractility, we investigated the mechanism regulating PDE expression and its potential role in mediating changes in agonist-induced constrictor and relaxation responsiveness in ASM following its heterologous beta2AR desensitization by prolonged exposure to cAMP-elevating agents. Isolated rabbit ASM tissues and cultured human ASM cells treated for 24 h with the receptor- or nonreceptor-coupled cAMP-stimulating agent, prostaglandin E(2) (PGE(2)) or forskolin, respectively, exhibited constrictor hyperresponsiveness to acetylcholine and impaired beta2AR-mediated relaxation and cAMP accumulation. These proasthmatic-like changes in ASM function were associated with upregulated PDE4 activity, reflective of increased transcription of the PDE4D5 isoform, and were prevented by pretreatment of the ASM with a PDE4 inhibitor. Extended studies using gene silencing and pharmacological approaches to inhibit specific intracellular signaling molecules demonstrated that the mechanism underlying PGE(2)-induced transcriptional upregulation of PDE4D5 involves PKA-dependent activation of G(i) protein signaling via the betagamma-subunits, the latter eliciting downstream activation of ERK1/2 and its consequent induction of PDE4D5 transcription. Collectively, these findings identify that beta2AR desensitization in ASM following prolonged exposure to cAMP-elevating agents is associated with proasthmatic-like changes in ASM responsiveness that are mediated by upregulated PDE4 expression induced by activated cross talk between the PKA and ERK1/2 signaling pathways.

Topics: Adrenergic beta-2 Receptor Agonists; Animals; Asthma; Butadienes; Cells, Cultured; Colforsin; Cyclic AMP; Cyclic AMP Response Element-Binding Protein; Cyclic AMP-Dependent Protein Kinases; Cyclic Nucleotide Phosphodiesterases, Type 4; Dinoprostone; Enzyme Induction; Humans; Isoquinolines; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Muscle Contraction; Muscle Relaxation; Nitriles; Pertussis Toxin; Phosphodiesterase 4 Inhibitors; Rabbits; Receptors, G-Protein-Coupled; Rolipram; Signal Transduction; Sulfonamides; Trachea