Alison S. Williams

Effect of Transforming Growth Factor-beta Receptor I Kinase Inhibitor 2,4-Disubstituted Pteridine (SD-208) in Chronic Allergic Airway Inflammation and Remodeling

Transforming growth factor (TGF)-β is a multifunctional regulator of cell growth and differentiation with both pro- and anti-inflammatory properties. We used an inhibitor of TGF-β receptor I (TGF-βRI) kinase, SD-208 (2,4-disubstituted pteridine, a ATP-competitive inhibitor of TGF-βRI kinase), to determine the role of TGF-β in airway allergic inflammation and remodeling. Brown-Norway rats sensitized and repeatedly exposed to ovalbumin (OVA) aerosol challenge were orally administered SD-208 twice daily, before each of six OVA exposures to determine the preventive effects, or only before each of the last three of six OVA exposures to investigate its reversal effects. SD-208 (60 mg/kg) reversed bronchial hyperresponsiveness (BHR) induced by repeated allergen exposure, but it did not prevent it. SD-208 prevented changes in serum total and OVA-specific IgE, but it did not reverse them. SD-208 had both a preventive and reversal effect on airway inflammation as measured by major basic protein-positive eosinophils and CD2+ T-cell counts in mucosal airways, cell proliferation measured by 5-bromo-2′-deoxyuridine expression in airway smooth muscle (ASM) cells and epithelial cells, and goblet cell hyperplasia induced by repeated allergen challenges. There was a significant decrease in intracellular Smad2/3 expression. SD-208 did not significantly decrease the increased ASM thickness induced by allergen exposure. These findings support a proinflammatory and proremodeling role for TGF-β in allergic airway inflammation. Inhibition of TGF-βRI kinase activities by SD-208 may be a useful approach to the reversal of BHR and to the prevention and reversal of inflammatory and remodeling features of chronic asthma.

Attenuation of Ozone-Induced Airway Inflammation and Hyper-Responsiveness by c-Jun NH2 Terminal Kinase Inhibitor SP600125

Ozone has potent oxidizing properties, and exposure to ozone causes airway hyper-responsiveness (AHR) and lung inflammation. We determined the importance of c-Jun NH2 terminal kinase (JNK), a member of the mitogen-activated protein kinase pathway, in ozone-induced AHR and inflammation. SP600125 [anthra[1,9-cd] pyrazol-6 (2H)-one], a specific JNK inhibitor (30 mg/kg) or vehicle, was administered by intraperitoneal injection before and after ozone exposure (3 ppm for 3 h). SP600125 significantly reduced total cells, and neutrophils in bronchoalveolar fluid recovered at 20 to 24 h after exposure and inhibited ozone-induced AHR. Ozone exposure induced activation of JNK in the lung as measured by the expression of phosphorylated-c-Jun, an effect abolished by SP600125. Gene-microarray analysis revealed that ozone increased the expression of over 400 genes by more than 2-fold, including interleukin-6 (IL-6), CXCL1 (keratinocyte cytokine), and CCL2 (monocyte chemoattractant protein-1). SP600125 modulated the expression of a subset of 29 ozone-induced genes; IL-6 and CCL2 expression were further increased, whereas the expression of metallothionein 1, hemopexin, and mitogen-activated 3 kinase 6 was decreased in SP600125-treated ozone-exposed mice. Changes in mRNA for IL-6, CXCL1, and CCL2 were confirmed by real-time polymerase chain reaction. Ozone also decreased the expression of over 500 genes, with the most potent effect on angiopoietin-1. SP600125 modulated the expression of 15 of these genes, and in particular, SP600125 reversed ozone-induced decrease in expression of the redox-sensitive transcription factor, hypoxia-induced factor-1α. This study highlights an important role for JNK in response to oxidative stress through modulation of specific inflammatory and redox mediators. Inhibition of JNK with small molecule kinase inhibitors may be a means of reducing ozone-induced inflammation and AHR.

Complete inhibition of allergic airway inflammation and remodelling in quadruple IL‐4/5/9/13−/− mice

Background T‐helper type 2 (Th2)‐derived cytokines such as IL‐4, IL‐5, IL‐9 and IL‐13 play an important role in the synthesis of IgE and in the promotion of allergic eosinophilic inflammation and airway wall remodelling.

Role of cathepsin S in ozone-induced airway hyperresponsiveness and inflammation

Ambient ozone has been linked to the worsening of symptoms of patients with obstructive diseases such as chronic obstructive pulmonary disease (COPD) and asthma. We investigated the role of cathepsin S on ozone-induced airway hyperresponsiveness (AHR) and inflammation, using the selective cathepsin S inhibitor, Compound A. Balb/c mice were exposed to ozone at a concentration of 3 ppm or air for 3 h, following administration by gavage of Compound A or vehicle. Bronchoalveolar lavage (BAL) was performed 3 h and 20-24 h following exposure, AHR was measured at 20-24 h only. Ozone exposure, compared to air exposure increased BAL cathepsin S levels, AHR and BAL inflammatory cells. Compound A (30 mg kg(-1) p.o.) dosing compared to vehicle dosing inhibited ozone-induced AHR (-logPC100 vehicle: -0.70+/-0.12, n=8 vs. cathepsin S inhibitor: -1.30+/-0.06, P<0.001, n=8) at 20-24 h and BAL neutrophilia at 3 h and 20-24 h (P<0.05, n=6). Ozone exposure increased levels of BAL cytokines IL-6, TNF-alpha and IFN-gamma. Compound A reduced IL-6 at 3 h and 20-24 h (P<0.05, n=5) and TNF-alpha, at 20-24 h (P<0.05, n=6). These data indicate an important role for cathepsin S in the regulation of ozone-induced AHR and neutrophil cell recruitment and suggest that cathepsin S may be a target in the treatment of oxidative stress-induced AHR and inflammation.

Role of p38 mitogen-activated protein kinase in ozone-induced airway hyperresponsiveness and inflammation

Ozone is a potent oxidant and causes airway hyperresponsiveness and neutrophilia. To determine the role of p38 mitogen-activated protein kinase (MAPK) activation, we studied the effect of a p38alpha inhibitor SD-282 (Scios Inc, Fremont, CA USA) on ozone-induced airway hyperresponsiveness and neutrophilia. Balb/c mice received SD-282 (30 or 90 mg/kg i.p) or vehicle 1 h before exposure to either ozone (3 ppm, 3 h) or air. Three hours after exposure, lungs were analysed for cytokine levels and bronchoalveolar lavage was performed. Another set of mice were dosed 6 h after exposure and 1 h before assessing airway hyperresponsiveness. SD-282 (90 mg/kg) significantly inhibited ozone-induced airway hyperresponsiveness (-LogPC(150): SD-282: -1.73+/-0.14 vs. vehicle: -0.99+/-0.15, P<0.05). Bronchoalveolar lavage neutrophil numbers were time-dependently increased in vehicle-dosed, ozone-exposed mice, greatest at 20-24 h after exposure. SD-282 (30 and 90 mg/kg) significantly inhibited ozone induced neutrophil numbers at 3 h and 20-24 h after ozone SD-282 significantly inhibited ozone-induced increases in phosphorylated p38 MAPK expression, and in cyclooxygenase-2 (COX-2), interleukin-6 (IL-6) and IL-1beta but not MIP-1alpha gene expression. We conclude that p38 MAPK is involved in ozone-induced airway hyperresponsiveness and lung neutrophilia. Inhibition of p38 MAPK with small molecule kinase inhibitors may be a means of reducing ozone-induced inflammation and airway hyperresponsiveness.

Inhibition of citric acid- and capsaicin-induced cough by novel TRPV-1 antagonist, V112220, in guinea-pig

BackgroundCough reflex can be induced by the pepper extract capsaicin and by low pH in guinea-pig airways. Transient receptor potential vanniloid-1 (TPRV-1) is expressed in the sensory and afferent nerve fibres in airways.ObjectiveWe hypothesized that a novel pyridazinylpiperazine analog TPRV-1 inhibitor can effectively reduce cough reflex stimulated by citric acid and capsaicin.MethodsGuinea pigs were injected with specific TPRV-1 inhibitor, V112220, a pyridazinylpiperazine analog of N-(4-tertiarybutylphenyl)-4-(3-chloropyridin-2-yl) tetrahydropyrazine-1(2H)-carbox-amide (BCTC) (3 mg/kg) intra-peritoneally. One hour before cough response assessment. Coughs were recorded using a recorder system that identified cough sound and accompanying expiratory flows, distinct from sneezes. Guinea-pigs exposed to citric acid (0.4 M) and to capsaicin (10-4M) aerosols, in succession separately by 2 hours.ResultsV112220 significantly inhibited the number of coughs induced by citric acid (73 ± 11%, p < 0.01) and capsaicin (70 ± 9.4%, p < 0.05) compared to vehicle control.ConclusionA novel pyridazinylpiperazine analog TPRV-1 inhibitor can inhibit the cough reflex, induced by both low pH and capsaicin, suggesting that it could be clinically beneficial in treatment of cough.

Modulation of ozone-induced airway hyperresponsiveness and inflammation by interleukin-13

The present study aimed to determine whether the T-helper cell type 2-derived cytokines, interleukin (IL)-4 and -13, can modulate the lung response to ozone exposure. IL-13-/-, IL-4/13-/- and IL-13-overexpressing transgenic (Tg) mice were exposed to ozone (3u2005ppm; 3u2005h) or air. Wild-type (Wt) Balb/c mice and transgenic-negative littermates (IL-13Wt) were used as controls for gene-deficient and IL-13Tg mice, respectively. IL-4/13-/- and IL-13-/- mice developed a lesser degree of ozone-induced airway hyperresponsiveness (AHR) while IL-13Tg mice developed a greater degree of AHR compared with ozone-exposed wild-type or IL-13Wt mice, respectively. Ozone caused a time-dependent increase of bronchoalveolar lavage (BAL) neutrophils and macrophages in wild-type mice, maximal at 20–24u2005h, which was attenuated in the IL-13-/- and IL-4/13-/- mice. In IL-13Tg mice, there was a greater increase in BAL neutrophils after ozone exposure compared with IL-13Wt mice. Using quantitative real-time PCR, ozone-induced mRNA expression for IL-6 and keratinocyte chemokine was further enhanced in IL-13-/- and IL-4/13-/- mice, and was inhibited in IL-13Tg mice. Macrophage inflammatory protein (MIP)-3α/CCL20 expression was enhanced after ozone exposure in wild-type mice, inhibited in IL-13-/- and IL-4/13-/- mice, while in IL-13Tg mice it was enhanced. A similar pattern of expression was observed with lipopolysaccharide-induced cytokine (LIX/CXCL5/ENA-78) expression. In conclusion, interleukin-13 augments ozone-induced airway hyperresponsiveness and neutrophilic inflammation, possibly through modulation of certain cytokines induced by ozone exposure.

Gene and protein expression of protease-activated receptor 2 in structural and inflammatory cells in the nasal mucosa in seasonal allergic rhinitis.

Background Protease‐activated receptor 2 (PAR 2) has been shown to be responsible for trypsin and mast cell tryptase‐induced airway inflammation. Here, the present study aimed to explore the expression of PAR 2 in the nasal mucosa of seasonal allergic rhinitis (SAR).

Bronchial hyperresponsiveness and lung inflammation induced by allergic immune response and oxidative stress: role of innate and adaptive immune responses

Abstract. Airway response to the external environment forms the basis for many airway/lung diseases such as asthma and chronic obstructive airways disease (COPD) which are characterized by chronic airflow obstruction, chronic airway inflammation and tissue remodeling. Allergic inflammation and oxidative stress contribute to airways dysfunction and direct effects of these processes have been demonstrated in human and animal models. Airway responses to allergens and to environmental pollution such as ozone represent a mixture of innate and adaptive immune responses. Through the release of inflammatory cytokines and expression of co-stimulatory molecules, and antigen presentation, innate immune cells such as dendritic cells can interact with CD4 T-cells and direct their maturation program. The oxidant, ozone, can also utilize innate immune recognition pathways through toll-like receptors and activate a range of innate immune cells including dendritic cells (DCs), macrophages, and immune responses. Crosstalk between these two responses is likely to be common occurrence as exposure to oxidants is unavoidable in urban environments with high levels of pollution and allergic inflammation is a potential source of oxidants. The interaction of ozone and allergen offers interesting perspectives in unraveling further the interactions of innate and adaptive responses that could form the basis of chronic airway diseases of chronic airflow obstruction and inflammation.