Possible role of cyclic AMP phosphodiesterases in the actions of ibudilast on eosinophil thromboxane generation and airways smooth muscle tone.

1. The possible role of cyclic AMP phosphodiesterase (PDE) in the inhibitory actions of ibudilast on tracheal smooth muscle contractility and eosinophil thromboxane generation was investigated. 2. Ibudilast was a non-selective inhibitor of partially purified cyclic nucleotide PDE isoenzymes from pig aorta and bovine tracheal smooth muscle, exhibiting only moderate potency against bovine tracheal PDE IV (IC50 = 12 +/- 4 microM, n = 3). Similar or slightly lower potencies were displayed against PDEs I, II, III and V. In contrast, rolipram exhibited selectivity for PDE IV (3 +/- 0.5 microM, n = 3). 3. Ibudilast (IC50 = 0.87 +/- 0.37 microM, n = 3), like rolipram (IC50 = 0.20 +/- 0.04 microM, n = 3), was a more potent inhibitor of membrane-bound PDE IV from guinea-pig eosinophils than of partially purified PDE IV from bovine tracheal smooth muscle. The potency of ibudilast increased when the eosinophil enzyme was solubilised with deoxycholate and NaCl (IC50 = 0.11 +/- 0.05 microM, n = 3) or exposed to vanadate/glutathione complex (V/GSH) (IC50 = 0.11 +/- 0.02 microM, n = 3). The potency of rolipram was also increased by solubilization (IC50 = 0.012 +/- 0.003, n = 3) or V/GSH (IC50 = 0.012 +/- 0.003, n = 3). 4. In intact eosinophils, ibudilast (0.032 microM-20 microM) potentiated isoprenaline-induced cyclic AMP accumulation in a concentration-dependent manner, being approximately 20 fold less potent than rolipram. Little or no effect on basal cyclic AMP levels was observed with either compound. The cyclicAMP-dependent protein kinase activity ratio was significantly increased following incubation of eosinophils with either ibudilast (20 MicroM) or rolipram (20 MicroM) in the absence or presence of isoprenaline.5. Leukotriene B4 (300 nM)-induced thromboxane generation from guinea-pig eosinophils was inhibited by ibudilast (IC50 = 11.3 +/- 3.7 MicroM, n = 5) and rolipram (IC50 = 0.280 +/- 0.067 MicroM, n = 5) in a concentration-dependent manner.6. Ibudilast (10 nM-1 MicroM), whilst generally less potent than rolipram (1 nM- 1 MicroM), produced concentration-dependent relaxation of spasmogen (methacholine, histamine, LTD4)-induced tone in the guinea pig isolated tracheal strip. Ibudilast was less potent in reversing the methacholine (IC50 = 1.95 +/- 0.40 JM,n =6)-induced contraction than those of histamine (IC50 = 0.18 +/- 0.70 MicroM, n =6) or leukotriene D4(LTD4, IC50 = 0.12 +/- 0.05 MicroM, n = 6). Rolipram also exhibited a similar pattern of activity, although the difference in potency against methacholine (IC50 = 0.1 +/- 0.01 MicroM, n = 6) compared with the other two spasmogens, histamine (IC50 = 0.034 +/- 0.017 MicroM, n = 7) and LTD4 (IC50 = 0.026 +/- 0.008 MicroM, n = 7), was not as great.7. These results demonstrate that ibudilast, like rolipram, has several biological actions on the eosinophil and airways smooth muscle which may be attributed to inhibition of cyclic AMP PDE. These actions may account, at least in part, for the recently reported anti-asthma effects of ibudilast.

Techniques for drug delivery to the airways, and the assessment of lung function in animal models.

A common approach to understanding the mechanisms underlying clinical asthma and in new drug development is to mimic the disease in animal models. When developing animal models of pulmonary diseases, such as asthma, the experimentally induced disease may be characterized in terms of pathophysiological changes induced (e.g., inflammation, smooth muscle contraction) or by the indices of lung function that are effected by such changes. Similarly, the effects of drugs can be assessed in terms of the reversal of disease- or mediator-induced changes in lung function. Small animals, such as the guinea pig and rat, are commonly used for the assessment of lung function in models of pulmonary diseases, such as asthma, and to evaluate the effects of drugs. A variety of techniques, differing in their level of sophistication, has been developed to measure parameters of lung function in small laboratory animals. Simple techniques involve the visual assessment of the response of a conscious animal to bronchoconstriction induced by an inhaled spasmogen or antigen. This technique is rapid but gives results that are difficult to interpret in physiological terms. Bronchospasm can be better assessed in anesthetized, mechanically ventilated animals by recording bronchial tone as changes in either 1) ventilation circuit pressure or 2) air overflow as the lungs are inflated. These techniques are widely used but because they require surgical intervention they are not suited to long-term or repeat studies. In addition, they give only a limited indication of the physiological changes that affect airway caliber. To improve the models available, researchers have subsequently developed techniques that use the same physiological principles as some of the tests applied to the assessment of lung function in humans. These techniques allow the measurement of parameters of respiratory mechanics, such as lung compliance and airway resistance, that determine the relationship between pulmonary pressure changes and air flow into and out of the lungs. Continued development has resulted in models that use nonsurgical plethysmographic techniques. These allow the long-term or repeated measurement of lung function in conscious animals under minimal restraint. In the treatment of asthma, inhalation is the preferred route of administration of a drug as it allows rapid drug delivery to the site of action. Systemic effects are reduced, and the therapeutic dose is minimized. Drugs are generally inhaled as either nebulized liquids or dry-powder formulations. Because drug inhalation requires patient cooperation, techniques have been modified to allow drug delivery to the airways of experimental animals.(ABSTRACT TRUNCATED AT 400 WORDS)