Airway function, oedema, cell infiltration and nitric oxide generation in conscious ozone-exposed guinea-pigs: effects of dexamethasone and rolipram

Abstract

1. The effects of ozone inhalation (90 min, 2.15+/-0.05 p.p.m.) and their modification by dexamethasone (20 mg kg(-1)) or the phosphodiesterase-4 inhibitor, rolipram (1 mg kg(-1)), administered (i.p.) 24 and 0.5 h before and 24 h after ozone exposure were examined in conscious guinea-pigs. 2. Ozone caused an early-phase bronchoconstriction (EPB) as a fall in specific airways conductance (sG(aw)) measured by whole body plethysmography, followed at 5 h by a late-phase bronchoconstriction (LPB) and increased respiratory rate. Rolipram did not alter this profile but dexamethasone inhibited the EPB. 3. Airway hyperreactivity to inhaled histamine (1 mM, 20 s) occurred at 0.5, 2, 12, 24 and 48 h after ozone inhalation, the 2 h change being abolished by rolipram and dexamethasone. 4. Bronchoalveolar lavage fluid (BALF) macrophages, eosinophils and neutrophils were significantly (P<0.05) elevated at 12, 24 and 48 h after ozone exposure, the 48 h influx being significantly attenuated (P<0.05) by rolipram and dexamethasone. 5. BALF nitric oxide (NO) metabolites decreased 0.5 h after ozone exposure by 52%, recovered at 2 h and significantly increased at 12 (101%) and 24 h (127%). The elevated NO was unaffected by rolipram or dexamethasone. 6. Lung oedema, measured from wet/dry weight differences, was significant 12, 24 and 48 h after ozone exposure, the latter being significantly attenuated (P<0.05) by rolipram and dexamethasone. 7. Ozone exposure of guinea-pigs produced features common to COPD. Although rolipram and dexamethasone did not affect the airway function changes, they inhibited the inflammation, airway hyperreactivity and oedema.

Figure 1

The effect of exposure (90 min) to medi-air (0.35 l min⁻¹, 18.8 – 25% oxygen in nitrogen), or medi-air via an ozone generator (2.15±0.05 p.p.m.) with and without dexamethasone (20 mg kg⁻¹) or rolipram (1 mg kg⁻¹) treatment on lung function, in conscious guinea-pigs. Treatment was administered 24 and 0.5 h before exposure to ozone and 24 h afterwards. Each point represents the mean±s.e.mean (n=6) change in sG_aw expressed as a percentage of the baseline sG_aw values (sG_aw (sec⁻¹ cmH₂O⁻¹): medi-air=0.29±0.01; ozone=0.34±0.01). Negative values represent bronchoconstriction. The EPB is depicted from 0 – 1 h (white bar) and the LPB is shown from 5 – 50 h (black bar) after exposure to ozone. Significance of differences in changes of sG_aw after ozone exposure: from baseline values (black and white bars: P<0.05); or between medi-air exposure (^*P<0.05, ^**P<0.01 and ^***P<0.001); or with and without dexamethasone treatment (⁺P<0.05, ⁺⁺P<0.01 and ⁺⁺⁺P<0.001), were determined by analysis of variance (single factor), followed by a Student's t-test (paired, unpaired and unpaired, respectively). There was no significant (P>0.05) difference between the lung function in ozone exposed guinea-pigs with and without vehicle treatment (data not shown).

Figure 2

The effect of exposure (90 min) to medi-air (0.35 l min⁻¹), or medi-air via an ozone generator (2.15±0.05 p.p.m.) with and without dexamethasone (20 mg kg⁻¹) or rolipram (1 mg kg⁻¹) treatment on respiratory rate, in conscious guinea-pigs. Treatment was administered 24 and 0.5 h before exposure to ozone and 24 h afterwards. Each point represents the mean±s.e.mean (n=6) change in respiratory rate (breaths min⁻¹). Black bar denotes the significance of differences (P<0.05) between respiratory rate before and after exposure to ozone as determined by analysis of variance (single factor), followed by Student's unpaired t-test. There was no significant (P>0.05) difference between the respiratory rate in ozone exposed guinea-pigs with and without rolipram, dexamethasone or vehicle (data not shown) treatment.

Figure 3

The airways responsiveness of a nose-only exposure (20 s) to histamine (1 mM) before and (a) 0.5 h after exposure (90 min) to ozone (2.15±0.05 p.p.m.); (b) 0.5, 2, 12, 24, 48 and 72 h after exposure (90 min) to ozone, 2 h after exposure (90 min) to medi-air, and 2 h after ozone with vehicle, dexamethasone (dex, 20 mg kg⁻¹), or rolipram (1 mg kg⁻¹) treatment, in conscious guinea-pigs. Treatment was administered (i.p.) 24 and 0.5 h before exposure to ozone. Each point (n=6) represents the (a) mean±s.e.mean change in sG_aw expressed as a percentage of the baseline sG_aw values (sG_aw (sec⁻¹ cmH₂O⁻¹) before=0.37±0.02 and after=0.40±0.02); (b) mean±s.e.mean of individual peak change in sG_aw from baseline sG_aw values, 0 – 10 min following inhalation of histamine expressed as a percentage of the baseline, 24 h before and: 2 h after medi-air (before=0.27±0.01 and after=0.28±0.01); 0.5 h (before=0.37±0.02 and after=0.40±0.02), 2 h (before=0.31±0.01 and after=0.33±0.02), 12 h (before=0.33±0.02 and after=0.31±0.03), 24 h (before=0.34±0.03 and after=0.35±0.02), 48 h (before=0.32±0.01 and after=0.28±0.01), 72 h (before=0.31±0.01 and after=0.30±0.01) after ozone, and 2 h after ozone and vehicle (before=0.33±0.01 and after=0.32±0.01), dex (before=0.34±0.02 and after=0.33±0.03), or rolipram (before=0.33±0.02 and after=0.27±0.02) treatment. Negative values represent bronchoconstriction. Differences between before and after exposure to histamine were determined by analysis of variance (single factor), followed by a Student's (paired) t-test and denoted as ^*P<0.05 and ^**P<0.01.

Figure 4

Differential cell (macrophage, eosinophil and neutrophil) counts of bronchoalveolar lavage fluid (BALF) removed before (naïve) and after exposure (90 min) to medi-air (48 h), ozone (2.15±0.05 p.p.m.) (0.5, 2, 12, 24, 48 and 72 h), or ozone with vehicle, dexamethasone (20 mg kg⁻¹), or rolipram (1 mg kg⁻¹) treatment (48 h). Treatment was administered (i.p.) at 24 and 0.5 h before exposure to ozone and at 24 h afterwards. Each point represents the mean±s.e.mean (n=6) of the differential cells per BALF sample (×10⁶) removed from the left lung lobes, via the left bronchi. The significance of differences in airway cell influx compared with naïve guinea-pigs (^*P<0.05, ^**P<0.01 and ^***P<0.001), and ozone exposure and vehicle vs dexamethasone or rolipram treatment (++P<0.01 and +++P<0.001), as determined by analysis of variance (single factor), followed by Scheffe's post hoc analysis.

Figure 5

Airways levels of combined nitric oxide (NO) metabolites (nitrate and nitrite) recovered before (naïve) and after exposure (90 min) to medi-air (48 h), ozone (2.15±0.05 p.p.m.) (0.5, 2, 12, 24, 48 and 72 h), or ozone with vehicle, dexamethasone (20 mg kg⁻¹), or rolipram (1 mg kg⁻¹) treatment (48 h). Treatment was administered (i.p.) at 24 and 0.5 h before exposure to ozone and at 24 h afterwards. Each point represents the mean±s.e.mean (n=6) of the concentration of combined NO metabolites (μM) per 100 μl of BALF removed from the left lung lobes, via the left bronchi. ^*P<0.05, and ^**P<0.01 denotes the significance of differences in airway NO levels compared with naïve guinea-pigs, as determined by analysis of variance (single factor), followed by Scheffe's post hoc analysis.

Figure 6

Difference in wet and oven-dried lung weights before (naïve) and after exposure (90 min) to medi-air (48 h), ozone (2.15±0.05 p.p.m.) (0.5, 2, 12, 24, 48 and 72 h), or ozone with vehicle, dexamethasone (20 mg kg⁻¹), or rolipram (1 mg kg⁻¹) treatment (48 h). Treatment was administered (i.p.) at 24 and 0.5 h before exposure to ozone and at 24 h afterwards. Each point represents the mean±s.e.mean (n=6) of the difference in right lung lobe weights (grams), removed below the right bronchi, before (wet) and after oven drying. The significance of differences in wet/dry lung weights compared with naïve guinea-pigs (^*P<0.05 and ^***P<0.001), and ozone exposure and vehicle versus dexamethasone or rolipram treatment (++P<0.01), was determined by analysis of variance (single factor), followed by Scheffe's post hoc analysis.