INTRODUCTION

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Bronchial asthma is an inflammatory airway disease and associated with an increase in airway blood flow (1). Most of the blood perfusion in the conducting airways is through the bronchial artery (2) and the majority of blood flow is distributed to the subepithelial tissue, the principal site of asthma-associated airway inflammation. As antiinflammatory agents, inhaled glucocorticosteroids (GS) have become the first-line therapy for the treatment of moderate to severe asthma. Even though newer preparations have been tested and approved for use in asthma therapy, it is difficult to compare the potency of inhaled GS in vivo (3). The "McKenzie test," a skin patch test in which cutaneous vasoconstriction or skin blanching is measured in healthy subjects, has been used to rank the topical antiinflammatory potency of GS in humans (4). The sensitivity of the skin-blanching test was subsequently increased by quantitating blood flow plethysmographically or by ¹³³Xe washout (5, 6). However, these tests assess the effects of GS in the skin while the therapeutic target of inhaled GS is the airway mucosa and the relevant physiological parameter for the quantitation of inhaled GS potency is airway mucosal blood flow (aw).

The newer inhaled GS are more airway selective, have low oral bioavailability, and have increased uptake for and retention in lung tissue (3). They also have greater receptor affinity. Fluticasone propionate is among the most potent inhaled GS available today. We chose fluticasone propionate to determine the short-term effect of inhaled GS on aw in subjects with and without asthma.

	METHODS

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Ten asthmatic and 10 healthy adult nonsmokers participated in the study (Table 1). They received financial remuneration for their participation. The study protocol was approved by the Mount Sinai Medical Center (Miami Beach, FL) Institutional Review Board. The patients had mild to moderate asthma, were in a stable state, and had not used inhaled or systemic GS for at least 2 wk before the study. None of the subjects was taking antibiotics, oral antiinflammatory agents, or vasoactive medications, or had cardiovascular disease. The subjects with asthma were using on demand -adrenergic agents by inhalation.

Pulmonary function was measured with an Essential Medics Unit (model 620-0 Autobox DL; Yorba Linda, CA). The highest forced expired volume in 1 s (FEV₁) of three forced vital capacity maneuvers was determined and expressed as a percentage of the predicted value (7).

Airway Mucosal Blood Flow (aw)

A previously validated soluble inert gas uptake method was used to measure aw (1, 8, 9). The subjects were seated in front of a valve system that allowed them to inhale through a mouthpiece (with the nasal passage occluded by a noseclip) room air or a gas mixture from a Teflon bag containing 10% dimethyl ether (DME), 5% helium, balance oxygen, and to exhale into a rolling seal spirometer (model 842; Ohio Instruments, Houston, TX). The subjects inhaled room air to and then exhaled 500 ml from the total lung capacity (TLC) position, and then rapidly inhaled the same volume of the gas mixture from the Teflon bag. They held their breath for a predetermined duration and then exhaled into the spirometer through a critical flow orifice to standardize expiratory flow. The maneuver was performed with two breathhold times each of 5, 10, 15, and 20 s in random order. During exhalation, the instantaneous concentrations of DME, nitrogen, and helium were measured at the airway opening with a mass spectrometer (Perkin-Elmer, Pomona, CA) along with the expired gas volume. The mass spectrometer inlet was not heated, and no corrections were made for water pressure. The resulting overestimation of DME concentration by measuring it at the airway opening was considered to be negligible (approximately 0.3%). The mass spectrometer was also used to verify the gas concentration in the Teflon bag before inhalation of the gas mixture. From the expired nitrogen concentration curve, the end of phase I was determined. The helium-corrected decrease in the DME concentration over time was obtained by least-squares fit, using the two measurements per gas for each of the four breathhold times. This was done in the expired dead space volume fraction corresponding to phase I minus the most proximal 50 ml (DSF). From the helium-corrected DME slope multiplied by the DSF (DME), the mean DME concentration in the DSF (FDME) and the solubility coefficient for DME in blood and tissue (), aw was calculated by the Fick principle (aw = DME/ · FDME). aw was normalized for DSF and expressed as µl · min¹ · ml¹.

Protocol

The subjects were asked to come to the research laboratory on the morning of the study day without having had any coffee or other caffeinated drinks. They were asked to abstain from ingesting alcoholic beverages the night before. The subjects with asthma were asked not to use their inhaled medications for 12 h before the study. On arrival, subjects performed spirometry immediately followed by the measurement of aw. On Day 1, subjects inhaled 4 puffs of placebo (CFC propellant), using a large volume spacer. The measurements of aw and FEV₁ were repeated immediately thereafter and 1 h later. The subjects then inhaled 880 µg of fluticasone propionate (4 puffs of 220 µg each), using the same spacer, and the aw measurement was repeated 30, 60, and 90 min later. FEV₁ was remeasured 60 min after drug inhalation. On Days 2-5, after baseline spirometry and aw measurements, the subjects inhaled either 88 µg (2 puffs of 44 µg each), 220 µg (2 puffs of 110 µg each), 440 µg (2 puffs of 220 µg each), or 1,760 µg (8 puffs of 220 µg each) of fluticasone propionate chosen in random order. aw and FEV₁ were remeasured 60 min after drug inhalation. The 60-min time point was chosen for the dose-response assessment because in the time course experiment with 880 µg of fluticasone propionate, the mean aw values were of the same magnitude at this time.

Statistical Analysis

The unpaired and paired variates of the Student t test were used to make statistical comparisons between groups and within groups. Analysis of variance was used to determine the time course of fluticasone propionate-induced changes and to determine the dose-related effect of fluticasone propionate 60 min postinhalation. Data were expressed as means ± SE. A p value < 0.05 was considered significant.

	RESULTS

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Baseline aw was higher in subjects with asthma than in subjects without asthma. The 5-d average value was 55.1 ± 1.0 µl · min¹ · ml in subjects with asthma and 44.2 ± 1.1 µl · min¹ · ml¹ in subjects without asthma (p < 0.001). This translated into a 20% greater aw in subjects with asthma. There was only a small, insignificant day-to-day variation in baseline aw in subjects with asthma and in unaffected subjects.

Respiratory function as assessed by FEV₁ was also different between subjects with asthma and unaffected subjects at baseline (Table 1). The values averaged for the five experiment days were 2.34 ± 0.13 L in subjects with asthma and 3.22 ± 0.12 L in subjects without asthma (p < 0.01). As for aw, baseline FEV₁ was similar on individual experiment days and not significantly different.

Placebo had no effect on aw and FEV₁ in subjects with asthma; the pre- and post placebo values were 59.5 ± 1.0 and 56.1 ± 9.5 µl · min¹ · ml¹ for aw (p = NS), and 2.34 ± 0.13 and 2.32 ± 0.27 L for FEV₁ (p = NS). In unaffected subjects, placebo increased aw from 45.3 ± 5.7 to 53.2 ± 12.1 µl · min¹ · ml¹ (p = NS) and had no effect on FEV₁, 3.22 ± 0.12 versus 3.10 ± 0.22 L (p = NS).

The time course of fluticasone propionate-induced changes in aw revealed a maximum decrease 30 min after drug inhalation in subjects with and without asthma (Figure 1). Figure 1 reflects the effects of 880 µg of fluticasone propionate, as this dose was the most potent in both subject groups. aw returned toward baseline 90 min after drug inhalation. Both in absolute and relative terms, the maximum decrease in aw was greater in subjects with asthma than in unaffected subjects. Thirty minutes after drug inhalation, the mean aw decreased by 19.6 ± 3.0 µl · min¹ · ml¹ or 37% in subjects with asthma, and by 9.2 ± 3.3 µl · min¹ · ml¹ or 21% in subjects without asthma, respectively (p < 0.01 for both groups).

View larger version (15K): [in this window] [in a new window]   Figure 1.   Effect of 880 µg of fluticasone propionate on airway mucosal blood flow in 10 subjects without asthma and 10 subjects with asthma over a 90-min observation period. Mean values ± SE. BSL = baseline.

In subjects with asthma as well as in unaffected subjects, only the highest fluticasone propionate doses of 880 and 1,760 µg caused a significant decrease in aw (Figure 2); the decrease was greater in the subjects with asthma (p < 0.01). There was a direct correlation between the magnitude of the maximum fluticasone propionate (880-µg dose)-induced decrease in aw and baseline aw (Figure 3). In other words, the vasoconstrictor response was greater in the subjects with higher baseline aw irrespective of whether they were normal or asthmatic.

View larger version (17K): [in this window] [in a new window]   Figure 2.   Dose-related effect of fluticasone propionate on airway mucosal blood flow (aw) in subjects with and without asthma 60 min after drug inhalation. Mean values ± SE. n = 10 in each group for all doses except 1,760 µg, for which n = 9 in subjects without asthma and n = 8 in subjects with asthma.

View larger version (11K): [in this window] [in a new window]   Figure 3.   Fluticasone propionate (880 µg)-induced change in airway mucosal blood flow (aw) as a function of predrug aw (BSL aw) in subjects with and without asthma.

Fluticasone propionate had no effects on FEV₁ in subjects with or without asthma at any of the doses.

	DISCUSSION

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The findings of this study demonstrate that inhalation of the GS fluticasone propionate causes a transient decrease in aw. We interpret the decrease in aw as reflecting vasoconstriction in the airway mucosa. Other explanations such as a decrease in perfusion pressure (aortic pressure) or an increase in downstream hydrostatic pressure (right and left atrial pressure) are unlikely. The nadir of mean aw occurred 30 min after drug inhalation and mean aw had returned to near baseline values by 90 min. A significant decrease in mean aw was seen only with the 880- and 1,760-µg doses, with a tendency toward a decrease at the 440-µg dose. There was an insignificant difference between the 880- and 1,760-µg values, with a trend toward a lesser degree of vasoconstriction at the 1,760-µg dose. The significance of this observation is not clear. It may signify an attenuated vasoconstrictor action of fluticasone propionate at high doses. To test this possibility, the inhalation of even higher drug doses would have been required; this was not feasible. Not all subjects were available for the 1,760-µg inhalation, and this may explain the difference between the 880- and 1,760-µg effect. The study also confirmed our previous observation that aw is increased in patients with stable asthma (1) and showed that the vasoconstrictor response to inhaled fluticasone propionate is enhanced in them when compared with unaffected control subjects. Finally, a direct relationship was observed between the fluticasone propionate-induced decrease in aw and predrug aw.

The placebo inhaler consisted of CFC and did not contain propionate. However, we do not believe that the vasoconstrictive action of fluticasone propionate was due to the propionate residue. Miyahara and coworkers (10) reported that propionate potentiated contractions of human internal mammary arteries induced by sodium removal in vitro, but the concentration required for this effect was 20 nM. The total delivered dose of fluticasone propionate at the 1,760-µg dose was 4 nM, 10-15% of which was probably deposited in the lower airway. Assuming a mucosal tissue distribution volume of 50 ml, the highest possible mucosal concentration of fluticasone propionate would have been 10 nM, or two times less than the threshold value reported by Miyahara and coworkers (10).

Since McKenzie and Stoughton observed in 1962 (4) that locally applied GS cause skin blanching, other authors have confirmed, using quantitative physiological techniques, that GS can cause vasoconstriction in the skin (5, 6). The time course of vasoconstriction induced by GS topically applied to the skin typically is characterized by an onset after several hours and duration up to 72 h, depending on the rate of GS absorption (4, 11). For intradermally injected GS, the skin blanching starts between 60 and 90 min and fades in 6-8 h (11). In our study, inhaled fluticasone propionate-induced vasoconstriction in the airway mucosa had an earlier onset (30 min) and was of shorter duration (~ 90 min). The reason for the discrepant timing of GS-induced vasoconstriction in the airway and skin is not known. Possible explanations include physicochemical differences in the diffusion distances between the site of GS application and the site of GS action in the vessel wall, and differences in the clearance of GS from the tissue. Bende and colleagues (12) examined the effect of topical GS on nasal mucosal blood flow by ¹³³Xe washout. They found no decrease in blood flow at 2 h, the first measurement after drug administration, but may have missed transient vasoconstriction as demonstrated by our study.

The exact mechanism whereby GS cause vasoconstriction has not been clarified. The rapid onset of the response suggests a nongenomic mode of action. Several pieces of evidence support the hypothesis that GS-induced vasoconstriction involves noradrenergic neurotransmission. GS have been reported to enhance vasoconstrictor responses to adrenaline in the human hand (13) and to noradrenaline in rat mesenteric arterioles (14, 15). Those observations indicate that locally applied GS do not act by releasing noradrenaline from adrenergic nerve endings but rather by potentiating its physiological effect by upregulating postsynaptic adrenergic receptors, interfering with noradrenaline metabolism, or inhibiting presynaptic or postsynaptic noradrenaline uptake. The latter possibility is supported by the demonstration that GS inhibit the uptake of noradrenaline by cultured vascular smooth muscle cells, an effect seen within 20 min (16). Further investigations are needed to establish the precise mode of GS action in vasoconstriction.

The vasoconstrictor response to inhaled fluticasone propionate was greater in our subjects with asthma than in those without asthma. This could have been related simply to the lower preexisting vasomotor tone due to inflammatory vasodilation or to an upregulation of vasomotor responsiveness in the airway. The direct correlation between the magnitude of fluticasone propionate (880 µg)-induced decrease in aw and baseline aw is consistent with either of the two suggested mechanisms.

In conclusion, the demonstration that inhaled fluticasone propionate, a topical GS, causes transient, short-term vasoconstriction in the airway mucosa has pharmacodynamic and therapeutic implications. Determining quantitatively and noninvasively the vasoconstrictor response to inhaled GS in the airway could provide a more relevant assessment of the potency of topical GS designed for airway therapy than the McKenzie test. Although there is no direct proof that the vasoconstrictive and antiinflammatory activities of GS are closely correlated, it has been shown that GS that produce the most powerful skin blanching are also the most effective topical antiinflammatory agents (17). With respect to therapy, our study suggests that by causing vasoconstriction and possibly mucosal decongestion, inhaled GS may have an immediate beneficial effect in asthma. Furthermore, GS-induced vasoconstriction could enhance the action of inhaled bronchodilators by diminishing their clearance from the airway (20). Since the GS-induced vasoconstriction peaks between 30 and 60 min after drug inhalation, adjusting the dosing interval between inhaled GS and bronchodilators accordingly may be of clinical significance. Thus, the transient vasoactive effect of inhaled GS may have therapeutic benefits that are not related to their antiinflammatory activity.