Introduction

Reducing the negative environmental impact associated with the use of pressurized metered dose inhalers (pMDI) remains a priority for many stakeholders.1,2 The methacholine challenge test (MCT) is widely used in respiratory medicine, both as a diagnostic aid and as a research tool. In those who exhibit methacholine induced bronchoconstriction (MIB), it is common practice to reverse MIB by administering a bronchodilator, usually 200 mcg salbutamol, via spacer and pMDI.

The annual number of positive MCT in a small urban clinical pulmonary function lab is estimated at 30% of all MCT performed annually and retrospectively calculated to be 32% per 1000 tests performed.3 While this is a relative number, one need only reflect on the widespread clinical use of methacholine challenge testing4 to appreciate the environmental impact of pMDI reversal of MIB on a global scale. Add to that the use of MCT and salbutamol administration in the research setting5,6 and one realizes there is a need to consider alternative methods of reversing MIB. One possibility would be to use the Aerogen® Solo vibrating mesh nebulizer (VMN). This nebulizer is used for performing the volumetric method of MCT7 and as such it may be possible to extend the use of the nebulizer to include bronchodilator delivery after testing is complete thereby decreasing the negative environmental impact of pMDI propellant. The current investigation compared reversal of MIB with 200 mcg salbutamol delivered via pMDI or VMN as a first step in determining the feasibility of using the VMN for this purpose.

Methods Study Design

We performed a double blind, double dummy, placebo controlled, randomized, three-way crossover study. The research was reviewed and approved by the University of Saskatchewan Biomedical Research Ethics Board (BIO 3632) and registered with clinicaltrials.gov (#NCT05977699). Study visits were conducted between January 15, 2024 and November 25, 2024. Participants attended the lab on three occasions separated by a minimum of one week. Participants were fully informed about the purpose of the trial, in accordance with the Declaration of Helsinki. Consent from each participant was obtained at the first visit prior to the performance of any study procedures. Visits began with measurement of baseline lung function followed by heart rate (HR) measurement via commercial pulse oximeter and methacholine challenge testing (MCT). Immediately after MCT, HR was again measured, and one of three treatment regimens was administered; materials were purchased locally or supplied by the manufacturer:

Salbutamol 200 mcg via pressurized metered dose inhaler + LiteAire® MDI Holding chamber (“pMDI”) and placebo (0.5 mL saline) via VMN + Aerogen® Ultra aerosol reservoir with mouthpiece (“VMN”) Salbutamol 200 mcg (0.5 mL drawn from 1 mg/mL solution) via VMN and placebo via pMDI Placebo via pMDI and placebo via VMN (“placebo”)

Measurements of forced expiratory volume during the first second of exhalation (FEV1) and HR were performed at baseline and at 5, 10, 15, 30, 45 and 60 minutes following treatment administration.

Treatment Blinding

Double-blind, double dummy kits were prepared by one of the study investigators not involved in the collection of data. Twenty kits were prepared, labelled 1 through 20 and assigned sequentially to participants. Each kit contained blinded treatments labelled 1, 2 and 3 and treatments were administered in that order. Each treatment contained a pMDI (placebo or salbutamol) and approximately 0.6 mL salbutamol solution or normal saline in a 1 mL syringe of which exactly 0.5 mL was loaded into the VMN. pMDI treatments were delivered as 2 puffs (200 mcg salbutamol or 2 puffs matching placebo) via the Lite-Aire MDI Holding Chamber. VMN treatments consisted of nebulizing the 0.5 mL of salbutamol (ie. 200 mcg) or saline to completion. Half the sample was randomized to receive the pMDI first and half to receive the VMN first.

Participants

Individuals at least 18 years of age, in general good health, with baseline FEV1 ≥65% predicted and methacholine PD20 ≤800 mcg were eligible to participate in the study. Individuals that experienced a respiratory infection or allergen exposure (as applicable) within 4 weeks of Visit 1 were excluded. Current smokers as well as pregnant or breast-feeding females were also excluded. Stable doses of inhaled corticosteroid and leukotriene receptor antagonists were allowed. Beta2-agonist use was allowed but required an appropriate washout (6 hours) prior to each visit. Muscarinic antagonist use was not allowed. Use of other asthma and non-asthma medications known or suspected to influence airway hyperresponsiveness (AHR) to methacholine, including over-the-counter formulations, was assessed on a case-by-case basis.

Spirometry

Spirometry was performed according to ATS/ERS standards8 and used to determine baseline lung function (study eligibility at visit 1), baseline lung function stability (visits 2 and 3), response to methacholine (all visits) and salbutamol reversal of methacholine induced bronchoconstriction (all visits). Measurements included full flow volume loops at baseline (ie. FEV1 and FVC) but only FEV1 maneuvers during and immediately after MCT.

Methacholine Challenge

MCT was performed using the volumetric method7 and guided by recent recommendations for methacholine bronchoprovocation.9 Following baseline spirometry, participants inhaled to completion, 0.5 mL of normal saline via the VMN; nose clips were worn and a filter was placed on the expiratory port of the T-piece to prevent contamination of ambient air. FEV1 was measured at 30- and 90-seconds post-inhalation. At 5-minute intervals (start of one inhalation to the start of the next inhalation) 0.5 mL of doubling doses of methacholine (3 mcg–800 mcg) were administered until the lowest post methacholine FEV1 relative to the lowest post saline FEV1 had declined at least 17%.10 The methacholine PD20 (dose of methacholine leading to a decline in FEV1 of at least 20%) was interpolated11 or extrapolated12 as necessary.

Statistical Analysis

Comparison of treatment effect on FEV1 and HR at each post treatment timepoint were made using repeated measures analysis of variance (RM ANOVA). Mean data (±SD) are presented. Pairwise multiple comparisons were performed using a Bonferroni correction where RM ANOVA differences between treatments were significant at the 5% level.

Results Participants

Twenty-eight individuals were screened for the study. Eleven had a negative (>800 mcg) MCT at Visit 1 and therefore ineligible to continue in the study. One individual completed the first two study visits but was unable to complete the third study visit in a timely fashion and was subsequently withdrawn from the study. Sixteen individuals completed the study, there were no safety concerns or adverse events. Baseline characteristics of individuals who completed the study are detailed in Table 1.

Table 1 Participant Baseline Characteristics

Reversal of MIB – Effect of Treatment on FEV1 (Figure 1)

Mean FEV1(L) pre and post MCT did not differ between the three treatments when corrected for repeated measurements; the magnitude of MIB (decline in post MCT FEV1) was similar across groups (30.5%, 27.4% and 24.8% for placebo, pMDI and VMN, respectively). Salbutamol administration via pMDI and VMN resulted in significantly higher FEV1 at all timepoints post administration compared to placebo with no significant difference between the two methods of salbutamol delivery.

Figure 1 Mean FEV1 data before, immediately after and at pre-specified timepoints post salbutamol administration. No significant differences were found pre and post methacholine challenge testing. Administration of salbutamol by either method significantly increased FEV1 compared to placebo but not to each other at all timepoints following administration.

Treatment Effect on Heartrate (Table 2)

Prior to Bonferroni correction, RM ANOVA showed a statistically significant difference in HR between treatment groups at 45 minutes post treatment administration with pMDI being significantly lower than VMN (p = 0.04). After Bonferroni correction for repeated measures, the difference was no longer statistically significant.

Table 2 Mean HR in Beats per Minute (SD) for the Three Treatment Groups Before and After Treatment Administration

Discussion

Data generated from the current study show recovery of lung function from MIB occurs to a similar extent following inhalation of 200 mcg salbutamol delivered via a spacer device and either pMDI or VMN with both methods resulting in faster recovery compared to placebo.

On average, five minutes after salbutamol administration, FEV1 values were 96.3% and 94.5% of baseline with VMN and pMDI, respectively. Slight further numerical increases at 10 and 15 minutes were observed with little improvement thereafter for both VMN and pMDI. This level of recovery is consistent with that generally required to allow an individual to leave the testing facility following MCT; that is, FEV1 should be within 10% of baseline. By comparison, mean FEV1 recovery at five minutes following placebo was 81.0% of baseline and did not return to within 10% of baseline until 45 minutes post treatment. These data are consistent with the common practice of measuring lung function 10 minutes after bronchodilator administration but also suggest recovery may be sufficient as early as 5 minutes following treatment. Allowing untreated recovery as was observed after placebo administration would take too long and this would neither be practical nor ethical in a clinical or research setting, particularly if significant bronchoconstriction had been induced. While recovery will be influenced by individual characteristics and by the extent of bronchoconstriction induced, the current data suggest that a formal protocol for reversing MIB could be standardized to begin re-assessing lung function at 5 minutes post bronchodilator administration with subsequent measurements made at 5-minute intervals until FEV1 has recovered to within 10% of baseline.

Previous studies using methacholine challenge testing to compare bronchodilator efficacy between beta2-agonist formulations (eg. short- versus long-acting13) and between similar devices (eg. pMDI versus DPI14) have shown similar results. In general, these studies document the quick onset of bronchodilator efficacy with little difference between formulations and devices. However, there are no published data comparing the efficacy of a standard dose of salbutamol delivered via a VMN versus pMDI for reversing MIB.

The significance of these data relates to the need for healthcare practitioners and researchers to be responsible stewards regarding the use of products that negatively affect the environment, in this case, the use of pMDI inhalers to reverse MIB. Much attention has been given to making environmentally friendly choices surrounding inhaler use as treatment15 but little if any has been given to assessing or decreasing use for clinical diagnostic and research purposes. Given the widespread use of MCT, both clinically and in research, efforts to minimize the use of pMDIs for reversal of MIB should be sought. Administering bronchodilator via DPI is an option but DPIs, although not greenhouse gas emitters, also negatively impact the environment albeit to a lesser extent than pMDI inhalers.16 Moreover, the expense and wastefulness of using a new DPI inhaler for every MCT would be significant and impractical. Another option would be to use a jet nebulizer, perhaps even the same jet nebulizer as was used to deliver the methacholine. However, jet nebulizers require a source of driving pressure (ie. plastic tubing to connect to wall oxygen, a tank of compressed gas or a compressor) and jet nebulizers are much larger than the VMN used in the current investigation both of which may translate to greater plastic waste. In addition, jet nebulizers require a lengthy duration of nebulization to deliver a dose of medication. One approach is to run the nebulizer for a fixed duration of 10-minutes which is impractical for a clinical pulmonary function lab. Moreover, there is uncertainty in the dose delivered given the significant evaporative loss that occurs with jet nebulizers.17 By comparison, nebulization of 0.5 mL of methacholine solution via the VMN ranges from 91 to 166 seconds.7 Although the duration of nebulization of 0.5 mL salbutamol was not formally measured in the current study, in reflecting on the process, the delivery time was similar and this timeframe is little more than that required to administer 200 mcg salbutamol via pMDI.

A second application of using the VMN for delivering salbutamol to reverse MIB relates to the volumetric method of performing the MCT.7 This method uses the VMN without the Ultra reservoir to deliver methacholine and if the same nebulizer was used to deliver bronchodilator as was used to deliver methacholine, the use of the nebulizer in the overall performance of the MCT would be extended. This approach would not only decrease pMDI use but also improve cost-effectiveness. While the VMN is expensive relative to the cost of jet nebulizers currently in use for MCT, there are several additional benefits, including significant cost-savings in the amount of methacholine required to perform the test. The top concentration required using a jet nebulizer can be as much as 16 mg/mL whereas a top concentration of 2 mg/mL is required with the VMN. Moreover, a volume of 2 or 3 mL of each methacholine dose is required if using a jet nebulizer whereas this same volume would be sufficient to perform 4 to 6 tests with the VMN.18

Inhalation of beta2-agonists have the potential to induce cardiovascular events, including a dose-dependent increase in heartrate.19 The VMN is an efficient nebulizer with superior deposition compared to jet nebulizers20 suggesting inhaling salbutamol via the VMN might increase heartrate. On average, heartrate measurements were similar or numerically decreased after treatment administration with no significant difference between treatments. Individually, there were seven instances where heartrate increased 12 or more beats per minute after treatment (three after placebo, three after A-VMN and one after pMDI). Five of the seven occurred in two individuals and no signal or trend for increased heartrate after salbutamol administration via the VMN or pMDI was observed.

The current data open the door to another potential application of using the VMN and that is to deliver salbutamol for bronchodilator responsiveness testing (BDRT). The global environmental impact of pMDI use for BDRT is undoubtedly much greater than that of MCT for at least three reasons. First, the recommended dose of salbutamol is 400 mcg, double that used to reverse MIB. Second, in most cases, BDRT will precede MCT in the clinical diagnosis of asthma. Third, BDRT is routinely performed in the clinical workup of other respiratory conditions (eg. chronic obstructive lung disease).21 Additional research is required to explore this potential application.

A limitation of the current study was the use of separate VMN nebulizers to administer the methacholine and salbutamol but it was important to assess delivery of 200 mcg salbutamol via pMDI versus the VMN on reversing MIB without the potential of residual methacholine solution confounding the response. The residual volume remaining following nebulization of a solution with the VMN is small, reportedly less than 0.1 mL for a 3 mL volume.22 The small amount of methacholine solution that might remain after nebulization of a 0.5 mL starting volume would not be expected to significantly suppress the bronchodilating effect of salbutamol although this also requires further investigation.

One concern regarding the nebulization of medication is the potential for secondary exposure to those individuals sharing the ambient environment (eg. research staff, healthcare providers or caregivers). A recent study using 25 minutes of breath simulation estimated secondary exposure to be about 1% of a 2500 mcg nominal dose of salbutamol at a distance of about three feet when nebulization was performed with a jet nebulizer and a facemask.23 The least secondary exposure was observed when aerosol was delivered with the VMN via mouthpiece and a filter placed on the expiratory port. We did not use a filter on the expiratory port but given the small dose of salbutamol and the short nebulization time required to deliver the dose, we believe the risk of secondary exposure to be quite low.

Inhaler therapy is mainstay for treating respiratory illness but inhaler use, particularly pMDI inhaler use, contributes to global warming. Drug manufacturers are investigating alternative, more environmentally friendly propellants but until these inhalers become available, any effort to reduce the carbon footprint associated with the use of inhalers should be a priority. Based on the preliminary data generated from the current study, nebulization of bronchodilator using the VMN appears to be a safe and equally effective method to that of pMDI for reversing MIB with potential benefit for use in BDRT. Given the global, widespread use of MCT and BDRT in diagnosing and researching respiratory illness, the impact on the environment of finding an alternative to pMDI use for these purposes should be further explored.

Data Sharing Statement

De-identified raw data pertaining to the study endpoints may be made available upon request to the corresponding author.

Acknowledgments

We would like to acknowledge and thank all study participants for their valuable contribution to research.

Author Contributions

All authors made a significant contribution to the work reported, whether that is in the conception, study design, execution, acquisition of data, analysis and interpretation, or in all these areas; took part in drafting, revising or critically reviewing the article; gave final approval of the version to be published; have agreed on the journal to which the article has been submitted; and agree to be accountable for all aspects of the work.

Funding

The conduct of the research received both financial and in-kind support from Aerogen Ltd. No personal financial gain was realized by the authors.

Disclosure

Dr Brianne Philipenko reports personal fees from GSK, AstraZeneca, Sanofi, and Covis Pharma, outside the submitted work. The authors report no other conflicts of interest in this work.

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