This study assessed the cardiac safety of long-acting muscarinic receptor antagonists (LAMAs) used to treat chronic obstructive pulmonary disease (COPD) using an isolated heart model of ischemia/reperfusion injury. The study found that the LAMAs aclidinium, tiotropium, and umeclidinium administered during reperfusion increased myocardial infarct size, suggesting these drugs may exacerbate ischemia/reperfusion injury. Further studies are needed to determine the mechanisms underlying this exacerbated cardiac injury.

1. Introduction
 Chronic obstructive pulmonary disease (COPD) is the 3rd leading
cause of death worldwide (Qureshi, Sharafkhaneh and Hanania
2014). COPD is a progressive condition with irreversible airway
damage, and includes conditions such as emphysema and chronic
bronchitis (Daheshia 2005). Patients are often at a greater risk of
cardiovascular complications such as ischaemic heart disease, due
to lifestyle associated risk factors, such as smoking.
 COPD patients are prescribed bronchodilators to improve lung
function. Often, a combination of pharmacological therapies are
prescribed, including β2 adrenoceptor agonists and muscarinic
receptor antagonists to alleviate bronchoconstriction.
 Recent clinical studies have associated the use of muscarinic
receptor antagonists such as the short acting, ipratropium bromide
and the long acting, tiotropium bromide with an increased risk of
adverse cardiovascular effects, including myocardial infarction and
stroke in COPD patients (Singh, Loke and Furberg 2008; Hilleman et
al. 2009). Previous studies from our group have shown that
ipratropium exacerbates myocardial injury in pre-clinical models of
ischaemia/reperfusion injury (Harvey, Hussain and Maddock 2014).
 These clinical findings have prompted the urgent need to assess the
cardiac safety profile of long acting muscarinic receptor antagonists
(LAMAs), in experimental models of myocardial
ischaemia/reperfusion injury.
Aim
 To characterise the cardiac safety profiles of LAMAs in in-vitro
models of myocardial ischaemia/reperfusion injury.
Methods
 Briefly: Isolated perfused hearts were subjected to 35 min of
regional ischaemia followed by reperfusion for 120 minutes (figure
1). For the drug treated groups; either Glycopyrronium, Aclidinium,
Tiotropium or Umeclidinium (10μM-1nM) was administered
throughout the reperfusion period.
 At the end of the experiment, hearts were subjected to TTC staining
for infarct-to-risk ratio (%) analysis. Hearts that underwent
ischaemia were also stained with Evans blue.
Figure 1: The experimental protocols used for the Langendorff models. KH – Krebs-
Henseleit buffer.
Cardiac Safety Profiles of Long Acting Muscarinic Receptor Antagonists
Used in the Treatment of Chronic Obstructive Pulmonary Disease
Shabana Cassambai, Sadie Dean, Christopher J Mee, Afthab Hussain
Faculty Research Centre - Applied Biological and Exercise Sciences, Coventry University, CV1 5FB
Conclusion
This is the first pre-clinical study to suggest that the administration of LAMAs during reperfusion, exacerbates myocardial ischaemia/reperfusion injury in an
in-vitro isolated heart model. In order to determine the signalling mechanisms underlying this exacerbated myocardial injury; further studies examining the
activation of cell death pathways and molecular changes associated with drug administration are urgently required.
Results
Figure 2: Glycopyrronium Bromide (10μM-1nM) administration does not significantly increase
Infarct size to risk ratio. Mean ± SEM n=4, ****P<0.0001 vs. normoxia.
Figure 3 : Aclidinium Bromide administration significantly increases infarct development at
concentrations of 10μM, 10nM and 1nM. Mean ± SEM n=3, **** P<0.0001 vs. normoxia, ###
P<0.001 vs. IR, ## P<0.01 vs. IR, $ P<0.05 vs. IR+AB (10μM), € P<0.05 vs. IR+AB (1μM), ^^ P<0.01
vs. IR+AB (100nM).
Figure 4: Tiotropium Bromide administration during reperfusion results in a significant increase
in infarct development at all concentrations. Mean ± SEM n=3, **** P<0.0001 vs. normoxia, ####
P<0.0001 vs. IR, ### P<0.001 vs. IR, ## P<0.01 vs. IR.
Figure 5: Administration of Umeclidinium Bromide (100nM, 10nM and 1nM) throughout
reperfusion increases infarct size to risk ratio (%). Mean ± SEM n=4, **** P<0.0001 vs. normoxia,
#### P<0.0001 vs. IR, €€€€ P<0.0001 vs. IR+UB (1μM).
References
Daheshia, M. (2005) 'Pathogenesis of Chronic Obstructive Pulmonary Disease (COPD)'. Clinical and Applied Immunology Reviews 5 (5), 339-351
Harvey, K. L., Hussain, A., and Maddock, H. L. (2014) 'Ipratropium Bromide-Mediated Myocardial Injury in in Vitro Models of Myocardial ischaemia/reperfusion'. Toxicological Sciences : An Official Journal of the Society of Toxicology 138 (2), 457-467
Hilleman, D. E., Malesker, M. A., Morrow, L. E., and Schuller, D. (2009) 'A Systematic Review of the Cardiovascular Risk of Inhaled Anticholinergics in Patients with COPD'. International Journal of Chronic Obstructive Pulmonary Disease 4, 253-263
Qureshi, H., Sharafkhaneh, A., and Hanania, N. A. (2014) 'Chronic Obstructive Pulmonary Disease Exacerbations: Latest Evidence and Clinical Implications'. Therapeutic Advances in Chronic Disease, 2040622314532862
Singh, S., Loke, Y. K., and Furberg, C. D. (2008) 'Inhaled Anticholinergics and Risk of Major Adverse Cardiovascular Events in Patients with Chronic Obstructive Pulmonary Disease: A Systematic Review and Meta-Analysis'. JAMA - Journal of the
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