Distinct skeletal muscle molecular responses to pulmonary rehabilitation in chronic obstructive pulmonary disease: a cluster analysis

Abstract

Background: Pulmonary rehabilitation (PR) is a cornerstone in the management of chronic obstructive pulmonary disease (COPD), targeting skeletal muscle to improve functional performance. However, there is substantial inter-individual variability in the effect of PR on functional performance, which cannot be fully accounted for by generic phenotypic factors. We performed an unbiased integrative analysis of the skeletal muscle molecular responses to PR in COPD patients and comprehensively characterized their baseline pulmonary and physical function, body composition, blood profile, comorbidities, and medication use.

Methods: Musculus vastus lateralis biopsies were obtained from 51 COPD patients (age 64 ± 1 years, sex 73% men, FEV₁ , 34 (26-41) %pred.) before and after 4 weeks high-intensity supervised in-patient PR. Muscle molecular markers were grouped by network-constrained clustering, and their relative changes in expression values-assessed by qPCR and western blot-were reduced to process scores by principal component analysis. Patients were subsequently clustered based on these process scores. Pre-PR and post-PR functional performance was assessed by incremental cycle ergometry and 6 min walking test (6MWT).

Results: Eight molecular processes were discerned by network-constrained hierarchical clustering of the skeletal muscle molecular rehabilitation responses. Based on the resulting process scores, four clusters of patients were identified by hierarchical cluster analysis. Two major patient clusters differed in PR-induced autophagy (P < 0.001), myogenesis (P = 0.014), glucocorticoid signalling (P < 0.001), and oxidative metabolism regulation (P < 0.001), with Cluster 1 (C1; n = 29) overall displaying a more pronounced change in marker expression than Cluster 2 (C2; n = 16). General baseline characteristics did not differ between clusters. Following PR, both 6 min walking distance (+26.5 ± 8.3 m, P = 0.003) and peak load on the cycle ergometer test (+9.7 ± 1.9 W, P < 0.001) were improved. However, the functional improvement was more pronounced in C1, as a higher percentage of patients exceeded the minimal clinically important difference in peak workload (61 vs. 21%, P = 0.022) and both peak workload and 6 min walking test (52 vs. 8%, P = 0.008) upon PR.

Conclusions: We identified patient groups with distinct skeletal muscle molecular responses to rehabilitation, associated with differences in functional improvements upon PR.

Keywords: Chronic obstructive pulmonary disease; Cluster analysis; Exercise training; Muscle plasticity; Peripheral muscle dysfunction.

Figure 1

Molecular network and network‐constrained hierarchical clustering of molecular rehabilitation responses. (A) Network‐constrained clusters revealing eight distinct processes (P), as indicated with different colours. (B) Molecular rehabilitation responses. The literature‐based molecular network is indicated as lines between molecular markers. Circles represent mRNA markers; pentagons represent protein markers. Numbers correspond to individual markers as depicted in Supporting Information, Table S1.

Figure 2

Hierarchical clustering of patients. (A) Silhouette scores for n clusters (mean ± SEM). Clustering based on raw molecular rehabilitation responses (marker values; i.e. no data reduction) is indicated in grey; clustering based on eight process scores is indicated in black. (B) Patients individual silhouette coefficient values per cluster. (c) Dendrogram and clustered heatmap of individual marker rehabilitation responses (Z‐scores) and process scores.

Figure 3

Associations between molecular rehabilitation responses. Each square displays a scatter plot and regression line of the processes indicated on the x and y axis, for both Cluster 1 and Cluster 2. Differences between correlations were tested by a Fisher's r‐to‐z transformation and indicated with a red outline (P < 0.05) or orange outline (P < 0.1). Pearson correlation coefficients per cluster are depicted for differential correlations and when P < 0.1. ^# P < 0.1, ^* P < 0.05, ^** P < 0.01, ^*** P < 0.001.

Figure 4

Functional rehabilitation responses per cluster. Individual rehabilitation‐induced changes. (A) Distance (metres) walked in 6 min walk test (6MWT), n = 28/15. (B) Peak load (W) on a cycle ergometer test, n = 23/14. Data expressed as mean ± SD. (C) Percentage of patients with a change in 6MWT, peak load, or both, exceeding the minimal clinically important difference (MCID; 6MWT: 25 m, peak load: 10 W). ^* P < 0.05, ^** P < 0.01, ^*** P < 0.001, indicating significance of within‐group rehabilitation responses or significance of differences between indicated groups.