Do epigenetic events take place in the vastus lateralis of patients with mild chronic obstructive pulmonary disease?

Abstract

Muscle dysfunction is a major comorbidity in Chronic Obstructive Pulmonary Disease (COPD). Several biological mechanisms including epigenetic events regulate muscle mass and function in models of muscle atrophy. Investigations conducted so far have focused on the elucidation of biological mechanisms involved in muscle dysfunction in advanced COPD. We assessed whether the epigenetic profile may be altered in the vastus lateralis of patients with mild COPD, normal body composition, and mildly impaired muscle function and exercise capacity. In vastus lateralis (VL) of mild COPD patients with well-preserved body composition and in healthy age-matched controls, expression of DNA methylation, muscle-enriched microRNAs, histone acetyltransferases (HTAs) and deacetylases (HDACs), protein acetylation, small ubiquitin-related modifier (SUMO) ligases, and muscle structure were explored. All subjects were clinically evaluated. Compared to healthy controls, in the VL of mild COPD patients, muscle function and exercise capacity were moderately reduced, DNA methylation levels did not differ, miR-1 expression levels were increased and positively correlated with both forced expiratory volume in one second (FEV1) and quadriceps force, HDAC4 protein levels were increased, and muscle fiber types and sizes were not different. Moderate skeletal muscle dysfunction is a relevant feature in patients with mild COPD and preserved body composition. Several epigenetic events are differentially expressed in the limb muscles of these patients, probably as an attempt to counterbalance the underlying mechanisms that alter muscle function and mass. The study of patients at early stages of their disease is of interest as they are a target for timely therapeutic interventions that may slow down the course of the disease and prevent the deleterious effects of major comorbidities.

Figure 1. Global percentage of methylated DNA in the vastus lateralis of mild COPD patients and healthy controls.

Mean values and standard deviation of global percentage of methylated DNA in the vastus lateralis did not differ (n.s., non-significant) between patients and controls.

Figure 2. Levels of expression of muscle-enriched microRNAs in the vastus lateralis of mild COPD patients and healthy controls.

Mean values and standard deviation (relative expression) of miR-1 (A) expression was upregulated (**: p<0.01) in the vastus lateralis of mild COPD patients compared to healthy controls, whereas expression levels of miR-133 (B), miR-206 (C), and miR-486 (D) did not differ (n.s., non-significant) in the lower limb muscles between the study groups.

Figure 3. Levels of expression of muscle-enriched microRNAs in the vastus lateralis of mild COPD patients and healthy controls.

Mean values and standard deviation (relative expression) of miR-27a (A), miR-29b (B), and miR-181a (C) did not differ (n.s., non-significant) in the lower limb muscles between the study groups.

Figure 4. Significant correlations between expression levels of miR-1 in the muscles of the COPD patients and both FEV₁ and QMVC.

Among the COPD patients, significant positive correlations were observed between the physiological parameters FEV₁ and QMVC and expression levels of miR-1 in their vastus lateralis. Note that only 12 patients were plotted in the graphs as miR-1 expression levels could not be obtained for technical reasons in one of the patients.

Figure 5. Protein levels of total acetyl-lysine protein content and mRNA levels of the HTA p300 in muscles of both COPD patients and controls.

Mean values and standard deviation of total protein acetyl-lysine/vinculin protein loading control as measured in optical densities (OD) using arbitrary units (a.u.) (A) and HTA p300 (B), levels in the vastus lateralis did not differ (n.s., non-significant) between the two study groups.

Figure 6. Protein levels of HDACs shown to play a role in muscle dysfunction in muscles of both patients and controls.

Mean values and standard deviation of protein HDAC3/vinculin protein loading control (A) HDAC6/vinculin protein loading control as measured in optical densities (OD) using arbitrary units (a.u.) (B), and SIRT1/vinculin protein loading control (C) levels in the vastus lateralis did not differ (n.s., non-significant) between the two study groups, while HDAC4/vinculin protein loading control (D) protein levels were significantly greater (**: p<0.01) in the patients than in the controls.

Figure 7. Protein levels of transcription factors shown to be involved in myogenesis and muscle repair in muscles of both patients and controls.

Mean values and standard deviation of total MEF2C/vinculin protein loading control as measured in optical densities (OD) using arbitrary units (a.u.) (A), MEF2D/vinculin protein loading control (B), and YY1/vinculin protein loading control (C) levels in the vastus lateralis did not differ (n.s., non-significant) between patients and control subjects.

Figure 8. mRNA expression of SUMO-2 and SUMO-3 in the vastus lateralis of mild COPD patients and controls.

Mean values and standard deviation (relative expression) of SUMO-2 (A) and SUMO-3 (B) in the vastus lateralis did not differ (n.s., non-significant) between patients and control subjects.