Elsevier

Free Radical Biology and Medicine

Volume 79, February 2015, Pages 91-108
Free Radical Biology and Medicine

Original Contribution
Oxidative stress, redox signaling pathways, and autophagy in cachectic muscles of male patients with advanced COPD and lung cancer

https://doi.org/10.1016/j.freeradbiomed.2014.11.006Get rights and content

Highlights

  • Muscle dysfunction and wasting add prognosis value in COPD and malignancies.

  • Oxidative stress and proteolysis contribute to muscle dysfunction and wasting in COPD.

  • Common biological features are shown in COPD and lung cancer (LC) cachectic muscles.

  • Redox signaling pathways and sarcomere disruption present in COPD and LC cachexia.

  • Key structural and functional proteins were similarly decreased in cachectic muscles.

  • Findings offer future avenues for research in human cachexia.

Abstract

Muscle dysfunction and wasting are predictors of mortality in advanced COPD and malignancies. Redox imbalance and enhanced protein catabolism are underlying mechanisms in COPD. We hypothesized that the expression profile of several biological markers share similarities in patients with cachexia associated with either COPD or lung cancer (LC). In vastus lateralis of cachectic patients with either LC (n=10) or advanced COPD (n=16) and healthy controls (n=10), markers of redox balance, inflammation, proteolysis, autophagy, signaling pathways, mitochondrial function, muscle structure, and sarcomere damage were measured using laboratory and light and electron microscopy techniques. Systemic redox balance and inflammation were also determined. All subjects were clinically evaluated. Compared to controls, in both cachectic groups of patients, a similar expression profile of different biological markers was observed in their muscles: increased levels of muscle protein oxidation and ubiquitination (p<0.05, both), which positively correlated (r=0.888), redox-sensitive signaling pathways (NF-κB and FoxO) were activated (p<0.05, all), fast-twitch fiber sizes were atrophied, muscle structural abnormalities and sarcomere disruptions were significantly greater (p<0.05, both). Structural and functional protein levels were lower in muscles of both cachectic patient groups than in controls (p<0.05, all). However, levels of autophagy markers including ultrastructural autophagosome counts were increased only in muscles of cachectic COPD patients (p<0.05). Systemic oxidative stress and inflammation levels were also increased in both patient groups compared to controls (p<0.005, both). Oxidative stress and redox-sensitive signaling pathways are likely to contribute to the etiology of muscle wasting and sarcomere disruption in patients with respiratory cachexia: LC and COPD.

Introduction

Chronic Obstructive Pulmonary Disease (COPD) is a highly prevalent condition that represents a major cause of death worldwide [1], [2]. Importantly, quadriceps muscle dysfunction [3], [4] and reduced muscle mass, as measured by mid-thigh cross-sectional area, were shown to be good predictors of COPD prognosis and mortality [5]. Furthermore, acute exacerbations and comorbidities, which are characteristic features of patients with COPD and contribute to muscle function and mass loss [6], have a significant impact on their exercise tolerance and quality of life (QoL), thus imposing a substantial economic burden in our societies [7]. Several pathways leading to muscle atrophy (ubiquitin-dependent protein catabolism) and mitochondrial dysfunction were shown to be enhanced in the limb muscles of patients with stable COPD and during exacerbations [6], [8]. On the other hand, cachexia is also particularly prevalent in advanced malignancies, leading to progressive functional impairment and poor QoL in the patients [9]. Its prevalence has been reported to be as high as 60 to 80% in patients with lung and gastrointestinal tumors [9], [10], [11]. Additionally, underlying COPD is also a major risk factor for lung cancer development, thus the two diseases may coincide in the same patient [12], [13].

Although the etiology of COPD muscle dysfunction and wasting in the context of chronic respiratory conditions and lung malignancies has not been yet fully elucidated, mechanisms such as oxidative stress [14], [15], [16], systemic inflammation [10], [11], [17], [18], metabolic disturbances and nutritional abnormalities [19] are part of its etiology. Furthermore, ubiquitin-proteasome appears to be the major proteolytic system involved in muscle protein degradation in catabolic states including cancer-induced cachexia models [20], [21] and COPD, in both limb [6], [8], [22], [23] and respiratory [24] muscles of patients with normal and altered body composition. Furthermore, several redox-signaling pathways also seem to regulate muscle protein breakdown in COPD and cancer cachexia models [6], [8], [20], [22], [23], [24]. However, whether oxidative stress and redox-signaling pathways may similarly drive muscle wasting and protein loss in cachectic patients of two distinct etiologies, a chronic condition (e.g. COPD) and a disease of more rapid progression (e.g. lung cancer) needs to be answered. Moreover, it remains to be identified whether the expression profile of several biological markers exhibit common features in different cachectic conditions such as those associated with COPD and lung cancer. Additionally, most of currently available information arises from studies conducted on animal and cell models, and despite that recent investigations have demonstrated alterations in key mechanisms of muscle function and mass maintenance in patients with oncologic cachexia [10], [11], [25], [26], [27], a paucity of data on human cachexia still exists.

In the study, the general hypothesis was to examine whether biological features that have been so far demonstrated to be involved in the muscle dysfunction and wasting of patients with COPD are expressed in a similar fashion in the lower limb muscles of patients with lung cancer cachexia. A more specific hypothesis was two-fold on the basis that in atrophying limb muscles of patients with two distinct respiratory conditions: 1) a significant rise in oxidative stress levels and expression of redox-signaling pathways may take place in their muscles in a similar fashion, and 2) other biological events including inflammatory cytokines, autophagy and proteolytic markers, contractile proteins, and both muscle structure and ultrastructural abnormalities known to be involved in muscle dysfunction and wasting may also bear resemblance in the two conditions. Accordingly, our primary objective was to comprehensively explore in vastus lateralis of patients with either severe COPD or lung cancer cachexia the expression levels of the following biological markers: 1) redox balance and inflammation, 2) redox-signaling pathways, 3) autophagy markers (autophagosomes), 4) ubiquitin-proteasome system markers and myostatin, 5) muscle contractile and functional proteins levels, 5) muscle fiber types and structure, and 6) muscle sarcomere ultrastructural architecture including mitochondrial density. Systemic inflammation and oxidative stress levels were also evaluated in the blood of the two groups of cachectic patients. A group of healthy sedentary subjects was also recruited for the purpose of the investigation and both patients and control individuals were clinically and functionally evaluated. Potential correlations between the different study variables including oxidative stress and proteolytic markers were also examined as a secondary objective.

Section snippets

Methods

(See the online data supplement for detailed information on all the methodologies used in the study).

Clinical characteristics

Table 1 illustrates all clinical and functional variables of the study groups. Age did not significantly differ among the study subjects. Compared to healthy controls, BMI and FFMI were significantly reduced in both groups of cachectic patients, and BMI was also lower in COPD than in cancer cachectic patients. COPD patients had very severe airflow obstruction, while cancer patients exhibited mild-to-moderate airflow limitation. COPD patients exhibited functional signs of severe emphysema

Discussion

(See the online data supplement for details on the main findings)

In view of the reported findings, the study hypothesis has been confirmed. Cachectic patients from two different respiratory conditions exhibited similar biological features in their lower limb muscles and blood. Despite that most of the molecular events analyzed in the current study have been previously shown in muscles of patients with different degrees of COPD severity and body composition, the novelty herein is based on the

Conclusions

Oxidative stress and redox-sensitive signaling pathways are likely to contribute to the etiology of muscle wasting and sarcomere disruption in patients with cachexia of two distinct respiratory conditions of high prevalence such as lung cancer and COPD. Contractile myosin loss and ultrastructural abnormalities such as enhanced sarcomere disruption are also characteristic phenotypic features of the cachectic muscles in both advanced COPD and lung cancer. These findings may offer insight into

Competing interests declared by all the authors

None.

Sources of funding

This study has been supported by CIBERES; FIS 05/2458; FIS 11/02029; FIS 12/02534; SAF-2011-26908; 2009-SGR-393; SEPAR 2007 & 2009; FUCAP 2011; FUCAP 2012; and Marató TV3 (MTV3-07-1010) (Spain). Dr. Esther Barreiro was a recipient of the ERS COPD Research Award 2008.

Authors’ Contributions

Conception and design: JR, JG, EB; Patient assessment and recruitment and sample collection: DAR, PA, JB, SPG; Ultrastructural assessment: JLL; Molecular biology analyses: EPV, EB; Statistical analyses and data interpretation: EPV, EB; manuscript drafting and intellectual input: EPV, JR, JG, EB; manuscript writing final version: EB.

Acknowledgments

The authors are grateful to Mr. Francisco Sanchez and Ms. Mònica Vilà-Ubach for their assistance with the ultrastructural analyses, identification and measurements of sarcomere damage areas and mitochondrial sizes, and counting of autophagosomes in the muscle specimens, and to Ms. Mireia Admetlló and Ms. Yolanda Torralba for their help with the patient reports and clinical information. We are also extremely thankful to Mr. Sergi Mojal for his valuable help with the statistical analyses of the

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