Inefficient skeletal muscle oxidative function flanks impaired motor neuron recruitment in Amyotrophic Lateral Sclerosis during exercise

www.nature.com/scientificreports OPEN Received: 3 January 2017 Accepted: 19 April 2017 Published: xx xx xxxx Inefficient skeletal muscle oxidative function flanks impaired motor neuron recruitment in Amyotrophic Lateral Sclerosis during exercise F. Lanfranconi1, A. Ferri1,2, G. Corna1, R. Bonazzi1, C. Lunetta3, V. Silani4,5, N. Riva6, A. Rigamonti7, A. Maggiani8, C. Ferrarese1,9 & L. Tremolizzo1,9 This study aimed to evaluate muscle oxidative function during exercise in amyotrophic lateral sclerosis patients (pALS) with non-invasive methods in order to assess if determinants of reduced exercise tolerance might match ALS clinical heterogeneity. 17 pALS, who were followed for 4 months, were compared with 13 healthy controls (CTRL). Exercise tolerance was assessed by an incremental exercise test on cycle ergometer measuring peak O2 uptake (VO2peak), vastus lateralis oxidative function by near infrared spectroscopy (NIRS) and breathing pattern (VE peak). pALS displayed: (1) 44% lower VO2peak vs. CTRL (p < 0.0001), paralleled by a 43% decreased peak skeletal muscle oxidative function (p < 0.01), with a linear regression between these two variables (r2 = 0.64, p < 0.0001); (2) 46% reduced VEpeak vs. CTRL (p < 0.0001), achieved by using an inefficient breathing pattern (increasing respiratory frequency) from the onset until the end of exercise. Inefficient skeletal muscle O2 function, when flanking the impaired motor units recruitment, is a major determinant of pALS clinical heterogeneity and working capacity exercise tolerance. CPET and NIRS are useful tools for detecting early stages of oxidative deficiency in skeletal muscles, disclosing individual impairments in the O2 transport and utilization chain. Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder involving alpha motor neurons and abnormal recruitment of motor units, due to lesions in the corticospinal pathways. The resulting clinical manifestations of asthenia, spasticity, and amyotrophy harshly impair functional independence of patients and their quality of life. ALS patients (pALS) have a heterogeneous onset, with increasing fatigability that may begin with impaired activation of limbs, or with dysphagia or dysarthria when the bulbar district is affected first, and with a final failure of respiratory muscles. The appearance of ALS, from the earliest phases of the disease, also typically consists of reduced exercise tolerance until there is complete restriction of activities of daily living1–3. The characteristic heterogeneity in exercise tolerance of pALS is related to both the pathologic pattern of motor unit recruitment, and muscle impairment due to disuse of potentially healthy muscles3, 4. The reduced exercise tolerance in pALS (i.e., the capacity to maintain workloads ranging from habitual activities to rehabilitation exercises) has been associated with mitochondrial dysfunction, both as a direct pathogenic mechanism and as a factor contributing to the exercise limitation5, 6. Furthermore, the degree of exercise intolerance in pALS might correlate with the reduction in the number and effectiveness of functional mitochondria able to guarantee an adequate O2 extraction at the skeletal muscles5. During the early stages (less than 9 months from disease onset), pALS show no evidence of mitochondrial dysfunction. However, this is clearly present with increasing severity and when the disease is finally identified by clinical disability scales7. Nevertheless, a 1 School of Medicine and Surgery and Milan Center for Neuroscience (NeuroMI), University of Milano-Bicocca, Milano, Italy. 2Clinical Exercise Science Research Program, Institute of Sport, Exercise and Active Living (ISEAL), Victoria University, Melbourne, Australia. 3NEuroMuscular Omnicentre (NEMO), Fondazione Serena Onlus, Milano, Italy. 4Department of Neurology and Laboratory Neuroscience – IRCCS Istituto Auxologico Italiano, Pioltello, Italy. 5Department of Pathophysiology and Transplantation, Dino Ferrari Centre, Università of Milan, Milano, Italy. 6 San Raffaele Hospital, Milano, Italy. 7Alessandro Manzoni Hospital, Lecco, Italy. 8Italian Academy of Osteopathic Medicine (AIMO), Saronno, Italy. 9Neurology Unit, “San Gerardo” Hospital, Monza, Italy. Correspondence and requests for materials should be addressed to F.L. (email: ) Scientific Reports | 7: 2951 | DOI:10.1038/s41598-017-02811-z 1 www.nature.com/scientificreports/ subclinical mitochondrial dysfunction may be present in the early stages of ALS and undetectable in resting conditions, only to become apparent with additional external and environmental stress such as exercise8. In pALS this phenomenon has not been fully characterized in term of the underlying governing dynamics of oxidative metabolism, and specifically by assessing skeletal muscle oxidative function during exercise. By evaluating the balance between O2 delivery and O2 extraction at the skeletal muscle level (fulfilling the principle of mass conservation as settled by Fick equation), it is possible to obtain an indirect measure of mitochondrial function. Near infrared spectroscopy (NIRS) is a non-invasive method that can be used to provide an estimation of skeletal muscle fractional O2 extraction during exercise in healthy participants or patients9–12. According to Ryan and colleagues13, NIRS is a useful technique in order to verify oxidative metabolism impairment at the mitochondrial level in the skeletal muscles of pALS. However, although NIRS has been used to assess skeletal muscle oxidative function, the use of low-intensity exercise in this study means that impairments in maximal mitochondrial function may have been missed. Compared to Ryan et al. report, we thought to consider an incremental exercise in order to determine the putative impairment of the maximal power of oxidative phosphorylation pathway. Indeed, the expression of integration and efficiency of our body systems is revealed only by exercise, being the resting condition a state where metabolic resources are essentially underused. An efficient chain of transport and utilization of O2 from ambient air to mitochondrial level is individually determinable and can be considered our oxidative reserve to face exercise, unless a disease compromises this wealth. In fact, during moderate- or high-intensity physical activities carried out for several minutes, such as activities of daily living in pALS, the oxidative metabolism becomes the prevalent mechanism responsible for adenosine triphosphate (ATP) resynthesis necessary to sustain skeletal muscle functions. In addition, there is often impairment of the respiratory system in neuromuscular conditions, such as ALS. Nonetheless, even individuals with a well-advanced disease can have a ventilatory reserve sufficient to cope with maximal exercise6. In this regard, the respiratory system is able to adapt and triggers mechanisms of spontaneous compensatory respiratory plasticity that preserve breathing ca (...truncated)