Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder characterized by progressive dysfunction in the motor system (van Es et al., 2017). The impairments are irreversible, which start from the weakness of the first affected site to paralysis, dysphagia, and eventually respiratory failure. Thus, patients are significantly limited in their ability to exercise (Chen et al., 2015). Recent identifications of pathophysiological mechanisms of ALS potentially provide new insights, including autonomic dysregulation for heart rate (HR) variability and hypermetabolism for respiratory decompensation (Hardiman et al., 2017; Ahmed et al., 2018). Moreover, a growing body of evidence has identified consistent mitochondrial abnormalities in neuromuscular tissues of patients with ALS and mouse models (Dupuis et al., 2004; Al-Sarraj et al., 2014; Jankovic et al., 2021). Though the pathogenesis of ALS is different from the metabolic myopathy, such as mitochondrial disease, dysfunction of mitochondria has been linked to muscle weakness in ALS: (1) the deposition of pathological protein of ALS such as TAR DNA-binding protein 43 (TDP-43) impairs the mitochondrial structure, which further induces death of muscular cells through devastating mitophagy, excitotoxicity, and oxidative stress (Kodavati et al., 2020). The consequent loss of actin and myosin filaments limits effective contractions of single-muscle fibers as the sliding filament theory, functionally manifesting as muscle weakness in patients (Miller et al., 2014). (2) Adenosine triphosphate (ATP) is the source of energy used to power muscle contraction. The intracellular-stored ATP is not enough for continuous muscular activities, hence the replenishment of ATP through mitochondrial respiration is substantial to high-intensity muscle movements (Baker et al., 2010). Overexpressing mutant TDP-43 of the ALS model decreases mitochondrial complex I activity, which further decreases oxygen utilization and ATP production via defective oxidative phosphorylation (Huang et al., 2020). The oxidative stress and inflammation in ALS aggravate the decrease in ATP regeneration and cause depletion of energy (Obrador et al., 2021). Thus, the insufficient energy supply induced by dysregulated mitochondria is linked to muscle fatigue and weakness in ALS. (3) In TDP-43 mouse models, distinct changes in mitochondrial dynamic and aggregation are reported (Kodavati et al., 2020). Mutated TDP-43 affects mitofusin 1 and 2 levels, causing fragmented mitochondrial morphology and muscle atrophy that further decreases muscular strength (Wang et al., 2013). (4) Dysregulated mitochondria are associated with dysfunction of ion channels (including Na⁺, K⁺, and Ca²⁺) in ALS (LoRusso et al., 2019). The abnormalities disturb the excitation-contraction coupling and decrease the excitability of muscle fibers that further induce muscle fatigue in patients (Allen et al., 2008). Thus, mitochondrial dysfunction has emerged as one of the significant reasons for the exercise limitation of ALS (Angelini and Siciliano, 2021). Cross-sectional studies and pilot trials of small size have revealed significant exercise intolerance in ALS (Sanjak et al., 1987; Siciliano et al., 2002; Mezzani et al., 2012; Braga et al., 2018). Further studies with more patients to explore its clinical value for the assessment of disease progression are required. The current methods for disease evaluation are mainly based on the disability degree, represented by the revised ALS Functional Rating Scale (ALSFRS-R) and the King’s College Staging System (KCSS) (Cedarbaum et al., 1999; Roche et al., 2012). Thus, the exploration of direct indicators for exercise capability associated with disease severity and prognosis could provide a novel prospective for disease evaluation (Sun et al., 2020). Detailed assessments of the systemic physiological responses to exercise could also give insights into potential aerobic rehabilitation in ALS.

Cardiopulmonary exercise testing (CPET) is recommended as the gold standard for objectively evaluating exercise physiology by the European Association for Cardiovascular Prevention and Rehabilitation and the American Heart Association (EACPR/AHA) (Guazzi et al., 2012, 2016). It can provide a comprehensive assessment in multiple relevant physiological systems, including the cardiovascular, pulmonary, and musculoskeletal systems (Fan and Jia, 2020). To quantitatively analyze exercise physiology, CPET non-invasively measures extensive variables reflecting different physiological responses under peak exercise stress (Arena and Sietsema, 2011). Oxygen uptake at peak exercise (VO₂ peak) is well recognized to represent the aerobic exercise capacity for the integrated function of the whole-body exercise capability. Other parameters separately represent the singular function of the system-targeted exercise physiology (i.e., HR recovery for the cardiovascular system, VE/VCO₂ slope for the pulmonary system) (Taivassalo et al., 2003; Crisafulli et al., 2018; Opina et al., 2019). The clinical value of CPET in cardiovascular and pulmonary diseases has been firmly established for preoperative evaluation and prognosis assessment (Grant et al., 2015; Guazzi et al., 2017). Recent studies have identified the reliable use of CPET in neurological diseases, including metabolic myopathies, stroke, multiple sclerosis, and Huntington’s disease; however, there is a lack of application of CPET for ALS so far (Jeppesen et al., 2003; Taivassalo et al., 2003; Ciammola et al., 2011; Heine et al., 2014, 2016; Fan and Jia, 2020). As a non-invasive tool that allows for quantitative assessment, CPET could supplement previous evaluations of functional disability and further provide a comprehensive profile of exercise physiology impairments for ALS.

We aimed to evaluate the exercise physiology impairments using CPET in patients with ALS compared with age- and sex-matched healthy controls. In addition, we explored their clinical values to assess disease severity and prognosis of ALS from a clinical physiology perspective in our prospective cohort.

This study performed comprehensive CPET measurements to investigate exercise physiology impairments in patients with ALS compared with age- and sex-matched healthy controls. This is a pioneering study to reveal the predictive values of CPET parameters for disease severity and prognosis of ALS in a prospective cohort. We provided comprehensive evidence for the application of CPET in ALS: our results quantify the significant decrease in exercise capacity in patients with ALS compared with strictly matched controls, including the overall VO₂ peak and the system-targeted HR peak and VE/VCO₂ slope. Furthermore, CPET parameters were significantly associated with disease severity scores, which showed the potential use of CPET as an objective marker for non-invasively quantifying disease impairment. Finally, typical CPET variables could distinguish patients with different prognoses well since the VO₂ peak, HR peak, and VE/VO₂ peak emerged as independent predictors of survival in ALS. Thus, CPET could provide novel evaluations from the perspective of exercise physiology in ALS and benefit not only for intervention timing in the clinic but also for stratification of patients in scientific research.

Among the CPET parameters, the VO₂ peak was identified as the most significant parameter of ALS in our study. As a well-recognized variable to represent the exercise capability, the VO₂ peak quantitatively reflects the exercise capacity of the whole body (Guazzi et al., 2012, 2016). We thoroughly identified the prevalence and range of VO₂ peak alterations in ALS compared with strictly matched controls (Figure 2). The significantly higher frequency of impaired exercise capacity in patients with ALS (44.95%) addressed the extensive functional decline. Furthermore, in our supplementary analysis, the VO₂ peak showed a significant decline in patients with a disease duration less than 12 months compared with healthy controls [16.23 (5.62) vs. 22.26 (7.09); p < 0.01; Supplementary Table 4]. Thus, the remarkable impairment in early phase patients emphasizes the sensitivity of exercise capacity for early disease impairment. Additionally, the positive correlation and survival analyses revealed the meaningful value of the VO₂ peak in the clinical practice of ALS. The significant associations between the VO₂ peak and most established clinical parameters of ALS (Table 2) suggest that it can be used as a comprehensive, objective, and quantitative measurement for monitoring disease severity and progression in ALS. As an indicator reflecting the whole-body physiological response, the VO₂ peak comprehensively summarizes the overall capacity for exercise affected by multiple systems (Guazzi et al., 2017). In addition, it is more objective as a quantitative index than traditional parameters based on self-reported symptoms (Fournier et al., 2020). Finally, our consistently positive results in univariable and multivariable survival analyses also highlight the prognostic role of the VO₂ peak for better outcomes in ALS. Given its repeatability and modifiability, the VO₂ peak has been used as a primary endpoint for non-invasively assessing the response to the treatment of physical exercise in several randomized clinical trials (Shulman et al., 2013; Wallace et al., 2019). Since improvements in the VO₂ peak after intervention have been observed in patients with inclusion body myositis and Charcot-Marie-Tooth disease, longitudinal studies on CPET in ALS are needed in the future to explore potential treatments from the perspective of exercise capacity to improve survival (Wallace et al., 2019).

In our study, CPET also identified the exercise physiology impairments in different systems to support the multisystem impairments in ALS from a functional perspective. We established direct relationships between the system-specific CPET parameters and the pathophysiological mechanism of the different systems affected in ALS (Guazzi et al., 2012, 2016). Additionally, we revealed that the decline in overall exercise capacity had multifaceted origins involving cardiovascular, pulmonary, and muscular impairment in ALS.

Conflicting data have been reported regarding cardiovascular disorders in ALS (Pereira et al., 2021). Our study identified the cardiac involvement of ALS with abnormal autonomic nervous function. We observed a significantly higher HR at rest in patients, which supports the continuous impairment of the sympathetic nervous system in ALS. Tanaka et al. (2013) recognized similar sympathetic hyperactivity with cardiac [¹²³I] MIBG scintigraphy and reasoned that they were related to sudden cardiac arrest in ALS. However, HR at peak exercise was significantly limited among our patients, which implies a dysregulation of the sympathetic nervous system in accommodating higher exercise levels. Since no significant difference was shown in HR recovery after exercise, we supposed that the parasympathetic nervous system was relatively reserved in ALS. Thus, cardiovascular impairment in ALS does not resemble the CPET pattern typical of infiltrative cardiomyopathy with decreased HR recovery (Patel et al., 2014). Furthermore, our positive correlation and survival analyses recognized that the HR peak was similar to the VO₂ peak for disease evaluation. This emphasizes the significant influence of the blunted HR response on overall exercise capacity in ALS.

For the pulmonary system, the significantly increased VE/VCO₂ slope suggests a ventilatory limitation in patients with ALS healthy controls in our study. Recognized as a measure of ventilatory efficiency, the VE/VCO₂ slope reflects the capacity of CO₂ elimination during the whole exercise process (Neder et al., 2017). The weakened respiratory muscles in ALS are known to cause decreased breathing activity, leading to ventilatory insufficiency (van Es et al., 2017). An increased VE/VCO₂ slope is typical in chronic obstructive pulmonary disease, another restrictive ventilatory disorder with a similar pathophysiological mechanism of pulmonary dysfunction to that in ALS (Neder et al., 2017). However, no significant difference in the survival of patients with different VE/VCO₂ slopes was observed in our study. We attributed this to the relatively mild respiratory dysfunction of patients in our cohort as the median value of the VE/VCO₂ slope in our patients did not reach the recommended cutoff value for ventilatory limitation (>30) (Guazzi et al., 2012). Further studies including patients with more severe respiratory insufficiency are required.

Regarding the muscular system, our results revealed no significant dysfunction of muscular oxygen utilization in patients with ALS compared with healthy controls. Considering that muscle dysfunction is indirect and secondary to neuronal impairment in ALS, it is reasonable that our CPET pattern is different from that seen in mitochondrial myopathies (Jeppesen et al., 2003). The VE/VO₂ peak was a comprehensive indicator of breathing economy. In the muscular disease with scarce impairment in the cardiopulmonary system, the higher VE/VO₂ peak has been reported to directly reflect the lower muscle oxidative capacity in the patients with mitochondrial myopathy compared with controls (Taivassalo et al., 2003). Besides, increased VE/VO₂ peak has also been identified in cardiopulmonary diseases for ventilatory inefficiency (i.e., heart failure) (Mejhert et al., 2002). Since ALS is a complex disease with the involvement of multiple systems, the interpretation of the VE/VO₂ peak could be a comprehensive indicator for impairments in both cardiopulmonary and muscular systems. Moreover, the significant correlation of clinical survival supported its potential use as a prognostic indicator in ALS (Table 4). Further studies including the detection of plasma lactate and biopsies of muscle tissues are needed to provide more direct evidence of muscular mitochondrial abnormalities in exercise intolerance of ALS.

Neurofilament light chain (NfL) was not significantly associated with typical CPET parameters in this study. This could be due to different pathogenic mechanisms represented by the two variables in ALS. NfL, the main by-product of neuroaxonal breakdown, mainly represents the degree of axonal damage and reflects the neurodegeneration process of ALS (Lu et al., 2015). In contrast, abnormal CPET parameters, the direct representative of exercise physiology impairments, mainly reflect the dysregulation of multiple systems under the exercise stressor (Guazzi et al., 2017). The negative correlations could be reasonable since the parameters stand for the pathophysiology of different dimensions in ALS. Further studies with a larger sample size and more elaborate study design are needed to get a better understanding of the issue.

This study has several limitations. First, since the standard CPET requires the adaptability to pedal the apparatus for the cycling test, we enrolled patients at the time of diagnosis to ensure functional availability. This might limit the application in late-stage patients with severe impairments. Furthermore, given the characteristics of CPET, we suggest conducting this evaluation in the early stage of patients with ALS. Second, this study was conducted in patients with sporadic ALS, independent replication of this study in those with familial ALS with a specific genetic background is essential to support a broader application of CPET in ALS. In addition, although this preliminary study first explored the clinical value of CPET in ALS, the present investigation was conducted in a single center with potential referral bias. Thus, multicenter studies involving larger numbers of participants are needed to confirm our findings in the future.

In summary, through the comprehensive assessment of CPET, we quantified the common and significant exercise physiology impairments in patients with ALS compared with matched controls. CPET variables were significantly associated with functional scores reflecting disease severity and progression. We demonstrated that patients with a higher VO₂ peak, HR peak, and a lower VE/VO₂ peak had better survival in ALS. Our findings of exercise physiology impairments also highlight the potential use of CPET in ALS for the non-invasive, quantitative, and objective evaluation of disease severity and clinical prognosis.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

The studies involving human participants were reviewed and approved by the institutional Ethics Committee of the Peking University Third Hospital. The patients/participants provided their written informed consent to participate in this study.

JH, DF, JF, WZ, and CR: concept and design. JH, JF, and WZ: drafting of the manuscript. DF, WZ, and CR: critical revision of the manuscript for important intellectual content. NL, LC, JH, and JF: statistical analysis. DF and JH: obtain funding. CR, PL, DL, LZho, and LC: administrative, technical, and material support. JH, DF, WZ, JF, CR, and PL: supervision. All authors: acquisition, analysis, or interpretation of data.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.