ViewpointPrinciples, insights, and potential pitfalls of the noninvasive determination of muscle oxidative capacity by near-infrared spectroscopyAlessandra Adami and Harry B. RossiterAlessandra AdamiRehabilitation Clinical Trials Center, Division of Respiratory and Critical Care Physiology and Medicine, Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, California and Harry B. RossiterRehabilitation Clinical Trials Center, Division of Respiratory and Critical Care Physiology and Medicine, Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, Torrance, CaliforniaPublished Online:24 Jan 2018https://doi.org/10.1152/japplphysiol.00445.2017This is the final version - click for previous versionMoreSectionsPDF (382 KB)Download PDF ToolsExport citationAdd to favoritesGet permissionsTrack citations ShareShare onFacebookTwitterLinkedInWeChat Skeletal muscle oxidative capacity is highly plastic, strongly associated with whole body aerobic capacity (16, 18) and state of health. Loss of muscle oxidative capacity is associated with physical inactivity, aging, and chronic disease (17) and has been implicated in the pathophysiology of obesity and diabetes (21). Evaluating these changes has traditionally been limited to invasive or costly assessments (biopsy or 31P MRS). To address this, Hamaoka and colleagues (23) developed an innovative, noninvasive approach using near-infrared spectroscopy (NIRS) to quantitatively measure muscle oxygen consumption (mV̇o2; 12) and use this to infer muscle oxidative capacity based on the mV̇o2 recovery rate constant (k) (later modified 26). This technique has subsequently been used to interpret relative differences in oxidative capacity across a wide range of muscles, ages and disease states (Fig. 1C). The purpose of this Viewpoint is to open a discussion on the principles, insights, and potential pitfalls of using NIRS to measure k and infer muscle oxidative capacity.Fig. 1.Muscle oxygen consumption (mV̇o2) recovery rate constant (k) by near-infrared spectroscopy. A and B: example of the oxidative capacity test by NIRS in the medial gastrocnemius of a 54 yr-old female. A: changes in the tissue saturation index (TSI) during dynamic exercise (EX, gray area) and subsequent intermittent arterial occlusions at rest. B: mV̇o2 recovery kinetics derived from the rate of change of TSI during intermittent arterial occlusions measured from A. mV̇o2 recovery data are fit to an exponential (dashed line) to estimate the recovery k. The time constant (τ) is the reciprocal of the rate constant k (τ = 1/k). C: summaries of current reports of the mV̇o2 recovery rate constant (k), which is proportional to oxidative capacity, in upper and lower limbs of adults in health and disease. A and B: redrawn with permission from Elsevier (1).Download figureDownload PowerPointPrinciples.First-order Michaelis-Menten enzyme kinetics dictates that mV̇o2 kinetics are directly proportional to muscle oxidative capacity (6, 20, 22). This concept is broadly supported when comparing across species during whole body exercise (24) and was specifically identified in the recovery k of single frog muscle fibers (r2 = 0.77; 33) (20). Such observations form the basis to infer muscle oxidative capacity from k in humans. Of note, this is distinct from the recovery k of pulmonary V̇o2 following exercise, which is dependent on both muscle and circulatory function. Isolated muscle cellular V̇o2 can be measured by NIRS during arterial occlusion from the changes in concentration of oxy- and deoxy-hemoglobin and myoglobin (10, 13), i.e., in the absence of blood flow, muscle deoxygenation occurs solely by O2 consumption. For this method, brief light-intensity muscle contractions are used to elicit an increase in mV̇o2, after which recovery k is assessed using a series of intermittent arterial occlusions (each 5–10 s, separated by 5–20 s of reperfusion; Fig. 1 Insights.The major advantage of NIRS-based muscle oxidative capacity estimation is its relative ease of application compared with muscle biopsy or 31P MRS. It is noninvasive, relatively inexpensive, short duration, and well tolerated. The isolated nature of the brief muscle contractions allows even functionally limited patients to perform the test. Assessment of different superficial limb muscle groups (plantar flexors, knee extenders, wrist flexors) or between limbs (e.g., for unilateral impairments) is highly feasible. The technique is particularly useful for assessing longitudinal change or interventional efficacy, such as following the response to training (7, 28, 30).In the past 5 years, the technique has found wide application in health (5, 28) and clinical populations (1–4, 8, 9, 27, 30, 34). Figure 1C shows k values across a wide range of muscle groups, age, and health status. These data reveal the extreme plasticity of relative muscle oxidative capacity (see Ref. 16), with a ~5-fold difference between muscles in patients with motor-complete spinal cord injury and endurance athletes. Evidence of the well-established age-associated decline in muscle oxidative capacity is seen among these cross-sectional studies in both upper and lower limb muscles. Also observed is the somewhat lower oxidative capacity of the wrist flexors compared with the vastus lateralis or gastrocnemius muscles across comparable groups, presumably reflecting the lower expression of oxidative type I muscle fibers in the forearm. Loss of muscle oxidative capacity (~25–45% vs. similar aged controls) is seen in COPD (GOLD class 3–4) and CHF (NYHA class I–III), a loss that appears consistent between upper and lower limbs.Potential pitfalls.As a major advantage of the NIRS approach is that it relies on mV̇o2 kinetics to estimate oxidative capacity, quantification of absolute mV̇o2 (which is complex by NIRS) is not necessary; only relative change in mV̇o2 over time is required. However, method relies on at least two competing assumptions and some technical limitations.Two key assumptions are 1) that mitochondrial oxidative enzymes are maximally activated by the brief contractions, ratifying the assumption of “functionally” first-order enzyme kinetics (21, 32, 33) and 2) that O2 concentration is not limiting to k (15, 33). Recent studies suggest that control of oxidative phosphorylation in human muscle is not first order (19). However, exercise rapidly activates mitochondrial enzymes (11, 19), and the recovery of this activation process is slow in relation to k (19). The NIRS approach relies upon brief contractions to release inhibition of mitochondrial enzyme activity such that linear proportionality exists between cellular oxidative capacity and k (33). An insufficient contraction-related stimulus could result in a low k that misrepresents the “true” oxidative capacity. Low activation may also reduce the confidence of the fitted curve and the modeled k. Although there appears to be no ordering effect of repeated measurements made during the same visit (1, 9, 27), we caution that poor test-retest reproducibility of k is found in participants with a low contraction-induced increase in mV̇o2 (1).Recovery k only reflects oxidative capacity when [O2] is abundant (33). As exercise and the imposed arterial occlusions required by the method reduce muscle Po2, care is required that [O2] does not become limiting. Haseler et al. (15) showed that PCr recovery was slowed during hypoxia compared with normoxia. For this reason it is recommended that NIRS estimation of oxidative capacity be preceded by a ~5 min arterial occlusion to identify the functional range of tissue O2 saturation (StO2). Subsequently, brief contractions and occlusions are metered such that StO2 remains high (1). Little data exist to determine whether this “ischemic preconditioning” acutely alters mitochondrial function or recovery k. Nevertheless, as StO2 is measured by NIRS itself, the assessor can administer the test so as to ensure that recovery k remains a reflection of the intrinsic intramuscular capacity for oxidation and independent of vascular function.There exist technical challenges with the NIRS assessment that also require consideration. Early attempts at NIRS-based mV̇o2 measurement identified that tissue hemoglobin often varies during arterial occlusion. This was attributed to residual pressure gradients causing movement of heme chromophores in and/or out of the NIRS field of view, even during arterial occlusion (26). Thus, if total hemoglobin is not constant, changes in deoxy-hemoglobin and myoglobin may result from not only O2 consumption but also hemo-concentration/dilution. To address this, Ryan et al. (26) developed a correction method for hemoglobin volume change based on the instantaneous relative oxygenation. Other studies have used spatially resolved spectroscopy (10) to estimate StO2, producing similar results (1). Nevertheless, failure to adequately control for hemoglobin changes during the brief arterial occlusions will influence the measured k.The technique relies upon complete occlusion of blood flow, such that changes in oxygenation reflect only mV̇o2: should partial occlusion occur (particularly relevant to measurements of the vastus lateralis in well-muscled or obese individuals), the result becomes misleading. This requirement effectively limits the application to limb muscles because respiratory or abdominal muscles cannot easily be subject to arterial occlusion.Other considerations for valid and reproducible application of the technique include that the skin and adipose tissue thickness be low enough that the diffused NIRS light can reach muscle, and sufficient intensity of light is received at the NIRS detector. Poor probe placement, large skinfold, or high skin melanin content can obfuscate these requirements.Overall, the test-retest reliability of k assessment by NIRS is good (coefficient of variation, ~10%; intraclass correlation coefficient range, 0.26–0.93; 1, 26, 31) and is typically not inferior to biopsy or 31P MRS methods. Both NIRS and 31P MRS have the added advantage that they sample a larger volume of (albeit superficially weighted) muscle than biopsy. But test-retest variability is somewhat large compared with the typical effect size of oxidative capacity loss observed in disease (Fig. 1C). For this reason it is recommended to average two to three repeat k measurements in the same individual to minimize variability and increase sensitivity (1, 9, 27).By meeting each of these conditions, a reliable estimate of relative muscle oxidative capacity, independent of macro- or microvascular (dys)function, can be inferred from k.Conclusion.Test-retest reliability is sufficient across several laboratories for muscle k assessment to be used as a noninvasive tool to assess the efficacy of interventions designed to ameliorate muscle mitochondrial impairment in patients with chronic disease. The ease of application of the method is a major benefit, but quality control procedures are needed to ensure measurement validity and to minimize error. Overall, the NIRS-based assessment of muscle k, originally developed by Hamaoka and colleagues, offers promise to simplify identification of relative changes in muscle oxidative capacity in both research and clinical settings.DISCLOSURESNo conflicts of interest, financial or otherwise, are declared by authors.AUTHOR CONTRIBUTIONSA.A. analyzed data; A.A. prepared figures; A.A. drafted manuscript; A.A. and H.B.R. edited and revised manuscript; A.A. and H.B.R. approved final version of manuscript.REFERENCES1. Adami A, Cao R, Porszasz J, Casaburi R, Rossiter HB. Reproducibility of NIRS assessment of muscle oxidative capacity in smokers with and without COPD. Respir Physiol Neurobiol 235: 18–26, 2017. doi:10.1016/j.resp.2016.09.008. Crossref | PubMed | ISI | Google Scholar2. Adami A, Corvino RB, Casaburi R, Cao R, Calmelat R, Porszasz J, Rossiter HB. Low oxidative capacity in skeletal muscle of both the upper and lower limbs in COPD patients. 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Adami, Division of Respiratory & Critical Care Physiology & Medicine, Los Angeles Biomedical Research Institute at Harbor-UCLA Medical Center, 1124 W Carson St., CDCRC Bldg., Torrance, CA 90502 (e-mail: [email protected]org). Download PDF Previous Back to Top Next FiguresReferencesRelatedInformation Cited ByEpinephrine iontophoresis attenuates changes in skin blood flow and abolishes cutaneous contamination of near-infrared diffuse correlation spectroscopy estimations of muscle perfusionMiles F. Bartlett, Alberto Palmero-Canton, Andrew P. Oneglia, Julissa Mireles, R. Matthew Brothers, Cynthia A. Trowbridge, Dustin Wilkes, and Michael D. Nelson24 February 2023 | American Journal of Physiology-Regulatory, Integrative and Comparative Physiology, Vol. 324, No. 3Comparing muscle V̇o2 from near-infrared spectroscopy desaturation rate to pulmonary V̇o2 during cycling below, at, and above the maximal lactate steady stateRafael de Almeida Azevedo, Jonas Forot, Guillaume Y. Millet, and Juan M. 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Barstow16 May 2019 | Journal of Applied Physiology, Vol. 126, No. 5Enhanced Local Skeletal Muscle Oxidative Capacity and Microvascular Blood Flow Following 7-Day Ischemic Preconditioning in Healthy Humans9 May 2018 | Frontiers in Physiology, Vol. 9Last Word on Viewpoint: Principles, insights, and potential pitfalls of the noninvasive determination of muscle oxidative capacity by near-infrared spectroscopyAlessandra Adami and Harry B. Rossiter24 January 2018 | Journal of Applied Physiology, Vol. 124, No. 1Commentaries on Viewpoint: Principles, insights, and potential pitfalls of the noninvasive determination of muscle oxidative capacity by near-infrared spectroscopy24 January 2018 | Journal of Applied Physiology, Vol. 124, No. 1Equipment, measurements and quality control More from this issue > Volume 124Issue 1January 2018Pages 245-248 Copyright & PermissionsCopyright © 2018 the American Physiological Societyhttps://doi.org/10.1152/japplphysiol.00445.2017PubMed28684592History Received 16 May 2017 Accepted 5 July 2017 Published online 24 January 2018 Published in print 1 January 2018 Metrics