Prolonged Increase in Glutamate Whole Body and Intracellular Production in Older Adults with COPD and Healthy Controls Post-Resistance Exercise

Abstract

Background:

Exercise training is essential for pulmonary rehabilitation in chronic obstructive pulmonary disease (COPD), yet patient responsiveness varies widely. We previously observed metabolic disturbances in amino acids critical for muscle health—such as glutamate, glutamine, branched-chain amino acids (BCAAs), and taurine—in COPD patients after an endurance exercise session, possibly related to increased energy demands and oxidative stress. However, the impact of resistance exercise on these metabolic pathways remains unclear.

Methods:

We measured plasma concentration, whole-body production (WBP), and intracellular production of glutamate, glutamine, BCAAs, and taurine using stable isotope pulse techniques in 24 COPD and 25 healthy older participants. Measurements were obtained before, and at 1 and 24 hours after, a resistance exercise session.

Results:

At baseline, COPD participants exhibited lower WBP of glutamine, taurine, and BCAAs compared to healthy participants (p < 0.05). Resistance exercise increased WBP of glutamate by 37–42%, glutamine by 9–10%, and intracellular glutamate production by 37–40%, while decreasing WBP of taurine by 7% (all p < 0.0001). These effects persisted at 24 hours post-exercise (p < 0.05). Although WBP of BCAAs remained unchanged, plasma leucine and isoleucine levels decreased by 16% and 13%, respectively, in COPD participants post-exercise (p < 0.05).

Conclusions:

A single resistance exercise session alters glutamate-related metabolism for at least 24 hours in healthy and COPD participants. A high BCAA clearance is likely required to rapidly upregulate glutamate production in COPD to meet increased energy demands, but this occurs at the cost of lowering plasma levels of BCAA necessary for muscle anabolism.

Graphical Abstract

1. Introduction

Chronic Obstructive Pulmonary Disease (COPD) is a prevalent and debilitating respiratory condition characterized by persistent airflow limitation and systemic features, including muscle function impairment and exercise intolerance [1,2]. Regular physical activity and exercise are recommended to manage COPD symptoms and systemic features, contributing to overall improvement in patients’ health status. Exercise training is considered the cornerstone of pulmonary rehabilitation. Numerous studies have highlighted the effectiveness of pulmonary rehabilitation in improving exercise tolerance in patients with COPD. However, several studies have reported heterogeneous individual responses, with the proportion of non-responders among COPD patients ranging from 30% to 50% [3,4]. Although responsiveness to exercise training in COPD patients primarily depends on disease severity and treatment regimen, other factors may also influence outcomes.

Exercise training in COPD generally consists of a combination of endurance and resistance exercise sessions. We previously demonstrated that endurance exercise alters the metabolism of specific amino acids—glutamate, glutamine, alanine, and branched-chain amino acids (BCAAs)—in COPD patients [5]. These alterations may be related to increased energy requirements [6][7], the protein catabolic effects of exercise [6], or other unidentified factors. We observed perturbations in glutamate and related amino acid metabolism both at rest and in response to a single endurance exercise session. For example, COPD patients exhibited lower post-exercise muscle glutamate and glutamine concentrations and elevated plasma alanine levels, suggesting enhanced exercise-induced nitrogen efflux [5][8,9]. Metabolic perturbations were still present 60 minutes after endurance exercise in COPD patients [10]. Glutamate, glutamine, and BCAAs have closely interrelated metabolic pathways. They respond to exercise and play crucial roles in muscle metabolism and function, including protein anabolism, regulation of oxidative stress, glucose homeostasis, nitrogen balance, and energy metabolism [11,12]. Furthermore, glutamate is instrumental in amino acid biosynthesis [13] and pH modulation [14]. Additionally, glutamate contributes to preserving high-energy phosphates during exercise, glutathione synthesis [8], cell proliferation, glycogenesis, the early lactate response [15], and ammonia buffering, thereby maintaining acid-base equilibrium. Taurine plays an important role in defending against free radical-mediated damage after exercise [15,16][17]. Therefore, a thorough understanding of the metabolism of glutamate, glutamine, BCAAs, and taurine, as well as their metabolic interactions in response to exercise, is vital for COPD management, particularly for patients who rely on exercise as part of their disease management strategy.

We previously observed alterations in amino acid concentrations and whole-body protein kinetics in response to endurance exercise using stable isotope methods [18]. However, it remains unclear whether resistance exercise affects the intracellular metabolism of glutamate-related amino acids differently in COPD patients compared to healthy individuals and, if so, how long these changes persist post-exercise. Therefore, the present study aims to determine whether participants with stable COPD, as a group, exhibit a different metabolic response to a standardized resistance exercise protocol than do healthy participants. We examined the acute metabolic responses of glutamate, glutamine, BCAAs, and taurine to a high-intensity resistance exercise session in participants with COPD versus healthy participants. We used a novel stable isotope pulse method at three specific time points: before exercise, 1 hour post-exercise, and 24 hours post-exercise. Alanine kinetics were not measured due to interference from its isotope label in other metabolic pathways. Compartmental modeling was used to assess the distribution of the amino acids of interest in the accessible (extracellular) and inaccessible (intracellular) compartments, the flows between these compartments, and their intracellular production [18,19]. We hypothesized that whole-body and intracellular production rates—but not concentrations—of glutamate, glutamine, and BCAAs increase immediately post-exercise, particularly in participants with COPD, and that these changes persist for at least 24 hours. Our long-term goal is to identify metabolic biomarkers that could predict non-response or low response to exercise rehabilitation in COPD, enabling the development of targeted nutritional approaches to optimize responsiveness to pulmonary rehabilitation.

2. Material and methods

2.1. Subjects

This was a prospective observational study comparing amino acid kinetics in participants with chronic obstructive pulmonary disease (COPD) and healthy participants before and after a resistance exercise session. We recruited a cohort of 24 older adults diagnosed with moderate to severe airflow obstruction (Stages II–IV) according to the Global Initiative for Chronic Obstructive Pulmonary Disease (GOLD) guidelines [20], and a healthy control group of 25 older adults. Participants were recruited through advertisements in local hospitals and communities, with all data collected at the Clinical Research Unit of the Center for Translational Research on Aging and Longevity at Texas A&M University. All COPD participants had a clinically confirmed diagnosis, a forced expiratory volume in 1 second (FEV₁) <70% of predicted, and had not experienced an exacerbation or respiratory tract infection within 4 weeks prior to the study. An assessment of medical history and medication use was conducted to evaluate comorbidities. Exclusion criteria included untreated metabolic or renal diseases (e.g., insulin-dependent diabetes), malignancy, recent surgery, and systemic corticosteroid use within 4 weeks before study commencement. Participants provided written informed consent, and the study was approved by the Institutional Review Board of Texas A&M University. All participants completed the study.

2.2. Anthropometrics, Body Composition, and Disease Characteristics

Participants’ body weight and height were measured using a digital beam scale and stadiometer, while fat mass and fat-free mass were determined via dual-energy X-ray absorptiometry (DXA) (Hologic QDR 4500, Version 12.7.3.1, Hologic, Bedford, USA) with participants in a supine position. All DXA procedures were performed consistently by a single researcher. We assessed post-bronchodilator forced expiratory volume in one second (FEV₁) and forced vital capacity (FVC) via spirometry, as well as the severity of dyspnea using the Modified Medical Research Council scale [21]. The Charlson Comorbidity Index Score (CCIS) was used to evaluate coexisting comorbidities [22], with a comprehensive medical history obtained through physical examinations, interviews, questionnaires, and medical charts. Habitual dietary intake was assessed using a 24-hour dietary recall [23].

2.3. Resistance Exercise Protocol and Muscle Function Testing

Participants underwent an isolated limb resistance exercise session on a KinCom isokinetic dynamometer (Isokinetic International, Chattanooga, USA), a device used to measure muscle strength and perform resistance exercises. Before the exercise session, a familiarization session ensured proper anatomical alignment and exercise training. Four exercises were performed on the dynamometer: right and left arm flexion-extension, and right and left leg flexion-extension, incorporating both concentric and eccentric contractions. Following a 10-repetition warm-up, each limb completed three sets of eight maximal voluntary contractions at 60°/sec, with two-minute rest periods between sets. Participants’ vital signs and fatigue/dyspnea scores were monitored using the Borg Scale, with additional oxygen supplied or rest periods extended if oxygen saturation fell below 85%. The exercise protocol lasted approximately 50 minutes. Leg muscle strength was assessed using the same dynamometer at 60°/sec [24]. After a warm-up, maximal force was determined through five maximal extension-flexion cycles, each followed by a 10-second rest period.

2.4. Assessment of Amino Acid Kinetics Using Stable Isotope IV Pulse

In the early morning, with participants in a postabsorptive state, a peripheral line was inserted into a dorsal hand vein or a lower arm vein. To simulate direct arterial sampling, the hand was placed in a controlled hot box maintained at an internal temperature of 60°C [25]. Following baseline blood sampling for enrichment, an intravenous tracer pulse consisting of 17 amino acid isotopes was administered [23][26][27]. This manuscript is the first in a series aimed at thoroughly evaluating the metabolic changes of six amino acids using stable isotope tracers: L-glutamate [1,2-¹³C₂] (9.17 mg), L-glutamine [¹;⁵N₂] (7.47 mg), L-leucine [¹³C₆] (5.04 mg), L-valine [¹³C₅] (3.96 mg), L-isoleucine [1-¹³C] (4.96 mg), and taurine [1,2-¹³C₂] (3.43 mg), selected because their metabolic pathways are closely interrelated and known to be affected by acute exercise. Arterialized-venous blood samples were collected just before pulse administration (background) and at 10, 20, 30, 60, and 120 minutes post-administration.

This stable isotope regimen was conducted over three consecutive study days to measure: Study Day 1, Baseline amino acid kinetics (pre-exercise). Study Day 2: Amino acid kinetics 1 hour after the acute resistance exercise protocol. Study Day 3: Amino acid kinetics 24 hours post-exercise from the acute resistance exercise protocol conducted on Study Day 2.

To ensure uniform metabolic conditions, all three metabolic procedures were performed in the early morning with participants in a postabsorptive state (fasting from 10 PM the previous evening). Participants were instructed to refrain from intense exercise for three days prior to the resistance exercise session and until completion of the 24-hour post-exercise stable isotope regimen. Additionally, participants were asked to maintain their habitual dietary intake, as assessed during the screening visit, throughout this period.

2.5. Biochemical Analysis and Calculations

Arterialized-venous blood was collected in lithium-heparinized tubes (Becton Dickinson Vacutainer System, Franklin Lakes, USA), placed on ice to minimize enzymatic reactions, and centrifuged to obtain plasma. Plasma aliquots were frozen at −80°C either immediately or after the addition of 0.1 volume of a 33.3% w/w trichloroacetic acid solution in water, followed by vortexing to denature proteins. Tracer enrichments [tracer:tracee ratio (TTR)] and amino acid concentrations in acid-deproteinized plasma supernatants were measured by liquid chromatography-tandem mass spectrometry (LC-MS/MS) [23]. Supplemental Table S1 describes the collisionally induced dissociation reactions (transitions) monitored by MS/MS to measure tracers and tracees. Transitions were selected to minimize background from naturally occurring ¹³C and to eliminate crosstalk due to label exchange, such as the conversion of L-glutamate [1,2-¹³C₂] to L-glutamine [1,2-¹³C₂] by glutamine synthetase, which could interfere with the measurement of the tracer L-glutamine [¹;⁵N₂], as both are heavy by two mass units. The LC-MS/MS method used can distinguish between ¹³C₂-glutamine and ¹;⁵N₂-glutamine. Plasma concentrations of glucose and high-sensitivity C-reactive protein (hs-CRP) were measured as markers of glucose intolerance and chronic inflammation, respectively, using a Cobas C111 Analyzer with standard kits (Roche Diagnostics, Mannheim, Germany).

2.6. Compartmental and Statistical Analysis

The measured TTR decay was transformed, and an analysis of covariance (ANCOVA) linear multi-regression analysis was performed with the covariates lean mass, group, age, sex, study day, time, GOLD stage, FEV₁:FVC ratio, pack-years, and ln(time). The estimated ln(TTR) was converted back by exponentiation, and the resulting TTR decay curve was fitted to obtain compartmental estimates [28]. This curve was fitted using a population pharmacology approach, where all participant data were analyzed with a two-exponential formula (TTR = aEXP(-k₁t) + bEXP(-k₂t)) in GraphPad Prism for the two groups [28]. The curve-fitting parameters a, b, k₁, and k₂ were used to calculate predicted whole-body production, perform compartmental analysis, and determine 95% confidence intervals for the measured parameters [28]. Compartmental analysis, as previously described [28][29], enabled the calculation of tracee fluxes between compartments using data from the tracer pulse method. The conversion of an amino acid to a metabolite was calculated by multiplying the whole-body production (WBP) of the product amino acid by the estimated ratio of the TTR of the product to that of the substrate [28].

The compartmental framework (Supplemental Figure 1) visualizes the distribution of a tracee in two principal cellular spaces: the accessible (extracellular) domain, labeled Q₁, and the inaccessible (intracellular) domain, labeled Q₂ [19]. Q₂ (and other pools) represents the space where small molecules reside. In this model, the appearance of tracee in the accessible plasma pool is termed Whole-Body Production (WBP). The flow of molecules from Q₁ to Q₂ is denoted F₂,₁, which equals the flow from Q₂ to Q₁ (F₁,₂). Intracellular amino acid production is represented by F₀,₂. Fractional loss (Frac_loss) is the proportion of the intracellular Q₂ pool lost irreversibly, calculated as the ratio of F₀,₂ (irreversible loss) to the total input (F₀,₂ + F₂,₁). Irreversible loss is then computed as F₀,₂ multiplied by fractional loss. Intracellular protein breakdown delivers amino acids via the U₂ entry and is disposed of via the F₀,₂ exit, with recycling contributing to turnover, as observed in continuous infusion protocols. The formulas used are provided in Supplemental Table S2 and described previously [30]. GraphPad Prism also calculated the standard error and confidence intervals for the derived variables.

Data were presented as means and standard deviations (SD) for baseline and exercise characteristics, and as means and estimated differences with 95% confidence intervals for all metabolic data. Data were examined for lognormal distribution; if lognormal, they were log-transformed prior to statistical testing. Group differences in categorical variables were assessed using Fisher’s exact test, while continuous variables were analyzed with an unpaired t-test or a repeated-measures mixed-effects model (RM-ANOVA) using the restricted maximum likelihood (REML) approach and Tukey’s multiple comparisons test. Due to the exploratory nature of the study, power analyses were not conducted prior to the study. The significance level was set at p < 0.05, and all reported values were compared to baseline metabolic data (pre-resistance exercise) unless otherwise specified. All statistical analyses were performed using GraphPad Prism (Version 10.0.3, GraphPad Software Inc., San Diego, USA).

3. Results

3.1. General, clinical, and body composition characteristics

The COPD and healthy participant groups were comparable in age, sex, and body mass index (BMI) (Table 1). However, higher values were found in COPD for smoking history and pack years (p<0.0001), but lower values for habitual physical activity (p=0.0001). Clinically, the COPD participants demonstrated moderate to severe airflow obstruction based on their FEV₁ and GOLD stage, and low-grade systemic inflammation as reflected by elevated plasma CRP levels (p<0.0001). The COPD participants had on average 0.30 exacerbations in the past year and a Charlson comorbidity index of 1.58, reflecting a population with comorbidities. No differences were observed in lean or fat mass between the groups, although a trend toward higher values for visceral adipose tissue was observed in COPD (p=0.078).

Table 1 -.

Baseline and exercise characteristics of the healthy and COPD groups

	Healthy (n=25)	COPD (n=24)	p-value
General characteristics
Sex (m/f)	14/11	15/9	0.426
Age (Yr)	70.7 ± 5.8	68.3 ± 10.3	0.320
BMI (kg/m2)	28.6 ± 6.7	29.6 ± 7.5	0.642
Smoking pack years	5.9 ± 15.6	44.1 ± 30.6	<0.0001
Years smoked	4.4 ± 10.7	30.6 ± 15.4	<0.0001
Habitual physical activity (score)	214.6 ± 77.9	132.3 ± 56.0	0.0001
Clinical characteristics
FEV1 (%pred)	105.6 ± 16.4	47.3 ± 16.2	<0.0001
FEV1 / FVC ratio (%pred)	81.6 ± 5.6	61.1 ± 10.8	<0.0001
GOLD stage	0.0 ± 0.0	2.6 ± 1.0	<0.0001
O2 saturation (%)	97.8 ± 1.2	95.8 ± 2.7	0.0003
Charlson comorbidity index (score)	0.3 ± 0.5	1.6 ± 0.8	<0.0001
COPD exacerbations (nr in last year)	0.0 ± 0.0	0.3 ± 0.7	<0.0001
Systolic blood pressure (mmHg)	134.6 ± 16.7	136.7 ± 16.8	0.756
Diastolic blood pressure (mmHg)	79.1 ± 6.7	79.8 ± 6.6	0.795
CRP (mg/L)	1.3 ± 1.5	7.6 ± 10.0	<0.0001
Glucose (mmol/L)	5.6 ± 0.7	5.5 ± 1.1	0.739
Body composition characteristics
Lean mass total (g)	49603 ± 11949	47988 ± 11497	0.632
Lean mass extremities (g)	19622 ± 5298	18884 ± 5254	0.627
Fat mass (%)	32.7 ± 8.1	35.5 ± 9.1	0.251
Visceral adipose tissue (g)	699 ± 327	902 ± 452	0.078
Exercise characteristics
Total work arms (J)	3235 ± 1210	2278 ± 933	0.006
Total work legs (J)	5832 ± 2142	3861 ± 1222	0.0006
Total work extension plus flexion (J)	8909 ± 3297	5919 ± 2080	0.0005
Average work extension + flexion (J)	373 ± 136	263 ± 89	0.002

Data are expressed as means with SD. Comparisons were performed using the unpaired Student’s t-test for continuous variables and Fisher’s exact test for categorical variables. Statistically significant values (p<0.05) are highlighted in bold. FEV1: Forced Expiratory Volume in one second. FVC: Forced Vital Capacity. hs-CRP: High sensitivity C-Reactive Protein.

3.2. Exercise characteristics

All participants completed the resistance exercise protocol. The COPD participants performed lower total resistance work than the healthy participants (p<0.006), due to 30% lower work by the arms (p=0.006) and 34% lower work by the legs (p=0.0006) (Table 1). As a measure of exercise-induced muscular fatigue, COPD participants but not healthy participants showed lower peak torque for leg extension at 1 hour post-exercise by 7% (p=0.036) and at 24 hours post-exercise by 7% (p = 0.045) as compared to pre-exercise values (data not shown).

3.3. Plasma concentrations in response to Resistance exercise

Plasma CRP was higher in COPD as compared to healthy participants at baseline (p<0.0001) (Table 1) and remained higher throughout the study (p=0.0007). Exercise did not change CRP in COPD (Δ 1hr (mean [95% CI]): −0.52 [−5.39, 4.35] mg/L, p=0.949; Δ 24hr: 0.03 [−2.93, 2.99] mg/L, p=0.997) and healthy group (Δ 1hr: −0.03 [−0.64, 0.59] mg/L, p=0.206; Δ 24hr: 1.27 [0.88, 1.66] mg/L, p=0.432) as compared to pre-exercise values. Also, plasma creatine kinase did not change in response to exercise in COPD (Δ 1hr (mean [95% CI]): 6.56 [−6.49, 19.61] U/L, p=0.210; Δ 24hr: 8.34 [−26.07, 42.76] U/L, p=0.745) and healthy group (Δ 1hr: −0.22 [−26.72, 26.29] U/L, p=0.878; Δ 24hr: 6.99 [−28.10, 42.07] U/L, p=0.633) as compared to pre-exercise values.

Overall, there were no differences in plasma concentrations of glutamate, glutamine, and the BCAAs—leucine, isoleucine, and valine—between the COPD and healthy participants (Table 2). However, plasma taurine levels were lower in COPD (p=0.0009). Resistance exercise did not alter the plasma concentrations of glutamate and glutamine. However, it decreased the plasma concentrations of leucine (p=0.004), isoleucine (p<0.001), and valine (p=0.014), with significant decreases immediately post-exercise in leucine (p=0.001) and isoleucine (p=0.014) in the COPD group only, with their levels returning to baseline values within 24 hours post-exercise. In contrast, plasma taurine levels increased in healthy subjects between 1 and 24 hours post-exercise (p=0.011).

Table 2 -.

Plasma concentrations of the healthy and COPD groups before, 1h and 24h after resistance exercise

		Baseline Mean	1h after exercise Mean	△ Predicted vs baseline	p	24h after exercise Mean	△ Predicted vs baseline	p	△ Predicted 24h vs 1h	p	p-time p-group p-time*group
Glutamate	H	44.12 [31.54, 56.71]	44.91 [24.80, 65.02]	−0.72 [−6.61, 5.17]	0.954	42.68 [28.02, 57.34]	−1.52 [−6.80, 3.75]	0.770	−0.80 [−6.69, 5.09]	0.944	0.120 0.229 0.514
	C	36.43 [43.35, 29.50]	32.26 [38.34, 26.18]	−4.14 [−9.59, 1.31]	0.172	35.27 [41.72, 28.82]	−1.49 [−6.94, 3.95]	0.790	2.65 [−2.80, 8.09]	0.481
Glutamine	H	535.0 [565.1, 504.8]	524.7 [557.9, 491.6]	−3.06 [−29.41, 23.29]	0.959	527.9 [556.6, 499.2]	−6.54 [−30.18, 17.09]	0.787	−3.48 [−29.87, 22.90]	0.947	0.742 0.771 0.989
	C	527.8 [571.4, 484.2]	527.3 [566.6, 488.0]	−2.20 [−26.63, 22.23]	0.975	527.9 [564.9, 490.8]	−4.48 [−28.90, 19.95]	0.900	−2.27 [−26.70, 22.16]	0.973
Taurine	H	39.90 [42.18, 37.62]	41.25 [45.07, 37.44]	2.20 [−1.76, 6.17]	0.385	44.43 [48.61, 40.24]	4.47 [0.89, 8.06]	0.011	2.27 [−1.71, 6.25]	0.366	0.062 <0.001 0.180
	C	35.25 [38.03, 32.48]	36.53 [39.30, 33.76]	1.34 [−2.37, 5.04]	0.667	35.92 [38.99, 32.84]	0.59 [−3.11, 4.30]	0.923	−0.74 [−4.45, 2.97]	0.882
Leucine	H	93.80 [105.5, 82.10]	85.21 [95.61, 74.82]	−4.98 [−15.17, 5.21]	0.476	93.94 [104.9, 82.96]	−0.48 [−9.63, 8.67]	0.991	4.50 [−5.71, 14.71]	0.546	0.004 0.364 0.262
	C	91.88 [107.5, 76.28]	76.56 [87.53, 65.59]	−14.56 [−24.02, −5.11]	0.00 1	83.07 [90.62, 75.52]	−4.07 [−13.53, 5.39]	0.562	10.49 [1.03, 19.95]	0.026
Isoleucine	H	54.8 [62.21, 47.39]	48.98 [54.43, 43.51]	−5.89 [−13.09, 1.31	0.130	56.57 [63.11, 50.03]	1.12 [−5.34, 7.59]	0.910	7.02 [−0.19, 14.23]	0.058	<0.001 0.484 0.850
	C	59.92 [72.36, 47.48]	52.10 [59.85, 44.36]	−8.06 [−14.74, −1.37]	0.01 4	55.72 [61.02, 50.42]	−0.61 [−7.29, 6.08]	0.974	7.45 [0.77, 14.13]	0.025
Valine	H	170.8 [190.4, 151.1]	153.6 [171.8, 135.5]	−14.75 [−31.96, 2.46]	0.108	169.1 [186.4, 151.8]	−3.37 [−18.83, 12.09]	0.862	11.38 [−5.87, 28.62]	0.262	0.014 0.397 0.982
	C	160.8 [185.1, 136.5]	145.8 [161.8, 129.8]	−13.43 [−29.41, 2.55]	0.117	152.3 [163.2, 141.3]	−1.66 [−17.64, 14.32]	0.967	11.77 [−4.21, 27.75]	0.190

Data are expressed in pM and as means and estimated differences with 95% CI. Comparisons were performed using RM-ANOVA and Tukey’s Multiple Comparisons test. Healthy (H) and COPD (C) subjects. Statistically significant values (p<0.05) are highlighted in bold.

3.4. Whole body production data in response to Resistance exercise

Both healthy and COPD participants demonstrated comparable overall values for WBP of glutamate, but lower WBP of glutamine (p<0.0001), taurine (p<0.0001), leucine (p=0.005), isoleucine (p=0.011), and valine (p<0.0001) in those with COPD (Table 3).

Table 3 -.

Whole body production of amino acids of the healthy and COPD groups before, 1h and 24h after resistance exercise

		Baseline Mean	1h after exercise Mean	△ Predicted vs baseline	p	24h after exercise Mean	△ Predicted vs baseline	p	△ Predicted 24h vs 1h	p	p-time p-group p-time*group
Glutamate	H	216.4 [197.7, 235.1]	296.3 [270.2, 322.4]	79.9 [46.22, 113.6]	<0.0001	281.1 [257.0, 305.2]	64.7 [31.67, 97.73]	0.00 04	−15.2 [−49.51, 19.11]	0.660	<0.0001 0.377 0.969
	C	205.4 [185.6, 225.2]	291.0 [262.2, 319.8]	85.6 [45.58, 125.6]	<0.00 01	271.1 [244.4, 297.8]	65.7 [25.93, 105.5]	0.0003	−19.9 [−61.21, 21.41]	0.495
Glutamine	H	401.2 [386.2, 416.2]	442.2 [424.6, 459.8]	41.00 [16.56, 65.44]	0.0003	436.4 [419.9, 452.9]	35.20 [11.12, 59.28]	0.002	−5.80 [−30.48, 18.88]	0.846	<0.0001 <0.0001 0.749
	C	370.7 [359.8, 381.6]	405.1 [393.2, 417.0]	34.40 [12.46, 56.34]	0.0007	395.4 [383.4, 407.4]	24.70 [2.342, 47.06]	0.026	−9.70 [−32.04, 12.64]	0.565
Taurine	H	23.2 [22.3, 24.2]	21.7 [20.8, 22.6]	−1.58 [−2.91, − 0.25]	0.015	21.4 [20.6, 22.3]	−1.80 [−3.11, − 0.49]	0.00 4	−0.22 [−1.56, 1.12]	0.921	<0.0001 <0.0001 0.993
	C	20.8 [20.1, 21.5]	19.3 [18.6, 19.9]	−1.53 [−2.73, − 0.33]	0.008	19.0 [18.3, 19.6]	−1.84 [−3.07, − 0.61]	0.001	−0.31 [−1.54, 0.92]	0.824
Leucine	H	93.4 [89.1, 97.8]	91.0 [86.6, 95.5]	−2.40 [−9.24, 4.44]	0.689	93.2 [88.9, 97.6]	−0.20 [−6.90, 6.50]	0.997	2.20 [−4.69, 9.09]	0.734	0.308 0.005 0.945
	C	89.7 [86.2, 93.2]	86.2 [82.9, 89.6]	−3.40 [−9.68, 2.74]	0.389	88.3 [84.8, 91.8]	−1.39 [−7.69, 4.91]	0.863	2.08 [−4.22, 8.38]	0.719
Isoleucine	H	13.4 [7.9, 18.9]	15.0 [9.3, 20.7]	1.59 [−5.82, 8.99]	0.869	13.8 [8.0, 19.6]	0.43 [−6.84, 7.70]	0.989	−1.16 [−8.08, 5.76]	0.918	0.911 0.011 0.929
	C	9.9 [7.6, 12.2]	9.9 [7.6, 12.3]	0.02 [−6.25, 6.28]	>0.999	9.7 [7.4, 11.9]	−0.23 [−6.57, 6.10]	0.996	−0.25 [−6.60, 6.09]	0.995
Valine	H	83.1 [78.8, 87.4]	80.2 [75.6, 84.7]	−2.92 [−9.15, 3.31]	0.515	83.1 [78.9, 87.2]	−0.03 [−6.16, 6.10]	>0.999	2.89 [−3.39, 9.17]	0.527	0.229 <0.0001 0.830
	C	77.9 [75.1, 80.7]	74.8 [72.1, 77.5]	−3.11 [−8.69, 2.47]	0.391	75.9 [73.1, 78.7]	−2.01 [−7.68, 3.66]	0.683	1.10 [−4.56, 6.76]	0.892

Data are expressed in pmol/min and as means and estimated differences with 95% CI. Comparisons were performed using RM-ANOVA and Tukey’s Multiple Comparisons test. Healthy (H) and COPD (C) subjects. Statistically significant values (p<0.05) are highlighted in bold.

Resistance exercise increased WBP of glutamate (p<0.0001) and glutamine (p<0.0001). The increases observed in WBP glutamate (Figure 1a) and glutamine (Figure 1b) were present at 1-hour post-exercise for both COPD and healthy groups (all p<0.001) and remained elevated at 24 hours (all p<0.05). Resistance exercise did not change the WBP of the individual BCAAs leucine, isoleucine, and valine in both the COPD and healthy groups (Table 3, Figure 2a-2c). The decrease in WBP of taurine (Table 3, Figure 3) remained significant in both cohorts between 1hr (p<0.05) and 24hr (p<0.004) post-exercise.

Fig. 1.

Whole body production (WBP) in response to a high-intensity resistance exercise session of glutamate (left panel) and glutamine (right panel) in healthy and COPD participants. Data are displayed as mean (95%CI). Comparisons between the results at baseline (open white bar), 1-hour post-exercise (striped red bar) and 24-hour post-exercise (open green bar) were performed using RM-ANOVA and Tukey’s Multiple Comparisons test. Statistically significant values are p values <0.05.

Fig. 2.

Whole body production (WBP) in response to a high-intensity resistance exercise session of leucine (left panel), isoleucine (middle panel), and valine (right panel) in healthy and COPD participants. Data are displayed as mean (95%CI). Comparisons between the results at baseline (open white bar), 1-hour post-exercise (striped red bar) and 24-hour post-exercise (open green bar) were performed using RM-ANOVA and Tukey’s Multiple Comparisons test. Statistically significant values are p values < 0.05.

Fig. 3.

Whole body production (WBP) in response to a high-intensity resistance exercise session of taurine in healthy and COPD participants. Data are displayed as mean (95%CI). Comparisons between the results at baseline (open white bar), 1-hour post-exercise (striped red bar) and 24-hour post-exercise (open green bar) were performed using RM-ANOVA and Tukey’s Multiple Comparisons test. Statistically significant values are p values < 0.05.

3.5. Compartmental analysis

In the Supplemental tables, the compartmental data for glutamate (Supplemental Table S2), glutamine (Supplemental Table S4), leucine (Supplemental Table S5), isoleucine (Supplemental Table S6), valine (Supplemental Table S7), and taurine (Supplemental Table S8) are provided. Notably, the COPD participants had smaller compartment sizes for glutamate (Q₁: p=0.036, Q₂: p<0.0001), taurine (Q₁: p=0.012, Q₂: p=0.021), leucine (Q₂: p=0.020), isoleucine (Q₁: p<0.0001, Q₂: p<0.0001) but not for glutamine or valine. Resistance exercise increased the compartment sizes only for glutamate (Q₁: p<0.0001, Q₂: p<0.0001) due to an increase at 1hr post-exercise (both p<0.05) which remained elevated at 24hr post-exercise (p<0.05) in both groups. The fluxes were lower in COPD for glutamine (F₂₁: p<0.05, F₀₂: p<0.05), taurine (F₂₁: p<0.0001, F₀₂: p<0.0001), leucine (F₀₂: p<0.001), isoleucine (F₂₁: p<0.0001, F₀₂: p<0.05) and valine (F₀₂: p<0.05). Resistance exercise increased the fluxes for glutamate (F₂₁: p<0.0001, F₀₂: p<0.0001). F₀₂ only for glutamate remained elevated between 1 and 24 hours post-exercise in both groups (p<0.0001). Fracloss was not different between the groups and did not change in response to exercise.

4. Discussion

In the present study, the high-intensity resistance exercise session immediately increased whole-body productions of glutamate and glutamine, and reduced that of taurine in both COPD and healthy participants. Substantial increases were also found in the accessible and inaccessible glutamate compartment sizes and intracellular glutamate flux in both groups. These changes persisted 24 hours post-exercise. Although WBP of the BCAAs remained unchanged, plasma concentrations of leucine and isoleucine decreased immediately post-exercise specifically in the COPD group. The COPD participants studied were characterized by muscle weakness, low-grade systemic inflammation, and physical inactivity, despite a relatively low prevalence of COPD exacerbations in the preceding year.

4.1. Prolonged increase in whole body and intracellular glutamate production following resistance exercise session

A rapid, substantial (~40%) increase in glutamate WBP persisted 24 hours post-exercise in COPD patients, despite 29–34% less limb work performed, and prolonged exercise induced muscle fatigue was observed in the COPD group only. This response likely reflects a metabolic adaptation to both the heightened energy demands of exercise and the inherent metabolic disturbances characteristic of COPD [31]. COPD is associated with mitochondrial dysfunction and oxidative stress, which may impair energy metabolism [32]. The large increase in glutamate production might reflect a compensatory mechanism to enhance substrate availability for the tricarboxylic acid cycle, thereby supporting ATP production in skeletal muscle. Additionally, glutamate plays a role in the synthesis of glutathione, an important antioxidant, which may help counteract the increased oxidative stress in COPD participants. Glutamate’s role in ATP production via the tricarboxylic acid (TCA) cycle is crucial for meeting energy requirements during exercise, particularly in COPD participants with pre-existing metabolic disturbances [8]. While there was a rise in glutamate WBP, the plasma concentration of glutamate did not exhibit a parallel increase, which suggests enhanced cellular uptake to meet energy needs even though a third less work was performed by the COPD participants. Increased uptake of glutamate has previously been reported in healthy individuals immediately following intense endurance exercise [33]. The enhanced glutamate production could also be a result of increased proteolysis or transamination reactions, reflecting muscle protein breakdown or amino acid re-utilization for energy production [6], which is particularly relevant in COPD participants, who often exhibit muscle wasting and altered protein metabolism.

4.2. Increased glutamine production in response to resistance exercise

Although glutamine WBP was lower in the COPD than in healthy group throughout the study, both groups exhibited an (~10%) increase in glutamine WBP post-exercise, which is likely related to the increased glutamate production in response to exercise as glutamate is for a large part converted to glutamine, mainly in muscle via glutamine synthetase [34,35]. In COPD participants, the increased glutamine production may also serve to support immune function and nitrogen balance, as glutamine is a key substrate for rapidly dividing cells and is involved in nucleotide synthesis [36]. Given that COPD is associated with systemic inflammation, the augmented glutamine production post-exercise might reflect an adaptive response to support tissue repair processes. The consequences of the increased glutamine production after exercise for other roles like the immune system and gut function [33,34] remain unclear.

4.3. Prolonged reduction in taurine production following resistance exercise

Taurine WBP, compartment sizes, and intracellular kinetics were consistently lower in COPD patients. Resistance exercise has been linked to elevated taurine levels in healthy individuals [16,37]. Although we did not find a significant increase in plasma taurine in COPD post resistance exercise as we observed previously in COPD after an endurance exercise [5], we noted an increase in taurine plasma concentration in the healthy participants at 24 hours post-exercise, which may be attributed to the heightened muscle efflux of amino acids observed post-exercise [38]. Taurine is also known for its antioxidant properties and role in calcium homeostasis within muscle cells [17,38,39]. The ~7% reduction in taurine production in COPD participants may contribute to increased oxidative stress and impaired muscle contractility, exacerbating exercise intolerance and muscle fatigue [39]. The inability to elevate taurine production post-exercise in COPD could therefore reflect a diminished adaptive capacity in muscle tissue, possibly due to impaired synthesis pathways or increased utilization of taurine to combat oxidative stress.

Taurine's plasma concentration following resistance exercise did not mirror that of WBP as WBP of taurine decreased immediately post-exercise indicating enhanced intracellular taurine disposal. This may also explain the previously observed reduction in muscle taurine concentration after exercise in COPD [5] indicating enhanced taurine need as increased oxidative stress and compromised redox balance are present in these participants, particularly during exercise [40]. As there is contradictory evidence that muscle taurine levels are impacted by exercise in healthy individuals, more research needs to be done on the location of taurine production/clearance as related to muscle.

4.4. Resistance exercise does not further alter whole-body productions and intracellular fluxes of BCAAs but reduces plasma concentrations in COPD

Overall, WBP and intracellular kinetics of the 3 individual BCAAs were lower in COPD than in healthy participants. Additionally, the compartmental pool sizes of leucine were smaller in COPD. BCAAs are known to serve as a source of ATP during periods of stress [41,42]. Post-exercise, plasma leucine and isoleucine decreased by 16% and 13%, respectively, in COPD despite stable WBP, indicating increased catabolism of leucine and isoleucine in muscle, likely mediated by branched-chain keto acid dehydrogenase, resulting in an increased alanine and glutamine release [11].

This shift towards BCAA catabolism for energy production, driven by impaired carbohydrate and lipid metabolism [48], may contribute to muscle wasting and weakness commonly observed in COPD, as muscle proteins are broken down to supply energy [11,43]. Additionally, the decreased plasma BCAA levels may affect protein synthesis signaling pathways, such as mTOR, potentially impairing muscle protein synthesis and recovery post-exercise [11,44].

4.5. Overall physiological significance of findings

Our findings demonstrate that glutamate plays a crucial role in energy metabolism, particularly in COPD participants with compromised metabolic function. Given glutamate’s pivotal role in the tricarboxylic acid (TCA) cycle, this increased availability is essential for ATP production, potentially helping to offset the impaired oxidative capacity that is characteristic of COPD [15,43]. The enhanced glutamate and glutamine production may be adaptive responses to support energy metabolism and antioxidant defenses, whereas the reduced taurine production and increased BCAA catabolism could contribute to muscle fatigue and wasting. Additionally, the rise in glutamine production that we observed post-exercise supports muscle recovery and immune function, potentially aiding in the recovery process for COPD participants who experience greater exercise-induced muscle fatigue [43,45]. The role of glutamine in maintaining nitrogen balance may also contribute to mitigating muscle breakdown after intense exercise. Therefore, our data suggest that the sustained upregulation of glutamate and glutamine production beyond 24 hours post exercise occurred at the expense of maintaining plasma BCAA levels. This indicates that the enhanced BCAA clearance required to meet the enhanced glutamate and glutamine production, exceeded BCAA production resulting in reduced plasma BCAA levels. This is of importance as BCAAs are needed to promote muscle building by stimulating muscle protein synthesis muscles and suppressing protein breakdown. The reduction in taurine production observed in our study is particularly noteworthy, given taurine’s roles in antioxidant defense and cellular osmoregulation [43]. This decrease in taurine could exacerbate oxidative stress and impair exercise tolerance, resulting in increased muscle fatigue and slower recovery in COPD participants. Collectively, these findings underscore the complex metabolic adaptations in COPD and emphasize the importance of addressing these alterations to improve exercise capacity and functional performance in this population.

4.6. Potential clinical implications of findings

Previous studies have demonstrated that short-term alterations in amino acid metabolism during exercise correlate strongly with, and may predict, long-term changes [46]. The metabolic response to a single exercise session therefore may provide predictive information about the potential benefits of resistance exercise within pulmonary rehabilitation for these participants, and whether adjustments, such as different training intensity or targeted nutritional support, are needed for optimal outcomes. The observed alterations in amino acid metabolism, especially the increased catabolism of BCAAs and reduced taurine production, indicate a need for early interventions to prevent muscle wasting and weakness. Incorporating BCAAs and glutamine into dietary plans may support muscle preservation and recovery via improved protein metabolism and direct mTORC1 activation [47–50]. Previous studies have also shown that increasing systemic glutamine availability through supplementation enhances glycogen synthesis and ammonia clearance [51]. Interestingly, glutamine supplementation in older adult women has a positive effect on glycemic control and oxidative stress reduction [52]. These findings collectively suggest a potential role of glutamine in metabolic regulation and recovery rather than direct enhancement of exercise performance. Whether there is a role for taurine supplementation to protect against antioxidants and enhance exercise tolerance [38,53] remains to be studied in COPD.

The study underscores the importance of metabolic monitoring as part of routine care for COPD participants. The prolonged muscle fatigue experienced by COPD participants post-exercise highlights the need for optimized exercise prescriptions and recovery strategies in pulmonary rehabilitation programs. The complex metabolic changes observed in COPD participants also necessitate a multidisciplinary approach to care, involving collaboration between pulmonologists, dietitians, physiotherapists, and other specialists to address the multifaceted needs of these participants. Comprehensive metabolic profiling on an individual level is important for personalized medicine given the distinct metabolic responses among COPD participants and between males and females [27,28]. Larger studies are needed to examine whether certain clinical characteristics are associated with particular metabolic responses to exercise. This could lead to reduction of metabolic perturbations to exercise rehabilitation on an individual level through personalized approaches such as tailored exercise programs and targeted protein/amino acid supplementation. Particularly, studies in emphysema are of clinical importance as these patients often exhibit reduced muscle glutamate levels and disturbances in glutamate kinetics, and are prone to muscle wasting which is associated with decreased plasma branched-chain amino acid concentrations [8,44]. Participants recovering from an acute exacerbation of COPD (AECOPD), who often experience poor muscle health and exercise impairment, have recently been shown to exhibit disturbances in plasma amino acids levels (elevated glutamine, glutamate and reduced BCAA levels) compared to stable COPD participants [54]. A combined score of these amino acid disturbances was associated with an increased risk of acute exacerbation of ventilatory function in COPD. Therefore, participants with emphysema and those recovering from AECOPD might be at high risk for exercise induced perturbations in glutamate related metabolism, which warrants targeted nutritional modulation particularly during exercise rehabilitation programs. By targeting specific disturbances in metabolic pathways, personalized nutritional and exercise interventions may enhance muscle function, reduce oxidative stress, and improve overall exercise tolerance.

4.7. Limitations of the study

This study assessed amino acid metabolic markers at only three time points (pre-exercise, 1 hour, and 24 hours post-exercise), limiting insights into long-term (>24 hours) metabolic changes. While most published studies so far have examined muscle fractional synthesis rates (FSR) in response to high-intensity exercise in healthy subjects up to 24 hours post exercise, we anticipated that metabolic responses in COPD patients would have been normalized within 24 hours due to lower exercise intensity. Moreover, the study’s design and participant burden precluded extended assessments. Future longitudinal studies with later time points are needed to fully capture long-term metabolic dynamics in COPD which will increase the understanding of how these metabolic alterations evolve and resolve. Randomizing study order was not feasible given the exercise-response focus, and whole-body tracers for a muscle-specific stimulus may not fully reflect muscle-specific metabolism, potentially confounded by non-muscle tissue contributions. Combining muscle measurements with advanced isotopomics could refine these insights, though multiple biopsies within 24 hours may be too invasive for this population at risk of muscle wasting and dysfunction. Plasma amino acid concentrations remained stable during the recovery phase of exercise (Supplemental Figure 2), suggesting a steady state of the tracee. No residual label above the background from previous label administration was observed, in line with our previous studies [23,55]. Important to note is that background enrichment was subtracted from the enrichment measured in the samples after pulse administration. Due to the pulse design, it was not possible to determine whether the WBP was in a steady state throughout the 2 hour measurement periods. However, since WBP is largely influenced by the area under the curve (AUC) in the first 10 min after pulse administration, our data primarily reflect WBP during this time frame. We lack information on WBP dynamics after the 1 hour period as this would have required an additional isotope pulse. No prior power analysis was conducted due to the novel multitracer pulse method and lack of prior data, potentially affecting interpretation of non-significant findings. Although the used sample size aligns with metabolic isotope study norms, larger cohorts could enhance generalizability. Studies in the fed-state may explore exercise-nutrition interactions.

Despite these limitations, the novel stable isotope pulse approach to assess the metabolic kinetic response to exercise offers strengths, enabling simultaneous measurements of multiple metabolites and pathways without priming or extensive sampling, and using small isotope doses within a 2-hour window. The compartmental analysis allows for the measurement of intracellular kinetics, which is particularly important because whole body appearance rates in plasma do not always accurately reflect intracellular appearance rates [19,56].

5. Conclusions

Our study demonstrated that a single session of resistance exercise already induces substantial and long-lasting metabolic adaptations in COPD—such as increased glutamate production and reduced plasma BCAA levels. These changes as well as the previously observed metabolic disturbances in response to endurance exercise [5,8,9] should be considered when developing a training protocol as part of pulmonary rehabilitation. Further research is warranted on the potential impacts of targeted amino acid modulation to restore pre-existing and exercise-induced metabolic perturbations so that nutritional deficiencies will be reduced and the physiological response to exercise improved in COPD. The ultimate goal is to enhance the effectiveness of pulmonary rehabilitation among COPD participants and reduce the incidence of non-responders to rehabilitation programs, thereby positively influencing the quality of life of these participants.

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Abbreviations:

BCAA

Branched-Chain Amino Acid(s)

CI

Confidence Interva

COPD

Chronic Obstructive Pulmonary Disease

DXA

Dual-Energy X-ray Absorptiometry

FEV1

Forced Expiratory Volume in 1 Second

FVC

Forced Vital Capacity

GOLD

Global Initiative for Chronic Obstructive Lung Disease

hs-CRP

High Sensitivity C-Reactive Protein

LC-MS/MS

Liquid Chromatography–Triple Quadrupole Mass Spectrometry

TCA

Tricarboxylic Acid

TTR

Tracer to Tracee Ratio

WBP

Whole Body Production

Data availability

The dataset presented in this article are not readily available because they are part of an ongoing study. Requests to access the datasets should be directed to the corresponding author up-on reason able request.