# Metabolic Reprogramming for Body Adaptation and Inflammatory Control in Eccentric Damaging Exercise: Comprehensive Molecular Insights From Repeated Downhill Running

Amir Mohammad Malvandi, Gianluca Vernillo, Veronica Sansoni, Martina Faraldi, Chiara Verdelli, Giuseppe Coratella, Giorgio Varesco, Vianney Rozand, Krystian Wochna, Laurent Mourot, et al.

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Amir Mohammad Malvandi, Gianluca Vernillo, Veronica Sansoni, Martina Faraldi, Chiara Verdelli, et al. . Metabolic Reprogramming for Body Adaptation and Inflammatory Control in Eccentric Damaging Exercise: Comprehensive Molecular Insights From Repeated Downhill Running. MedComm, 2025, 6 (11), 10.1002/mco2.70480. . hal-05459975

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LETTER OPEN ACCESS

# Metabolic Reprogramming for Body Adaptation and Inflammatory Control in Eccentric Damaging Exercise: Comprehensive Molecular Insights From Repeated Downhill Running

Amir Mohammad Malvandi¹ | Gianluca Vernillo² | Veronica Sansoni¹ | Martina Faraldi¹ | Chiara Verdelli¹ |
Giuseppe Coratella² | Giorgio Varesco³ | Vianney Rozand⁴ | Krystian Wochna⁵ | Laurent Mourot⁴ |
Giovanni Lombardi¹,⁵

¹Laboratory of Experimental Biochemistry & Advanced Diagnostics, IRCCS Ospedale Galeazzi-Sant'Ambrogio, Milano, Italy | ²Department of Biomedical Sciences For Health, Università Degli Studi di Milano, Milano, Italy | ³Department of Medicine, Université de Montreal, Montreal, Canada | ⁴Université Marie et Louis Pasteur, SINERGIES, Besançon, France | ⁵Chair of Sport Kinesiology, Poznań University of Physical Education, Poznań, Poland

**Correspondence:** Amir Mohammad Malvandi (Amirmohammad.Malvandi@grupposandonato.it)

**Received:** 30 December 2024 | **Revised:** 2 October 2025 | **Accepted:** 14 October 2025

Dear Editor,

Exercise triggers metabolic adaptations that enhance cellular function and resilience, primarily through energy-demanding processes. It stimulates key pathways like glycolysis, the tricarboxylic acid cycle (TCA), and oxidative phosphorylation, which are vital for muscle contraction and recovery. Studies on specific metabolites indicate that aerobic exercise enhances oxidative metabolism, whereas anaerobic and resistance exercises increase lactate production and stimulate muscle protein synthesis. Eccentric exercise, such as downhill running, leads to unique metabolic changes, marked by increased muscle damage markers such as creatine kinase (CK), myoglobin (MB), and lactate dehydrogenase (LDH); and inflammation (Cox-2, iNOS). Although targeted studies have made progress, understanding the overall molecular dynamics of eccentric exercise remains limited [1]. The liquid chromatography with high-resolution mass spectrometry (HRMS)-based untargeted total small-molecule profiling provides detailed insights into these changes [2], potentially guiding strategies to reduce muscle damage and enhance recovery [3].

This study investigated the biochemical and metabolic responses to repeated bouts of eccentric exercise, specifically downhill running, revealing distinct molecular trends between the first and second bouts (Figure 1A). The experimental protocol was effective in inducing muscle damage, as evidenced by a significant

reduction in maximum voluntary isometric contraction (MVIC) 24 h post-exercise (Figure 1A). These data were confirmed at the molecular level, as a notable increase in CK activity and MB concentrations was observed after exercise, with a significant reduction in the second bout (Figure 1B), indicative of reduced muscle damage—a hallmark of the repeated bout effect (RBE). LDH activity increased immediately postexercise in both bouts, with a faster recovery observed in the second bout, suggesting reduced cellular stress and faster recovery during the second bout (Figure 1B). Inflammatory markers showed minimal changes, with IL-6 briefly detected postexercise and TNFα levels remaining stable (Figure 1B), indicating a controlled inflammatory response potentially influenced by prior adaptation [4].

High-resolution small-molecular profiling revealed profound metabolic shifts associated with exercise-induced fatigue and recovery (Figure 1C). In this study, we focused on comparing *immediate pre-* and postexercise samples across both bouts, which demonstrated distinct metabolic responses, underscoring the adaptive nature of repeated eccentric stress. The volcano plot analysis identified significant metabolites that underwent substantial changes postexercise (Figure 1C), with pathway enrichment analysis revealing key metabolic shifts. The first bout was characterized by alterations in linoleate metabolism, hexose phosphorylation, fatty acid (FA) activation, aminosugar metabolism, and de novo FA biosynthesis (Figure 1C). These

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© 2025 The Author(s). *MedComm* published by Sichuan International Medical Exchange & Promotion Association (SCIMEA) and John Wiley & Sons Australia, Ltd.
<figure><img src="image_3.png" /><figcaption>FIGURE 1 | Legend on next page.</figcaption></figure>
pathways reflect an acute metabolic response involving energy
utilization, lipid metabolism, and structural remodeling to coun-
teract exercise-induced damage.

The second bout continued to exhibit some of these metabolic
perturbations. Still, it introduced new adaptive pathways com-
pared with the first bout, including Omega-3 FA metabolism,
protein metabolism, and saturated FAs β-oxidation (Figure 1C),
revealing metabolic rewiring states in the body. The emergence
of Omega-3 FA metabolism highlights an increased reliance
on anti-inflammatory and membrane-stabilizing mechanisms,
consistent with a protective adaptation that mitigates muscle
damage. The activation of protein metabolism observed in serum
may indirectly reflect enhanced muscle remodeling and turnover
(Figure 1C); however, this interpretation should be made with
caution, as serum-based measurements do not directly repre-
sent changes occurring within skeletal muscle tissue [5]. The
upregulation of saturated FAs β-oxidation further underscores
a shift toward lipid-based energy metabolism, a characteristic
adaptation in repeated exercise bouts to optimize endurance and
recovery efficiency [6].

A detailed comparison of serum molecular profiles before and
after both exercise sessions revealed a distinctive pattern of
molecular dynamics resulting from the RBE. Enrichment analy-
sis of metabolic pathways indicated linoleate metabolism as the
most significantly modified, followed by the carnitine shuttle,
glycerophospholipid metabolism, 3-oxo-10R-octadecatrienoate β-
oxidation, and FA activation. These findings highlight the
progressive reprogramming at the systemic level of metabolic
networks, favoring lipid metabolism and energy efficiency upon
repeated exposure to eccentric exercise stress.

Examining the participants' serum molecular dynamics pro-
vides deeper insights into these adaptations. The persistence of
alterations in pathways such as 3-oxo-10R-octadecatrienoate β-
oxidation and FA activation suggests a sustained demand for
lipid metabolism, possibly reflecting a long-term adaptation to
eccentric exercise. The detection of ethyl-2-hydroxy-3-(3-indolyl)
propanoate glucoside, a derivative of tryptophan metabolism,

underscores the interplay between muscle function and central
nervous system activity (Figure 1D). Tryptophan metabolism
can influence serotonin synthesis, thereby linking metabolic
adaptations to the regulation of neuromuscular fatigue and
psychological stress resilience.

Additionally, the increased levels of LysoPE (0:0/20:0) immediately after exercise in serum may indirectly indicate the role of lipid metabolism in preserving muscle membrane integrity and facilitating post-exercise recovery. LysoPEs contribute to membrane remodeling and generate signaling molecules essential for managing inflammation and muscle repair (Figure 1D). The upregulation of N-[(4-hydroxy-3-methoxyphenyl)methyl]octanamide, associated with the phenylpropanoid pathway, can suggest an interaction between muscle activity and gut health, reinforcing the concept of a gut-muscle axis influencing recovery and metabolic balance.

Further, our systemic molecular profiling revealed the possi-
ble presence of exogenous metabolites in participants' serum,
such as 2-benzamido-N-(1-[(naphthalen-1-yl)amino]-1-oxobutan-
2-yl)benzamide, N-(3-acetamidopropyl)pyrrolidin-2-one, α- or
β-triticene, and piperidine, indicating potential alterations in
gut permeability induced by exercise. These changes high-
light the comprehensive impact of eccentric exercise on the
entire body, as metabolic shifts extend beyond muscle tissue
to encompass broader physiological systems. The elevation
of N(2)-phenylacetyl-L-glutamate reflects active detoxification
pathways, highlighting the body's effort to clear metabolic waste
products generated during physical exertion [7].

These findings suggest that repeated eccentric exercise pro-
motes a reorganization of metabolic pathways, favoring lipid
metabolism, efficient energy utilization, and enhanced recovery
processes at the systemic body level. The identified metabolites
and pathways provide valuable serum biomarkers for monitoring
exercise adaptation and recovery. The shift toward Omega-3 FA
metabolism and the persistence of lipid-based energy utilization
can potentially be targeted by nutritional interventions, such
as Omega-3 supplementation, that could further enhance these

**FIGURE 1** | (A) Experimental design, sampling time-points, and analysis. The exercise consisted of two downhill bouts of running (30 min; 10 km·h⁻¹; -20%) interspersed by 4 weeks. On the right side of this part, the graph presents the maximum voluntary isometric contraction (MVIC) measurements before (pre) and at different time-points after the bouts. Green represents the first bout, and orange represents the data for the second bout. (B) Serum biochemical evaluations of the participants. Data are presented in mean ± SD for the first bout in green squares, and the orange state for the second bout. (C) Global molecular profiling of serum of two consecutive eccentric damaging exercise bouts; at left of this panel, heatmap illustration of clustered (Euclidean-Ward) identified molecular features in the group (first is first bout, second is second bout). Data are presented as the mean of all participants in each group. On the upper right, Vulcano plots of fold change (logarithmic) of molecular feature against *p*-value (logarithmic) for the first and second bouts (post vs. pre); GSEA-Mummichog integrated metabolic pathway enrichment analysis for the first bout. Levels of top hits identified metabolites for the bout. At the bottom of the central panel, top hits resulting from ANOVA are presented, demonstrating the metabolic dynamics of body adaptation to the repeated bout of eccentric damaging exercise. At the bottom right, the enrichment analysis, performed using the Mummichog algorithm, is presented. (D) Adaptive response to damaging eccentric exercise. This panel shows molecules that are observed to reduce or increase in participants after 1 month from the first bout of damaging eccentric exercise, returning to initial levels after the second bout of exercise. This adaptive response can indirectly help muscle regeneration and overall body resilience. (6R,7S)-Epoxy-TD, (6R,7S)-epoxy-tetradecadienoic acid; 2-PBA, 2-Phenylbutyric acid; BNABB, N-benzoyl-N-(4-aminobutyl)benzamide; Cis4-10:1 MG(10:0), Cis-4-decenoic acid monoglyceride (10:0); CK, creatine kinase; EHIPG, ethyl 2-hydroxy-3-(3-indolyl)propanoate carboxylate; GHP, glutathione hydroperoxide; GLA, gamma-linolenic acid; HMPMO, N-(3-hydroxyphenyl)malonamic acid; HRMS, high-resolution mass spectrometry; IL-6, interleukin-6; LDH, lactate dehydrogenase; LysoPE (20:0/0:0), lysophosphatidylethanolamine with a 20:0 fatty acid chain and no second fatty acid chain; LysoPE, lysophosphatidylethanolamine; MB, myoglobin; N-AC-LP amid, N-acetyl-L-cysteine-lipid amid; NAE, N-acylethanolamine; OPA, (Z)-2-octylpent-2-enedioic acid; PhAc-L-Glu, phenylacetic acid-L-glutamate; TNFα, tumor necrosis factor-α. Created in BioRender. Lombardi (2025). https://BioRender.com/b2hkbpg.
adaptations. Moreover, the observed gut-derived metabolites
highlight the importance of considering gut health in designing
personalized recovery strategies [8].

Considering key and significantly observed molecules from our
analysis, we can hypothesize and suggest that enzymes linked
to observed exercise-induced metabolic perturbations are poten-
tial new serum biomarkers for assessing physiological stress
and tissue damage from intense physical activity. Tryptophan
hydroxylase (TPH), indoleamine 2,3-dioxygenase (IDO), and
tryptophan 2,3-dioxygenase (TDO), associated with ethyl 2-
hydroxy-3-(3-indolyl)propanoate glucoside and tryptophan, may
reflect disruptions in serotonin and kynurenine pathways during
prolonged exercise, which are linked to fatigue, inflammation,
and oxidative stress. Lysophospholipase (LYPLA) and phospho-
lipase A2 (PLA2), tied to LysoPE (0:0/20:0), could indicate
membrane phospholipid breakdown in muscle or erythrocytes
under mechanical or oxidative strain. The detoxification enzymes
catechol-O-methyltransferase COMT and uridine diphosphate-
glucuronosyltransferases (UGT), involved in metabolizing N-
[(4-Hydroxy-3-methoxyphenyl)methyl]octanamide and 4',7-Di-
O-methylcatechin, may highlight catecholamine clearance and
antioxidant responses critical to recovery. Glutamine synthetase
(GS) and glutaminase (GLS), linked to N(2)-phenylacetyl-L-
glutamate and glutamylhydroxyproline, could signal shifts in
nitrogen balance and collagen turnover during muscle repair
or catabolism. Enzymes like Acyl-CoA dehydrogenase (ACAD)
and enoyl-CoA hydratases (ECH), acting on cis-4-Decenoic acid,
might reflect mitochondrial dysfunction in FA oxidation during
endurance overload, while FA desaturase 2 (FADS2)/elongase
gamma-linoleic acid) could denote altered anti-inflammatory
lipid mediator synthesis. Histidine decarboxylase (HDC) and
histidase, which are linked to histidine, may contribute to
histamine-driven inflammation or impaired carnosine synthesis
in muscle fatigue. Lastly, monoglyceride lipase (MGLL), linked
to MG (18:0/0:0/0:0), may indicate dysregulated lipolysis during
energy depletion. These enzymes provide a multifaceted serum
biomarker panel to assess metabolic strain, tissue damage, and
recovery needs in response to damaging exercise, facilitating
the optimization of training loads or the mitigation of injury
risks.

In conclusion, this study presents a high-resolution systemic
molecular landscape of metabolic adaptation to repeated eccen-
tric exercise. The findings reinforce the role of lipid metabolism
in facilitating recovery, reducing muscle damage, and improving
exercise efficiency. Understanding these molecular adaptations
provides a foundation for optimizing exercise strategies, recovery
interventions, and the development of potential biomarkers for
monitoring of body physiological status. Our findings reflect
systemic circulating signatures, their temporal proximity to mus-
cle contraction, and the dominant role of muscle in exercise
metabolism, supporting a biologically meaningful link to mus-
cular responses. However, since our conclusions are limited by
serum molecular profiling at a relatively small cohort, further
research incorporating direct muscle tissue analysis and larger
cohorts is needed to expand on these insights and establish
broader applications of these findings.

Author Contributions

A.M.M. designed the project, performed the experiments, analyzed the data, and wrote the manuscript. G.V. designed the project, performed the experiments, and edited the manuscript. V.S., M.F., and C.V. performed biochemical experiments. G.C., G.V., V.R., K.W., and L.M. performed the exercise experiments. G.L. designed the project, edited the manuscript, and supervised the project. All authors have read and approved the final manuscript.

Acknowledgments

The authors thank the participants for their time and effort and Léo
Vanini's valuable technical assistance during the data acquisition. We
express our gratitude for the kind contributions from the Laboratory of
Analysis of Galeazzi-Saint Ambrogio Hospital, Milan, Italy.

Open access funding provided by BIBLIOSAN.

Funding

This research was supported by the Ultra-Endurance Sports Science & Medicine Grant Program. This work was supported by the Italian Ministry of Health "Ricerca Corrente."

Ethics Statement

The institutional ethical committee approved the study (RCB number ID-RCB: 2019-A03012-55). All participants were informed and provided consent to participate in this study. The protocol was designed in compliance with the Helsinki Declaration.

Conflicts of Interest

The authors declare no conflicts of interest.

Data Availability Statement

The Supporting Information for this work, which is available on the journal's website, provides a detailed description of our study design and the Materials and Methods used. The raw data of HRMS analysis are available online at https://doi.org/10.5281/zenodo.14204023.

References

1. M. Hargreaves and L. L. Spriet, "Skeletal Muscle Energy Metabolism During Exercise," *Nature Metabolism* 2, no. 9 (2020): 817-828.

2. A. M. Malvandi, E. Halilaj, M. Faraldi, et al., "Enhanced Molecular Release From Elderly Bone Samples Using Collagenase I: Insights Into Fatty Acid Metabolism Alterations," *Journal of Translational Medicine* 22, no. 1 (2024): 143.

3. Y. He, Y. Zhang, J. Lai, et al., "Plasma Metabolomics Dataset of Race-Walking Athletes Illuminating Systemic Metabolic Reaction of Exercise," *Scientific Data* 12, no. 1 (2025): 448.

4. A. Nedergaard, M. A. Karsdal, S. Sun, and K. Henriksen, "Serological Muscle Loss Biomarkers: An Overview of Current Concepts and Future Possibilities," *Journal of Cachexia, Sarcopenia and Muscle* 4, no. 1 (2013): 1-17.

5. R. Sartori, V. Romanello, and M. Sandri, "Mechanisms of Muscle Atrophy and Hypertrophy: Implications in Health and Disease," *Nature Communications* 12, no. 1 (2021): 33.

6. T. D. Heden, J. M. Johnson, P. J. Ferrara, et al., "Mitochondrial PE Potentiates Respiratory Enzymes to Amplify Skeletal Muscle Aerobic Capacity," *Science Advances* 5, no. 9 (2019): eaax8352.

7. J. A. Hawley, S. C. Forster, and E. M. Giles, "Exercise, the Gut Microbiome and Gastrointestinal Diseases: Therapeutic Impact and Molecular Mechanisms," *Gastroenterology* 169, no. 1 (2025): 48-62.
8. S. Sato, K. A. Dyar, J. T. Treebak, et al., "Atlas of Exercise Metabolism Reveals Time-Dependent Signatures of Metabolic Homeostasis," *Cell Metabolism* 34, no. 2 (2022): 329-345.

**Supporting Information**

Additional supporting information can be found online in the Supporting Information section.

[Supplement information file 1](<https://www.example.com>) "mco270480-sup-0001-SuppMat.docx"