Author Identifiers

Georgios Mavropalias
ORCID: 0000-0001-7753-5693

Date of Award

2020

Degree Type

Thesis

Degree Name

Doctor of Philosophy

School

School of Medical and Health Sciences

First Advisor

Professor Ken Nosaka

Second Advisor

Professor Anthony Blazevich

Abstract

Both the acute damage and resulting adaptations induced by eccentric exercise have been well documented, but less is known about their underpinning mechanisms. It is possible that muscle extracellular matrix (ECM), including both collagenous and costameric (e.g. integrins) components, plays a major role in both damage and adaptation, but this has not been extensively investigated. Eccentric cycling (ECC) was chosen as a model to investigate these mechanisms, since it consists only of eccentric muscle actions, and can induce typical characteristics of muscle damage such as prolonged decreases in muscle function and delayed onset muscle soreness (DOMS); while ECC training has been shown to induce increases in muscle function and muscle mass, even at low intensities. The purpose of this PhD project was to investigate the muscle damage and adaptations induced by the ECC in relation to collagen breakdown and integrin signalling pathways, which may shed light on the connection between ECM-mediated muscle changes.

This research project consisted of three studies. In Study 1, high- (HI) and low-intensity (LI) ECC with the same mechanical work were compared for changes in muscle function and DOMS, while the repeated bout effect was examined after subsequent HI ECC. Eleven men performed HI-ECC (1 min × 5 at 20% of peak power output: PPO) for two bouts separated by 2 weeks (H-H), and nine men performed LI-ECC (4 min × 5 at 5% PPO) for the first bout and then HI-ECC for the second bout (L-H). At 24 h after the first bout, both groups showed similar decreases in maximal isometric (70° knee angle, -10.6±11.8%) and isokinetic (-11.0±8.2%) knee extensor (KE) torque, and squat (-7.7±10.4%) and counter-movement (-5.9±8.4%) jump heights (P<0.05). Changes in KE torque and jump heights were smaller after the second than first bout for both groups (P<0.05). Increases in plasma creatine kinase (CK) activity were small, and no significant changes in vastus lateralis (VL) or intermedius thickness nor ultrasound echo-intensity were observed in any group or timepoint. KE soreness during palpation was greater (P<0.01) in H-H (peak: 4.2±1.0) than L-H (1.4±0.6) after the first bout, but greater in L-H (3.6±0.9) than H-H (1.5±0.5) after the second bout. The greater DOMS after HI-ECC than LI-ECC may hypothetically result from greater ECM damage and inflammation.

In Study 2, plasma CK activity, hydroxyproline (Hyp) and cell-free DNA (cfDNA) concentrations were measured before, immediately after and 24 – 72 h after HI-ECC and LI-ECC, and examined in relation to changes in maximum voluntary isometric contraction strength (MVIC) and DOMS. The participants were the same as those in Study 1. Plasma CK activity increased at 24 h (135%), and Hyp concentration increased (40–53%, P<0.05) at 24–72 h post-ECC without differences between groups. cfDNA increased immediately after only for HI-ECC (2.12-fold, P<0.001) but not after LI-ECC, with a significant difference between the groups (P=0.0017). DNA-methylation analysis indicated that the detected cfDNA was not derived from skeletal muscle. In relation to the MVIC and DOMS changes shown in Study 1, no significant correlations were observed between the magnitude of changes in CK, Hyp, cfDNA, MVIC and DOMS. However, significant correlations were found between the cfDNA increase at immediately post-ECC and peak heart rate during ECC (r=0.513, P=0.025). These results suggest that the increased plasma cfDNA concentration after HI-ECC was not associated with muscle damage but with physiological load. Increases in plasma hydroxyproline concentration, which were similar between groups, indicate increased post-ECC collagen turnover.

In Study 3, changes in the expression of integrin – integrin-linked kinase (ILK) – rapamycin-insensitive companion of mTOR (RICTOR) protein complex in VL before and after 8 weeks of HI-ECC training were examined along with possible associations to changes in muscle function and VL cross-sectional area (CSA) from pre- to post-training. Eleven young men completed 24 sessions of ECC with progressive increases in intensity and duration, resulting in a 2-fold increase in work from the first 3 sessions (75.4±14.1 kJ) to the last 3 sessions (150.7±28.4 kJ). Lower-limb lean mass (dual-energy X-ray absorptiometry), VL CSA (extended field-of-view ultrasonography), lower-limb static strength measured on the ergometer (breaking strength against the force provided by the crank pushing the foot), peak and average cycling power output were measured, and VL micro-biopsy samples were obtained before and after training. Significant (P<0.05) increases in integrin-β1 (1.64-fold) and RICTOR (2.99-fold) expressions as well as the phosphorylated-to-total ILK1 ratio (1.70-fold) were observed from pre- to post-training. Lower-limb, thigh, and trunk lean mass increased (2.8–5.3%, P<0.05), and CSA increased (13.3±9.0%, P<0.001) from pre- to post-training. Increases in static strength (18.1±10.8%) and both peak (8.6±10.5%) and average power output (7.4±8.3%) were also evident (P<0.05). However, no significant Pearson (r) or Spearman (ρ) correlations were evident between the increases in integrin-β1D or RICTOR expression and increases in CSA (r=-0.184, ρ=-0.327), static strength (r=-0.272, ρ=-0.055) or PPO (r=-0.106, ρ=-0.391) were observed. These results suggest that ECC training increased the integrin-ILK-RICTOR expression, but the increases were not associated with the post-training increases in muscle function or CSA.

The findings from the three studies provide new insights into the mechanisms underlying muscle damage and adaptations induced by eccentric exercise. The Study 1 results showed that even low-intensity eccentric muscle actions affected the ECM, as indicated by DOMS, which has been shown by previous studies to be more associated with connective tissue damage and inflammation. ECC did not induce extensive muscle fibre damage, as indicated by the small increases in plasma CK activity. It is interesting that the increases in plasma Hyp concentration were similar between HI-ECC and LI-ECC, despite a significant difference in DOMS magnitude. ECC training increased integrin-ILK-RICTOR expression, but it was not associated with the increased CSA or muscle function. It seems possible that the ECM is involved in muscle damage and adaptations, but this was not endorsed by the present studies. It is necessary to investigate further histological changes in ECM after the initial and repeated bouts of eccentric exercise, how eccentric exercise induces ECM remodelling, and how this affects changes in muscle architecture including muscle CSA or mass and muscle function.

Access Note

Chapters 2, 3, 4 and 5 are not available in this version of the thesis.

Available for download on Sunday, August 24, 2025

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