Participants

Twenty-four healthy but sedentary individuals (5 men, 19 women) volunteered to participate in the study. Participants were recruited from a convenience sample of staff and community members in and around Edith Cowan University (Perth, Australia), which contributed to the predominance of female participants. Participant activity was assessed using the international physical activity questionnaire (IPAQ) and the total activity time for each participant was converted to MET-minutes per week (Committee 2005). Those who were categorized as inactive or minimally active, without neuromuscular or musculoskeletal disorders, injuries, or other medical conditions that could prevent safe exercise participation, and were not on medication that could influence exercise capacity, were accepted into this study. This study was approved by the Edith Cowan University Human Research Ethics Committee (approval number: 2021–03011-KIRK) and conformed to the Declaration of Helsinki. Participants gave their written informed consent and confirmed that they were physically capable of participating in the study, including the testing procedures.

Twenty-four participants commenced the study, of which two (1 male, 1 female) withdrew due to work and personal reasons unrelated to the study. Thus, the final sample size was 22. The sample size was based on the study by Katsura et al. (2019), which reported large effect sizes (>0.8) for changes in outcome measures similar to those of the present study after 8 weeks of bodyweight eccentric exercise training. The final sample size (n=22) was considered to be adequate to detect possible changes in the outcome measures in the present study. Participants were 50 ± 10 (mean ± SD, range: 32–69) years old, with a height of 174.9 ± 1.8 (172.5–176.5) cm and body mass of 82.7 ± 9.0 (69.9–90.4) kg for men and 165.8 ± 5.9 (157–182) cm and 76.5 ± 13.2 (50.9–104.5) kg for women, respectively.

Study design

Participants were familiarized with all testing procedures prior to study commencement in a single session which included demonstration and practice of the physical fitness tests described below. Upon study commencement, participants completed a 2-week control period followed by a 4-week daily exercise intervention period. The 2-week control period was used to mitigate a possible learning effect on outcome measures that is regularly observed in physical fitness tests (Hopkins 2000). Outcome measures described below were taken at three time points, before (PRE-1) and after the control period (PRE-2) as well as after the 4-week intervention (POST) between 1 and 2 days after the last training session. At each testing session, participants were asked to arrive fasted for blood tests and body composition measurement.

To monitor daily activity, each participant was provided with a Charge 5 FitBit watch (Fitbit, San Francisco CA, USA) to wear for the study’s duration. Each participant was given a unique email address, which was used by the investigator to track daily steps. Participants were instructed not to change their physical activity other than performing the exercise program provided. They were also asked not to change their diet during the experimental period.

Exercise program

Following the 2-week control period, participants completed a low-intensity exercise program (Table 2), adapted from the study by Katsura et al. (2019), in which the eccentric phase of each movement was emphasized by modulating movement velocity. Each day, participants completed one set of chair squat, chair recline, wall push-up, and heel drop exercises for 10 repetitions each using a 5-s eccentric (lowering) phase with 1-s concentric (raising) phase. Participants were allowed to choose the time at which they completed the exercises, and the exercises could be performed together or spread throughout the day. Once 10 repetitions could be easily completed for two consecutive exercise sessions (i.e., the exercise provided an RPE score: < 5/10), participants were advised to progress to a more difficult version of the exercise, which then replaced the previous exercise. Several versions of each exercise were provided to participants that progressed in difficulty (Table 2).

Before program commencement, participants were familiarized with the four initial exercises as well as their progressions and a sheet describing the exercises was provided (the program exercises can be found in Online Resource 1). Participants also performed the first exercise session under supervision to ensure that correct technique was used. A password-protected online spreadsheet was used by the participants to log their exercise sessions. This spreadsheet could be accessed by the lead investigator, who monitored exercise adherence throughout the study. All participants were followed up via email after the first week of training to check for any issues or uncertainties surrounding the exercise program.

Body composition, heart rate, and blood pressure

Participants had their height and body mass recorded using a stadiometer and a calibrated scale, respectively. A whole-body dual-energy absorptiometry (DEXA, Hologic Discovery X, USA) scan was then performed to assess body composition (e.g., lean body mass (LBM), fat mass, and relative body fat percentage). Participants were asked to lay supine on the center of the bed ensuring all body parts were inside the scanning zone. Upon scan completion, participants had their heart rate (HR), systolic blood pressure (SBP), and diastolic blood pressure (DBP) measured by automatic sphygmomanometer (HEM-7122; Omron Healthcare Co., Ltd, Japan).

Physical fitness testsIsometric mid-thigh pull (IMTP)

The IMTP was used to assess maximal lower body strength (Comfort et al. 2019). Each participant was asked to stand on a set of Pasport force plates (Pasco, Roseville, USA) (one under each foot) attached to a portable IMTP rig (Vald, Newstead, Australia). To assume the correct pull position, each participant was instructed to stand close to the bar so that the thighs made contact, bend the knees while keeping an upright torso, and to grip the bar (with the aid of lifting straps). Minor adjustments were made to the rack height and participant position so that the bar was between the mid-point of the thigh and the iliac crest with the hips and knees slightly bent (125º–145º and 120º–150º, respectively) and the torso upright (Haff et al. 2015; Beckham et al. 2018) to ensure the strongest force output was produced. A warm-up consisting of 3-s pulls at 50%, 75%, then 90% of perceived maximum effort was performed, in which participants were instructed to “stand up” while maintaining grip of the bar (thus producing torque at both the knee and hip). Attempts were separated by 1 min of rest. After the warm-up, participants completed 3 maximal attempts, with a 1-min rest between efforts. The average of the three attempts was used for analysis (Dos' Santos et al. 2017; Comfort et al. 2019).

Squat (SJ) and countermovement jump (CMJ)

Each participant was instructed to place their hands on their hips with feet hip-width apart while standing on a force platform (Pasport Force Platform PS-2141; Pasco scientific, CA, USA). For the SJ, participants were instructed to crouch down to a self-selected squat depth (Petronijevic et al. 2018). Once in position, a countdown of “3, 2, 1 jump” was given and on “jump,” participants jumped upwards as high as possible and landed back onto the force platform (Kotani et al. 2022). A 3-s hold of the bottom position was used to eliminate any countermovement. If a countermovement was visually observed by the researcher, the repetition was repeated. For the CMJ, participants were instructed to freely flex the hip, knee, and ankle joints while keeping the hands on the hips. Participants were given three maximal attempts in each jump and the best score was used for analysis. The rest time between jumps was 20 s.

Handgrip strength (HG) test

Each participant was asked to stand and hold a hand grip dynamometer (Jamar Smart Hand Dynamometer; Patterson Medical, IL, USA) with the shoulder adducted and neutrally rotated, elbow extended at ~180° and the wrist between 0° and 30° extension and between 0° and 15° ulnar deviation (Innes 1999). Once in position, the participant was asked to apply maximal force for 3 s and the maximal force measured in kg was recorded (Innes 1999). Three trials for each hand were completed, alternating hands and allowing 30 s between attempts. The best of the three attempts for each hand was used for further analysis.

Push-up endurance test

Participants were given 2 min to perform as many push-ups as possible. For men, push-ups were conducted on the hands and feet and for women, push-ups were completed on the hands and knees. Participants were instructed to maintain a ~180º hip angle, with hands spaced shoulder width apart and aligned with the participants eyes. In each repetition, the body was lowered so that a 90° angle formed between the upper and the lower arm at the elbow and with the hands aligned with the chest before returning to the starting position. Participants were reminded to maintain a flat back and to avoid flexion or extension at the lumber spine throughout the test. Where correct technique was not adhered to, the repetition was not counted as valid. The test was terminated if any body part other than the feet (or knees) and hands contacted the ground, if the technique faltered following a warning, or upon the participant’s own volition. The number of push-ups successfully completed at test end was recorded and used for analysis.

Sit-up endurance test

Participants were instructed to assume a supine position with the knees bent to 90°. During each repetition, the participants hands ran along the top of the upper leg until the fingertips reached the kneecap, before returning to the start position. Participants were instructed to avoid pulling on their clothes to pull the torso upward and to avoid sudden movement of the neck, shoulder, and hips to generate momentum. One repetition was completed every 3 s to a cadence. The test was terminated if the participant fell behind the cadence for 3 consecutive repetitions, or alternatively if they completed the maximum of 100 repetitions. The number of sit-ups successfully completed at test end was recorded and used for analysis.

3-minute step-up test

Each participant put on a heart rate monitor (H10, Polar Electro, Finland) and rested for 5 min in a seated position. During the final minute, heart rate (HR) was averaged and used as the resting value (HRpre) for the 3-min step test. Following the HR measurement, the participant was asked to step up and down on a box of adjustable height to a metronome (24 steps/min) for 3 min. Step height was adjusted using the equations Hf = 0·189Ih for females and Hf = 0·192Ih for males, where Hf is the step height and Ih is the height of the participant (Culpepper and Francis 1987). At the end of each minute of the test, HR and rating of perceived effort (RPE; 1–10 scale; displayed in Appendix I) were recorded. Upon test completion, the participant was asked to sit down and then remain seated for 1 min, during which time HR and RPE (1–10 scale) were recorded 5 s and 1 min after sitting (HRpost, HRpost,1-min). The change in HR (HRpre to HRpost) over the 3-min step-up test was used as an indicator of cardiovascular fitness. A recovery score was also calculated using the change in HR from HRpre to HRpost,1-min. Additionally, RPE was used to assess perceived exertion experienced from the 3-min step test.

Sit-and-reach (S&R) flexibility test

Each participant was asked to sit on the floor with legs together, knees extended, and with shoes removed, the soles of the feet placed against the edge of a sit-and-reach box (Figure Finder Flex Tester, Novel Products Inc, USA). The feet were located at 22.9 cm on the scale for all attempts. With arms outstretched, the participant reached forward while sliding their hands along the measuring scale as far as possible without bending the knees (Hartman and Looney 2003). To ensure accurate scoring the stretch was held at maximal range of motion for 2 s before the score was recorded to the nearest 0.5 cm (Hartman and Looney 2003). The best score from three trials was used for analysis.

Blood sampling and analyses

Participants reported to the laboratory in the morning after 10–12 hours fasting. Blood samples were collected from the antecubital vein using four separate vacutainers: plasma preparation (PPT), serum separating (SST), ethylenediaminetetraacetic acid (EDTA), and fluoride/oxalate tubes (BD Biosciences, Franklin Lakes, New Jersey, USA) (Candlish 2023). After collection, the EDTA tube was labeled, refrigerated, and shipped for HbA1c analysis. The remaining PPT, SST, and fluoride/oxalate tubes were placed at room temperature for 10 min to allow clotting to occur before being centrifuged (Heraeus Multifuge 3 SR, Thermo Fisher Scientific, USA) at 3000 g for 10 min, and aliquoting the supernatant into microtubes for storage at −80 °C before subsequent analysis.

Fluoride/oxalate samples were used for determination of glucose concentrations, while SST samples were used to measure insulin, fructosamine, triglyceride, total cholesterol, low-density lipoprotein cholesterol (LDL), high-density lipoprotein cholesterol (HDL), high-sensitivity C-reactive protein (hsCRP), and HbA1c was assessed using EDTA. Procollagen type 1 N-terminal propeptide (P1NP) and C-terminal telopeptide of type I collagen (CTX-1) were measured to assess bone metabolism using SST and PPT, respectively. Plasma glucose and serum insulin HDL, LDL triglyceride, total cholesterol, and hsCRP were analyzed using the Alinity ci-series clinical chemistry and immunoassay integrated analyzer (Abbott Laboratories, Chicago, Illinois, USA) using a combination of photometric, potentiometric, and biotin interference free-chemiluminescent detection technologies. Fructosamine and HbA1c analyses were performed using the Roche Cobas c501 and Cobas c513 chemistry analyzers, respectively (Roche Diagnostics, Basel, Switzerland). Briefly, fructosamine is a colorimetric assay and HbA1c is a turbidimetric inhibition immunoassay standardized and traceable to the IFCC (International Federation of Clinical Chemistry and Laboratory Medicine) reference method free from interference with most known HbA1c variants. Laboratory analyses were conducted by PathWest Laboratory Medicine WA in Perth, Western Australia, according to standard accredited clinical laboratory procedures.

Questionnaires

Participants were asked to complete a 36-item short form survey (SF-36) and 6-point subjective vitality scale (SVS) to assess physical and mental wellbeing (McHorney et al. 1993). SF-36 physical and mental health scores were calculated using the RAND scoring instructions (RAND n.d.). The sum of the 6-point vitality scale for six questions was calculated with a higher score indicating a better condition.

At the end of the study, participants completed an exit survey which asked how they felt (stronger, fitter and healthier) after the 4-week exercise intervention as well as whether they enjoyed the program. For each question, a five-point Likert scale was used (1 = strongly disagree, 2 = disagree, 3 = neither agree nor disagree, 4 = agree, 5 = strongly agree). The mean of these responses was then calculated and reported. Participants were also asked whether they planned to exercise in future using the following scoring: 1 = yes, using the same eccentric-biased regime, 2 = yes, but doing my own thing, 3 = no, and 4 = unsure. Finally, participants were asked “how likely are you to recommend this exercise program to a friend or family member”? Scoring for this question ranged between 1 and 10, with 10 being the strongest recommendation score. At 4 weeks post-intervention, participants were asked whether they were currently exercising, and if so, what type of exercise they were performing.

Statistical analyses

Intraclass correlation coefficient (ICC) estimates and their 95% confident intervals were calculated based on a mean-rating (k = 2), absolute-agreement, and one-way mixed-effects model using the two measures in the control period to establish the test–retest reliability of the outcome measures. ICC estimates were interpreted as poor (<0.5), moderate (0.5 and 0.75), good (0.75 and 0.9), and excellent (>0.90) (Koo and Li 2016).

The data were assessed for assumptions of normality by a Shapiro–Wilk test and for sphericity by a Mauchly’s sphericity test. In instances where normality was violated, a Friedman test was used. A one-way repeated-measures analysis of variance was used to test for changes in the dependent variables over the three time points. In the case of a significant time effect, a Holm’s sequential Bonferroni correction was performed to compare the values between time points (Holm 1979). Eta squared values (η2) were also reported as a measure of factor variation size, which were interpreted as small (η2 = 0.01), medium (η2 = 0.06), and large (η2 = 0.14) effect (Lakens 2013). The significance level was set to p ≤ 0.05.

Additionally, the changes from PRE-1 to PRE-2 (control period) were compared to the changes from PRE-2 to POST (training period) using a linear mixed-effects model (LMM). The LMM was specified with the change scores as the dependent variable, the period (control vs. intervention) as a fixed effect, and subject as a random effect to account for the repeated-measures design. This model allowed for the assessment of the intervention effect while controlling for individual variability and potential confounders. This comparison aimed to ensure that any significant changes observed during the intervention period were not present during the control period, thereby supporting the causal effect of the training program. To reject the null hypothesis, the following conditions had to be simultaneously met: 1) ICC in the control period was expected to exceed 0.70, indicating moderate reliability, 2) the one-way repeated-measures ANOVA had to detect a statistically significant effect of time, demonstrating that there were differences between the time points, 3) post hoc comparisons with Holm’s sequential Bonferroni correction had to identify significant differences between PRE-2 vs. POST, and 4) the LMM had to reveal a significant fixed effect of time (control vs. intervention) on the change scores, confirming that the changes observed during the intervention period were significantly different from those observed during the control period.

To explore whether age or body mass influenced the training outcomes, an additional LMM analysis was performed with time point as a fixed factor and age and body mass as covariates. Interaction terms (time point × age and time point × body mass) were also included to determine whether changes in outcome measures differed across individuals with varying age or body mass. This analysis was applied to all performance-based measures where significant time effects were detected.

A within-subject correlation analysis was performed to quantify the relationships between variables across time points. This analysis utilized the sum of squares and residual sum of squares outputs computed by the analysis of covariance (ANCOVA), as recommended by Bland and Altman (1995). By incorporating data collected from each participant across multiple time points into a single correlation, this method allowed for the computation of an overall correlation between variables over time. The strength of relationship was quantified as trivial (r < 0.1), small (r = 0.10–0.29), moderate (r = 0.3–0.49), large (r = 0.5–0.69), very large (r = 0.7–0.89), and nearly perfect (r ≥ 0.9) (Cohen 2013). All statistical testing was performed using Jamovi version 2.3.21 (Jamovi project, 2018). Data are presented as mean ± standard deviation (SD).