Participants
Forty-two nonsmoking men and women (18 to 54 years) were recruited from the student and faculty population of a local university, as well as the surrounding community, via advertisement through the university website, local community newspaper, and word-of-mouth. Participants were eligible for inclusion into the study if they were low-to-moderate risk as defined by the ACSM and sedentary. Participants were considered sedentary if they reported not participating in at least 30 min of moderate intensity physical activity on at least three days of the week for at least three months [3]. Participants were also eligible for inclusion into the study if they verbally agreed to continue previous dietary habits and not perform additional exercise beyond that required for the present study. Exclusionary criteria included evidence of cardiovascular pulmonary, and/or metabolic disease as determined by medical history questionnaire. This study was approved by the Human Research Committee at Western State Colorado University. Each participant signed an informed consent form prior to participation.
Baseline, midpoint, and post-program experimental testing procedures
Measurements of all primary and secondary outcome variables were obtained both before and after the exercise training intervention. Additionally, a measure of the primary outcome variable (maximal oxygen uptake – VO2max) was also obtained at midpoint. Secondary outcome variables consisted of resting heart rate and blood pressure, and basic anthropometric measures including height, weight, waist circumference, and skinfolds. Fasting blood lipid and blood glucose measurements were also performed. All measurements were obtained by following standardized procedures as outlined elsewhere [3]. Procedures for each measurement are also briefly described below. Prior to testing participants refrained from all food and drink other than water for 12 h. Participants were also instructed to refrain from strenuous exertion 12 h prior to testing. All post-program testing took place within 1 to 4 days of the last exercise training session.
Resting heart rate and blood pressure measurement
The procedures for assessment of resting heart rate and blood pressure outlined elsewhere were followed [3]. Briefly, participants were seated quietly for 5 min in a chair with a back support with feet on the floor and arm supported at heart level. Resting heart rate was obtained via manual palpation of radial artery in the left wrist and recording the number of beats for 60 s. The left arm brachial artery blood pressure was measured using a sphygmomanometer in duplicate and separated by 1-min. The mean of the two measurements was reported for baseline and post-program values.
Anthropometric measurements
Participants were weighed to the nearest 0.1 kg on a medical grade scale and measured for height to the nearest 0.5 cm using a stadiometer. Percent body fat was determined via skinfolds [3]. Skinfold thickness was measured to the nearest ± 0.5 mm using a Lange caliper (Cambridge Scientific Industries, Columbia, MD). All measurements were taken on the right side of the body using standardized anatomical sites (three-site) for men and women. These measurements were performed until two were within 10 % of each other. Waist circumference measurements were obtained using a cloth tape measure with a spring loaded-handle (Creative Health Products, Ann Arbor, MI). A horizontal measurement was taken at the narrowest point of the torso (below the xiphoid process and above the umbilicus). These measurements were taken until two were within 0.5 mm of each other.
Fasting blood lipid and blood glucose measurement
All fasting lipid and blood glucose analyses were collected in duplicate and performed at room temperature. The mean of the two measurements was reported for baseline and post-program values. Participants’ hands were washed with soap and rinsed thoroughly with water, then cleaned with alcohol swabs and allowed to dry. Skin was punctured using lancets and a fingerstick sample was collected into heparin-coated 40 μl capillary tube. Blood was allowed to flow freely from the fingerstick into the capillary tube without milking of the finger. Samples were then dispensed immediately onto commercially available test cassettes for analysis in a Cholestech LDX System (Alere Inc., Waltham, MA) according to strict standardized operating procedures. The LDX Cholestech measured total cholesterol, high density lipoprotein (HDL) cholesterol, low density lipoprotein (LDL) cholesterol, triglycerides, and blood glucose in fingerstick blood. A daily optics check was performed on the LDX Cholestech analyzer used for the study. Independent studies have provided data to indicate that the Cholestech LDX system has excellent reproducibility with standard clinical laboratory measurement of plasma lipids and lipoproteins [12, 13] and meets the National Cholesterol Education Program Adult Treatment Panel III (NCEP-ATP) criteria for accuracy and reproducibility [14].
Maximal exercise testing
Participants completed a modified-Balke, pseudo-ramp graded exercise test (GXT) on a power treadmill (Powerjog GX200, Maine). Participants walked or jogged at a self-selected pace. Treadmill incline was increased by 1 % every minute until the participant reached volitional fatigue. Participant HR was continuously recorded during the GXT via a chest strap and radio-telemetric receiver (Polar Electro, Woodbury, NY, USA). Expired air and gas exchange data were recorded continuously during the GXT using a metabolic analyzer (Parvo Medics TrueOne 2.0, Salt Lake City, UT, USA). Before each exercise test, the metabolic analyzer was calibrated with gases of known concentrations (14.01 ± 0.07 % O2, 6.00 ± 0.03 % CO2) and with room air (20.93%O2 and 0.03 % CO2) as per the instruction manual. Volume calibration of the pneumotachometer was done via a 3-Litre calibration syringe system (Hans-Rudolph, Kansas City, MO, USA). The last 15 s of the GXT were averaged – this was considered the final data point. The closest neighbouring data point was calculated by averaging the data collected 15 s immediately before the last 15 s of the test. The mean of the two processed data points represented VO2max. Maximal HR was considered to be the highest recorded HR in beats per minute (bpm) during the GXT. Participant heart rate reserve (HRR) was determined by taking the difference between maximal HR and resting HR.
Determination of ventilatory thresholds
Determination of both the first ventilatory threshold (VT1) and second ventilatory threshold (VT2) were made by visual inspection of graphs of time plotted against each relevant respiratory variable (according to 15 s time-averaging). The criteria for VT1 was an increase in VE/VO2 with no concurrent increase in VE/VCO2 and departure from the linearity of VE. The criteria for VT2 was a simultaneous increase in both VE/VO2 and VE/VCO2 [15]. All assessments were done by two experienced exercise physiologists. In the event of conflicting results, the original assessments were reevaluated and collectively a consensus was agreed upon.
Randomization and exercise intervention
After the completion of baseline testing, participants were randomized to a non-exercise control group or one of two exercise training groups according to a computer generated sequence of random numbers that was stratified by sex (Fig. 1). This was a double-blind research design in that participants were unaware of the group to which they had been assigned. Likewise, the researchers specifically responsible for testing and supervision of exercise sessions were unaware of the group to which participants had been allocated. Participants randomized to the exercise training groups performed 12weeks of exercise training according to one of two exercise intensity regimens: 1) a relative percent method was used in which intensity was prescribed according to percentages of HRR (HRR group), or 2) a threshold-based method (ACE-3ZM) was used in which intensity was prescribed according to VT1 and VT2 as recommended by ACE in its three-zone model [11]. The exercise prescription details for each training group over the course of the 12weeks training period is presented in Fig. 1.
Each group performed a similar frequency and duration of exercise training. All exercise training sessions for each treatment group were performed on a treadmill. Overall, the exercise prescription was intended to fulfill the consensus recommendation of 150 min/week [4]. The exercise prescription for exercise intensity method differed between treatment groups. The HRR group was prescribed exercise intensity according to a percentage of HRR. Conversely, the ACE-3ZM group was prescribed exercise intensity according to ventilatory threshold. In both exercise training groups a target heart rate (HR) coinciding with either the prescribed HRR or prescribed VT (Fig. 1) was used to establish a specific exercise training intensity for each exercise session. In the ACE-3ZM group target HR for each training zone (Fig. 1) was established in the following manner:
-
Wk 1–4 (HR < VT1): target HR = HR range of 10–15 bpm just below VT1
-
Wk 5–8 (HR ≥ VT1 to < VT2): target HR = HR range of 10–20 bpm above VT1 and below VT2
-
Wk 9–12 (HR ≥ VT2): target HR = HR range of 10–15 bpm just above VT2
Exercise training was progressed according to recommendations made elsewhere by the ACSM [3] and ACE [11]. Polar HR monitors (Polar Electro Inc., Woodbury, NY) were used to monitor HR during all exercise sessions. Researchers adjusted treadmill workload accordingly during each exercise session to ensure actual HR responses aligned with target HR.
Statistical analyses
All analyses were performed using SPSS Version 22.0 (Chicago, IL) and GraphPad Prism 6.0. (San Diego, CA). Sample size was projected with change in VO2max as the main outcome variable. The means and standard deviation of a previous study [16] were examined and the effect size of this study was calculated. Assuming that a power of 0.90 was needed and the calculated effect size for change in VO2max was 0.8, it was determined that approximately 12 subjects would be needed for each of the three groups [17]. Further, we assumed there would be an approximate 20 % dropout rate based on findings from one of our previous exercise training studies [18]. Accordingly, we recruited and randomized an additional three participants to each of the exercise training groups to account for potential attrition.
Measures of centrality and spread are presented as mean ± standard deviation (SD). All baseline-dependent variables were compared using general linear model (GLM) ANOVA and, where appropriate, Tukey post hoc tests. Within-group comparisons were made using paired t-tests. Because baseline, 6weeks, and post-program data were available, the effect of exercise training on cardiorespiratory fitness (VO2max) was determined using repeated-measures GLM-ANOVA with exercise intensity method (HRR or ACE-3ZM) as the between-subjects factor. All other between-group 12weeks changes were analyzed using GLM-ANOVA and, where appropriate, Tukey post hoc tests. The assumption of normality was tested by examining normal plots of the residuals in ANOVA models. Residuals were regarded as normally distributed if Shapiro-Wilk tests were not significant [17].
Delta values (Δ) were calculated (post-program minus baseline value divided by baseline value) for percent change in relative VO2max (%) and participants were categorized as: ‘1’ = responders (% Δ > 5.9 %) or ‘0’ = non-responders (Δ ≤ 5.9 %) to exercise training using a day-to-day variability, within subject coefficient of variation (CV) criterion applied previously in the literature [6, 19]. Chi-square (χ2) tests were subsequently used to analyze the point prevalence of responders and non-responders to exercise training separated by exercise intensity group (i.e., HRR and ACE-3ZM) between baseline and post-program. The probability of making a Type I error was set at p ≤ 0.05 for all statistical analyses.