Participants
Nine boys were recruited from year 10 of a local secondary school. This sample size was based on the ability to detect a moderate to large mean difference for glucose and insulin outcomes based on previous work examining the acute effect of HIIT [21, 30, 31]. All participants were deemed able to participate in the study by completing an initial health questionnaire to exclude any metabolic or medical conditions that contradict exercise or are known to effect glucose metabolism. Following an explanation of the study procedures and the associated risks and benefits, parental consent and participant assent were obtained. Ethical approval was granted by the University of Exeter sport and health sciences ethics committee. One boy failed to complete the HIIT due to an unrelated illness, and one boy could not complete the training due to an unrelated injury. This left a sample of seven participants (14.3 ± 0.3 y) for analysis.
Study design
This study consisted of four laboratory visits, and 6 training sessions in the school setting, which took place over a three week period. Visits included an initial familiarisation visit and three experimental visits. Visits 1 and 2 consisted of baseline measures of aerobic fitness and the glucose and insulin response to a mixed meal tolerance test (MMTT) prior to undertaking the HIIT intervention (PRE). Visits 1 and 2 were separated by 3-5 days. Participants then completed 6 supervised HIIT sessions over a two week period, after which post training measures were assessed 20 h (visit 3; 20 h POST) and 70 h post-intervention (visit 4; 70 h POST).
Visit 1: Familiarisation and baseline fitness assessment
Stature and body mass were measured to the nearest 0.01 m and 0.1 kg, and used to calculate body mass index (BMI). BMI was used to classify participants as normal weight, overweight and obese, using validated age-specific percentile cut points [32]. Pubertal status was determined by self-assessment of the five stages of pubic hair development described by Tanner [33].
Participants were familiarised with the cycle ergometer (Lode Excalibur Sport, Groningen, Netherlands) and completed a combined ramp-incremental and supramaximal test to exhaustion to determine maximal oxygen uptake ( O2 max) and the gas exchange threshold (GET) [34]. Pulmonary gas exchange and heart rate were measured (Cortex Metalyzer III B, Germany) and O2 max was accepted as the highest 10 s average O2 during the ramp or supra-maximal test. Peak power (PP) was taken as the highest power output during the ramp test whilst maintaining a cadence > 60 revolutions.min-1. The GET was estimated at the point where the first disproportionate increase in VCO2 production compared to O2 and verified using the ventilatory equivalents for O2 and CO2.
Visits 2: Baseline metabolic assessment
Participants were driven to the laboratory and arrived at ~ 07:45 following a 12 h overnight fast. After 15 min of seated rest, participants provided a capillary blood sample for plasma glucose and insulin. At ~ 08:30 a MMTT was conducted which consisted of a commercially available fruit smoothie with 50 ml of double cream added, chocolate croissant with chocolate spread and a chocolate muffin (80 g of glucose, 68 g of fat, 7134 kJ). The meal was consumed over a 15 min period, after which capillary blood samples were taken at 30, 60, 120 min for assessment of plasma glucose and insulin. No other food was consumed and water was available ad libitum during visit 2 (PRE). This was recorded and subsequently replicated for the POST measures. Participants remained in the laboratory throughout the visit, completing sedentary activities such as reading, watching DVDs or playing computer games. Participants left the laboratory at ~ 15:00.
HIIT intervention
Participants performed a two week HIIT programme on a cycle ergometer (Monark 827e, Monark exercise AB, Sweden) with adjustments made to the handle bar and seat height for each participant. Training took place within a local secondary school and consisted of 3 supervised HIIT sessions per week. Sessions were carried out during the school lunch break. Each session started with a 3 min warm up of unloaded pedalling, followed by 8-10 one min intervals at 90 % of the PP achieved during the incremental ramp test performed during visit 1. Each interval was interspersed with 75 s of unloaded pedalling. This HIIT protocol was selected to mimic previous studies from our laboratory [18, 35, 36]. Sessions one and two consisted of 8 x 1 minute bouts, sessions three and four 9 x 1 minute bouts and sessions five and six 10 x 1 minute bouts. Participants were asked to maintain a self-selected cadence (70-95 revolutions.min-1) and were reminded of this during each session.
Visit 3 and 4: Post-training
The protocol outlined above for visit 2 was replicated the day after (20-POST) and three days (70-POST) after the last training session. 1 hour after completion of the MMTT during the 70- POST visit, participants completed a post intervention O2 max assessment as described in visit 1.
Standardisation of physical activity and diet
Physical activity was measured during the 48 h period prior to each experimental visit using a wrist worn accelerometer (GENEActiv, Activinsights, UK). For visit three this 48 h period included the final training session. Time spent performing, light, moderate and vigorous PA was determined using cut points previously validated in a paediatric population [37]. Participants were asked to avoid any structured physical activity outside of the training intervention and before any laboratory visits.
With supervision from their parents/guardians, a food diary was completed by each participant during the 48 h period preceding each experimental visit. Food diaries were assessed to estimate total energy and macronutrient content using commercially available software (CompEat Pro, Nutrition systems, UK). Participants were asked to replicate their diet during the 48 h preceding each experimental visit and if appropriate, to document any discrepancies.
Blood analyses
Fingertip capillary blood samples (~ 600 µL) were taken from a pre-warmed hand into a fluoride heparin coated and lithium heparin coated microvette (CB 300 tubes, Sarstedt Ltd, Leicester, UK) for plasma glucose and insulin determination, respectively. Both microvettes were centrifuged at 6000 revolutions.min-1 for 10 min. Plasma was separated for immediate analysis of glucose (YSI 2300 Stat Plus Glucose analyser, Yellow Springs, OH, USA) or stored at –80oC for later analysis of plasma insulin using an ELISA enzyme immunoassay kit (DRG Diagnostics, Germany). In our laboratory, the within batch coefficients of variation for the plasma insulin and glucose analyses were < 5%.
Data handling
Changes in plasma glucose and insulin during the postprandial period following the MMTT were quantified using total and incremental area under the curve (tAUC, iAUC) [38] calculated using the trapezium rule (GraphPad Prism, GraphPad, SanDiego, CA). tAUC is related to basal blood glucose and reflects the amplitude of change. iAUC more accurately describes the glycaemic response to MMTT, and the dynamic change over time, independent of baseline value. Fasting plasma glucose and insulin were used to calculate IR, IS and %b using using HOMA-IR [39], QUICKI [27] and FGIR [28], which have been validated for use in adolescents [40].
Statistical analysis
Descriptive statistics were calculated using SPSS (version 19.0, Chicago, USA) and presented as mean ± SD. Analysis of the HOMA, QUICKI, FGIR, fasting glucose and insulin, and tAUC and iAUC response to the MMTT across visits was performed using a one-way repeated-measures ANOVA, follow up comparisons between time points (PRE, 20 h-POST and 70 h-post) were only carried out if there was a significant main effect in the ANOVA. The Eta squared thresholds of 0.01, 0.06 and 0.14 were used to identify a small, moderate and large effect from the ANOVA analyses. Changes in aerobic fitness parameters were assessed by a paired sample t-test. The magnitude of the difference between variables of interest were explored using ES [41].
To understand the influence of different parameters on the effectiveness of the HIIIT intervention and understand the relationship between pre intervention health outcomes and the effects of HIIT intervention, Pearson’s correlations were performed between HOMA-IR, QUICKI, FGIR, V ̇O2 max and BMI at baseline (PRE) and change in HOMA-IR after the 2 week training period (20 h-POST). A significant correlation was accepted if P<0.05.