The Impact of 6 Weeks of Moderate- and Heavy-intensity Continous Training on Peak Oxygen Uptake, Plasma Extracellular Vesicles and Inflammatory Cytokines

This study is being carried out by the Exercise Metabolism and Inflammation Laboratory at the University of British Columbia campus in Kelowna, British Columbia. Participants who have met the inclusion criteria will first complete an informed consent form. Participants will also complete forms to determine their level of activity. Females will also complete a menstrual cycle questionnaire - but will not be required to start on a particular day in their cycle. Participants will have their anthropometrics recorded (height, weight, body composition), will provide a fasted blood sample, and undergo a maximal staged exercise test to determine their lactate threshold and their maximal oxygen uptake (VO2peak).

The blood test will be used to measure various markers. First, whole blood will be analyzed on a standard hematology analyzer to determine white blood cell counts, red blood cell counts, hemoglobin and platelets. Secondly, blood samples will be spun down to platelet poor plasma and stored for batch analysis. Analysis of plasma will include measurement of plasma cytokines: Interleukin (IL)-6, IL-10, IL-1 receptor agonist, tumor necrosis factor alpha (TNF-alpha) and plasma extracellular vesicles. Extracellular vesicles (EVs) will be isolated using size exclusion chromatography (SEC) and measured for size and concentration using tunable resistive pulse sensing (Izon Exoid, Izon USA). EV phenotype and origin will be determined using nano-flow cytometry (CytoFlex S, Beckman Coulter, USA). EV micro ribonucleic acid (miRNA) will be determined by extracting miRNA from isolated and concentrated EVs (high speed centrifugation of 25,000 x g for 1 hour at 20 degrees Celsius). Extracted miRNA will be measured using PCR (BioRad, USA).

The maximal exercise will occur on a stationary cycle ergometer (Lode Excalibur, Lode, The Netherlands). The ergometer is adjusted to be ideal for each participant. Participants are fitted with a chest strap heart rate monitor (Polar, Finland) and fitted with a face mask to collect to determine the amount of O2 consumption, and CO2¬ production. Lactate measures will be collected from fingertip capillary blood and measured with a lactate analyzer (Nova, USA) during the last 30 seconds of each stage, as is a rating of perceived exertion (RPE). The protocol involves remaining stationary on the ergometer for 2 minutes. Following baseline measures, participants will complete 2 minutes of warm up at 50 watts (W), and then 3-minute stages at consecutively higher workloads (20W increases for females, 30W increases for males). Participants cycle at 60 - 80 revolutions per minute (rpm) for as long as they can. Once they fall below, or cannot maintain a minimum of 50rpm, the test is terminated.

Following the test, the lactate threshold determined as a weighted combination of the log-log method, and Dmax method will be calculated. Weighting consists of 75% of the log-log value and 25% of the Dmax value as calculated in the "lactater" R package. Using their lactate thresholds, specific exercise prescriptions are determined.

Group randomization is completed using a 2:2:1 ratio of moderate intensity continuous training (MICT), heavy intensity continuous training (HICT) and control (CTRL). A power calculation using an alpha of 0.05, power of 0.8 and a medium effect size using changes in VO2peak within the literature suggest that 52 participants will need to complete the study.