The Impact of a 48-hour Fast with or Without Exercise on Immune Cell Metabolism and Glycemic Control in Healthy Active Adults

Immune cell functions will be assessed with the combination of whole blood cultures treated with lipopolysaccharide (with or without interleukin-10) and the subsequent measurement of cytokine secretion, immune cell counts, and phenotyping via flow cytometry, both before and after 48 hours fast (with and without exercise).

Investigate whether exercise performed during a 48-hour fast can reduce the temporary glucose intolerance typically seen after fasting, both in controlled and free-living conditions. Glucose tolerance will be evaluated by measuring capillary glucose levels before, and at 30, 60, and 120 minutes after a mixed meal tolerance test.

Systolic and diastolic blood pressure, as well as heart rate, will be measured using an automatic blood pressure device at baseline (Day 1), 24 hours into the fast (Day 2), and at the end of the 48-hour fast (Day 3) in both conditions (fasting alone and fasting with exercise).

Examine whether performing exercise at the beginning and during a fast can influence the ketone levels achieved after 24 and 48 hours of fasting. Capillary beta-hydroxybutyrate (a ketone body) will be measured via finger pricks using the Freestyle Neo ketone test.

Explore whether adding exercise during fasting could intensify compensatory movement behaviors, as indicated by changes in sedentary behavior, light physical activity, and moderate to vigorous physical activity, measured using an accelerometer (ActivPAL).

Hunger will be assessed using a 10-cm visual analogue scale at several time points: in the morning at the start of the fast, at 12:00 and 18:00 on the same day, at the same times the following day, and just before breaking the fast 48 hours later.

Sleep quality will be assessed using a 10-cm visual analog scale based on the Richards-Campbell Sleep Questionnaire (2000). Measurements will be taken at baseline the morning before starting the fast, after 24 hours of fasting, and again after 48 hours of fasting.

The Feeling Scale (FS) developed by Hardy and Rejeski (Hardy \& Rejeski, 1989) will be used to measure affective valence when exercise is performed in the fed (Baseline) and fasted state (24h fasted). Specifically, FS will be measured before, during (e.g., at 10, 20, 30, 40, and 50 minutes as well as during intervals 1,2,3,4, and 5 at the end of the intervals as well as during the passive recovery) and following the two exercises conditions (post 5 minutes in a seated position).

Remembered enjoyment will be measured with the original Physical Activity Enjoyment Scale (PACES; Kendzierski \& DeCarlo, 1991) 10 minutes following each supervised exercise session (i.e., fed and fasted).

Rate of perceived exertion will be measured during both exercise sessions (i.e., fed and fasted) at 10, 20, 30, 40, and 50 minutes as well as during intervals 1,2,3,4, and 5 at the end of the intervals.

Glucose homeostasis will be evaluated using a continuous glucose monitor (Freestyle Libre 2, Abbott) under both conditions-fasting alone and fasting combined with exercise. Key markers of glucose homeostasis will be assessed based on Battelino et al. (2022; Lancet Diabetes Endocrinol), and will include average glucose levels, glycemic variability (SD), and time spent below (≤ 3.9 mmol/L), within (3.9-10.0 mmol/L), and above range (≥ 10.0 mmol/L).