Acute Effects of Caffeine on Mitochondrial Function in Peripheral Blood Mononuclear Cells

This randomized, double-blind, placebo-controlled crossover study will evaluate the acute effects of caffeine consumption on substrate utilization, resting energy metabolism, and mitochondrial function in peripheral blood mononuclear cells (PBMCs) in healthy young adults. Approximately 10 healthy men and women aged 18 to 35 years will participate. Each participant will complete two experimental visits, receiving caffeine in one visit and placebo in the other, in randomized order. The two visits will be separated by a 48-hour washout period. Participants will attend the laboratory after an overnight fast of approximately 14 hours and will abstain from caffeine, alcohol, and intense physical activity before each visit. Resting gas exchange will be measured before ingestion and 1 and 3 hours after caffeine or placebo administration. Blood samples will be collected at the same time points to isolate PBMCs and obtain plasma for metabolic analyses. Mitochondrial function will be assessed by high-resolution respirometry, and mitochondrial dynamics and morphology will be evaluated using protein analyses and transmission electron microscopy. The study will determine whether acute caffeine consumption alters resting substrate utilization and mitochondrial responses in PBMCs in healthy young adults.

Inclusion Criteria: * Healthy adults aged 18 to 65 years. * Body mass index between 18.5 and 27 kg/m². * Non-smokers. * Habitual caffeine intake of 200 mg/day or less. * No use of medications or supplements known to affect energy metabolism. * Able and willing to provide written informed consent. Exclusion Criteria: * Chronic metabolic, cardiovascular, or inflammatory disease. * Use of antioxidant supplements or ergogenic substances within the 4 weeks before participation. * Known hypersensitivity to caffeine or history of adverse effects after caffeine consumption.