Stress is an established risk factor for negative health outcomes. Salivary cortisol and testosterone concentrations increase in response to acute psychosocial stress. It’s crucial to reduce stress for health and well-being through evidence-based interventions. Body-mind interventions such as meditation and Tai Chi have shown reduced cortisol levels but mixed results in testosterone concentration after stress. To address this research gap, we conducted a pilot randomized controlled trial to examine the modulating effects of a short-term (seven 20-minute sessions) mindfulness meditation on testosterone and cortisol in response to acute stress. Using one form of mindfulness meditation – Integrative Body-Mind Training (IBMT) and an active control–relaxation training (RT), we assessed salivary cortisol and testosterone concentrations at three stages of stress intervention – rest, stress, and an additional 20-min IBMT or RT practice. We found increased cortisol and testosterone concentrations after acute stress in both groups, but testosterone rise was not associated with cortisol rise. Moreover, an additional practice immediately after stress produced higher testosterone concentrations in the IBMT group than the RT group, whereas cortisol concentration increased in the RT group than in the IBMT group at the same time point. These findings indicate that brief mindfulness intervention modulates a dual-hormone profile of testosterone and cortisol in response to acute stress presumably via the co-regulation of hypothalamus-pituitary-adrenal and hypothalamus-pituitary-testicular axes.