Mitochondria are central to cellular metabolism, fate determination, and homeostasis, with their homeostasis being vital for cell function and survival. In stem cell senescence, mitochondrial dysfunction is a key factor in aging across species and is closely associated with age-related pathologies. Advances in understanding mitochondrial homeostasis in stem cells offer opportunities for developing therapies to rejuvenate aged stem cells and improve tissue health. This overview highlights the pivotal roles of mitochondria in both healthy and aging states, emphasizing recent progress in deciphering the regulatory processes that preserve mitochondrial homeostasis. In addition, we explore the role of mitochondrial homeostasis on stem cell senescence and discuss potential future research directions. Understanding the link between mitochondrial homeostasis and stem cell senescence has important implications for promoting longevity.