Mitochondria play fundamental roles in multiple processes in eukaryotic cells, including energy production, calcium homeostasis, intermediate metabolism, endogenous reactive oxygen species production, cell signaling, and apoptosis. In order to meet different demands of cell types and tissues, cells modulate mitochondrial function through biogenesis, degradation, dynamic fusion and fission events. Stem cells are defined by two key properties: self-renew and pluripotency. Studied show that the metabolism of mitochondria is closely related to the fate decision and the development of stem cells. Here, we review mitochondrial morphology, structure, function, and metabolic changes in the process of maintaining pluripotency of stem cells, and in the process of directed differentiation.