Mitochondria are highly dynamic organelles, which not only provide energy and material basis for cells, but also regulate cell proliferation, migration, differentiation, and apoptosis. Cell fate is regulated by mechanical cues from the microenvironment. Recent studies have shown that energy metabolism is regulated by mechanical cues. Mitochondria can act as mechanical sensors and hubs that connect the mechanics and metabolism to regulate cell fate. A deep understanding of the relationship between the mechanical microenvironment and mitochondrial metabolism provides sufficient guidance for promoting tissue regeneration and treating diseases. In this review, the progression in mitochondrial mechanobiology is mainly introduced and its potential applications in tissue regeneration and disease treatment are explored.