Approximately 25% of diabetic patients suffer from diabetic lower-extremity ulcer throughout their lives and 7%-20% of patients will eventually need an amputation despite standard care treatment. The development of new therapies to treat diabetic wounds is urgent. In this study, we used cryopreserved living micronized amnion (300-600?μm) to treat wounds in diabetic mice. Post-thaw micronized amnion retained high cell viability, as well as intact cell morphology and membrane structure. When transplanted onto the wounds of db/db mice, the cryopreserved living micronized amnion greatly promoted wound healing in diabetic mice mainly by secreting growth, inflammation, and chemotaxis-related factors that regulated macrophage migration and phenotype switch, recruited CD34(+) progenitor cells, and increased neovascularization. In addition, the micronized amnion matrix can exist in the dermis and serve as a long-term dermal scaffold. These results demonstrated the potential of the cryopreserved living micronized amnion as a ready-to-use living dermal substitute that addresses multiple defective physiological processes of impaired wounds to treat diabetic ulcers and other chronic wounds in clinics.