Background and Purpose: Diabetic wound affects the health and safety of the people with diabetes. FGF21 has been shown to promote diabetic wound healing, but its low half-life and transdermal rate limit its use. This study was used to investigate the role of TDP1-FGF21 in promoting the diabetic wound healing. Experimental Approach: To increase the transdermal rate of FGF21, a cell-penetrating peptide, TDP1 was N-terminally fused to FGF21 to produce a fusion protein, TDP1-FGF21. TDP1-FGF21 was expressed in a prokaryotic expression system and purified by chromatography. To evaluate whether TDP1-FGF21 could regulate the macrophage polarization to promote diabetic wound healing, the levels of histopathological changes, macrophages polarization, and protein expression were assessed after the application of TDP1-FGF21-containg hydrogel to the wound models of diabetic mice. To confirm whether TDP1-FGF21 could regulate the macrophage polarization in vitro, the levels of macrophage polarization, macrophages metabolic reprogramming, mitochondria and the protein expression were assessed after the application of TDP1-FGF21. Key Results: TDP1-FGF21 was purified by chromatography. TDP1 effectively promoted the transition of diabetic wounds from the inflammatory stage to the proliferative stage by enhancing the transdermal efficiency of FGF21. Both FGF21 and TDP1-FGF21 had the same effect on promoting macrophage polarization. Further mechanistic investigation showed that both TDP1-FGF21 and FGF21 could increase the expression of p-AMPK, SIRT1, and PGC-1a compared with untreated diabetic wounds. Conclusion and Implications: The findings suggested that TDP1 could enhance the transdermal efficiency of FGF21. TDP1-FGF21 could improve the diabetic wound healing through regulating macrophage polarization by targeting the AMPK/SIRT1/PGC-1a pathway.