Introduction and Objective: Diabetic foot ulcers complicated by lower limb ischemia are common complications in the person with diabetes, leading to non-traumatic amputations. Currently, there are no specific drugs, and treatment is limited to surgical intervention. This study explores the therapeutic effects and mechanisms of multi-cell fusion-based 3D-printed bioengineered skin, incorporating engineered exosomes, for treating diabetic chronic wounds with lower limb ischemia.Methods: A multifunctional hybrid 3D bioprinting platform combining inkjet and extrusion was used to create complex structures, including free-form 3D shapes with embedded micro-vessels. Exosomes from MSCs stimulated by the GLP-1 end product-GLP-1(32-36) were incorporated into hydrogel for bio-3D printing. Therapeutic effects have been demonstrated in T2DM mice and porcine wound models. Single-cell proteomic was employed to investigate the mechanisms of exosome-loaded “artificial skin”.Results: In vivo, GLP-1(32-36) treated exosome-loaded “artificial skin” (32-36AS) significantly improved blood flow, accelerated wound healing, and promoted collagen production. Single-cell proteomic analysis showed that 32-36AS especially promoted rigidity-sensing contractions in ECs and attenuate mouse dermal fibroblast (MDFs) and keratinocytes (KCs) mediated scar formation. Depletion of cytoskeletal proteins, including tropomyosins (Tpm2.1 and 3.1) would block the 32-36AS mediated angiogenesis and wound healing. In vitro, GLP-1(32-36) engineered exosomes enhanced tube formation and migration of ECs and fibroblasts. They also improved vascular structures, increasing vessel length, connectivity, and diameter distribution, forming a dense fibrovascular network.Conclusion: This study develops multicellular vascularized 32-36AS with bio-3D printing, providing a foundation for clinical application in treating diabetic foot ulcers with lower limb ischemia.
Y. Zhang: None. M. Gao: None. S. Zhang: None. Y. Wan: None. C. Zheng: None.
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