Abstract: BACKGROUND: Exosomes, as an important mediator of intercellular communication, have been widely used in tissue repair and regeneration. Exosome-loaded hydrogels can significantly improve the stability and bioavailability of exosomes, thereby enhancing therapeutic efficacy.
OBJECTIVE: To investigate the effects of fibroblast-exosome-loaded hydrogels on endothelial cell function recovery and wound repair in diabetic rats.
METHODS: Exosomes isolated from human skin fibroblasts were added to PF-127 hydrogels to prepare fibroblast-exosome-loaded PF-127 hydrogels. (1) Cellular experiment: A suspension of third-generation human umbilical vein endothelial cells was cultured in four groups: a control group incubated with 5 mmol/L glucose; a high glucose group incubated with 50 mmol/L glucose; a high glucose + hydrogel group incubated with 50 mmol/L glucose and PF-127 hydrogel, and a high glucose + exosome-loaded hydrogel group incubated with 50 mmol/L glucose and exosome-loaded PF-127 hydrogel. Cell proliferation was assessed by EdU staining. Cell migration was assessed by wound healing assay and Transwell assay. Cell angiogenesis was assessed by tube formation assay. Intracellular ferroptosis was assessed by western blot assay and transmission electron microscopy. (2) Animal experiment: Twenty-four SD rats were randomly divided into four intervention groups. The control group (n=6) underwent a 1-cm diameter full-thickness skin defect on the back without any treatment. The diabetes group (n=6) underwent a 1-cm diameter full-thickness skin defect after the establishment of a type 1 diabetes model without any treatment. Type 1 diabetes models were established in the diabetes + hydrogel group (n=6) and the diabetes + exosome-loaded hydrogel group (n=6). A 1-cm diameter full-thickness skin defect was made on the back. PF-127 hydrogel and PF-127 hydrogel loaded with fibroblast exosomes were injected for treatment, respectively. The drugs were administered twice a week for a total of 3 weeks. Wound healing was observed during the treatment period. After the administration, samples were collected and analyzed by hematoxylin-eosin staining and CD31 immunohistochemical staining.
RESULTS AND CONCLUSION: (1) Cell experiment: High glucose treatment inhibited the proliferation, migration, and tube formation of human umbilical vein endothelial cells, inducing ferroptosis. PF-127 hydrogel loaded with fibroblast exosomes significantly enhanced the proliferation, migration, and tube formation of human umbilical vein endothelial cells treated with high glucose, inhibiting ferroptosis. (2) Animal experiment: The wound closure rate in the diabetes + exosome-loaded hydrogel group was faster than that in the diabetes and diabetes + hydrogel groups. Hematoxylin-eosin staining showed that the wound healing quality in the diabetes group and the diabetes + hydrogel group was poor, while the wound healing quality in the diabetes + exosome-loaded hydrogel group was good, but not as good as that in the control group. CD31 immunohistochemical staining showed that angiogenesis in the wounds in the diabetes group and the diabetes + hydrogel group was less than that in the control group and the diabetes + exosome-loaded hydrogel group. (3) These results indicate that PF-127 hydrogel loaded with fibroblast exosomes accelerates wound repair in diabetic rats by inhibiting ferroptosis and promoting endothelial cell function recovery.

Key words: fibroblast-derived exosome, PF-127 hydrogel, ferroptosis, endothelial cell, human umbilical vein endothelial cell, diabetic wound repair, biomaterial