Post-transcriptional regulation of diabetic wound healing by junctional adhesion molecule A/miR-106b axis.

Abstract

Despite advancements in molecular science and biomaterial technology, the mechanisms underlying the impaired healing of diabetic wounds remain unclear. In this study, we investigated the post-transcriptional regulation of diabetic wound healing using JAM-A. Mouse wound models, hematoxylin and eosin staining analysis, and scratch wound assays were used to investigate the effects of JAM-A 3'-UTR on the re-epithelialization of diabetic wounds, whereas RNA pulldown, microRNA-seq, and bioinformatics analyses were performed to identify key miRNA players and predict their target genes. In situ hybridization, immunohistochemistry, western blotting, and polymerase chain reaction (PCR) were used to confirm the alternative splicing of JAM-A 3'-UTR in diabetic conditions. CCK-8 proliferation assays, scratch wound assays, PCR, western blotting, and dual-luciferase assays were performed to study the changes in cell proliferation and migration induced by miR-106b-5p modification and confirm the target gene. JAM-A 3'-UTR accelerated re-epithelialization in diabetic mouse wounds. Shortened splicing was found in the 3'-UTR of JAM-A under diabetic conditions, leading to the excessive release of miR-106b-5p while the promoter of miR-106b was activated. Furthermore, upregulated miR-106b-5p over-activated cell proliferation and inhibited cell migration in diabetic wound keratinocytes by suppressing the target gene PTEN/TIAM1 and regulating the AKT and RAC1 pathways, thereby impairing wound re-epithelialization. We identified alternative splicing of JAM-A 3'-UTR in diabetic conditions, which caused the excessive release of miR-106b. Upregulation of miR-106b reduced the expression of its target genes, PTEN and TIAM1, which led to hyperactive proliferation and impaired migration of keratinocytes, thereby dysregulating wound re-epithelialization.