Chronic wounds represent a major and rising health and economic burden worldwide. There is a continued search toward more effective wound therapy. We found significantly reduced microRNA-132 (miR-132) expression in human diabetic ulcers compared with normal skin wounds and also in skin wounds of leptin receptor-deficient (db/db) diabetic mice compared with wild-type mice. Local replenishment of miR-132 in the wounds of db/db mice accelerated wound closure effectively, which was accompanied by increased proliferation of wound edge keratinocytes and reduced inflammation. The pro-healing effect of miR-132 was further supported by global transcriptome analysis, which showed that several inflammation-related signaling pathways (e.g., NF-κB, NOD-like receptor, toll-like receptor, and tumor necrosis factor signaling pathways) were the top ones regulated by miR-132 in vivo. Moreover, we topically applied liposome-formulated miR-132 mimics mixed with pluronic F-127 gel on human ex vivo skin wounds, which promoted re-epithelialization. Together, our study showed the therapeutic potential of miR-132 in chronic wounds, which warrants further evaluation in controlled clinical trials.

Insufficient knowledge about the molecular pathology of diabetic foot ulcer (DFU) impedes the development of effective wound treatment. Circular RNAs (circRNAs) are a novel class of RNA recently discovered to be widely expressed and have important biological functions; however, their role in skin wound healing remains largely unexplored. In this study, we investigated the role of circRNAs in DFU. Approach: CircRNA expression was profiled in normal wounds (NWs) and DFUs by microarray analysis, and hsa_circ_0084443 was identified as differentially expressed. The circularity and subcellular localization of hsa_circ_0084443 were characterized by northern blotting, real-time PCR, and fluorescence in situ hybridization. Cell migration, cell growth, and the transcriptome of human primary keratinocytes were analyzed after overexpression or RNA interference of hsa_circ_0084443. Results: hsa_circ_0084443 is downregulated in NWs compared with intact skin, and its level is higher in DFUs than NWs. We confirmed its circularity and presence in the cytoplasm of human epidermal keratinocytes. We showed that hsa_circ_0084443 reduced motility while enhancing the growth of keratinocytes. Furthermore, we identified a gene network with the potential to mediate the biological effect of hsa_circ_0084443. Innovation: CircRNAs have a functional role and a potential clinical significance in skin wound healing. Conclusions: We identified hsa_circ_0084443, a circRNA downregulated during NW healing, as a negative regulator of keratinocyte migration. Higher levels of hsa_circ_0084443 were detected in DFU samples, suggesting that it plays a role in pathology. These findings pave the way to understanding the functional role of circRNAs in human skin wound healing.

Persistent and impaired inflammation impedes tissue healing and is a characteristic of chronic wounds. A better understanding of the mechanisms controlling wound inflammation is needed. In this study, we show that in human wound-edge keratinocytes, the expressions of microRNA (miR)-17, miR-18a, miR-19a, miR-19b, and miR-20a, which all belong to the miR-17w92 cluster, are upregulated during wound repair. However, their levels are lower in chronic ulcers than in acute wounds at the proliferative phase. Conditional knockout of miR-17w92 in keratinocytes as well as injection of miR-19a/b and miR-20a antisense inhibitors into wound edges enhanced inflammation and delayed wound closure in mice. In contrast, conditional overexpression of the miR-17w92 cluster or miR-19b alone in mice keratinocytes accelerated wound closure in vivo. Mechanistically, miR-19a/b and miR-20a decreased TLR3-mediated NF-kB activation by targeting SHCBP1 and SEMA7A, respectively, reducing the production of inflammatory chemokines and cytokines by keratinocytes. Thus, miR-19a/b and miR-20a being crucial regulators of wound inflammation, the lack thereof may contribute to sustained inflammation and impaired healing in chronic wounds. In line with this, we show that a combinatory treatment with miR-19b and miR-20a improved wound healing in a mouse model of type 2 diabetes.

An increasing number of studies reveal the importance of long noncoding RNAs (lncRNAs) in gene expression control underlying many physiological and pathological processes. However, their role in skin wound healing remains poorly understood. Our study focused on a skin-specific lncRNA, LOC105372576, whose expression was increased during physiological wound healing. In human nonhealing wounds, however, its level was significantly lower compared with normal wounds under reepithelialization. We characterized LOC105372576 as a nuclear-localized, RNAPII-transcribed, and polyadenylated lncRNA. In keratinocytes, its expression was induced by TGF-β signaling. Knockdown of LOC105372576 and activation of its endogenous transcription, respectively, reduced and increased the motility of keratinocytes and reepithelialization of human ex vivo skin wounds. Therefore, LOC105372576 was termed “wound and keratinocyte migration-associated lncRNA 1” (WAKMAR1). Further study revealed that WAKMAR1 regulated a network of protein-coding genes important for cell migration, most of which were under the control of transcription factor E2F1. Mechanistically, WAKMAR1 enhanced E2F1 expression by interfering with E2F1 promoter methylation through the sequestration of DNA methyltransferases. Collectively, we have identified a lncRNA important for keratinocyte migration, whose deficiency may be involved in the pathogenesis of chronic wounds.

Chronic wounds represent a major and growing health and economic burden worldwide. A better understanding of molecular mechanisms of normal as well as impaired wound healing is needed to develop effective treatment. Herein we studied the potential role of long noncoding RNA LOC100130476 in skin wound repair. LOC100130476 is an RNA polymerase IIeencoded polyadenylated transcript present in both cytoplasm and nucleus. We found that its expression was lower in wound-edge keratinocytes of human chronic wounds compared to normal wounds of healthy donors and intact skin. In cultured keratinocytes, LOC100130476 expression was induced by TGF-b signaling. By reducing LOC100130476 expression with antisense oligos or activating its transcription with CRISPR/Cas9 Synergistic Activation Mediator system, we showed that LOC100130476 restricted the production of inflammatory chemokines by keratinocytes, while enhancing cell migration. In line with this, knockdown of LOC100130476 impaired re-epithelization of human ex vivo wounds. Based on these results, we named LOC100130476 wound and keratinocyte migrationeassociated long noncoding RNA 2 (WAKMAR2). Moreover, we identified a molecular network that may mediate the biological function of WAKMAR2 in keratinocytes using microarray. In summary, our data suggest that WAKMAR2 is an important regulator of skin wound healing and its deficiency may contribute to the pathogenesis of chronic wounds.Abbreviations: DFU, diabetic food ulcer; lncRNA, long noncoding RNA; QRT-PCR, quantitative real-time reverse transcriptase PCR; VU, venous ulcer; WAKMAR 2, wound and keratinocyte migrationeassociated long noncoding RNA 2