Circular RNA signatures of human healing and non-healing wounds

BackgroundAlthough the widespread expression of circular RNAs (circRNAs) has only been recognized recently, increasing evidence has suggested their important roles in health and disease. To identify clinically relevant circRNAs with potential for wound diagnosis and therapy, an in-depth characterization of circRNA expression in human healing and non-healing wounds is a prerequisite that has not been attained yet. MethodsWe collected wound-edge biopsies through the healing process of healthy donors and in chronic non-healing venous ulcers (VU). Paired total RNA- and small RNA-sequencing were performed to profile circRNAs, protein-coding mRNAs, and microRNA expression. We analyzed the co-expression relationship between circRNAs and mRNAs with weighted correlation network analysis (WGCNA) and constructed circRNA-microRNA-mRNA networks. For the circRNAs surfaced in the in-silico analysis, after validating their expression with RT-PCR and sequencing, we silenced hsa-CHST15_0003 and hsa-TNFRSF21_0001 expression in keratinocytes with siRNAs and studied their function with transcriptomic profiling and live-cell monitoring. ResultsOur study unravels the dynamically changed expression patterns of circRNAs during human skin wound healing and their abnormal expression signature in VU, which are presented as a searchable web resource (http://130.229.28.87/shiny/circRNA_wholebiopsy-shinyApp/). In silico analysis deciphers the circRNA-miRNAs-mRNA networks specific to the inflammatory and proliferative phases of wound repair and VU, the biological processes that circRNAs are involved, and the circRNAs that could act as miRNAs sponge in human wounds. Importantly, we found that hsa-CHST15_0003 and hsa-TNFRSF21_0001, two circRNAs upregulated in VU, hampered keratinocyte migration while promoting proliferation through modulating gene networks underpinning these cellular processes. ConclusionBy integrating circRNA, mRNA, and miRNA expression profiles in a unique collection of clinical samples, we identify the circRNAs that are relevant to human wound healing physiology and pathology. This study paves the way to decipher the functional significance of circRNAs in tissue repair.