Single-cell RNA-seq and bulk-seq identify RAB17 as a potential regulator of angiogenesis by human dermal microvascular endothelial cells in diabetic foot ulcers

Hengyu Du; Shenghong Li; Jinqiang Lu; Lingzhi Tang; Xiao Jiang; Xi He; Jiaji Liang; Xuan Liao; Taixing Cui; Yuesheng Huang; Hongwei Liu

doi:10.1093/burnst/tkad020

Single-cell RNA-seq and bulk-seq identify RAB17 as a potential regulator of angiogenesis by human dermal microvascular endothelial cells in diabetic foot ulcers

Burns Trauma. 2023 Aug 18:11:tkad020. doi: 10.1093/burnst/tkad020. eCollection 2023.

Authors

Affiliations

¹ Department of Plastic Surgery of the First Affiliated Hospital of Jinan University, Institute of New Technology of Plastic Surgery of Jinan University, Key Laboratory of Regenerative Medicine of Ministry of Education, Guangzhou 510630, P.R. China.
² Dalton Cardiovascular Research Center, Department of Medical Pharmacology and Physiology, University of Missouri, School of Medicine, 134 Research Park Dr, Columbia, MO 65211, USA.
³ Institute of Wound Repair and Regeneration Medicine, Southern University of Science and Technology School of Medicine, and Department of Wound Repair, Southern University of Science and Technology Hospital, Shenzhen, Guangdong, 518055, China.

Abstract

Background: Angiogenesis is crucial in diabetic wound healing and is often impaired in diabetic foot ulcers (DFUs). Human dermal microvascular endothelial cells (HDMECs) are vital components in dermal angiogenesis; however, their functional and transcriptomic characteristics in DFU patients are not well understood. This study aimed to comprehensively analyse HDMECs from DFU patients and healthy controls and find the potential regulator of angiogenesis in DFUs.

Methods: HDMECs were isolated from skin specimens of DFU patients and healthy controls via magnetic-activated cell sorting. The proliferation, migration and tube-formation abilities of the cells were then compared between the experimental groups. Both bulk RNA sequencing (bulk-seq) and single-cell RNA-seq (scRNA-seq) were used to identify RAB17 as a potential marker of angiogenesis, which was further confirmed via weighted gene co-expression network analysis (WGCNA) and least absolute shrink and selection operator (LASSO) regression. The role of RAB17 in angiogenesis was examined through in vitro and in vivo experiments.

Results: The isolated HDMECs displayed typical markers of endothelial cells. HDMECs isolated from DFU patients showed considerably impaired tube formation, rather than proliferation or migration, compared to those from healthy controls. Gene set enrichment analysis (GSEA), fGSEA, and gene set variation analysis (GSVA) of bulk-seq and scRNA-seq indicated that angiogenesis was downregulated in DFU-HDMECs. LASSO regression identified two genes, RAB17 and CD200, as characteristic of DFU-HDMECs; additionally, the expression of RAB17 was found to be significantly reduced in DFU-HDMECs compared to that in the HDMECs of healthy controls. Overexpression of RAB17 was found to enhance angiogenesis, the expression of hypoxia inducible factor-1α and vascular endothelial growth factor A, and diabetic wound healing, partially through the mitogen-activated protein kinase/extracellular signal-regulated kinase signalling pathway.

Conclusions: Our findings suggest that the impaired angiogenic capacity in DFUs may be related to the dysregulated expression of RAB17 in HDMECs. The identification of RAB17 as a potential molecular target provides a potential avenue for the treatment of impaired angiogenesis in DFUs.

Keywords: Angiogenesis; Diabetic foot ulcers; Diabetic wound healing; Human dermal microvascular endothelial cells; RAB17; Single-cell RNA-seq.