Liu Yiling, Wang Peng, Li Jingting, Chen Lei, Shu Bin, Wang Hanwen, Liu Hengdeng, Zhao Shixin, Zhou Junli, Chen Xiaodong, Xie Julin
Department of Burn and Wound Repair Surgery, The First Affiliated Hospital of Sun Yat-sen University, No. 58 Zhongshan 2 Road, Guangzhou 510080, China.
Institute of Precision Medicine, The First Affiliated Hospital, Sun Yat-Sen University, No. 58, Zhongshan 2 Road, Guangzhou 510080, China.
Burns Trauma. 2025 Mar 4;13:tkae065. doi: 10.1093/burnst/tkae065. eCollection 2025.
Diabetic foot ulcer (DFU) is one of the most common and complex complications of diabetes, but the underlying pathophysiology remains unclear. Single-cell RNA sequencing (scRNA-seq) has been conducted to explore novel cell types or molecular profiles of DFU from various perspectives. This study aimed to comprehensively analyze the potential mechanisms underlying impaired re-epithelization of DFU in a single-cell perspective.
We conducted scRNA-seq on tissues from human normal skin, acute wound, and DFU to investigate the potential mechanisms underlying impaired epidermal differentiation and the pathological microenvironment. Pseudo-time and lineage inference analyses revealed the distinct states and transition trajectories of epidermal cells under different conditions. Transcription factor analysis revealed the potential regulatory mechanism of key subtypes of keratinocytes. Cell-cell interaction analysis revealed the regulatory network between the proinflammatory microenvironment and epidermal cells. Laser-capture microscopy coupled with RNA sequencing (LCM-seq) and multiplex immunohistochemistry were used to validate the expression and location of key subtypes of keratinocytes.
Our research provided a comprehensive map of the phenotypic and dynamic changes that occur during epidermal differentiation, alongside the corresponding regulatory networks in DFU. Importantly, we identified two subtypes of keratinocytes: basal cells (BC-2) and diabetes-associated keratinocytes (DAK) that might play crucial roles in the impairment of epidermal homeostasis. BC-2 and DAK showed a marked increase in DFU, with an inactive state and insufficient motivation for epidermal differentiation. BC-2 was involved in the cellular response and apoptosis processes, with high expression of TXNIP, IFITM1, and IL1R2. Additionally, the pro-differentiation transcription factors were downregulated in BC-2 in DFU, indicating that the differentiation process might be inhibited in BC-2 in DFU. DAK was associated with cellular glucose homeostasis. Furthermore, increased CCL2 + CXCL2+ fibroblasts, VWA1+ vascular endothelial cells, and GZMA+CD8+ T cells were detected in DFU. These changes in the wound microenvironment could regulate the fate of epidermal cells through the TNFSF12-TNFRSF12A, IFNG-IFNGR1/2, and IL-1B-IL1R2 pathways, which might result in persistent inflammation and impaired epidermal differentiation in DFU.
Our findings offer novel insights into the pathophysiology of DFU and present potential therapeutic targets that could improve wound care and treatment outcomes for DFU patients.
糖尿病足溃疡（DFU）是糖尿病最常见且复杂的并发症之一，但其潜在的病理生理学机制仍不清楚。已开展单细胞RNA测序（scRNA-seq）从多个角度探索DFU的新型细胞类型或分子特征。本研究旨在从单细胞角度全面分析DFU再上皮化受损的潜在机制。
我们对来自人类正常皮肤、急性伤口和DFU的组织进行scRNA-seq，以研究表皮分化受损和病理微环境的潜在机制。拟时间和谱系推断分析揭示了不同条件下表皮细胞的不同状态和转变轨迹。转录因子分析揭示了角质形成细胞关键亚型的潜在调控机制。细胞-细胞相互作用分析揭示了促炎微环境与表皮细胞之间的调控网络。激光捕获显微镜联合RNA测序（LCM-seq）和多重免疫组化用于验证角质形成细胞关键亚型的表达和定位。
我们的研究提供了一张表皮分化过程中发生的表型和动态变化的综合图谱，以及DFU中相应的调控网络。重要的是，我们鉴定出两种角质形成细胞亚型：基底细胞（BC-2）和糖尿病相关角质形成细胞（DAK），它们可能在表皮稳态受损中起关键作用。BC-2和DAK在DFU中显著增加，处于非活性状态且表皮分化动力不足。BC-2参与细胞反应和凋亡过程，TXNIP、IFITM1和IL1R2表达较高。此外，促分化转录因子在DFU的BC-2中下调，表明DFU中BC-2的分化过程可能受到抑制。DAK与细胞葡萄糖稳态相关。此外，在DFU中检测到CCL2 + CXCL2 +成纤维细胞、VWA1 +血管内皮细胞和GZMA + CD8 + T细胞增加。伤口微环境的这些变化可通过TNFSF12-TNFRSF12A、IFNG-IFNGR1/2和IL-1B-IL1R2途径调节表皮细胞的命运，这可能导致DFU中持续的炎症和表皮分化受损。
我们的研究结果为DFU的病理生理学提供了新的见解，并提出了潜在的治疗靶点，有望改善DFU患者的伤口护理和治疗效果。
