State Key Laboratory of Genetic Engineering, Collaborative Innovation Center for Genetics and Development, School of Life Sciences, and Human Phenome Institute, Fudan University, Shanghai, China.
Greater Bay Area Institute of Precision Medicine (Guangzhou), School of Life Sciences, Fudan University, Guangzhou, China.
Br J Dermatol. 2024 Nov 18;191(6):964-978. doi: 10.1093/bjd/ljae286.
Skin fibrosis is the typical pathological manifestation of systemic sclerosis (SSc) and localized scleroderma (LS); it has an unclear aetiology and few effective treatments. Although excessive collagen secretion by fibroblasts is the primary cause of skin fibrosis, evidence has suggested that vascular damage is the initiating event and that various cell types, including fibroblasts, work together to contribute to the pathogenesis of skin fibrosis.
To explore the relationship between vascular endothelial cell lesions and immune cell infiltration, along with the interactions between various cell types within the fibrotic skin ecosystem.
Single-cell RNA sequencing was performed on skin biopsies from three healthy donors and seven patients with SSc. Additional data from three patients with localized scleroderma available in the Gene Expression Omnibus (GSE160536) were integrated by Harmony. CellChat (version 1.5.0) was used to analyse the cell communication network. A Transwell® assay and a bleomycin (BLM) mouse model were used to explore the role of atypical chemokine receptor 1 (ACKR1; 'Duffy antigen') in immune cell infiltration. Milo single-cell Western blot was used to show fibroblast subcluster activation.
A total of 62 295 cells were obtained and subpopulations of stromal and immune cells identified. Interaction network analysis found that multiple chemokines secreted by macrophages, pericytes and proinflammatory fibroblasts could bind with ACKR1, which was highly expressed by endothelial cells in lesional skin. The Transwell® assay revealed that overexpression of ACKR1 in human umbilical vein endothelial cells facilitated leucocyte infiltration following treatment with interleukin-8. BLM mice showed enhanced ACKR1 expression, massive immune cell infiltration and skin fibrosis that was attenuated by ACKR1 inhibition. Furthermore, infiltrated macrophages expressing high levels of transforming growth factor (TGF)-β1 or platelet-derived growth factor B (PDGFB) could activate secreted frizzled-related protein 2 (SFRP2)/asporin (ASPN)+ fibroblasts to contribute to the excessive accumulation of extracellular matrix. It was also found that the SOX4-ASPN axis plays an important role in the TGF-β signalling cascade and the aetiology of skin fibrosis.
Our results reveal that high expression of ACKR1 by endothelial cells in fibrotic skin tissue promotes immune cell infiltration and that SFRP2/ASPN+ fibroblasts synergize to exacerbate skin fibrosis.
皮肤纤维化是系统性硬化症（SSc）和局限性硬皮病（LS）的典型病理学表现；其病因尚不清楚，治疗方法也很少。虽然成纤维细胞过度分泌胶原是皮肤纤维化的主要原因，但有证据表明血管损伤是起始事件，各种细胞类型（包括成纤维细胞）共同作用导致皮肤纤维化的发病机制。
探讨血管内皮细胞损伤与免疫细胞浸润的关系，以及纤维化皮肤生态系统中各种细胞类型之间的相互作用。
对 3 名健康供体和 7 名 SSc 患者的皮肤活检组织进行单细胞 RNA 测序。通过 Harmony 整合了 Gene Expression Omnibus（GSE160536）中另外 3 名局限性硬皮病患者的数据。使用 CellChat（版本 1.5.0）分析细胞通讯网络。通过 Transwell®测定和博来霉素（BLM）小鼠模型探讨非典型趋化因子受体 1（ACKR1；“达菲抗原”）在免疫细胞浸润中的作用。Milo 单细胞 Western blot 用于显示成纤维细胞亚群的激活。
共获得 62295 个细胞，鉴定出基质和免疫细胞亚群。相互作用网络分析发现，巨噬细胞、周细胞和促炎成纤维细胞分泌的多种趋化因子可与内皮细胞中高表达的 ACKR1 结合。Transwell®测定显示，白细胞介素-8 处理后，人脐静脉内皮细胞中 ACKR1 的过表达促进白细胞浸润。BLM 小鼠表现出 ACKR1 表达增强、大量免疫细胞浸润和皮肤纤维化，ACKR1 抑制可减轻皮肤纤维化。此外，浸润的巨噬细胞表达高水平的转化生长因子（TGF）-β1 或血小板衍生生长因子 B（PDGFB）可激活分泌卷曲相关蛋白 2（SFRP2）/无翅型 MMTV 整合位点家族成员 2（ASPN）+成纤维细胞，导致细胞外基质过度积累。还发现 SOX4-ASPN 轴在 TGF-β信号级联和皮肤纤维化发病机制中起重要作用。
我们的研究结果表明，纤维化皮肤组织中内皮细胞高表达 ACKR1 可促进免疫细胞浸润，SFRP2/ASPN+成纤维细胞协同作用可加重皮肤纤维化。
