Objectives Prior work demonstrates that co-cultured macrophages and fibroblasts from patients with systemic sclerosis (SSc) engage in reciprocal activation. However, the mechanism by which these cell types communicate and contribute to fibrosis and inflammation in SSc is unknown. Methods Fibroblasts were isolated from skin biopsies obtained from 7 SSc patients or 6 healthy age and gender-matched control subjects following written informed consent. Human donor-derived macrophages were cultured with exosomes isolated from control or SSc fibroblasts for an additional 48 h. Macrophages were immunophenotyped using flow cytometry, qRT-PCR and multiplex. For mutual activation studies, exosome-activated macrophages were co-cultured with SSc or healthy fibroblasts using Transwells. Results Macrophages activated with dermal fibroblast-derived exosomes from SSc patients upregulated surface expression of CD163, CD206, MHC Class II and CD16 and secreted increased levels of IL-6, IL-10, IL-12p40 and TNF compared with macrophages incubated with healthy control fibroblasts (n = 7, p< 0.05). Exosome-stimulated macrophages and SSc fibroblasts engaged in reciprocal activation, as production of collagen and fibronectin was significantly increased in SSc fibroblasts receiving signals from SSc exosome-stimulated macrophages (n = 7, p< 0.05). Conclusion In this work, we demonstrate for the first time that human SSc dermal fibroblasts mediate macrophage activation through exosomes. Our findings suggest that macrophages and fibroblasts engage in cross-talk in SSc skin, resulting in mutual activation, inflammation, and ECM deposition. Collectively, these studies implicate macrophages and fibroblasts as cooperative mediators of fibrosis in SSc and suggest therapeutic targeting of both cell types may provide maximal benefit in ameliorating disease in SSc patients.

中文翻译：

---

人真皮成纤维细胞来源的外泌体在系统性硬化症中诱导巨噬细胞活化

目标 先前的工作表明，来自系统性硬化症 (SSc) 患者的共培养巨噬细胞和成纤维细胞会相互激活。然而，这些细胞类型在 SSc 中沟通并导致纤维化和炎症的机制尚不清楚。方法 在签署书面知情同意书后，从 7 名 SSc 患者或 6 名年龄和性别匹配的健康对照受试者的皮肤活检中分离出成纤维细胞。将人类供体来源的巨噬细胞与从对照或 SSc 成纤维细胞分离的外泌体一起培养另外 48 小时。使用流式细胞术、qRT-PCR 和多重分析对巨噬细胞进行免疫表型分析。对于相互激活研究，使用 Transwell 将外泌体激活的巨噬细胞与 SSc 或健康成纤维细胞共培养。结果 与健康对照巨噬细胞相比，SSc 患者真皮成纤维细胞衍生的外泌体激活的巨噬细胞上调 CD163、CD206、MHC II 类和 CD16 的表面表达，并分泌更高水平的 IL-6、IL-10、IL-12p40 和 TNF成纤维细胞(n＝7，p＜0.05)。外泌体刺激的巨噬细胞和SSc成纤维细胞参与相互激活，因为在接收来自SSc外泌体刺激的巨噬细胞的信号的SSc成纤维细胞中胶原和纤连蛋白的产生显着增加（n=7，p＜0.05）。结论在这项工作中，我们首次证明人 SSc 真皮成纤维细胞通过外泌体介导巨噬细胞活化。我们的研究结果表明，巨噬细胞和成纤维细胞在 SSc 皮肤中进行串扰，导致相互激活、炎症和 ECM 沉积。总的来说，这些研究表明巨噬细胞和成纤维细胞是 SSc 纤维化的协同介质，并表明针对这两种细胞类型的治疗可能为改善 SSc 患者的疾病提供最大益处。