Mesenchymal stem cells (MSCs) activate the endogenous immune regulatory system, inducing a therapeutic effect in recipients. MSCs have demonstrated the ability to modulate the differentiation of myeloid cells toward a phagocytic and anti-inflammatory profile. Allogeneic, adipose-derived MSCs (ASCs) have been investigated for the management of complex perianal fistula, with darvadstrocel being the first ASC therapy approved in Europe in March 2018. Additionally, ASCs are being explored as a potential treatment in other indications. Yet, despite these clinical advances, their mechanism of action is only partially understood. Freshly isolated human monocytes from the peripheral blood were differentiated in vitro toward M0 non-polarized macrophages (Mphs), M1 pro-inflammatory Mphs, M2 anti-inflammatory Mphs, or mature dendritic cells (mDCs) in the presence or absence of ASCs, in non-contact conditions. The phenotype and function of the differentiated myeloid populations were determined by flow cytometry, and their secretome was analyzed by OLINK technology. We also investigated the capacity of ASCs to modulate the phenotype and function of terminally differentiated M1 Mphs. The role of soluble factors interleukin (IL)-6 and prostaglandin E2 (PGE2) on the ability of ASCs to modulate myeloid cells was assessed using neutralization assays, CRISPR/Cas9 knock-down of cyclooxygenase 2 (COX-2), and ASC-conditioned medium assays using pro-inflammatory stimulus. Co-culture of monocytes in the presence of ASCs resulted in the polarization of Mphs and mDCs toward an anti-inflammatory and phagocytic phenotype. This was characterized by an increase in phagocytic receptors on the cell surface of Mphs (M0, M1, and M2) and mDCs, as well as modulation of chemokine receptors and reduced expression of pro-inflammatory, co-stimulatory molecules. ASCs also modulated the secretome of Mphs and mDCs, demonstrated by reduced expression of pro-inflammatory factors and increased expression of anti-inflammatory and reparative factors. Chemical inhibition of PGE2 with indomethacin abolished this modulatory effect, whereas treatment with a neutralizing anti-IL-6 antibody resulted in a partial abolishment. The knock-down of COX-2 in ASCs and the use of IL-1β-activated ASC-conditioned media confirmed the key role of PGE2 in ASC-mediated myeloid modulation. In our in vitro experimental settings, ASCs failed to modulate the phenotype and function of terminally polarized M1 Mphs. The results demonstrate that ASCs are able to modulate the in vitro differentiation of myeloid cells toward an anti-inflammatory and reparative profile. This modulatory effect was mediated mainly by PGE2 and, to a lesser extent, IL-6.

中文翻译：

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人脂肪间充质干细胞将骨髓细胞调节为抗炎和修复表型：IL-6 和 PGE2 的作用


间充质干细胞（MSC）激活内源性免疫调节系统，在受体中产生治疗效果。间充质干细胞已被证明能够调节骨髓细胞向吞噬细胞和抗炎细胞分化。同种异体脂肪来源的间充质干细胞 (ASC) 已被研究用于治疗复杂性肛周瘘，darvadstrocel 是 2018 年 3 月在欧洲批准的第一个 ASC 疗法。此外，ASC 正在探索作为其他适应症的潜在治疗方法。然而，尽管取得了这些临床进展，但它们的作用机制仅被部分了解。在存在或不存在 ASC 的情况下，从外周血中新鲜分离的人单核细胞在体外分化为 M0 非极化巨噬细胞 (Mphs)、M1 促炎性 Mphs、M2 抗炎性 Mphs 或成熟树突状细胞 (mDC)。非接触条件。通过流式细胞术测定分化的髓系群体的表型和功能，并通过OLINK技术分析其分泌组。我们还研究了 ASC 调节终末分化 M1 Mphs 表型和功能的能力。使用中和测定、环氧合酶 2 (COX-2) 的 CRISPR/Cas9 敲除和 ASC- 评估可溶性因子白细胞介素 (IL)-6 和前列腺素 E2 (PGE2) 对 ASC 调节骨髓细胞能力的作用。使用促炎刺激物进行条件培养基测定。在 ASC 存在的情况下共培养单核细胞导致 Mph 和 mDC 向抗炎和吞噬表型极化。 其特征是 Mphs（M0、M1 和 M2）和 mDC 细胞表面吞噬细胞受体的增加，以及趋化因子受体的调节和促炎共刺激分子表达的减少。 ASC 还调节 Mphs 和 mDC 的分泌组，这通过促炎因子表达减少和抗炎和修复因子表达增加来证明。用吲哚美辛对 PGE2 进行化学抑制消除了这种调节作用，而用中和性抗 IL-6 抗体治疗则导致部分消除。 ASC 中 COX-2 的敲除以及 IL-1β 激活的 ASC 条件培养基的使用证实了 PGE2 在 ASC 介导的骨髓调节中的关键作用。在我们的体外实验环境中，ASC 未能调节末端极化 M1 Mphs 的表型和功能。结果表明，ASC 能够调节骨髓细胞的体外分化，使其具有抗炎和修复功能。这种调节作用主要由 PGE2 介导，其次是 IL-6。