Kobatake Tetsuya, Miyamoto Yoshiyuki, Fujihara Yuko, Saijo Hideto, Hoshi Kazuto, Hikita Atsuhiko
Department of Sensory and Motor System Medicine, Graduate School of Medicine, The University of Tokyo, Hongo 7-3-1, Bunkyo-ku, Tokyo 113-8655, Japan.
Division of Dentistry and Oral Surgery, Mitsui Memorial Hospital, Kanda-Izumi-cho 1, Chiyoda-ku, Tokyo, 101-8643, Japan.
Regen Ther. 2025 Jan 24;28:421-430. doi: 10.1016/j.reth.2025.01.009. eCollection 2025 Mar.
Elucidation of the paracrine interaction between chondrocytes and macrophages is useful for understanding the mechanisms of cartilage regeneration. Extracellular vesicles are granular substances with a diameter of approximately 150 nm, surrounded by a phospholipid bilayer membrane. In recent years, research has been conducted on clinical applications of extracellular vesicles. It has been shown that macrophages promote cartilage maturation, and macrophages acquire anti-inflammatory properties through cartilage, but the detailed mechanism of paracrine action involving extracellular vesicles remains unclear. Therefore, we focused on the effect of chondrocyte-derived extracellular vesicles on changes in macrophage characteristics.
Macrophages induced with granulocyte-macrophage colony stimulating factor (M1-like macrophages) and auricular chondrocytes were co-cultured using cell culture inserts and exosome inhibitors, and the expression of macrophage markers were analyzed. Next, extracellular vesicles separated from auricular chondrocytes were added to in vitro macrophage culture medium, and time-lapse observations of macrophage uptake of auricular chondrocyte-derived extracellular vesicles were performed. In addition, the effects of extracellular vesicles on the expression of macrophage markers were also analyzed.
The expression of CD206, an M2 macrophage marker, was increased in macrophages due to the paracrine effect of chondrocytes, and CD206 expression was further increased by pharmacological inhibition of chondrocyte-derived exosomes. It was shown that chondrocyte-derived extracellular vesicles were taken up by macrophages and promoted the production of interleukin-10, an anti-inflammatory cytokine while reducing CD206 expression.
Auricular chondrocyte-derived extracellular vesicles promoted the production of interleukin-10 in bone marrow M1-like macrophages but reduced CD206 expression.
阐明软骨细胞与巨噬细胞之间的旁分泌相互作用有助于理解软骨再生机制。细胞外囊泡是直径约为150纳米的颗粒物质，被磷脂双分子层膜包围。近年来，人们对细胞外囊泡的临床应用进行了研究。研究表明，巨噬细胞可促进软骨成熟，且巨噬细胞通过软骨获得抗炎特性，但涉及细胞外囊泡的旁分泌作用的详细机制仍不清楚。因此，我们重点研究了软骨细胞衍生的细胞外囊泡对巨噬细胞特性变化的影响。
使用细胞培养插入物和外泌体抑制剂将经粒细胞-巨噬细胞集落刺激因子诱导的巨噬细胞（M1样巨噬细胞）与耳廓软骨细胞共培养，并分析巨噬细胞标志物的表达。接下来，将从耳廓软骨细胞中分离出的细胞外囊泡添加到体外巨噬细胞培养基中，并对巨噬细胞摄取耳廓软骨细胞衍生的细胞外囊泡进行延时观察。此外，还分析了细胞外囊泡对巨噬细胞标志物表达的影响。
由于软骨细胞的旁分泌作用，巨噬细胞中M2巨噬细胞标志物CD206的表达增加，并且通过药物抑制软骨细胞衍生的外泌体可进一步增加CD206的表达。结果表明，软骨细胞衍生的细胞外囊泡被巨噬细胞摄取，促进了抗炎细胞因子白细胞介素-10的产生，同时降低了CD206的表达。
耳廓软骨细胞衍生的细胞外囊泡促进了骨髓M1样巨噬细胞中白细胞介素-10的产生，但降低了CD206的表达。
