Luo Qianting, Li Xingyang, Zhong Wenchao, Cao Wei, Zhu Mingjing, Wu Antong, Chen Wanyi, Ye Zhitong, Han Qiao, Natarajan Duraipandy, Pathak Janak L, Zhang Qingbin
Department of Temporomandibular Joint, Affiliated Stomatology Hospital of Guangzhou Medical University, Guangdong Engineering Research Center of Oral Restoration and Reconstruction, Guangzhou Key Laboratory of Basic and Applied Research of Oral Regenerative Medicine, Guangzhou 510182, China.
Jiangmen Central Hospital, Affiliated Jiangmen Hospital of Sun Yat-sen University, Jiangmen 529030, China.
Regen Biomater. 2021 Dec 13;9:rbab075. doi: 10.1093/rb/rbab075. eCollection 2022.
Dicalcium silicate (CaSiO, CS) has osteogenic potential but induces macrophagic inflammation. Mitochondrial function plays a vital role in macrophage polarization and macrophagic inflammation. The mitochondrial function of CS-treated macrophages is still unclear. This study hypothesized: (i) the CS modulates mitochondrial function and autophagy in macrophages to regulate macrophagic inflammation, and (ii) CS-induced macrophagic inflammation regulates osteogenesis. We used RAW264.7 cells as a model of macrophage. The CS (75-150 μg/ml) extract was used to analyze the macrophagic mitochondrial function and macrophage-mediated effect on osteogenic differentiation of mouse bone marrow-derived mesenchymal stem cells (BMSCs). The results showed that CS extract (150 μg/ml) induced TNF-α, IL-1β and IL-6 production in macrophages. CS extract (150 μg/ml) enhanced reactive oxygen species level and intracellular calcium level but reduced mitochondrial membrane potential and ATP production. TEM images showed reduced mitochondrial abundance and altered the mitochondrial morphology in CS (150 μg/ml)-treated macrophages. Protein level expression of PINK1, Parkin, Beclin1 and LC3 was upregulated but TOMM20 was downregulated. mRNA sequencing and KEGG analysis showed that CS-induced differentially expressed mRNAs in macrophages were mainly distributed in the essential signaling pathways involved in mitochondrial function and autophagy. The conditioned medium from CS-treated macrophage robustly promoted osteogenic differentiation in BMSCs. In conclusion, our results indicate mitochondrial dysfunction and autophagy as the possible mechanism of CS-induced macrophagic inflammation. The promotion of osteogenic differentiation of BMSCs by the CS-induced macrophagic inflammation suggests the potential application of CS in developing immunomodulatory bone grafts.
硅酸二钙（CaSiO，CS）具有成骨潜力，但会引发巨噬细胞炎症。线粒体功能在巨噬细胞极化和巨噬细胞炎症中起着至关重要的作用。CS处理的巨噬细胞的线粒体功能仍不清楚。本研究假设：（i）CS调节巨噬细胞中的线粒体功能和自噬以调节巨噬细胞炎症，以及（ii）CS诱导的巨噬细胞炎症调节成骨作用。我们使用RAW264.7细胞作为巨噬细胞模型。CS（75 - 150μg/ml）提取物用于分析巨噬细胞线粒体功能以及巨噬细胞对小鼠骨髓间充质干细胞（BMSC）成骨分化的介导作用。结果表明，CS提取物（150μg/ml）诱导巨噬细胞产生TNF-α、IL-1β和IL-6。CS提取物（150μg/ml）提高了活性氧水平和细胞内钙水平，但降低了线粒体膜电位和ATP产生。透射电镜图像显示，CS（150μg/ml）处理的巨噬细胞中线粒体丰度降低且线粒体形态改变。PINK1、Parkin、Beclin1和LC3的蛋白水平表达上调，但TOMM20下调。mRNA测序和KEGG分析表明，CS诱导的巨噬细胞中差异表达的mRNA主要分布在线粒体功能和自噬相关的关键信号通路中。CS处理的巨噬细胞的条件培养基有力地促进了BMSC的成骨分化。总之，我们的结果表明线粒体功能障碍和自噬是CS诱导巨噬细胞炎症的可能机制。CS诱导的巨噬细胞炎症对BMSC成骨分化的促进作用表明CS在开发免疫调节性骨移植方面具有潜在应用价值。
