Macrophage-derived neurotrophin-3 promotes heterotopic ossification in rats
Macrophage-derived neurotrophin-3 promotes heterotopic ossification in rats
复制标题
巨噬细胞源性神经营养素 3 促进大鼠异位骨化
DOI：
10.1038/s41374-019-0367-x
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发表时间：
2020-01-02
影响因子：
5
通讯作者：
Zhang, Zhongmin
中科院分区：
文献类型：
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作者：
Zhang, Jie;Wang, Liang;Zhang, Zhongmin
异位骨化（HO）是一种由创伤性损伤或遗传疾病导致的使人衰弱的病症，其潜在机制仍不明确。最近，我们已证实神经营养因子 - 3（NT - 3）的表达及其通过介导血管内皮细胞的内皮 - 间充质转化（EndMT）促进异位骨化形成的作用。本研究探究了NT - 3在大鼠受伤跟腱异位骨化形成过程中对周围间充质细胞的作用及其潜在来源。我们采用跟腱切断术在体内诱导异位骨化形成，并培养原代肌腱来源干细胞（TDSCs）以在体外研究介导成骨作用的潜在机制。此外，利用RAW264.7细胞来确定NT - 3的来源。在损伤部位，神经生长因子（NGF）、脑源性神经营养因子（BDNF）、NT - 3和NT - 4及其酪氨酸蛋白激酶（Trk）受体以及p75受体的mRNA水平均升高。NT - 3和TrkC的诱导程度最高。泛Trk抑制剂GNF5837中和NT - 3诱导的效应后，骨/软骨相关基因的表达降低，而注射NT - 3则促进了异位骨化的形成，损伤部位骨/软骨相关标志物的mRNA水平升高。在体外，NT - 3通过激活ERK1/2和PI3K/Akt信号通路加速了TDSCs的成骨分化和矿化。此外，在整个异位骨化形成过程中，在损伤部位观察到NT - 3与巨噬细胞（包括M1和M2巨噬细胞）共定位，体外研究表明活化的巨噬细胞介导了NT - 3的分泌。另外，在体内和体外均观察到血清或上清液中NT - 3浓度升高。用载有氯膦酸盐的脂质体消耗巨噬细胞可减少异位骨化的形成以及NT - 3的分泌和mRNA表达。我们的研究表明，巨噬细胞来源的NT - 3可能通过ERK1/2和PI3K/Akt信号通路促进异位骨化形成和TDSCs的成骨作用，这可能为未来异位骨化的治疗方向提供新的见解。
Heterotopic ossification (HO) is a debilitating condition that results from traumatic injuries or genetic diseases, for which the underlying mechanisms remain unclear. Recently, we have demonstrated the expression of neurotrophin-3 (NT-3) and its role in promoting HO formation via mediating endothelial-mesenchymal transition (EndMT) of vascular endothelial cells. The current study investigated the role of NT-3 on the surrounding mesenchymal cells and its potential origin throughout HO formation at injured Achilles tendons in rats. We used an Achilles tenotomy to induce HO formation in vivo and cultured primary tendon-derived stem cells (TDSCs) to investigate the underlying mechanisms mediating the osteogenesis in vitro. Furthermore, RAW264.7 cells were employed to identify the origin of NT-3. The mRNA levels of NGF, BDNF, NT-3, and NT-4 and their tyrosine protein kinase (Trk) receptors as well as p75 receptor were elevated at injury sites. NT-3 and TrkC showed the highest induction. Neutralization of the NT-3-induced effects by the pan-Trk inhibitor GNF5837 reduced the expression of bone/cartilage-related genes while injection of NT-3 promoted HO formation with elevated mRNA of bone/cartilage-related markers at injured sites. In vitro, NT-3 accelerated osteogenic differentiation and mineralization of TDSCs through activation of the ERK1/2 and PI3K/Akt signaling pathways. Moreover, the colocalization of NT-3 and macrophages, including M1 and M2 macrophages, was observed in injured sites throughout HO formation, and in vitro studies demonstrated that activated macrophages mediated the secretion of NT-3. In addition, an increasing concentration of serum or supernatant NT-3 was observed both in vivo and in vitro. Depletion of macrophages with clodronate-loaded liposomes reduced HO formation as well as secretion and mRNA expression of NT-3. Our study suggests that macrophage-derived NT-3 may promote HO formation and osteogenesis of TDSCs via the ERK1/2 and PI3K/Akt signaling pathways, which may provide new insights for the therapeutic directions of HO in the future.