Department of Cardiology and Echocardiography, The First Hospital of Jilin University, Changchun, China.
Department of Orthopedic Trauma, The First Hospital of Jilin University, Changchun, China.
Am J Physiol Cell Physiol. 2020 Jan 1;318(1):C73-C82. doi: 10.1152/ajpcell.00273.2019. Epub 2019 Oct 2.
Our objective was to investigate the role of primary cilia in low-magnitude, high-frequency vibration (LMHFV) treatment of MC3T3-E1 osteoblasts (OBs). We used chloral hydrate (CH), which has a well-characterized function in chemically removing primary cilia, to elucidate the role of primary cilia in LMHFV-induced OB osteogenic responses through cell viability assay, Western blot analysis, real-time quantitative RT-PCR, and histochemical staining methods. We observed a significant, 30% decrease in the number of MC3T3-E1 OBs with primary cilia (reduced from 64.3 ± 5%) and an approximately 50% reduction in length of primary cilia (reduced from 3 ± 0.8 μm) after LMHFV stimulation. LMHFV stimulation upregulated protein expression of the bone matrix markers collagen 1 (COL-1), osteopontin (OPN), and osteoclacin(OCN) in MC3T3-E1 OBs, indicating that LMHFV induces osteogenesis. High-concentration or long-duration CH exposure resulted in inhibition of MC3T3-E1 OB survival. In addition, Western blot analysis and RT-PCR revealed that CH treatment prevented LMHFV-induced osteogenesis. Furthermore, decreased alkaline phosphate activity, reduced OB differentiation, mineralization, and maturation were observed in CH-pretreated and LMHFV-treated OBs. We showed that LMHFV induces morphological changes in primary cilia that may fine-tune their mechanosensitivity. In addition, we demonstrated the significant inhibition by CH of LMHFV-induced OB mineralization, maturation, and differentiation, which might reveal the critical role of primary cilia in the process.
我们的目的是研究初级纤毛在低幅度、高频率振动（LMHFV）治疗 MC3T3-E1 成骨细胞（OBs）中的作用。我们使用水合氯醛（CH），它在化学去除初级纤毛方面具有良好的功能，通过细胞活力测定、Western blot 分析、实时定量 RT-PCR 和组织化学染色方法，阐明初级纤毛在 LMHFV 诱导 OB 成骨反应中的作用。我们观察到，LMHFV 刺激后，具有初级纤毛的 MC3T3-E1 OB 的数量显著减少了 30%（从 64.3±5%减少），初级纤毛的长度减少了约 50%（从 3±0.8μm 减少）。LMHFV 刺激上调了 MC3T3-E1 OB 中骨基质标志物胶原 1（COL-1）、骨桥蛋白（OPN）和骨钙素（OCN）的蛋白表达，表明 LMHFV 诱导成骨。高浓度或长持续时间的 CH 暴露导致 MC3T3-E1 OB 存活受到抑制。此外，Western blot 分析和 RT-PCR 显示，CH 处理可阻止 LMHFV 诱导的成骨作用。此外，在 CH 预处理和 LMHFV 处理的 OB 中观察到碱性磷酸酶活性降低、OB 分化、矿化和成熟减少。我们表明，LMHFV 引起初级纤毛的形态变化，可能微调其机械敏感性。此外，我们证明了 CH 对 LMHFV 诱导的 OB 矿化、成熟和分化的显著抑制作用，这可能揭示了初级纤毛在该过程中的关键作用。
