Department of Regenerative Medical Science, School of Pharmaceutical Sciences, Jilin University, Changchun, 130021, People's Republic of China.
Alihe Community Health Service Center, Elunchun Banner, Inner Mongolia Autonomous Region, Inner Mongolia, 165450, People's Republic of China.
Toxicology. 2020 Apr 30;436:152429. doi: 10.1016/j.tox.2020.152429. Epub 2020 Mar 7.
Excessive systemic uptake of inorganic fluorides causes disturbances of bone homeostasis. The mechanism of skeletal fluorosis is still uncertain. This study aimed to study the effect of fluoride on osteocyte-driven osteoclastogenesis and probe into the role of PTH in this process. IDG-SW3 cells seeded in collagen-coated constructs were developed into osteocyte-like cells through induction of mineral agents. Then, osteocyte-like cells were exposed to fluoride in the presence or absence of parathyroid hormone (PTH). Cell viability and their capacity to produce receptor activator of nuclear factor kappa-B ligand (RANKL), osteoprotegerin (OPG) and sclerostin (SOST) were detected by MTT and Western blot assays, respectively. Finally, a transwell coculture system using osteocyte-like cells seeded in the low compartment, and osteoclast precursors added in the inserts was developed to observe the osteocyte-driven osteoclasogenesis response to fluoride with or without PTH, and the expression of molecules involved in this mechanism were measure by real time RT-PCR. Results showed that osteocytes withstood a toxic dose of fluoride, and yet PTH administration significantly reduced osteocytes viability. PTH amplified the effect of fluoride on the expression of osteoclastogenesis-related molecules in osteocyte, but did not enlarged the stimulating effect of fluoride on osteoclastogenesis drove by osteocyte coculture. Gene expression levels of TRAP, RANK, JNK and NFAtc1 significantly increased in fluoride affected osteoclast precursor cocultured with osteocyte-like cells. The impact of fluoride on osteocyte-driven osteoclast differentiation was stronger than that of PTH. In conclusion, osteocyte played a pivotal role on the mechanism underlying fluoride-affected osteoclastogenesis in which RANK-JNK-NFATc1 signaling pathway was involved, and PTH had a significant impact in this process.
过量的无机氟化物摄取会导致骨内稳态紊乱。骨氟中毒的发病机制尚不清楚。本研究旨在研究氟化物对破骨细胞生成的影响，并探讨甲状旁腺激素（PTH）在此过程中的作用。将 IDG-SW3 细胞接种于胶原包被的构建物中，通过矿化剂诱导分化为成骨细胞样细胞。然后，将成骨细胞样细胞在有无甲状旁腺激素（PTH）的情况下暴露于氟化物中。通过 MTT 和 Western blot 检测细胞活力及其产生核因子κB 受体激活剂配体（RANKL）、骨保护素（OPG）和硬化素（SOST）的能力。最后，建立了一个使用低室中成骨细胞样细胞接种和插入物中添加破骨细胞前体的 Transwell 共培养系统，以观察有无 PTH 存在时氟化物对成骨细胞驱动的破骨细胞生成的影响，并通过实时 RT-PCR 测量涉及该机制的分子的表达。结果表明，成骨细胞能耐受氟化物的毒性剂量，但 PTH 给药显著降低了成骨细胞的活力。PTH 放大了氟化物对成骨细胞中破骨细胞生成相关分子表达的影响，但没有放大氟化物对成骨细胞共培养驱动的破骨细胞生成的刺激作用。在与成骨细胞样细胞共培养的破骨细胞前体中，TRAP、RANK、JNK 和 NFAtc1 的基因表达水平显著增加。氟化物对成骨细胞驱动的破骨细胞分化的影响强于 PTH。总之，成骨细胞在氟化物影响破骨细胞生成的机制中起着关键作用，其中涉及 RANK-JNK-NFATc1 信号通路，而 PTH 在这一过程中具有重要影响。
