Center for Craniofacial Regeneration, Dept. of Oral Biology, University of Pittsburgh School of Dental Medicine, Pittsburgh, PA, USA.
Center for Craniofacial Regeneration, Dept. of Oral Biology, University of Pittsburgh School of Dental Medicine, Pittsburgh, PA, USA; McGowan Institute for Regenerative Medicine, University of Pittsburgh, Pittsburgh, PA, USA.
Free Radic Biol Med. 2020 Jun;153:103-111. doi: 10.1016/j.freeradbiomed.2020.04.008. Epub 2020 Apr 21.
Reactive Oxygen Species (ROS) are a natural byproduct of oxygen metabolism. At physiological levels, ROS regulate multiple cellular processes like proliferation, migration, and differentiation. Increased levels of ROS are associated with pathological conditions, such as inflammation and vascular calcification, where they elicit cytotoxic effects. These contrasting outcomes of ROS have also been reported in osteogenic precursor cells. However, the role of ROS in committed osteogenic cells has not been investigated. Cytotoxic and physiologic effects have also been demonstrated for extracellular phosphate (P). Specifically, in committed osteogenic cells P stimulates their major function (mineralization), however in osteogenic precursors and endothelial cells P cytotoxicity has been reported. Interestingly, P cytotoxic effects have been associated with ROS production in the pathological vascular mineralization. In this study, we investigated a molecular mechanistic link between elevated P and ROS production in the context of the mineralization function of committed osteogenic cells. Using committed osteogenic cells, 17IIA11 odontoblast-like cell and MLO-A5 osteoblast cell lines, we have unveil that P enhances intracellular ROS production. Furthermore, using a combination of mineralization assays and gene expression analyses, we determined that P-induced intracellular ROS supports the physiological mineralization process. In contrast, the exogenous ROS, provided in a form of HO, was detrimental for osteogenic cells. By comparing molecular signaling cascades induced by extracellular ROS and P, we identified differences in signaling routes that determine physiologic versus toxic effect of ROS on osteogenic cells. Specifically, while both extracellular and P-induced intracellular ROS utilize Erk1/2 signaling mediator, only extracellular ROS induces stress-activated mitogen-activated protein kinases P38 and JNK that are associated with cell death. In summary, our results uncovered a physiological role of ROS in the P-induced mineralization through the molecular pathway that is distinct from ROS-induced cytotoxic effects.
活性氧 (ROS) 是氧代谢的自然产物。在生理水平下，ROS 调节多种细胞过程，如增殖、迁移和分化。ROS 水平升高与炎症和血管钙化等病理状况有关，在这些情况下，ROS 会产生细胞毒性作用。ROS 在成骨前体细胞中也有报道过这种相反的作用。然而，ROS 在成骨细胞中的作用尚未被研究过。细胞毒性和生理作用也已在细胞外磷酸盐 (P) 中得到证实。具体来说，在成骨细胞中，P 刺激它们的主要功能（矿化），然而在成骨前体细胞和内皮细胞中，已经报道了 P 的细胞毒性。有趣的是，在病理性血管钙化中，P 的细胞毒性作用与 ROS 的产生有关。在这项研究中，我们研究了在成骨细胞矿化功能的背景下，升高的 P 与 ROS 产生之间的分子机制联系。使用成骨细胞，17IIA11 牙本质样细胞和 MLO-A5 成骨细胞系，我们发现 P 增强了细胞内 ROS 的产生。此外，通过矿化测定和基因表达分析的组合，我们确定 P 诱导的细胞内 ROS 支持生理矿化过程。相比之下，外源性 ROS，以 HO 的形式提供，对成骨细胞是有害的。通过比较细胞外 ROS 和 P 诱导的分子信号级联，我们确定了决定 ROS 对成骨细胞产生生理或毒性作用的信号途径的差异。具体来说，虽然细胞外和 P 诱导的细胞内 ROS 都利用 Erk1/2 信号转导介质，但只有细胞外 ROS 诱导与细胞死亡相关的应激激活丝裂原激活的蛋白激酶 P38 和 JNK。总之，我们的结果揭示了 ROS 在 P 诱导的矿化中的生理作用，通过与 ROS 诱导的细胞毒性作用不同的分子途径。
