Low‑frequency pulsed electromagnetic field inhibits RANKL‑induced osteoclastic differentiation in RAW264.7 cells by scavenging reactive oxygen species

Abstract

Bone homeostasis is a dynamic balance maintained by bone formation and resorption. An increase in the number and activity of osteoclasts leads to excessive bone resorption, which in turn results in bone disease, including osteoporosis. Therefore, inhibiting the differentiation and activity of osteoclasts is important for maintaining bone mass. Several studies have revealed that the use of a low-frequency pulsed electromagnetic field (PEMF) is an effective method to treat osteoporosis. However, its exact mechanism remains to be fully clarified. Therefore, the present study was designed to examine the effects that PEMF exerts on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclastogenesis and intracellular reactive oxygen species (ROS) production in RAW264.7 cells. The viability of cells was determined using a Cell Counting Kit-8 assay, and gene and protein expression were investigated via reverse transcription-quantitative polymerase chain reaction and western blot analyses. Furthermore, microscopy was performed to detect the levels of intracellular ROS and tartrate-resistant acid phosphatase (TRAP). Following the culture of RAW264.7 cells with RANKL (50 ng/ml) for 4 days (3 h/day) under PEMF (75 Hz, 1 mt) exposure, it was observed that PEMF had an inhibitory effect on RANKL-induced osteoclastic differentiation. Multinucleated osteoclast formation, the activity of TRAP and the expression of osteoclastogenesis-associated genes, including cathepsin K, nuclear factor of activated T cells cytoplasmic 1 and TRAP, were significantly reduced by PEMF. Furthermore, PEMF effectively decreased the generation of intracellular ROS during osteoclastic differentiation. In addition, the results demonstrated that ROS are the key factor in osteoclast differentiation and formation. Reducing intracellular ROS with diphenylene-iodonium chloride significantly inhibited RANKL-induced osteoclast differentiation. Taken together, the results of the present study demonstrated that PEMF may inhibit RANKL-induced osteoclastogenesis by scavenging intracellular ROS. These results may provide the groundwork for future PEMF clinical applications in osteoclast-associated bone disease.

Post-exposure Effects of PEMF on ROS levels in H2O2-treated Glioblastoma Cell Line

“…In addition, this result is also supported by the studies of Osera et al [4] and Falone et al [5], showing that improving antioxidant response effect of short-and repeated PEMF pre-conditioning in drug-sensitive (SH-SY5Y) and drugresistant (SK-N-BE(2)) human neuroblastoma cell lines against pro-oxidant challenge by restraining H2O2-induced ROS production and increasing Mn-dependent superoxide dismutase (MnSOD) activity. The inhibitory effect of PEMF on intracellular ROS levels was also shown to inhibit osteoclastic differentiation [7,10]. Taken together, our findings could lay the framework for future research into the processes underlying the effects of post-exposed PEMFs at different doses, frequencies, and exposure conditions on intracellular ROS levels in neuronal signaling pathways, allowing us to reach a more thorough conclusion.…”

Translational Insights into Extremely Low Frequency Pulsed Electromagnetic Fields (ELF-PEMFs) for Bone Regeneration after Trauma and Orthopedic Surgery

“…In certain bone modeling phases, ROS may activate osteoclasts while inhibiting osteoblasts [63]. By inducing ROS, ELF-PEMF thus may favor osteoclastogenesis, an assumption supported by the studies of Pi and Zhang [64,65]. However, there are also several other reports showing ELF-PEMF inhibitory effects on osteoclastogenesis [66][67][68].…”

Novel Nanohydroxyapatite (nHAp)-Based Scaffold Doped with Iron Oxide Nanoparticles (IO), Functionalized with Small Non-Coding RNA (miR-21/124) Modulates Expression of Runt-Related Transcriptional Factor 2 and Osteopontin, Promoting Regeneration of Osteoporotic Bone in Bilateral Cranial Defects in a Senescence-Accelerated Mouse Model (SAM/P6). PART 2

“…This observation is in good agreement with the study of Pi et al, who showed that osteoclastogenesis decreased after the exposition of murine monocyte-macrophage lineage (RAW 264.7) to the low-frequency pulsed electromagnetic field and was associated with the lowered activity of TRAP. 37 In order to test the pro-osteogenic effectiveness of the scaffolds, we performed a bilateral cranial defect model in senescence-accelerated mouse prone 6 (SAM/P6) and evaluated the process of bone healing. Since SAM/P6 represents the model of senile osteoporosis with low bone mass and reduced bone mineral density, verification of clinical effectiveness using this model is fully justified.…”