Pulsed electromagnetic fields promote bone formation by activating the sAC–cAMP–PKA–CREB signaling pathway

Abstract

The application of pulsed electromagnetic fields (PEMFs) in the prevention and treatment of osteoporosis has long been an area of interest. However, the clinical application of PEMFs remains limited because of the poor understanding of the PEMF action mechanism. Here, we report that PEMFs promote bone formation by activating soluble adenylyl cyclase (sAC), cyclic adenosine monophosphate (cAMP), protein kinase A (PKA), and cAMP response element-binding protein (CREB) signaling pathways. First, it was found that 50 Hz 0.6 millitesla (mT) PEMFs promoted osteogenic differentiation of rat calvarial osteoblasts (ROBs), and that PEMFs activated cAMP-PKA-CREB signaling by increasing intracellular cAMP levels, facilitating phosphorylation of PKA and CREB, and inducing nuclear translocation of phosphorylated (p)-CREB. Blocking the signaling by adenylate cyclase (AC) and PKA inhibitors both abolished the osteogenic effect of PEMFs. Second, expression of sAC isoform was found to be increased significantly by PEMF treatment. Blocking sAC using sAC-specific inhibitor KH7 dramatically inhibited the osteogenic differentiation of ROBs. Finally, the peak bone mass of growing rats was significantly increased after 2 months of PEMF treatment with 90 min/day. The serum cAMP content, p-PKA, and p-CREB as well as the sAC protein expression levels were all increased significantly in femurs of treated rats. The current study indicated that PEMFs promote bone formation in vitro and in vivo by activating sAC-cAMP-PKA-CREB signaling pathway of osteoblasts directly or indirectly.

Sinusoidal Electromagnetic Fields Increase Peak Bone Mass in Rats by Activating Wnt10b/β-Catenin in Primary Cilia of Osteoblasts

“…The solenoid was 27 cm long and 14 cm in diameter, in which EMF signals of high uniformity could be generated, thus ensuring all cells were subjected to the same intensity of EMFs . The EMF generator used to treat the experimental rats was based on our same patent, and could provide a homogenous range of EMFs in which all rats were exposed to same EMFs in a cage . The EMFs, including static magnetic fields, continuous (sinusoidal, triangular, and square waveforms), and PEMFs, could be produced within the range of frequencies at 5 to 100 Hz and intensities at 0.0 to 5.0 mT.…”

“…(18) The EMF generator used to treat the experimental rats was based on our same patent, and could provide a homogenous range of EMFs in which all rats were exposed to same EMFs in a cage. (20) The EMFs, including static magnetic fields, continuous (sinusoidal, triangular, and square waveforms), and PEMFs, could be produced within the range of frequencies at 5 to 100 Hz and intensities at 0.0 to 5.0 mT. The generator was calibrated by the staff at the Department of Magnetic Fields at the Institute of Modern Physics in Lanzhou, China.…”

“…(18) This role of the primary cilia as the cellular sensors of EMFs was further identified by our finding that the promoting effects were achieved by activating bone morphogenetic protein (BMP-2)/ Smad1/5/8 signaling pathway through the BMP type-2 receptor (BMPRII), which was localized at the bases of primary cilia. (19) Besides, PEMFs were also found to promote bone formation by activating the soluble adenylyl cyclase (sAC)/cAMP/protein kinase A (PKA)/cAMP response element-binding protein (CREB) signaling pathway, (20) and sAC and p-PKA were both localized at the primary cilia of osteoblasts. (21) These results indicate that primary cilia may play an indispensable role in PEMF-stimulated osteogenesis by controlling the distribution and expression of certain important receptors and signaling proteins.…”

Novel functions of the primary cilium in bone disease and cancer

“…Previous investigators have shown that cAMP signaling is localized in the primary cilium and plays a role in the dynamic modulation of ciliary length (Besschetnova et al, 2010). Blocking cAMP signaling with adenylate cyclase or PKA inhibitors abolished the osteogenic effect of PEMFs Wang, Li, et al, 2018;Wang, Pu, et al, 2018). In addition, phosphorylated PKA was also found to localize in the primary cilium; this protein is preferentially localized to the entire cilium and sometimes to the ciliary base and/or tip in osteoblasts Shi, Xie, et al, 2017).…”

“…PEMFs activate cAMP/PKA/CREB signaling by increasing intracellular cAMP levels, facilitating the phosphorylation of PKA and CREB, and inducing the nuclear translocation of phosphorylated CREB both in vitro and in vivo. Blocking cAMP signaling with adenylate cyclase or PKA inhibitors abolished the osteogenic effect of PEMFs Wang, Li, et al, 2018;Wang, Pu, et al, 2018).…”