The effects of chrysin on osteoclastogenesis in RAW264.7 murine macrophages and bone metabolism in Sprague Dawley rats

Abstract

Bone is regarded as a metabolically active tissue that is constantly being reformed and resorbed by osteoblasts and osteoclasts. Abnormal increases in osteoclast productivity can result in bone deterioration. Receptor activator of nuclear factor-kappa B (RANK) signalling is critical for function and formation of osteoclasts. This study aimed to investigate the potential beneficial effects on bone from chrysin, a bioactive phytochemical. Western blotting was carried out to assess the impact of chrysin on the RANK signalling pathways. Immunofluorescence was conducted to determine how chrysin effects the translocation of nuclear factor kappa B (NFκB) to the nucleus. Micro-computed tomography (microCT) was utilised to assess the tibia bones in Sprague-Dawley rats. The parameters evaluated of the whole bone, length, volume, surface area, bone volume fraction, segmented surface density, and the thickness, number, and spacing of trabecular bone. The parameters evaluated for the midpoint of the bone, volume, area, bone volume fraction, segmented surface density, thickest and thinnest part of the bone midpoint. The IC50 (14µM) was determined in a pilot study performed previously. Western blotting revealed no significant changes in the phosphorylation of mitogen-activated protein kinases (MAPKs), specifically ERK, JNK, and P38, and NFκB signally pathways when treated to chrysin. Immunofluorescence investigation showed a significant inhibition in the amount of NFκB that translocated to the nucleus from the cytoplasm when treated to chrysin, when compared to the positive RANK ligand (RANKL) control. Micro-CT analysis of tibial bones indicated no significant differences in most of the parameters analysed except when an unpaired T-test was performed comparing the control group and the chrysin treated group. Chrysin significantly increased the area, volume, and segmented bone density of the midpoint of the bones. Overall, the findings suggest that chrysin inhibits osteoclastogenesis via the NFκB signalling pathway, by inhibiting the translocation from the cytoplasm to the nucleus. These findings highlight chrysin’s potential as a viable therapeutic agent in bone degenerative disorders.