A novel RANKL‐targeted flavonoid glycoside prevents osteoporosis through inhibiting NFATc1 and reactive oxygen species. Issue 5 (21st May 2021)

• Record Type: Journal Article
• Authors: Hong, Guoju; Chen, Zhenqiu; Han, Xiaorui; Zhou, Lin; Pang, Fengxiang; Wu, Rishana; Shen, Yingshan; He, Xiaoming; Hong, Zhinan; Li, Ziqi; He, Wei; Wei, Qiushi
• Is Part Of: Clinical and translational medicine. Volume 11:Issue 5(2021)
• Page Start: n/a
• Page End: n/a
• Publication Date: 2021-05-21
• Subjects: NFAcT1 -- osteoclast -- osteoporosis -- RANKL -- Robinin -- ROS; Clinical medicine -- Periodicals; Medicine, Experimental -- Periodicals; Medical innovations -- Periodicals; Molecular biology -- Periodicals; Pathology, Molecular -- Periodicals; 616.027
• DOI: 10.1002/ctm2.392
• Languages: English
• ISSNs: 2001-1326

Abstract

Abstract: Background and purpose: Osteoporosis is characterized by excessive bone resorption due to enhanced osteoclast activation. Stimulation of nuclear factor of activated T cells 1 (NFATc1) and accumulation of reactive oxygen species (ROS) are important mechanisms underlying osteoclastogenesis. Robinin (Rob) is a flavonoid glycoside that has shown anti‐inflammatory and antioxidative effects in previous studies, but little is known about its effects on bone homeostasis. The purpose of our research was to investigate whether Rob could prevent bone resorption in ovariectomized (OVX) mice by suppressing osteoclast production through its underlying mechanisms. Methods: The docking pose of Rob and RANKL was identified by protein‐ligand molecular docking. Rob was added to bone marrow macrophages (BMMs) stimulated by nuclear factor‐κB (NF‐κB) ligand (RANKL). The effects of Rob on osteoclastic activity were evaluated by positive tartrate resistant acid phosphatase (TRAcP) staining kit and hydroxyapatite resorption assay. RANKL‐induced ROS generation in osteoclasts was detected by H2 DCFDA and MitoSox Red staining. The classic molecular cascades triggered by RANKL, such as NF‐κB, ROS, calcium oscillations, and NFATc1‐mediated signaling pathways, were investigated using Fluo4 staining, western blot, and quantitative real‐time polymerase chain reaction. In addition, an OVX mouse model mimicking estrogen‐deficient osteoporosis was created to evaluate the therapeutic effects of Rob in vivo . Results: Computational docking results showed that Rob could bind specifically to RANKL's predicted binding sites. In vitro, Rob inhibited RANKL‐mediated osteoclastogenesis dose‐dependently without obvious cytotoxicity at low concentrations. We also found that Rob attenuated RANKL‐induced mitochondrial ROS production or enhanced activities of ROS‐scavenging enzymes, and ultimately reduced intracellular ROS levels. Rob abrogated the RANKL‐induced mitogen‐activated protein kinase (MAPK) and NF‐κB signaling pathways, and subsequently blocked NFATc1 signaling and TRAcP expression. In addition, Rob inhibited osteoclast proliferation by downregulating the expression of osteoclast target genes ( Acp5, Cathepsin K, Atp6v0d2, Nfact1, c‐Fos, and Mmp9 ) and reducing Ca 2+ oscillations. Our in vivo results showed that Rob reduced bone resorption in OVX animal model by repressing osteoclast activity and function. Conclusions: Rob inhibits the activation of osteoclasts by targeting RANKL and is therefore a potential osteoporosis drug. Abstract : In this manuscript, our research is performed to investigate whether Robinin, a novel flavonoid glycoside, could prevent bone resorption in ovariectomized (OVX) mice by suppressing osteoclast production via its underlying mechanisms. Our findings suggest that Rob abrogates the RANKL‐induced MAPK and NF‐κB pathway, attenuates RANKL‐induced reactive oxygen species (ROS) production and subsequently blocks NFATc1 signaling and TRAcP expression. Our in vivo results show that Rob reduces bone loss in OVX mice by repressing osteoclast activity and function.